Post on 09-Jun-2018
Optimisation of High‐frequency Jet Ventilation for the
Management of Respiratory Distress Syndrome in
Preterm Babies using a Preterm Lamb Model
Gabrielle Christine Musk
BSc BVMS Cert VA Dipl ECVAA
This thesis is presented for the degree of Doctor of Philosophy of
The University of Western Australia
School of Women’s and Infants’ Health
Faculty of Medicine and Dentistry
2011
Thesis Abstract
High‐frequency jet ventilation (HFJV) is a lung protective ventilation strategy used for
the prevention and treatment of ventilator induced lung injury in preterm infants.
Despite its widespread use in neonatal intensive care units there is little data to
support the patient management algorithms that are currently utilised. The strategies
for alveolar recruitment during HFJV rely upon incrementing positive end‐expiratory
pressure (PEEP) and delivering occasional conventional mechanical ventilator (CMV)
breaths, but the impact of these recruitment manoeuvres on pulmonary blood flow,
oxygenation, ventilation and lung injury is largely unknown. This thesis investigates the
parameters that must be selected during HFJV and their impact upon pulmonary blood
flow, physiological changes during ventilation and post mortem lung injury in a
preterm lamb model. The first study examined the effect of incrementing PEEP during
HFJV and found that alveolar recruitment was achieved by incrementing PEEP up to 12
cmH2O without detrimental effects on physiological parameters. The following 3
studies examined the delivery of CMV breaths during HFJV to compare the effect of 2
different CMV breath inspiratory times, peak inspiratory pressures and frequencies. A
shorter inspiratory time CMV breath, a CMV breath delivered to a peak inspiratory
pressure (PIP) above the HFJV breaths and CMV breaths delivered less frequently
provided the most physiological benefit with the least evidence of harm while
adequately ventilating and oxygenating the preterm lambs. The final study compared
what we determined to be optimal HFJV strategy (based upon the results of the
preceding studies) with optimal high‐frequency oscillatory ventilation and an open
lung gentle CMV strategy. In our preterm lamb model of respiratory distress syndrome
we demonstrated little difference in physiological benefit and adverse effects between
these 3 lung protective ventilation strategies.
The results of these studies contribute to the sparse data on HFJV and have provided
fundamental information that will enable a more evidence based approach to clinical decision
making in the neonatal intensive care unit. Future work in this area should focus on the target
population and incorporate randomised controlled trials comparing HFJV to other ventilatory
strategies.
Statement of Candidate Contribution
Chapters 4, 5, 6, 7 and 8 are presented in a manuscript format as they are at various
stages of the publication process. My contribution, and that of my supervisors and co‐
authors, is detailed below:
Chapter 4: High Positive End‐Expiratory Pressure during High Frequency Jet
Ventilation Improves Oxygenation and Ventilation in Preterm Lambs.
Gabrielle C Musk, Graeme R Polglase, J Bert Bunnell, Carryn J McLean, Ilias Nitsos, Yong
Song and J Jane Pillow.
I was involved in the anaesthesia of the pregnant ewe, delivery of the preterm lamb
fetus, and subsequent ventilation. I was also responsible for data collection during the
ventilation period and sample analysis. Graeme Polglase and Ilias Nitsos performed
surgical instrumentation of the fetus prior to delivery, Bert Bunnell assisted in the
ventilator management of the lamb following delivery, Carryn McLean assisted in post
mortem of the lambs and Yong Song performed the q PCR for pro‐inflammatory
cytokines. Graeme Polglase also assisted with pulmonary blood flow waveform
analyses. I was responsible for manuscript preparation. Jane Pillow was involved with
all aspects of the study in a supervisory capacity.
Chapter 5: The Impact of Conventional Breath Inspiratory Time during High‐
frequency Jet Ventilation in Preterm Lambs.
Gabrielle C Musk, Graeme R Polglase, Yong Song, and J Jane Pillow.
I was involved in the anaesthesia of the pregnant ewe, delivery of the preterm lamb
fetus, and subsequent ventilation. I was also responsible for data collection during the
ventilation period and sample analysis. Graeme Polglase assisted with delivery of the
preterm lamb fetus and subsequent ventilation. Yong Song performed the q PCR for
pro‐inflammatory cytokines. I was responsible for manuscript preparation. Jane Pillow
was involved with all aspects of the study in a supervisory capacity.
Chapter 6: The Effect of Conventional Breath Peak Inspiratory Pressure during High‐
frequency Jet Ventilation in Preterm Lambs.
Gabrielle C Musk, Graeme R Polglase and J Jane Pillow.
I was involved in the anaesthesia of the pregnant ewe, delivery of the preterm lamb
fetus, and subsequent ventilation. I was also responsible for data collection during the
ventilation period and sample analysis. Graeme Polglase assisted with delivery of the
preterm lamb fetus and subsequent ventilation. Yong Song performed the q PCR for
pro‐inflammatory cytokines. I was responsible for manuscript preparation. Jane Pillow
was involved with all aspects of the study in a supervisory capacity.
Chapter 7: Alveolar Recruitment with Five or Twenty Conventional Mechanical
Ventilator Breaths per minute during High‐frequency Jet Ventilation in Preterm
Lambs.
Gabrielle C Musk, Graeme R Polglase and J Jane Pillow.
I was involved in the anaesthesia of the pregnant ewe, delivery of the preterm lamb
fetus, and subsequent ventilation. I was also responsible for data collection during the
ventilation period and sample analysis. Graeme Polglase assisted with delivery of the
preterm lamb fetus and subsequent ventilation. Yong Song performed the q PCR for
pro‐inflammatory cytokines. I was responsible for manuscript preparation. Jane Pillow
was involved with all aspects of the study in a supervisory capacity.
Chapter 8: A Comparison of High‐frequency Jet Ventilation and High‐frequency
Oscillatory Ventilation with Conventional Mechanical Ventilation in Preterm Lambs.
Gabrielle C Musk, Graeme R Polglase, J Bert Bunnell, Ilias Nitsos, David Tingay, J Jane
Pillow.
I was involved in the anaesthesia of the pregnant ewe, delivery of the preterm lamb
fetus, and subsequent ventilation. I was also responsible for data collection during the
ventilation period and sample analysis. Graeme Polglase and Ilias Nitsos performed
surgical instrumentation of the fetus prior to delivery, Bert Bunnell and David Tingay
assisted in the ventilator management of the lamb following delivery and data
collection, Yong Song performed the q PCR for pro‐inflammatory cytokines. Graeme
Polglase assisted with pulmonary blood flow waveform analysis. I was responsible for
manuscript preparation. Jane Pillow was involved with all aspects of the study in a
supervisory capacity.
Jane Pillow's supervisory role extended to all aspects of the project including study
design, conduct and critical review of data analysis and written interpretation.
Graeme Polglase and Karen Simmer contributed to critical review of each manuscript.
It is not possible to have each co‐author sign this declaration, in which case, my co‐
ordinating supervisor J Jane Pillow has signed to confirm that these statements are an
accurate reflection of the contributions of each author.
Gabrielle Christine Musk
J Jane Pillow
Publications arising from this Thesis
Musk GC, Polglase GR, Bunnell JB, McLean CJ, Nitsos I, Song Y and Pillow JJ 2011 High
Positive End‐Expiratory Pressure during High‐Frequency Jet Ventilation Improves
Oxygenation and Ventilation in Preterm Lambs. Pediatric Research 69(4):319‐324
(Chapter 4).
Musk GC, Polglase GR, Song Y and Pillow JJ The Impact of Conventional Breath
Inspiratory Time during High‐frequency Jet Ventilation in Preterm Lambs. Submitted to
Neonatology April 2011 (Chapter 5).
Musk GC, Polglase GR, and Pillow JJ The Effect of Conventional Breath Peak Inspiratory
Pressure and Frequency during High‐frequency Jet Ventilation in Preterm Lambs.
Manuscript in preparation (Chapters 6 and 7).
Musk GC, Polglase GR, Bunnel JB, Nitsos I, Tingay D and Pillow JJ A Comparison of High‐
frequency Jet Ventilation and High‐frequency Oscillatory ventilation with Conventional
Mechanical Ventilation in Preterm Lambs. Manuscript in preparation (Chapter 8).
Acknowledgements
This work would not have been possible without the contribution and support of many
people. I have been fortunate to receive scientific guidance, intellectual inspiration
and moral support from my supervisors, colleagues, family and friends. Jane Pillow
had the idea for these studies and a vision for their clinical application in neonatal
intensive care units. She has been an incredible supervisor who leads by example
when it comes to being thorough, concise, articulate, scientifically watertight and
generous with her time. She’s taught me a lot and I will never forget that I am a
passive splitter who sometimes likes to use as many words as possible to simply say
something. Graeme Polglase deserves mention for his unrelenting mission to train me
out of putting 2 spaces after a full stop. If I had a dollar for every time he commented
on that I’d be mortgage free. Karen Simmer made me realise it wasn’t unreasonable to
arrive in another timezone more than 6 hours before a conference is due to start.
Thank you to you all for supervising me – in your own unique ways.
Bert Bunnell developed the ventilator at the centre of this work and is an inspiration to
me. He leads a formidable team of dedicated, enthusiastic and kind people who
mirror his ethos in life. It has been a pleasure and a privilege to be a small part of the
evolution of High‐frequency Jet Ventilation.
Being a vet, working with sheep and based in a maternity hospital took some getting
used to but I very quickly felt settled in at the School of Women’s and Infants’ Health
on the second floor of A block at King Edward Memorial Hospital. Maz Schneider,
Bevelynn Ibrahim and Catherine Arresse in particular have made me feel right at home
and I miss that feeling of family immensely. John Newnham can certainly tell a good
story and I have listened in awe to many of his. Thank you John.
The daily grind of my PhD was filled with humour and friendship thanks to Carryn
McLean, David Cruise, Richard GB Dalton and Joe Derwort. Carryn has great taste in
music (especially Mossimo Park) and necklaces, and was my most constant and reliable
moral support through the frustrating times. Cruisey looks great in undersized overalls
and bakes a mean brownie. I was just sorry that he was only really around for a year.
Richard came from Tasmania to Forrest St and after a road trip to Darkan on day 1 we
became firm friends. Joe couldn’t quite get me out of my cappuccino habit, despite his
efforts in promoting the long macchiato.
Office life was always a giggle with Chris (AKA Toots) and his Style Guide, and Irving.
Irving made sure I didn’t burn down the lab on weekends and Toots made sure that
PRONG’s purse strings were tight.
Ilias has taught me about removing endogenous peroxidase activity, the importance of
blind intubations and has been a stalwart colleague and friend. He’s gone back to
Melbourne now but there’ll never be another Nossie in my life.
My family have all helped make student life less impoverished than it could have been.
My house has become a lovely home thanks to Mum and the pruning jobs are shorter
each year thanks to Dad. My nieces fill it with fun and Harry guards it as his castle.
Harry warrants his own mention – a finer canine companion does not exist.
Lastly, it’s a big thank you to my wonderful friends (that’s you Fraser and Karen) who
have been so supportive. Thank you!
List of Abbreviations
ANOVA analysis of variance
BPD bronchopulmonary dysplasia
CMV conventional mechanical
ventilation
cmH2O centimeters of water
CO2 carbon dioxide
ΔP delta P (PIP-PEEP)
ETT endotracheal tube
FiO2 fractional inspired oxygen
concentration
HFV high-frequency ventilation
HFJV high-frequency jet
ventilation/ventilator
HFOV high-frequency oscillatory
ventilation/ventilator
h hour
Hz Hertz
IgG immunoglobulin G
IL interleukin
iNOS inducible nitric oxide
synthetase
i.v. intravenous
kg kilogram
min minute
mL milliliters
mmHg millimeters of mercury
MPO myeloperoxidase
nm nanometer
O2 oxygen
OI Oxygenation Index
Paw mean airway pressure
PaCO2 partial pressure of carbon
dioxide in arterial blood
PaO2 partial pressure of oxygen in
arterial blood
PAP pulmonary arterial blood
pressure
PBF pulmonary blood flow
PEEP positive end-expiratory
pressure
PIP peak inspiratory pressure
PmvO2 partial pressure of oxygen in
mixed venous blood
psi pounds per square inch (1psi =
70 cmH2O)
PVR pulmonary vascular resistance
RDS respiratory distress syndrome
s seconds
SABP systemic arterial blood
pressure
SD standard deviation
SEM standard error of the mean
SpO2 oxyhemoglobin saturation
measured by a pulse oximeter
tI inspiratory time
tE expiratory time
UVC unventilated control
VT tidal volume
µg microgram
1
TABLE OF CONTENTS
1 LITERATURE REVIEW .......................................................................... 5
1.1 Introduction ........................................................................................................ 5
1.2 Stages of Lung Development .............................................................................. 6
1.2.1 Strategies for Inducing Preterm Lung Maturation .................................... 10
1.3 Respiratory Consequences of Premature Birth ................................................ 12
1.3.1 Respiratory Distress Syndrome ................................................................. 12
1.3.2 Bronchopulmonary Dysplasia ................................................................... 13
1.3.3 Respiratory Outcomes of Bronchopulmonary Dysplasia .......................... 15
1.4 Respiratory Management of Preterm Babies .................................................. 16
1.4.1 Non-invasive Therapies ............................................................................. 17
1.4.2 Indications for Mechanical Ventilation ..................................................... 17
1.4.3 Adjunctive Treatments .............................................................................. 26
1.4.4 Surfactant .................................................................................................. 28
1.4.5 Postnatal Corticosteroids .......................................................................... 30
1.5 Side Effects of Positive Pressure Ventilation .................................................... 32
1.5.1 Ventilator Induced Lung Injury ................................................................. 32
1.5.2 Haemodynamic Consequences of Positive Pressure Ventilation ............. 36
1.5.3 Central Nervous System Consequences of Positive Pressure Ventilation 41
1.6 Assessment of Lung Injury ................................................................................ 44
2
1.6.1 Bronchoalveolar Lavage Fluid ................................................................... 46
1.6.2 Lung Tissue ................................................................................................ 48
1.7 Lung Protective Ventilation Strategies ............................................................. 50
2 HIGH-FREQUENCY VENTILATION ....................................................... 54
2.1 High-frequency Ventilation .............................................................................. 54
2.1.1 Gas Mixing during High-frequency Ventilation ......................................... 54
2.1.2 Mechanical Properties of the Lung and High-frequency ventilation ........ 56
2.1.3 Airway Pressure Waveforms during High-frequency Ventilation ............. 58
2.1.4 Modes of High-frequency Ventilation ...................................................... 60
2.2 Using a High-frequency Jet Ventilator ............................................................. 61
2.2.1 The Role of the Conventional Ventilator .................................................. 64
2.2.2 The Role of the High-frequency Jet Ventilator ......................................... 71
2.2.3 Monitoring during High-frequency Jet Ventilation ................................... 73
2.3 High-Frequency Jet Ventilation in the Clinical Environment ........................... 74
2.3.1 High-frequency Jet Ventilation as a Rescue Therapy ............................... 75
2.3.2 HFJV used Early in the Management of Respiratory Distress Syndrome . 76
2.3.3 Clinical Strategies ...................................................................................... 78
2.4 Summary........................................................................................................... 80
3 GENERAL METHODOLOGY ................................................................ 83
3.1 Animal Breeding and Welfare .......................................................................... 83
3
3.1.1 Nutrition .................................................................................................... 84
3.1.2 General Anaesthesia and Instrumentation ............................................... 84
3.1.3 Caesarean Delivery .................................................................................... 87
3.2 Ventilator Set-up .............................................................................................. 88
3.3 Data Collection ................................................................................................. 89
3.3.1 Pulmonary Arterial Blood Pressure and Blood Flow Measurements ....... 90
3.4 Euthanasia and Post Mortem ........................................................................... 92
3.4.1 Cell Population of Bronchoalveolar Lavage Fluid ..................................... 93
3.4.2 Bronchoalveolar Lavage Protein Assay ..................................................... 94
3.4.3 Immunohistochemistry ............................................................................. 94
3.4.4 Qualitative Polymerase Chain Reaction .................................................... 95
3.4.5 Myeloperoxidase Activity in Lung Tissue .................................................. 96
3.5 Statistical Analyses ........................................................................................... 96
3.6 References ........................................................................................................ 98
4 HIGH POSITIVE END-EXPIRATORY PRESSURE DURING HIGH-
FREQUENCY JET VENTILATION IMPROVES OXYGENATION AND
VENTILATION IN PRETERM LAMBS ................................................. 121
4
5 THE IMPACT OF CONVENTIONAL BREATH INSPIRATORY TIME
DURING HIGH-FREQUENCY JET VENTILATION IN PRETERM LAMBS 153
6 THE IMPACT OF CONVENTIONAL BREATH PEAK INSPIRATORY
PRESSURE DURING HIGH-FREQUENCY JET VENTILATION IN PRETERM
LAMBS ........................................................................................... 181
7 ALVEOLAR RECRUITMENT WITH FIVE OR TWENTY CONVENTIONAL
MECHANICAL VENTILATOR BREATHS PER MINUTE DURING HIGH-
FREQUENCY JET VENTILATION IN PRETERM LAMBS ....................... 207
8 A COMPARISON OF HIGH-FREQUENCY JET VENTILATION WITH HIGH-
FREQUENCY OSCILLATORY VENTILATION AND CONVENTIONAL
MECHANICAL VENTILATION IN PRETERM LAMBS ........................... 233
9 DISCUSSION .................................................................................... 271
10 APPENDIX ....................................................................................... 279
5
1 Literature Review
1.1 Introduction
High-frequency jet ventilation (HFJV) is a novel mode of high-frequency ventilation
offering the potential for lung protective ventilation (1, 2). In the United States of
America, HFJV is a common ventilation modality employed in the clinical setting,
especially for the management of premature babies with respiratory distress
syndrome (RDS). In Australia, only one maternity hospital has access to HJFV.
Babies can be oxygenated and ventilated with HFJV but the incidence of
haemodynamic, respiratory and neurological morbidity is largely unknown. Published
data presents conflicting information with regards to the incidence of adverse
outcomes in a clinical environment and to date there are no controlled studies
exploring the optimal settings for HFJV. Despite 25 years of clinical application, HFJV is
more often employed as a ‘rescue’ ventilation strategy rather than a first line
treatment option. It is possible that if HFJV is used earlier in the clinical course, the
outcome statistics may improve. To use this therapeutic tool to its full potential, a
thorough understanding of the pathophysiology of respiratory diseases, the clinical
indications for HFJV, the unique features of this mode of ventilation, and the
equipment required to safely and effectively apply HFJV is required. This review will
examine information in the scientific literature with relevance to each of these points.
6
1.2 Stages of Lung Development
Human lung development can be subdivided into three chronological periods: early
embryonic, fetal and postnatal (3). The fetal period is further subdivided into three
phases which describe the morphology of the airways and airspaces (pseudoglandular,
canalicular and saccular) as described in Table 1-A. The final phase of lung
development is alveolarisation which commences before birth, and continues
thereafter.
The formation of sufficient gas exchange surface area and pulmonary vasculature
capable of transporting CO2 and O2 through the lungs is vital for survival. Gas exchange
will only occur if inspired alveolar air and pulmonary arterial blood flow are contiguous
to each other and is more efficient if the surface area is greater and the membrane
between air and blood is thinner. Complete development of these components of the
respiratory unit is fundamental to normal postnatal respiratory function. Given the
developmental steps involved in maturation of the lungs, it is no surprise that there is
an inverse relationship between gestational age and the incidence of respiratory
morbidity and mortality. Infants born prior to the alveolar phase of development will
have fewer functional gas exchange units, reduced surfactant, a less compliant chest
wall and an immature pulmonary capillary network. These infants are therefore more
likely to struggle with the transition to air breathing.
7
Table 1-A Stages of prenatal and postnatal structural lung development
Phase Postconceptional Age
Length: terminal bronchiole to pleura
Structure
Embryonic 0-7 weeks <0.1 mm Budding from the foregut
Pseudoglandular 8-16 weeks 0.1 mm Airway division commences and terminal bronchioles formed
Canalicular 17-27 weeks 0.2 mm 3 generations of respiratory bronchioles; primitive saccular formation with type I and type II epithelial cells; capillarisation
Saccular 28-35 weeks 0.6 mm Transitional saccules formed; true alveoli appear
Alveolar >36 weeks 11 mm Terminal saccules formed; true alveoli appear
Postnatal 2 months 175 mm 5 generations of alveolar ducts; alveoli form with septation
Early childhood 6-7 years 400 mm Airways remodelled; alveolar sac budding occurs
Adapted from Bhutani (4).
Aside from maturation of the lung parenchyma, upper airway development is also
essential for normal gas exchange at birth. The patency of the upper airways is under
complex control and allows for conduction, humidification, warming and filtering of air
during inspiration and expiration (4). Airway development is essentially complete by
8
the end of the canalicular period of lung development, so viable preterm babies should
have sufficiently developed conducting airways.
Furthermore, clearance of fetal lung fluid must occur before normal gas exchange can
occur. Fetal lung fluid is produced during the canalicular stage of lung development
and creates a hydrostatic pressure within the respiratory system. The 3 to 5 cmH2O
exerted by the fetal lung fluid contributes to structural development of the lungs and
ensures that lung volume remains approximately equivalent to the functional residual
capacity of the fetal lung (4). Clearance of fetal lung fluid normally occurs during the
transition to air breathing at birth but if a baby is born prematurely, lung fluid may not
clear without intervention.
Recent Australian data reports the mean gestational age for all babies as 38.8 weeks
and the proportion of babies born at term (37-41 w) as 90.9 % (5). The mean
gestational age for all preterm births was 33.2 w (0.9 % of births were at 20-27 w
gestation, 0.8 % were at 28-31 w and 6.5 % were at 32-36 w) (5). In Western Australia
between 2002 and 2004, 8.4 % of births were preterm (< 37 w) (6). The highest
mortality rate occurred in the most immature babies and decreased as gestational age
increased (Table 1-B).
9
Table 1-B Birth and death statistics by gestational age, Western Australia 2002-2004
Gestational
Age (weeks)
Total births
Neonatal
deaths
Post-natal
deaths
Total
deaths
Death rate
(%)
20-27 613 80 8 88 14.4
28-32 933 21 5 26 2.8
33-36 4744 17 15 32 0.7
37-43 68705 48 66 114 0.2
<37 6290 118 28 146 2.3
Adapted from Newnham et al. (6)
Data from a Californian hospital in the 1990s described a similar pattern where mean
hospital stay was longer for infants born at an earlier gestational age (7). Survival
without BPD or retinopathy of prematurity is also strongly associated with maturation:
whereas only 35 % of infants born at 24 w survive without either of these morbidities,
this figure increases to almost 78 % at 26 w (8).
While the statistics describing the incidence of preterm birth reveal that a relatively
small proportion of babies are born at a stage prior to alveolarisation, it is these babies
that require the highest level of care. In 2005, 2.44 % of live births in Australia and
New Zealand were admitted to a neonatal intensive care unit and 91.1 % required
assisted ventilation (9). The major indication for assisted ventilation in babies born
less than 32 w was RDS (72.4 %) and the duration of ventilation increased with
10
decreasing gestational age. From this particular cohort of babies, the median length of
stay in hospital (the survivors) was 133 d if born at 23 w, 118 d at 24 w, and 106 d at 25
w. The length of stay steadily decreased as gestational age increased (9).
1.2.1 Strategies for Inducing Preterm Lung Maturation
Accelerated maturation of the lungs and the impact on the incidence and severity of
BPD have been widely investigated. Exposure to glucocorticoids, chorioamnionitis and
fetal stress in utero all contribute to decreasing the incidence of RDS and therefore
BPD (10, 11).
1.2.1.1 Antenatal Corticosteroids
Clinical and experimental observations of antenatal corticosteroid treatment
demonstrate that endogenous and exogenous glucocorticoids function primarily to
accelerate lung maturation by thinning alveolar walls and increasing lung gas volume
(8, 11). This functional maturation of the preterm fetal lung has led to the
administration of corticosteroids to women at risk of preterm delivery becoming
established therapy (10, 12, 13).
The optimal dose and timing of corticosteroid treatment should aim to stimulate
generalised lung maturation and surfactant synthesis. Human and animal studies have
been performed to investigate the effects of antenatal steroid administration and help
determine when to administer corticosteroids and how much to administer (12, 14-
16). Improvement in lung function and increases in surfactant production both
contribute to improved outcomes in preterm babies whose mothers have received
antenatal corticosteroids (12).
11
1.2.1.2 Chorioamnionitis
Intrauterine inflammation is associated with a reduced risk of RDS in some preterm
babies (17, 18). This realisation has prompted a vast number of animal studies
examining the impact of induced chorioamnionitis on lung maturation and surfactant
production (19-21) and evidence suggests that fetal lung inflammation causes marked
improvements in preterm lung function. These improvements are primarily caused by
increases in pulmonary surfactant but remodelling of the lung parenchyma also occurs
(20). Whether or not chorioamnionitis and corticosteroid therapy work together to
reduce the risk of RDS to an extent greater than either alone is unknown.
Most cases of chorioamnionitis in preterm and term labour are sub-clinical (22) which
makes diagnosis difficult. The most common organism identified is Ureaplasma
urealyticum (22, 23) and a number of animal studies have documented lung
maturation associated with this infection (21). Colonisation of the chorioamnion incites
a fetal inflammatory response which targets a range of organs and tissues in the fetus,
including the lungs (22). Injury subsequent to infection develops and structural
changes to the lungs occur. These changes may in fact be beneficial, at least in the
short term.
1.2.1.3 Fetal stress
The short and long term effects of fetal stress may be both beneficial and adverse. The
beneficial effects result from accelerated organ and tissue maturation which improve
the capacity of a patient to cope with preterm birth (24). This ability to cope may be
interpreted as fetal adaptation to unfavourable circumstances and is likely to be
12
associated with increased circulating cortisol and inflammatory mediators (24). The
manifestations of antenatal glucocorticoid therapy, chorioamnionitis and fetal stress
are comparable, albeit complex, and the balance between the beneficial and adverse
effects will vary according to gestational age and the presence of other pathology (24).
Nevertheless, fetal stress causes an increase in circulating cortisol and the release of
pro-inflammatory mediators which impact upon a number of organs, including the
lungs, to hasten maturation.
1.3 Respiratory Consequences of Premature Birth
1.3.1 Respiratory Distress Syndrome
Respiratory distress syndrome occurs as a result of both structural immaturity and
surfactant deficiency of the premature lung. The symptoms of RDS include
tachypnoea, chest wall retraction, cyanosis and characteristic radiological
abnormalities of the lungs. Clinical management invariably includes oxygen therapy
with or without ventilator support: 96 % of infants born at less than 28 w gestation in
Australasia in 2002, received supplemental oxygen (25). These extremely preterm
infants have a higher incidence of RDS and are hospitalised for longer compared to
those born after 28 w gestation (26). A range of ventilation techniques have been used
to support these preterm infants but mechanical ventilation has itself been implicated
as a cause of lung injury. Further, the combination of immature lung parenchyma and
lung injury may predispose the baby to bronchopulmonary dysplasia (BPD) (27).
13
1.3.2 Bronchopulmonary Dysplasia
Bronchopulmonary dysplasia (BPD) was first defined in 1967 by Northway et al (28) to
describe the clinical, radiological and pathological changes in infants with severe RDS.
The definition applied to babies who had been treated with prolonged mechanical
ventilation and warmed, humidified 80-100 % inspired oxygen concentrations. The
term BPD was coined to emphasise the involvement of all tissues of the developing
lung in the pathological process and represented the toxic manifestations of high
oxygen concentrations and mechanical ventilation on the developing lung
superimposed upon the healing phase of RDS. In 1968 a radiographic description was
published highlighting the distinctive features of this phenomenon (29). The
radiographic features were staged with grade 4 representing the worst case scenario:
symptomatic chronic lung disease with strands of pulmonary parenchymal density;
increased thoracic volume; and cardiomegaly which may gradually clear or progress to
cor pulmonale and death at less than 1 month of age (29). These two publications
marked the beginning of an era.
Despite advances in antenatal glucocorticoid therapy, surfactant treatment and gentle
‘lung protective’ ventilation techniques, the incidence of BPD is increasing (30),
prompting further investigation into its multifactorial pathophysiology. In 2001 a new
NIH consensus definition of BPD was proposed by Jobe and Bancalari (27) to include
categorisation of the severity of BPD (Table 1-C).
14
Table 1-C Diagnostic criteria for defining bronchopulmonary dysplasia
Gestational
Age
<32 weeks ≥ 32 weeks
Time point of
assessment
36 w PMA or discharge to home,
whichever comes first.
>28 d but < 56 d postnatal age or
discharge to home, whichever
comes first.
Treatment with >21 % oxygen for at least 28 d plus.
Mild BPD Breathing room air at 36 w PMA
or discharge, whichever comes
first.
Breathing room air by 56 d
postnatal age or discharge,
whichever comes first.
Moderate BPD Need* for < 30 % oxygen at 36 w
PMA or discharge, whichever
comes first.
Need* for < 30 % oxygen at 56 d
postnatal age or discharge,
whichever comes first.
Severe BPD Need* for ≥ 30 % oxygen and/or
positive pressure, (PPV or
NCPAP) at 36 w PMA or
discharge, whichever comes first.
Need* for ≥ 30 % oxygen and/or
positive pressure (PPV or
NCPAP) at 56 d postnatal age or
discharge, whichever comes
first.
Adapted from Jobe and Bancalari (27). BPD = bronchopulmonary dysplasia, NCPAP = nasal continuous
positive airway pressure, PMA = postmenstrual age; PPV = positive pressure ventilation. * A
physiological test confirming that the oxygen requirement at the assessment time point remains to be
defined. This assessment may include a pulse oximetry saturation range.
15
As mechanical ventilation may contribute to the development of an inflammatory
response resulting in altered alveolar and pulmonary vasculature development (27),
the identification of ventilatory strategies that minimise this damage and prevent the
progression of BPD is an important research goal.
1.3.3 Respiratory Outcomes of Bronchopulmonary Dysplasia
In the last 30 years, the chances of surviving the neonatal period as a premature baby
have improved (31). The introduction and evolution of assisted ventilation, surfactant
therapy and antenatal steroid administration are primarily responsible for this
progress (32). Improved survival of extremely low gestational age infants is frequently
followed by significant long term respiratory morbidity and has fiscal implications. The
cost of initial care for this population of neonates in the United States is estimated at
US$10.2 billion each year. Infants born between 24 and 26 weeks gestation account for
11.9 % of this bill (33).
There are limited reports of the long term respiratory complications of preterm birth in
the contemporary era. The survivors of extremely preterm birth (prior to 26 weeks
gestation) are only just reaching adulthood so data are limited to patients less than 20
years of age (34-36). Furthermore, the cohort of patients is small. Despite these
limitations, there is evidence that lung function is reduced as a result of surviving BPD
(36).
Earlier studies present data on patients of a similar age group (gestational age and age
in adulthood at the time of data collection) but from a different era. These patients
16
were born before the routine administration of antenatal corticosteroid therapy and
exogenous surfactant which makes comparisons inappropriate. The definitions of BPD,
the birth weight of the patients and the respiratory parameters that were assessed
vary. The data does however reflect, once again, that in adulthood, the survivors of
preterm birth have some degree of decreased lung function (37, 38).
Overall it appears that a significant number of adult survivors of moderate and severe
BPD may be left with residual functional and characteristic structural pulmonary
abnormalities. The limitation of these studies is a failure to demonstrate a correlation
between perinatal variables and the manifestations in adulthood. This lack of
correlation makes prognostication difficult and emphasises the importance of
exploring management strategies for these preterm babies that will minimise injury to
the preterm lung.
1.4 Respiratory Management of Preterm Babies
The management of preterm babies with RDS aims to reduce the incidence and
severity of BPD. This management goal is challenging given the complexity of the
disease process and the precarious balance between risk and benefit associated with
any therapy. The general treatment goals are to minimise ventilator induced lung
injury, minimise the inspired oxygen concentration, avoid infection and manage
nutritional and fluid requirements (39). Respiratory management of preterm babies
may be non-invasive (e.g. nasal continuous airway pressure (CPAP)) or invasive
(requiring endotracheal intubation). Invasive techniques range from supported
ventilation by CPAP to intermittent positive pressure ventilation (IPPV).
17
1.4.1 Non-invasive Therapies
Endotracheal intubation and the prophylactic administration of surfactant is effective
for the management of RDS in preterm infants (40). A comparison of early versus later
administration of surfactant has demonstrated reduced mortality, frequency of BPD,
and the risk of pneumothorax in ventilated preterm infants with RDS (41).
Endotracheal intubation however, is itself associated with considerable risk and if it
can be avoided, so will the associated complications. Non-invasive respiratory support
may therefore be considered for some infants. This form of respiratory support is a
continuous distending pressure, or CPAP. Continuous positive airway pressure is
applied using a conventional ventilator, bubble circuit or CPAP driver via a face mask,
nasopharyngeal tube, or nasal prongs (40). A Cochrane review published in 2002 found
that these methods of respiratory support reduced the rate of death or the need for
assisted ventilation and reduced the need for IPPV in preterm babies that were able to
breathe spontaneously (40). Subsequent randomised trials have failed to confirm that
CPAP is superior to intubation and mechanical ventilation when outcomes such as BPD
and death are considered (42, 43).
1.4.2 Indications for Mechanical Ventilation
Mechanical ventilation is indicated for any patient unable to breathe spontaneously
and achieve adequate gas exchange. Preterm babies have underdeveloped lungs and
respiratory muscles, insufficient endogenous surfactant production and secretion and
a highly compliant chest wall. Not surprisingly, they may require sophisticated
respiratory support in the form of mechanical ventilation via an endotracheal tube.
18
1.4.2.1 Goals of Mechanical Ventilation
The aims of any ventilation strategy include alveolar recruitment and stabilisation, the
acquisition and maintenance of appropriate arterial blood gas parameters and
minimising the impact of increased intra-thoracic pressure on cardiac output and lung
injury. Achieving just one of these aims may come at the expense of another so it is
essential to strike a balance between oxygenation, CO2 removal and the implications of
higher than normal airway pressures.
1.4.2.1.1 Oxygenation
Hypoxaemia is defined by either a low PaO2, low peripheral or arterial oxyhaemoglobin
saturation (SpO2 or SaO2) or low oxygen content of arterial blood (CaO2). Regardless of
the descriptor, hypoxaemia may result from hypoventilation, inspiration of a hypoxic
gas mixture and venous admixture. Venous admixture refers to the passage of blood
through the pulmonary circulation without oxygenation and occurs in the presence of
ventilation and perfusion mismatch, right to left shunting of blood and diffusion
impairment. Prevention and treatment of hypoxaemia should therefore be targeted to
decrease the impact of each of these factors.
Airway pressure
Airway pressure plays an important role in oxygenation insofar as preventing alveolar
collapse and maintaining an open lung. Lung volume recruitment to reduce shunting of
blood through the lungs is an established concept: in 1959 Mead and Collier showed
that periodic lung inflations were necessary to prevent a progressive fall in compliance
during mechanical ventilation (44). Lung volume recruitment is possible if the lung is
19
inflated beyond the pressure at which atelectatic lung begins to open, and is
maintained at a pressure above its closing pressure (45). In immature or injured lungs
where compliance is poor, these pressures may be high and the detrimental effects of
increased airway pressure must be considered. The balance between sufficient airway
pressure to prevent alveolar collapse and conservative enough airway pressure to
prevent alveolar overdistention may be difficult to achieve. Furthermore, the
haemodynamic implications must also be closely monitored and managed.
Inspired gas mixture
In preterm infants, oxygen supplementation is a balancing act between the treatment
of tissue hypoxia, pulmonary vasoconstriction, and patency of the ductus arteriosus
with lung injury, retinal damage and oxidative stress. Oxygen has therapeutic and toxic
characteristics and optimal oxygenation, especially in the first few weeks of life is often
under-emphasised. Hypoxanthine accumulates during hypoxia, and during re-
oxygenation, superoxide radicals are produced, leading to cell injury (46). Optimal
oxygen concentrations for resuscitation were reviewed recently and it is
recommended that babies of gestational age greater than 32 weeks should be
ventilated initially with 21 % oxygen. Babies of gestational age less than 32 weeks
should be ventilated initially with 21-30 % oxygen (47). Changes in SpO2 and heart rate
are often used to monitor oxygenation, especially in the acute period following
delivery and alterations to the fractional inspired oxygen concentration (FiO2) should
be made to target a SpO2 range. The consequences of various SpO2 targets have been
investigated and while there may be fewer complications in neonates that have been
20
managed to achieve a lower target range (48), the mortality rate must also be
considered. The current recommendation is a target SpO2 of 91-95% (49).
Automated oxygen delivery is an evolving technology that automatically adjusts the
FiO2 according to feedback information from a pulse oximeter. Preliminary studies
under routine clinical conditions showed this system increased the time in which SpO2
was maintained within a target range, decreased FiO2 and decreased the incidence of
hyperoxaemia without increasing severe hypoxaemia (50). This method of oxygen
delivery may be a useful tool to reduce exposure to hyperoxaemia and hypoxaemia
which may contribute to the development of BPD. Furthermore, this study is pertinent
as the optimal SpO2 range for the extremely preterm or very low birthweight infant
remains elusive (49, 51). There is, however, convincing evidence that there is an
association between oxidative stress and BPD (52).
While hypoxaemia usually stimulates more concern, judicious use of oxygen and
continuous, or at least regular, monitoring of oxygenation to minimise exposure to
high concentrations of oxygen is important. The optimal SpO2 is not known in
premature infants: 2008 data indicated lower mortality if SpO2 was kept below 93 %
and stable (53). More recent data demonstrates increased mortality in a lower SpO2
range (85-89 %) when compared to a higher SpO2 range (91-95%) (49). The incidence
of retinopathy of prematurity was lower in survivors from the lower SpO2 range group
but the higher mortality in this group raises question over the appropriate target range
for SpO2. Further investigations are currently being undertaken to determine the safest
SpO2 target for preterm infants. Results from the Neonatal Oxygenation Prospective
Meta-analysis Collaboration are expected in 2014 (54).
21
1.4.2.1.2 Removal of Carbon Dioxide
Normal cellular metabolism relies upon oxygen supply exceeding oxygen demand and
normal pH. The partial pressure of carbon dioxide in arterial blood (PaCO2) is directly
related to pH and both are maintained within a narrow range. Normal PaCO2 is 35 – 45
mmHg (4.6-5.5 kPa) and normal pH is 7.35 – 7.45.
Carbon dioxide production is a product of the carbonic anhydrase equation:
Equation 1-A
H+ + HCO3- ↔ H2CO3 ↔ H2O + CO2
Carbon dioxide is a byproduct of cellular metabolism and is eliminated by the kidneys
or the lungs. PaCO2 is a balance between carbon dioxide production and elimination.
During conventional mechanical ventilation (CMV), CO2 removal is directly related to
minute volume:
Equation 1-B
V’CO2 α V’min = fx VT
where V’CO2 = rate of carbon dioxide elimination; V’min = minute volume; f = ventilator
frequency or respiratory rate; and VT = tidal volume (alveolar).
A decrease in V’min due to either a decrease in f, VT, or both will decrease the
elimination of carbon dioxide (potentially causing hypercapnia). Conversely, an
increase in V’min accelerates CO2 elimination (potentially causing hypocapnia).
Equipment and physiological dead space will impact upon VT and therefore V’CO2.
22
During high-frequency ventilation, however, CO2 removal is more closely linked to the
VT:
Equation 1-C
V’CO2 α V’min = f x VT2 (55)
The side effects of abnormal CO2 levels are dose and time dependent. However, in
general, hypercapnia leads to cerebral vasodilation, an increase in intracranial
pressure, splanchnic vasodilation and hypotension. Hypocapnia causes cerebral and
splanchnic vasoconstriction and may compromise perfusion of important organs.
During growth and development, the effects of alterations in CO2 are potentially more
profound. Hypocapnia is associated with an increased risk of periventricular
leukomalacia in low birth weight infants (56, 57) while hypercapnia is a predictor of
severe intraventricular haemorrhage in very low birth weight infants (58).
Furthermore, fluctuations in PaCO2 are associated with worse neurodevelopmental
outcomes in extremely low birthweight infants (59).
Permissive hypercapnia refers to an acceptance of PaCO2 levels higher than the normal
range (usually ~ 45-55 mmHg) and has been promoted as a strategy to reduce cyclic
volutrauma and barotrauma associated with mechanical ventilation. Other beneficial
effects include maintenance of cardiovascular performance secondary to hypercapnic
stimulation of the sympathetic nervous system. Despite these potential benefits of
permissive hypercapnia, the impact upon neurological development must be
considered.
23
1.4.2.1.3 Minimising Iatrogenic Injury
Managing preterm babies with RDS is challenging and the outcome data demonstrates
that adverse effects may have implications into adulthood. Minimising iatrogenic injury
is essential to ensure the risk/benefit balance is in favour of the latter. The main risk
factors are IPPV, the administration of supplemental oxygen, the presence of a patent
ductus arteriosus and infection (chorioamnionitis, sepsis, pneumonia, meningitis) (39).
The main therapeutic options which may be associated with adverse side effects are
exogenous surfactant, caffeine, vitamin A, late corticosteroids, IPPV, oxygen, nutrition,
fluid therapy and prophylactic antibiosis (39). The interplay between these factors is
complex and the balance may be precarious.
1.4.2.2 Types of Ventilators
There are 3 main modes of ventilation employed in the management of RDS in
preterm babies. These modes are considered ‘gentle’ if used appropriately in these
patients with immature lungs that are poorly compliant and lacking in surfactant. The
decision making process regarding choice of ventilation strategy should be driven by
the individual patient’s pathophysiology but may also be influenced by availability of
equipment and past experience. Conventional mechanical ventilation (CMV) utilising
high respiratory frequencies and small tidal volumes, high-frequency oscillatory
ventilation (HFOV) and high-frequency jet ventilation (HFJV) have become established
as the ventilator modes most appropriate for the management of RDS in preterm
babies.
24
1.4.2.2.1 Conventional Mechanical Ventilation
There are a number of conventional mechanical ventilators available for use in a
neonatal intensive care unit (NICU). They can be broadly classified according to
whether or not they cycle between breaths when a set pressure or VT has been
delivered. These ventilators have a multitude of settings which include, but are not
limited to: time-cycled pressure-limited ventilation; flow-cycled pressure-limited
ventilation; pressure controlled ventilation; pressure support ventilation; volume
controlled ventilation; volume guarantee; pressure regulated volume control; volume
assured pressure support; synchronised, or patient triggered, ventilation; and assist
control (60). In the context of RDS these ventilators can be used to deliver gas which is
involved in gas exchange by bulk convection and diffusion.
1.4.2.2.2 High-frequency oscillatory ventilation
High-frequency oscillatory ventilation (HFOV) is a form of high-frequency ventilation
where gas is delivered at a rapid rate, with a tidal volume usually less than dead space
volume (61). HFOV generates individual breaths with either a piston and diaphragm
that is electromagnetically driven, a rotating ball and reverse jet, or a vibrating
diaphragm. However the oscillation is generated, both the exhalation and inhalation
phases are active (55). Adjustments to HFOV frequency and breath amplitude (∆P)
alter minute volume while changes to PEEP alter mean airway pressure (Paw), which in
combination with FiO2, determines oxygenation.
High-frequency oscillatory ventilation has increasingly been utilised in the clinical
management of preterm babies with RDS and has evolved over the last 3 decades.
25
Theoretical knowledge and clinical experience led to the recognition by Bryan and
Froese that using higher mean airway pressures than those measured during CMV is
essential to optimise oxygenation (45). Strategies aimed at achieving higher lung
volumes resulted in decreased FiO2 requirements, improved pulmonary mechanics and
less structural injury (62-64). The use of a high volume approach in clinical trials was
shown to be safe and effective and substantially decreased the incidence of BPD (65-
67) compared to earlier trials in which a high volume strategy was not protocolised.
The high volume strategy prioritises alveolar expansion over airway pressure and
creates mean airways pressures higher than those created during CMV (64, 68). Airway
pressure is only decreased after FiO2 is < 0.3 and when decreasing Paw does not
increase the FiO2 requirement. This link between higher Paw and improved
oxygenation formed the basis for studies focused on the high lung volume strategy.
The mechanisms of gas exchange and lung mechanics relevant to HFOV will be
discussed in Chapter 2.
1.4.2.2.3 High-frequency jet ventilation
High-frequency jet ventilation is also a form of high-frequency ventilation that has
been extensively utilised in NICUs. Individual breaths are generated by a flow-
interrupting pinch valve which is located outside the ventilator itself, and close to the
tracheal tube. A detailed description of HFJV and the ways it differs from HFOV will be
presented in Chapter 2.
26
1.4.3 Adjunctive Treatments
Management of preterm babies goes beyond ventilatory support: a multimodal
approach to their care is essential to minimise morbidity and mortality. Respiratory
support is just one aspect of treatment which includes adequate nutrition, appropriate
fluid therapy and pharmacological intervention (39). The overall aims are to limit lung
injury, avoid infection, provide optimal nutrition and carefully control fluid balance.
Caffeine is administered to stimulate spontaneous breathing and seems to be safe and
effective for the prevention of BPD and neurodevelopmental delays. A large
randomised controlled trial published in 2007 concluded that caffeine therapy for
apnoea of prematurity improved the rate of survival without neurodevelopmental
disability at 18 to 21 months in infants with very low birth weight (69). A recent study
assessing children at the age of 5 years, however, concluded that caffeine therapy as a
very low birth weight neonate was not associated with an improved rate of survival
without disability (70).
A number of studies demonstrate that vitamin A intake is inadequate in extremely low-
birthweight infants and that supplementation of vitamin A may reduce the risk of
chronic lung disease (71, 72). Vitamin A deficiency may promote chronic lung disease
by impairing lung healing, increasing the loss of cilia, increasing squamous-cell
metaplasia, increasing susceptibility to infection, and decreasing the number of alveoli
(73). While the short-term benefits of vitamin A supplementation are promising, the
long-term benefits are unclear (72) and as for caffeine treatment, the clinician must
always consider the risk and benefit of such treatment (39).
27
The administration of postnatal corticosteroids has been extensively investigated.
Halliday et al (2009) identified 47 randomised controlled trials of postnatal systemic
corticosteroid administration for the prevention of BPD (74, 75) and found a decreased
risk of death or BPD at 28 d and 36 w post menstrual age when corticosteroids were
used in the first week of life. The adverse effects that were identified included
gastrointestinal bleeding and intestinal perforation, hyperglycaemia, hypertension,
hypertrophic cardiomyopathy, growth failure, developmental delay, cerebral palsy and
abnormal neurological examination results (75). When corticosteroids were
administered later (> 7 days) the risk of BPD and death or BPD was reduced at both 28
d and 36 w (74). Once again, adverse effects included hyperglycaemia, hypertension,
infection and cerebral palsy (if the initial risk of BPD was low). Given these data it has
been suggested that systemic corticosteroids given after 7 d of life should be limited to
infants who cannot be weaned from mechanical ventilation. Furthermore, the dose
and duration of any such course should be minimised (39).
Diuretics have also been administered to preterm infants in an effort to improve lung
compliance. There are no known long term benefits from these therapies so while they
may be appropriate for infants with poor lung compliance or those who are difficult to
wean from a ventilator, it is prudent to use them transiently (39).
Nitric oxide (NO) is a selective pulmonary vasodilator and has been shown to improve
gas exchange and lung maturation in animal models (76-78). Nitric oxide is usually
delivered via the airways in an inhaled form into the inspiratory limb of the breathing
system. In a study investigating 3 doses of NO in term babies it was concluded that a
higher dose (80 ppm) did not seem to have any advantages over either 5 or 20 ppm
28
and caused an increase in methaemoglobin and NO2 levels (79). Its use for preterm
infants, however, has increased as the evidence suggests improved
neurodevelopmental and respiratory outcomes following treatment with inhaled NO
(80-83).
1.4.4 Surfactant
Surfactant is critical for lowering surface tension in the alveoli and prevents alveolar
collapse. It is usually produced by type II pneumocytes in time to promote alveolar
stability at term birth (84, 85). In utero the fetal lung is full of fluid which must be
cleared to enable the transition to air breathing at birth. The amount of fluid in the
lung after birth will be determined by the efficacy of fluid clearance mechanisms, the
length of labour, and whether or not delivery is vaginal or via caesarean section (84).
Delayed clearance of lung fluid will increase the potential for hypoxaemia and
hypercapnia. Furthermore the composition of the fluid also impacts upon the
transition to air breathing as fluid without surfactant, and therefore a high surface
tension, increases the risk of small airway obstruction and the pressure required to
open these airways (84).
In adult animal and human lungs, surfactant is a fluid composed of lipids and proteins,
the action of which is to lower the surface tension of an open air-water interface. It is
the saturated phosphatidylcholine species and surfactant protein B and C components
that give surfactant this unique property (85). Preterm infants with RDS have low levels
of surfactant that is deficient in these lipids and proteins. This deficiency promotes
alveolar collapse, instead of expansion, which contributes to the requirement for
mechanical ventilation.
29
The administration of exogenous surfactant soon after birth has become routine in the
clinical management of neonates with RDS and has yielded significant improvements in
morbidity and mortality (85-87). The most common method of administration is direct
intratracheal instillation (88) and the response to treatment has been described in
three phases: acute, lasting minutes; a more prolonged phase lasting hours; and
chronic, with effects lasting days or even weeks (85). The magnitude of the acute
response depends upon the biophysical properties of the surfactant and the extent of
distribution throughout the lung. Uniform distribution is ideal and whether or not it is
achieved is governed by surface activity, volume of surfactant, gravity, speed of
delivery of surfactant, ventilator settings and the presence of lung fluid within the lung
(85). Attention to these factors will optimise the delivery and efficacy of exogenous
surfactant.
Preterm lungs ventilated without surfactant rapidly develop epithelial disruptions in
the airways and pulmonary oedema as proteins from the vascular space enter the
airspaces (89). Oedema fluid will adversely affect lung mechanics and gas exchange
(90) and potentiate VILI. If surfactant is administered prior to the initiation of
mechanical ventilation it facilitates the movement of fluid out of the small airways and
into the saccules and interstitium, effectively increasing total lung capacity and
decreasing the inflation pressures required to deliver gas volume to the lung (91).
Surfactant is most often delivered together with mechanical ventilation. Ventilatory
mode may influence the distribution of surfactant to the lungs and the clinical
response of the patient. A rabbit study investigating the response to surfactant and
either CMV or HFJV following saline lavage induced lung injury demonstrated
30
improved gas exchange and reduced pulmonary right-to-left shunt in the animals in
which surfactant had been administered prior to HFJV (92). These results support the
hypothesis that the response to surfactant treatment in acute lung injury depends on
the mode of ventilation utilised after surfactant delivery. HFJV may facilitate the
delivery of surfactant to the distal alveoli, decrease alveolar dead space, support open
alveoli and improve gas exchange. This theory is not supported by any data and
warrants investigation in a controlled setting.
A pilot study of 28 newborn infants with RDS, meconium aspiration syndrome or
pneumonia who deteriorated in spite of optimal CMV and exogenous surfactant
therapy were treated with HFJV and continued surfactant therapy (93). The patients
that met the criteria for treatment with HFJV and additional surfactant showed
significant and sustained improvement in several respiratory variables. These results
suggested that the combination of HFJV and exogenous surfactant may be effective in
treating infants with more severe respiratory failure (93). Unfortunately this work has
not progressed and it remains that further investigation into the impact of specific
ventilation methods and strategies on the delivery, distribution and efficacy of
exogenous surfactants is required to optimise this therapy (88).
1.4.5 Postnatal Corticosteroids
The administration of corticosteroids to women at risk of preterm delivery induces
functional maturation of preterm fetal lungs (11) and has become established therapy
to improve the outcome for premature infants (12). The administration of postnatal
corticosteroids however is less encouraging. A recent review of several studies focused
on the risk/benefit balance of postnatal corticosteroids administered to premature
31
babies for prevention and treatment of BPD highlights the risk of long-term adverse
neurodevelopmental outcomes (94). Their recommendations are that systemic
administration of corticosteroids for prevention or treatment of BPD: (i) should not be
used during the first 4 days of life; (ii) is not indicated in the first 3 weeks of life nor (iii)
in extubated infants (nasal ventilation or oxygen therapy) (94). Furthermore, these
authors advise that the systemic administration of steroids should only be considered
after the first 3 weeks of life in very preterm infants to facilitate extubation. In this
scenario corticosteroids may help wean the infant from the ventilator and decrease
the requirement for oxygen supplementation. The justification for these suggestions is
based upon potential unfavourable neurocognitive outcomes associated with the use
of postnatal steroids (94). A policy statement from the American Academy of Pediatrics
was released in 2010 and is in a similar vein, highlighting the deficiency of data to
support the use of corticosteroids in the post natal period without careful
consideration of the potential risks. The Academy did, however, recommend that
consideration of a course of corticosteroids be given if the patient is ventilator-
dependent at 1-2 weeks of age.
The debate regarding the administration of postnatal steroids is not over: in light of
Australian data it appears that low-dose dexamethasone treatment after the first week
of life facilitates extubation thereby decreasing the days of intubation of ventilator
dependent very preterm and extremely low birth weight infants (95). Doyle et al
(2006) reached these conclusions after a randomised controlled trail of either
dexamethasone or saline was administered to 70 infants recruited from 11 centres at a
median age of 23 days. A 2 year follow up on these patients revealed that there was no
32
difference in the incidence of mortality or major disability at 2 years of age between
the treatment and the saline groups (96). Until further data are available it remains
that while dexamethasone administered after the first week of life may have short
term benefits, the long term implications are unknown.
1.5 Side Effects of Positive Pressure Ventilation
Positive pressure ventilation disrupts normal physiological processes and may interfere
with lung structure and cardiac output, and therefore pulmonary and systemic blood
flow. Furthermore, if blood flow to the brain is compromised and cerebral oxygen
delivery does not meet cerebral oxygen demand normal neurological development
may not occur. The side effects of IPPV will be divided into those effects pertaining to
the respiratory (ventilator induced lung injury), haemodynamic (pulmonary blood flow,
persistent pulmonary hypertension of the neonate and systemic blood flow) and
neurological systems.
1.5.1 Ventilator Induced Lung Injury
While mechanical ventilation has decreased mortality rates in neonates since its
introduction in the 1960s, at the same time it has created a new set of potentially life
threatening conditions for these patients (97). Ventilator induced lung injury is
considered an important risk factor for the development of BPD from barotrauma,
atelectatrauma, volutrauma and biotrauma (98, 99) and may have long term
implications for the patient. In preterm infants VILI may be attributed to a plethora of
mechanical ventilation strategies (65, 98, 100). The factors that contribute to the
deleterious effects of mechanical ventilation include the applied airway pressure, the
33
volumes changes associated with that airway pressure, the size of the patient, the
duration of ventilation, whether or not the thorax is open or closed and the presence
of pre-existing respiratory disease (98).
Barotrauma causes the leakage of air due to disruption of the airspace wall (98). The
term is used to describe the impact of pressure related dysfunction resulting from
exposure to persistent, elevated pressures (99). Alveolar distension during IPPV causes
damage to the pulmonary epithelial-endothelial barrier which allows air to shift into
the pulmonary interstitium (101). The consequences of this damage includes:
interstitial airleak; pneumothorax; subcutaneous emphysema; pneumomediastinum;
pneumoperitoneum and pneumopericardium (102). Furthermore, a complex cascade
of physiological events occurs including fluid and protein shifts across the blood-air
interface resulting in a high permeability pulmonary oedema (103). Gas exchange
becomes compromised as ventilation and perfusion (V/Q) mismatch develops and lung
compliance decreases. The mechanical stress associated with barotrauma may activate
humoral and cellular immunological responses leading to the release of pro-
inflammatory mediators and neutrophils (99). Fibrin is then deposited in the alveolar
membrane, increasing V/Q mismatch and decreasing compliance further.
Atelectatrauma refers to lung injury associated with atelectasis and the consequent
shear forces associated with repetitive opening of collapsed alveoli. This creates V/Q
mismatch and increases the inflation pressures required to recruit those areas of the
lung (91). According to LaPlace’s law, the pressure required to inflate a lung unit
depends on the initial radius of that unit. To achieve a certain volume change in larger
alveoli, the necessary pressure changes are much smaller compared to alveoli which
34
are collapsed or have a lower volume. It follows that the pressure needed to keep
alveoli open is lower at a higher functional residual capacity. The critical opening
pressure of a lung unit is therefore inversely proportional to the size of that unit so
higher pressures are required to recruit collapsed lung. The delivery of higher
pressures is potentially dangerous as these pressures may be transmitted to the
normal areas of lung, opening alveoli, promoting stretch, and causing over distension.
Furthermore, the opening of those collapsed units imposes large shear forces which
promotes alveolar disruption (91). Surfactant helps prevent alveolar collapse by
decreasing the surface tension of distal alveoli. The surface area of the alveolus will
however affect the efficacy of surfactant, especially if the surface area is smaller than
the total surface of surfactant molecules. The surfactant molecules will be squeezed
off the alveolar surface towards the alveoli and become inactive. When that lung unit
is inflated again, the surface is replenished with surfactant molecules, a proportion of
which will be ineffectual (104). Opening the lung, and keeping the lung open is the best
strategy for minimising atelectatrauma.
Volutrauma refers to lung injury associated with the static overdistention of the lung
as well as the cyclical delivery of high tidal volumes. By definition, it is independent of
the peak inspiratory pressure required for tidal volume delivery (98, 105). Volutrauma
can occur with only brief exposures to large tidal volumes (106) including as few as 6
consecutive large tidal volume breaths associated with the initiation of ventilation
after birth (107). Thus volutrauma is a risk for the preterm baby receiving IPPV during
postnatal resuscitation if tidal volumes are not monitored. Volutrauma not only causes
acute changes, but increases the injury associated with subsequent IPPV (106). At a
35
microscopic and a molecular level, volutrauma leads to a plethora of abnormalities:
alveolar epithelial cell damage; alveolar protein leakage; altered lymphatic flow;
hyaline membrane formation; inflammatory cell influx; decreased lung compliance and
altered surfactant structure and function (106, 108, 109).
While efforts should be made to minimise volutrauma, there are sparse data in the
literature documenting the effect of predetermined tidal volumes during IPPV. One
prospective randomised controlled trial in preterm infants compared 3 mL kg-1 and 5
mL kg-1 tidal volumes delivered during volume guarantee ventilation. The authors
hypothesised that the lower tidal volume breaths would be associated with less
inflammation and less BPD (110). Their results found the converse, that the lower tidal
volume breaths were associated with greater expression of inflammatory markers,
likely due to progressive atelectasis with the delivery of small tidal volumes in the
absence of an accompanying strategy to optimise distending lung volume. This study
highlights the importance of delivering a targeted ventilation strategy: adequate PEEP
should be used to prevent atelectatrauma and optimal tidal volumes should be used to
prevent volutrauma.
Biotrauma refers to lung injury associated with the release of pro-inflammatory
mediators. This occurs in addition to the lung parenchymal and airway epithelial
changes previously described and results in an increased concentration of cytokines in
the lung tissue, which in turn incites an inflammatory response (102, 111, 112).
Quantification of these pro-inflammatory mediators is particularly useful when
assessing lung injury in an experimental setting as it provides information about the
potential systemic effects of IPPV.
36
Barotrauma, atelectatrauma, volutrauma and biotrauma have been documented in
experimental studies in animals (98, 102, 111, 113) and while they may not be easy to
identify early in the pathogenesis of VILI in a clinical setting they are likely to occur to
some degree in response to mechanical ventilation. Controlled animal studies looking
directly at HFJV are scarce, but a study presented in 1990 was a turning point as it
reported less lung injury in a rabbit model of RDS when comparing high and low
volume strategies during HFJV and HFOV (114). Investigation and documentation of
injury associated with HFJV is therefore warranted to fully understand the impact of
this ventilation strategy on the lungs.
1.5.2 Haemodynamic Consequences of Positive Pressure Ventilation
Lowered right ventricular filling pressure, decreased central blood volume and a
consequent decrease in cardiac output are all side effects of IPPV (115). Animal studies
have demonstrated superior preservation of haemodynamic function during HFJV
when compared to CMV in dogs (115) and when compared to high-frequency positive
pressure ventilation in rabbits (116).
1.5.2.1 Pulmonary Blood Flow
A cat study comparing haemodynamic function during HFJV and HFOV aimed to
investigate the impact of Paw on cardiac output and pulmonary vascular resistance
(117). Despite demonstrating that increasing Paw caused a decrease in cardiac output
and an increase in pulmonary vascular resistance there was no cardiovascular
advantage of one strategy over the other. The major limitation of this study was the
37
range of Paw was low: 2-12 cmH2O. A bigger difference may have been demonstrated
if higher airway pressures were applied.
Kawahito et al (2000) studied HFJV and CMV in adult patients with healthy lungs to
compare the impact of each strategy on pulmonary perfusion and cardiac output
(118). They used transoesphageal echocardiography and found significantly decreased
pulmonary arterial pressure and left atrial pressure during HFJV. These findings
correlated with increases in cardiac output and ejection fraction in their cohort of
healthy patients. Their conclusions that there is a haemodynamic advantage of HFJV
over CMV attributable to lower PIP during HFJV warrant consideration. While the
haemodynamic implications of a ventilation strategy will be affected by the presence
of cardiovascular and pulmonary disease it is still noteworthy that their findings
convincingly demonstrate that HFJV interferes less with venous return and therefore
cardiac output. It is also important to note that the respiratory frequency during HFJV
was 3 Hz in their study. This is considerably lower than would be used in a preterm
infant with RDS.
High-frequency jet ventilation has also been investigated in children. Kocis et al (1992)
studied the impact of pulmonary vascular resistance on cardiac output in the setting of
changing Paw in the transition from CMV to HFJV (7). They found that decreasing Paw
by 50 % caused a 59 % reduction in pulmonary vascular resistance and a 25 % increase
in cardiac output. Their conclusions were that HFJV was a suitable option for their
patient cohort (children with respiratory failure following congenital heart surgery).
While their patients are different to the target population for HFJV in this thesis, these
38
cardiovascular data are nonetheless supportive of the use of HFJV in patients with
severe respiratory dysfunction.
Measurement of pulmonary blood flow is difficult in a clinical setting and given the
paucity of information on the impact of HFJV on pulmonary blood flow, it is prudent to
examine this in a controlled experimental environment. Soon after birth, neonates
undergo the transition from gas exchange across to the placenta to gas exchange
across the alveolar membrane. This transition requires establishment of a low
pressure pulmonary circulation and inflation of the lungs with air. If these patients
require mechanical ventilation, the intervention should not prevent this transition. For
this reason, and in light of the airway pressure required during mechanical ventilation,
animal studies are the only method by which this can be closely examined.
1.5.2.2 Persistent Pulmonary Hypertension of the Neonate
Persistent pulmonary hypertension of the neonate (PPHN) is defined as failure of the
pulmonary vasculature to relax (or dilate) at birth. The transition from fetal to neonatal
life is therefore complicated by right to left shunting of blood, resulting in V/Q
mismatch, venous admixture and hypoxaemia. Persistent pulmonary hypertension
occurs in approximately 1-2 newborns per 1000 live births and despite significant
improvements in treatment it is associated with substantial infant mortality and
morbidity (119). Diagnosis of PPHN can be made with echocardiography but a clinical
impression of the resistance in the pulmonary circulation versus the resistance in the
systemic circulation can be made by comparing pre- and post-ductal SpO2. If
pulmonary vascular resistance is higher than systemic vascular resistance, right-to-left
39
shunting will occur through the ductus arteriosus. Pre-ductal (right forearm) SpO2 will
therefore be higher than post-ductal (lower extremities) SpO2.
Patients with PPHN may have adequate respiratory drive but can remain hypoxaemic
in spite of high FiO2. They may therefore require mechanical ventilation (77). The
suitability of HFJV for these patients has been investigated in a controlled prospective
clinical trial comparing HFJV and CMV. The authors found that in the acute period HFJV
improved oxygenation and ventilation without significantly increasing mortality.
Furthermore, HFJV may be a useful adjunct for stabilisation of the condition in
neonates with severe PPHN (120).
A potential complication of mechanical ventilation of patients with PPHN is a further
increase in pulmonary vascular resistance secondary to positive intrathoracic pressure.
Given the lower airway pressures during HFJV, it may have the least impact on
pulmonary vascular resistance and be the most appropriate ventilation strategy for
patients with PPHN.
Inhaled nitric oxide is widely used to manage PPHN (77, 119, 121). Nitric oxide is a
vasodilator and improves systemic oxygenation by decreasing the right to left shunting
of blood through the ductus arteriosus. It decreases the work of the right side of the
heart and prevents further right ventricular hypertrophy that occurs in response to the
increased pulmonary vascular resistance. The delivery of NO may be facilitated by HFJV
as the flow streaming of the inspired gases helps distribute it to the distal alveoli (122).
40
1.5.2.3 Systemic Blood Flow
While perfusion of the pulmonary vasculature is essential to ensure matching of
ventilation and perfusion and therefore normal gas exchange, changes in systemic
blood flow will determine oxygen delivery to the brain, organs and other tissues.
Assessment of cardiac output is paramount to ensure oxygen supply exceeds oxygen
demand. Animal and clinical studies documenting the impact of HFJV on systemic
blood flow consistently find that there is comparable or better preservation of
systemic arterial blood pressure or cardiac output during HFJV compared to both HFOV
and CMV (7, 123-125).
Intrathoracic pressure during IPPV is higher than during spontaneous ventilation. This
pressure will alter the flow, pressure and resistance of intrathoracic vascular structures
and potentially decrease venous return and cardiac output. These alterations may be
minimised if the increase in intrathoracic pressure is small and short lived.
Furthermore, if respiratory frequency and heart rate are comparable cardiac output
may not decrease (126). Whereas synchronisation of HFJV PIP with ventricular systole
has been proposed for optimisation of ventricular loading in adult patients it is unlikely
to be as applicable to neonates. However, as the lowest respiratory frequency during
HFJV is 3 Hz (though this is determined by the manufacturer) and the normal heart
rate of the neonate is between 2-3 Hz it may be possible to synchronise every second
beat.
Intrathoracic pressure variations during HFJV are smaller than during CMV and it has
been postulated that there will be less cyclical change in cardiac output as a
consequence (127). Sherry et al (1988) measured cardiac output during CMV and HFJV
41
in postoperative elective cardiac surgery patients to determine the impact of CMV or
HFJV at different PEEPs (0, 5, 10 cmH2O) and found that during HFJV with 0 cmH2O
PEEP there was little change in cardiac output. Increasing PEEP increased Paw and
intrathoracic pressure but did not significantly alter cardiac output during HFJV (127).
As premature infants undergo the transition to breathing air systemic arterial blood
pressure will increase. If this increase in blood pressure, and therefore perfusion of the
brain, organs and tissues, is inhibited oxygen delivery may not meet oxygen demand.
While the literature suggests that HFJV has relatively benign affects on cardiac output
it remains important to monitor continuously for evidence of anaerobic metabolism.
1.5.3 Central Nervous System Consequences of Positive Pressure Ventilation
Neurological injury associated with mechanical ventilation is assessed by the presence
of intraventricular haemorrhage (IVH) and periventricular leukomalacia (PVL). The
manifestations of this neurological injury vary in severity but cerebral palsy to some
degree is 25 to 30 times more likely to occur in infants weighing less than 1500 g (128).
Neurodevelopmental follow-up must also be performed in the first few years of life to
determine the consequences of any neurological injury sustained in the first few days
of life (129). The incidence of both IVH and PVL has a direct relationship to cerebral
palsy, intellectual impairment and visual disturbances (55). The causes of these lesions
are unclear but the following factors may predispose to both: asphyxia; severe
haemorrhage; septicaemia; patent ductus arteriosus; low arterial blood pressure;
impaired cardiac function; PaO2 and PaCO2 (130, 131).
42
1.5.3.1 Intraventricular Haemorrhage and Periventricular Leukomalacia
Intraventricular haemorrhage refers to bleeding into the cerebrospinal fluid filled
lateral ventricles of the brain. The majority of IVH occurs in the first few days of life
and the incidence of it is inversely related to gestational age and body weight at birth
(132, 133). It is graded from I to IV by ultrasonography (134) and grades III or IV are
considered severe (135). Periventricular leukomalacia is the most common ischaemic
brain injury in premature infants and occurs in the white matter adjacent to the lateral
ventricles.
Hypocapnia has been investigated as a cause of IVH and PVL but the results of various
studies do not demonstrate an absolute link between hyperventilation (hypocapnia)
and adverse neurological outcomes. Murase and Ishida (2005) found that hypocapnia
(PaCO2 < 3.3 kPa (25 mmHg)) in the first 48 h of life was significantly associated with
cerebral palsy and late-onset PVL. This suggests that any association of cerebral palsy
with early hypocapnia is limited to a minor subtype of PVL, or to infants with cerebral
palsy not related to PVL (130). Fujimoto et al (1994) found that hypocapnia, and other
complications, were associated with PVL and concluded that mechanical ventilation in
premature infants should be managed carefully to avoid a PaCO2 lower than about
2.67 kPa (20 mmHg) (131). Other studies have similarly described periods of
hypocapnia as contributing factors to a poor neurological outcome (56, 136).
The HIFI Study comparing HFOV and CMV demonstrated a significant increase in
severe IVH and PVL (and respiratory and haemodynamic complications) in patients
ventilated with HFOV (137). The authors speculated that the poor outcome results
were a consequence of failure to emphasise volume recruitment and maintenance in
43
the respiratory management of these infants (45). Although this study did not focus on
the PaCO2 targets for these patients and the impact of hypocapnia on cerebral blood
flow, it raised questions about whether the ventilation modality itself, or the blood gas
status of the patient, had an impact on adverse neurological outcome.
There are few clinical trials focusing on the neurological consequences in patients
managed with HFJV, and the results are inconsistent. Keszler et al (1997) documented
a randomised, controlled clinical trial of HFJV and CMV in 130 preterm infants between
700 and 1500 g born before 36 w post conceptional age. They found that HFJV reduced
the incidence of BPD at 36 w, decreased exposure to hypocapnia and reduced the risk
of grade III and IV IVH and/or PVL compared to CMV (138). Wiswell et al (1996) studied
infants born prior to 33 w gestation between 500 and 2000 g. In this randomised
controlled trial they aimed to ventilate infants to normocapnia and found that HFJV
was associated with a greater risk for adverse outcomes (grade IV IVH, PVL or death)
when compared to CMV (139). These authors went on to examine the role of
hypocapnia in the development of IVH and PVL during HFJV and found that the infants
with PVL had been hypocapnic for longer during the first day of life (140).
Direct comparison between HFJV and CMV has been attempted but the populations
are small (100, 120, 141). The results suggest HFJV is associated with lower mortality
but no significant differences were found in the incidence of IVH. There are multiple
limitations to these studies so to determine the cause of IVH and/or PVL the studies
need to target the at-risk population, include enough patients to give the results
power and incorporate long term pulmonary and neurodevelopmental follow up
assessment.
44
The mechanism by which hypocapnia affects the development of IVH and PVL remains
unclear. Whether the PaCO2 level or the duration of time spent at a low PaCO2 has a
causative effect warrants further investigation. Furthermore, it’s possible the blood
gas status created during mechanical ventilation has a greater impact on adverse
neurological outcomes than the ventilation modality itself. Understanding how to
manage a particular ventilator to achieve target blood gas ranges is essential to
maintain a balance between the risks and benefits of mechanical ventilation.
Initial HFJV strategies more often created hypocapnia for periods of time. The focus on
managing Paw to maintain oxygenation and concern about the adverse effects of high
PEEP kept ∆P high enough to create hypocapnia. As the strategies have evolved and
higher PEEP settings are used the incidence of hypocapnia may decrease. Optimal
volume HFJV strategies have been shown to improve oxygenation and decrease
exposure to hypocapnia which in turn reduces the risk of grade III and IV IVH and/or
PVL (142). Future studies should focus on the incidence of adverse neurological effects
in the face of normocapnia.
1.6 Assessment of Lung Injury
The role of inflammation in VILI is important in the pathogenesis of BPD (143, 144). It
may be initiated by a pulmonary fetal inflammatory response and be exacerbated by
mechanical ventilation and exposure to supplemental oxygen. The response is a
complex interaction between proteins that attract inflammatory cells (chemokines),
proteins that facilitate the transendothelial migration of inflammatory cells from blood
vessels (adhesion molecules), proteins that promote tissue damage (pro-inflammatory
45
cytokines and proteases) and proteins that modulate the process (e.g. anti-
inflammatory cytokines, binding proteins and receptor antagonists) (143).
Inflammation as a prelude to injury can be assessed in a number of ways: measuring
the protein content and assessing the cell population of bronchoalveolar lavage (BAL)
fluid; identifying the cellular infiltrate in lung tissue; assessing the expression of
messenger RNA (mRNA) for pro-inflammatory mediators; and measuring the products
of mRNA transcription. Non-invasive tests on BAL fluid are more appropriate in the
clinical setting and help identify infants at risk of BPD. In a research environment,
measurement of biomarkers in lung tissue, and BAL fluid, will further contribute to an
enhanced understanding of the significance of inflammation associated with
mechanical ventilation.
An appreciable increase in biomarkers may take many hours or days (144) which
makes it difficult to identify the initial injury pathways stimulated immediately after
birth. Injury is initiated on commencement of mechanical ventilation, when the lungs
are partially liquid-filled, surfactant deficient and partially aerated. The quantification
of expression of early response genes in the immediate postnatal period revealed that
VILI during the immediate newborn period can initiate changes in gene expression
within 15 minutes. This abnormal gene expression will potentiate inflammation and
promote abnormal lung development (145). Wallace et al concluded that connective
tissue growth factor (CTGF), cysteine-rich 61 (CYR61), early growth response factor 1
(EGR1), interleukin (IL) 1β, IL-6 and IL- 8 are likely to be useful biomarkers for VILI in
the newborn, particularly in the short term (111).
46
Recent work has focused on the identification of mediators of lung injury and
characterisation of their interaction with alveologenesis. This discovery has enabled
identification of the protective mechanisms specific to the mediator of injury which in
turn enables robust protection against lung disease (146). Transforming growth factor
(TGF) β1 was identified in the lungs of preterm infants and is involved in inflammatory
and repair processes encountered in acute and chronic lung disease (147). High initial
levels of TGF β1 persisted over time and were predictive of the need for oxygen
therapy at home. Minoo et al have recently concluded that fibroblast growth factor
(FGF) 10 offers a distinct protective effect by attenuating the TGF β1 pathway and that
FGF 10 treatment strategies may provide protection to neonatal and other forms of
lung diseases caused by TGF β1 (146).
1.6.1 Bronchoalveolar Lavage Fluid
Bronchoalveolar lavage fluid can be collected via a tracheal tube in a clinical setting
and provides information about the permeability of the alveolar membrane and the
infiltration of inflammatory cells into the alveoli.
1.6.1.1 Protein in Bronchoalveolar Lavage Fluid
Two barriers form the alveolar-capillary interface: the microvascular endothelium and
the alveolar epithelium. In the acute phase of lung injury there is an influx of protein
rich oedema fluid into the air spaces as a consequence of increased permeability of the
alveolar-capillary barrier (148). The importance of endothelial injury causing increased
vascular permeability and pulmonary oedema in acute RDS is well established (149).
However, the importance of epithelial injury to recovery from lung injury has become
47
better recognised (150) and the degree of alveolar injury is a predictor of outcome
(149).
The increase in capillary-alveolar permeability to plasma proteins can be quantified by
measuring BAL fluid total protein concentration (19, 106). An increase in BAL fluid
protein concentration is considered a global indicator of lung injury (106). The plasma
proteins that flood the alveoli contribute to the development of fibroproliferation
which in turn contributes to the risk of fibrosis. Efforts to monitor and reduce plasma
protein accumulation in the alveoli could benefit the patient (151).
The BAL procedure may be performed during sedation or anaesthesia, or post-mortem
in animals, and has been included in the overall assessment of lung injury in a number
of studies using a number of species. Measurement of total protein is an indicator of
lung injury but does not characterise the injury unless specific proteins or the actual
size of the protein is determined.
1.6.1.2 Inflammatory Cells in Bronchoalveolar Lavage Fluid
Extravasation of inflammatory cells due to endothelial and epithelial damage occurs as
a result of lung injury. Bronchoalveolar lavage fluid collected ante- or post mortem can
be examined to determine the number and composition of these inflammatory cells to
help determine the degree of lung injury. Bronchoalveolar lavage inflammatory cell
counts (neutrophils and mononuclear cells) increase in ventilated animals compared to
unventilated controls (106, 144, 152).
A recent review by Reynolds (2009) acknowledges that collection and analysis of BAL
fluid is a relatively non-invasive diagnostic test, but that it has limitations as a
48
diagnostic tool. In both a clinical and a research setting BAL fluid analysis contributes
to the understanding of disease processes but the cellular components of BAL fluid are
not strongly correlated with definitive diagnosis (153).
1.6.2 Lung Tissue
Analysis of lung tissue is primarily a research tool. The identification and localisation of
inflammatory cells will characterise alterations in lung tissue associated with
mechanical ventilation. The maturity of inflammatory cells aids in assessing the
duration of the inflammatory process while the location helps characterise the insult.
1.6.2.1 Inflammatory Cells in Lung Tissue
Localisation of inflammatory cells in the microanatomy of the lung is useful to further
characterise an inflammatory response occurring as a prelude to lung injury.
Immunohistochemical staining and histopathological analysis of targeted enzymes
such as inducible nitric oxide synthase (iNOS) and myeloperoxidase (MPO) indicate the
location of inflammatory cells and form part of the overall assessment of inflammation
(112). Positive MPO staining identifies neutrophils and mononuclear cells while
positive iNOS staining identifies macrophages. This differentiation helps age the
inflammatory infiltrate.
1.6.2.2 Gene Expression in Lung Tissue
Describing the pattern of gene expression and activation associated with lung injury
provides the most precise account of the initial mechanisms involved in lung injury. As
the greatest risk of injury may be during the period immediately after birth when the
49
lungs are particularly vulnerable, early response genes and growth factors are of
particular interest (108, 145, 154). The role of early response genes in VILI in the
preterm neonate was elucidated recently (145). Quantitative polymerase chain
reaction (qPCR) of mRNA for CTGF, CYR61, EGR1, IL-1β, IL-6 and IL-8 indicated that
resuscitation and mechanical ventilation at birth with high tidal volumes caused
upregulation of these genes compared to mechanical ventilation with low tidal
volumes. The authors concluded that these findings were indicative of more lung injury
from mechanical ventilation with low tidal volumes.
The precise role of specific mediators in the pathogenesis of VILI is not entirely
understood. The literature regarding the specific role of inflammatory mediators is
expanding, but is not comprehensive (155, 156). Inflammation and injury is a complex
process: initially gene expression is altered and these genes are subsequently
translated to specific proteins. Understanding the temporal properties of specific
genes and proteins enables appropriate analyses to demonstrate inflammation and
injury. Copland et al (2003) demonstrated that altered gene expression occurs before
demonstrable lung injury and that these alterations are time and stretch dependent
with characteristic spatial distributions (109). The choice of specific genes for
examination in the context of a particular study should be made carefully. Factors that
influence the suitability of particular genes include: the duration of the study; the
underlying pathophysiology of the lungs; and the ventilator strategy under
investigation.
50
1.7 Lung Protective Ventilation Strategies
Mechanical ventilation was originally achieved by creating a negative extra-thoracic
pressure to simulate the negative intra-thoracic pressure created during normal
inspiration. These iron lungs mimicked normal spontaneous ventilation insofar as the
respiratory rate, tidal volume and airways pressures were comparable. There were
practical problems unrelated to breathing that led to the development of IPPV. Overall
patient management was a lot easier and ventilation was more efficient with the
ventilator attached to a tracheal tube. The main causes of ventilator associated
morbidity arise from the positive pressure within the chest and the impact on lung
tissue, pulmonary circulation and systemic and cerebral blood flow. These morbidities
may also be described as volutrauma, atelectatrauma, biotrauma and barotrauma with
reference to the impact on the lungs themselves. High-frequency ventilators were
developed as an alternative to conventional positive pressure ventilation as the
concept of smaller breaths more often was believed to be associated with fewer
adverse side effects (157).
Various ventilation strategies have been developed to reduce the prevalence of BPD,
but despite these advances the risk of BPD is still high for very preterm babies (30, 34).
There are 3 gentle ventilation strategies available to the neonatologist in the clinical
realm: gentle CMV with relatively high respiratory rates and small tidal volumes, HFOV
and HFJV. Each of these strategies has evolved to achieve a similar broad aim which is
described by the open lung approach. The open lung approach was first documented
in 1992 and incorporates the following basic treatment principles (91, 158):
51
1. Open the whole lung with the necessary inspiratory pressure
2. Keep the lung open with PEEP above the closing pressure
3. Maintain optimal gas exchange at the smallest possible pressure amplitude to
optimise CO2 removal
This approach has changed the focus of ventilator management from targeting
physiological goals alone to protecting the lung from injury and decreasing the
cytokine modulation of the lung (98, 159, 160). The open lung approach can be applied
during CMV, HFOV and HFJV and aims to recruit and stabilise alveoli, minimise
atelectasis, and maximise gas exchange area without injury to the lung or compromise
to systemic or pulmonary blood flow (158).
Alveolar recruitment refers to the dynamic process of opening previously collapsed
lung units by increasing transpulmonary pressure (158). This pressure change is
primarily responsible for VILI (161, 162) and the effects of pressure should always be
monitored. The choice of recruitment manoeuvre will depend upon the individual
patient and the baseline ventilator mode. The delivery of a sustained high distending
pressure, an increase in PEEP and/or a transient increase in PIP will all facilitate
alveolar recruitment, but may also compromise blood flow and lung structure. Getting
the balance right is essential during lung protective ventilation.
Understanding hysteresis of the pressure-volume curve of lung inflation and deflation
will help attain this balance. The volume achieved on the deflation limb of the
pressure-volume curve is larger for the same distending pressure, compared to that
achieved on the inflation limb. The point of maximal curvature of the deflation
pressure-volume curve is the point at which the lowest pressure achieves optimal lung
52
volume and PaO2 (163). Targeting ventilation to this point by delivering a sustained
inflation followed by small tidal volumes (5-6 mL kg-1) with PEEP above the inflection
point of the pressure-volume curve has been demonstrated to minimise lung injury in
a rat model (164). Alveolar recruitment using a sustained inflation followed by small
breaths with a PEEP below the inflection point will also boost the ventilator to cycle
onto the deflation limb of the pressure-volume curve (165). These findings suggest
that if sufficient lung volume recruitment is achieved with a sustained inflation that
relatively low airway pressures can be used to maintain tidal ventilation with a lower
pressure cost (165).
53
54
2 High-frequency Ventilation
2.1 High-frequency Ventilation
2.1.1 Gas Mixing during High-frequency Ventilation
The mechanisms determining gas flow, gas mixing and airway pressure during high-
frequency ventilation (HFV) are fundamentally different to ventilation at respiratory
frequencies employed during CMV. HFV is characterised by ventilation with tidal
volumes smaller than dead space volume but adequate gas exchange can still occur
because the increased energy of the gas molecules at the high frequencies and flows
leads to augmented mixing of gas in the airways (166). The dynamics of gas flow
distribution during HFV involve a number a different mechanisms including bulk
convection, asymmetric velocity profiles, pendelluft, cardiogenic mixing, Taylor
dispersion and turbulence, molecular diffusion and collateral ventilation (166, 167)
(Figure 2-A).
55
Figure 2-A Gas Transport Mechanisms during High-frequency Ventilation. The gas transport mechanisms responsible for gas exchange during CMV (convection, convection and diffusion, and diffusion) are enhanced during HFV by seven mechanisms: turbulence; direct ventilation of proximal alveoli; turbulent flow with lateral convective mixing; pendelluft; gas mixing due to velocity profiles that cause a central stream of inspiratory gases along the airways and a stream of expiratory gas around this central stream; laminar flow with lateral transport by diffusion (Taylor dispersion); and collateral ventilation through non-airway connections between neighbouring alveoli (166).
High-frequency ventilators do not mimic normal breathing. Much smaller tidal volumes
are delivered at a much higher frequency and gas exchange occurs in a highly efficient
manner similar to that achieved by panting animals whereby:
Equation 2-A
V’CO2 = f x VT2
where V’CO2 = rate of CO2 elimination, f = ventilator frequency, and VT = tidal volume.
56
During normal breathing, effective or physiological dead space must be greater than or
equal to anatomical dead space (157). In the context of the equation above there must
be a lower limit on VT that will effectively provide alveolar ventilation. That lower limit
is related to the effective or physiologic dead space of the lungs according to:
Equation 2-B
VA = f x (VT – VD, physiol)
where VA = alveolar ventilation and VD, physiol = physiological dead space.
As VT approaches VD, physiol, direct ventilation of the alveoli approaches zero at
conventional breathing frequencies. However, when an animal pants, physiological
dead space becomes smaller than anatomic dead space (168). This was demonstrated
in 1915 by Henderson et al in an experiment demonstrating the jet stream of smoke
created during fast and shallow breathing through a tube. Thus, the extent to which a
smaller tidal volume can ventilate alveoli is balanced by an increased in breathing
frequency.
2.1.2 Mechanical Properties of the Lung and High-frequency ventilation
The interaction of resistance, compliance and inertance in the frequency range of high-
frequency ventilators will govern gas exchange. Resistance refers to the opposition to
flow and is determined by the dimensions of the airway, the viscosity of the gas and
whether flow is laminar or turbulent. Compliance is determined by assessing changes
in volume per unit of pressure and inertance is the pressure required to cause a
change in flow. The behaviour of the respiratory system during mechanical ventilation
is determined by these 3 components (169). They therefore give the respiratory
57
system a measure of impedance which will vary according to respiratory frequency.
Impedance is an important determinant of the efficiency of ventilation and is a global
term that encompasses compliance, resistance and inertance. Impedance therefore
represents a mechanical barrier to flow and as impedance increases, greater changes
in pressure are required to generate an equivalent flow (and VT) (170). As the
amplitude of the airway pressure waveform increases, the risk of barotrauma
increases, but if it is too small airway closure and alveolar collapse may ensue. It is
therefore essential to understand impedance throughout the respiratory system (from
the breathing system, the endotracheal tube, the airways and lung tissue) to
determine the magnitude of the pressure required to transmit gas to the alveoli during
high-frequency ventilation safely (170).
In vitro and in vivo studies found that a resonant frequency is observed below which
elastic behaviour of the lungs is dominant (impedance decreases with increasing
frequency) and above which inertial behaviour is dominant (impedance increases with
increasing frequency) (169). The resonant frequency of the lungs will vary according to
maturity and pathology and is inversely related to the square root of the compliance
and the inertance. In the overdamped lung (e.g. preterm lung), the corner frequency
(which is inversely proportional to the resistance and the compliance) defines the
frequency above which the pressure required to ventilate those lungs is minimised
(171). Hence optimal frequency for ventilation most likely falls between the corner
frequency and the resonant frequency. A study of 6 newborn infants with RDS found
the resonant frequency to range from 13 to 23 Hz (169) when an endotracheal tube
was in position. The impedance displayed compliance-like behaviour below the
58
resonant frequency and inertance-like behaviour above the resonant frequency.
Without an endotracheal tube the resonant frequency was higher (169). This study
found that endotracheal tube resistance was equivalent to ~ 50 % of the respiratory
system resistance distal to the tip of the endotracheal tube, whereas virtually all the
respiratory system compliance resided distal to the tip of the endotracheal tube (169).
If the frequency of ventilation is greater than the resonant frequency of a set of lungs,
overriding the resonant frequency requires less pressure and energy compared to a
mechanical frequency that is less than the resonant frequency. The smaller the lungs
are, the higher the resonant frequency: resonant frequency of adult lungs is ~ 4 Hz,
while that of premature infants lungs may be as high as 40 Hz (157). High-frequency
ventilators deliver breaths up to 780 times per minute so come considerably closer to
the resonant frequency of immature lungs than conventional mechanical ventilators
do. This property alone is important when taking into account the potential for side
effects associated with positive pressure ventilation.
2.1.3 Airway Pressure Waveforms during High-frequency Ventilation
In the presence of normal lung compliance, the amplitude of the airway pressure
waveform is greatly attenuated during HFV as gas passes at rapid rates through the
rigid endotracheal tube (Figure 2-B) (55). During CMV, both peak and mean airway
pressures are transmitted to the distal airways and alveoli. During high-frequency
oscillatory ventilation (HFOV) the amplitude of the pressure waveform diminishes but
the Paw is sustained, while during high-frequency jet ventilation (HFJV) both the peak
and mean airway pressure are attenuated at the distal airways and alveoli. This gives
59
HFJV a distinct theoretical advantage when considering the pressure cost of
ventilation.
Figure 2-B Pressure amplitudes generated at the ventilator are attenuated during HFV as high velocity gas passes through the rigid endotracheal tube. At slow respiratory rates during conventional ventilation this does not occur (55).
The impact of lung compliance on ventilator performance has been investigated
extensively for CMV and HFOV (170, 171). In vitro studies of HFOV have demonstrated
that if compliance is reduced, oscillation of alveolar pressure is large and coupled with
a concomitant reduction in VT (170). Furthermore, the clinical significance of changes
in compliance on the inspiratory pressures required to maintain normocapnia are best
assessed as the alveolar pressure cost per unit of ventilation. In a poorly compliant
lung, which has a higher corner frequency than a healthy recruited lung, the lowest
pressure cost of ventilation is achieved at higher frequencies.
60
2.1.4 Modes of High-frequency Ventilation
The important differences between HFOV and HFJV are summarised (Table 2-A). These
differences give rise to the potential for physiological advantages of one strategy over
the other: during HFJV the Paw tends to be lower because less of the respiratory cycle
is spent in inspiration; HFJV enhances mucociliary clearance by combining fast
inspirations with relatively slow, passive exhalations (tI:tE ratio may be as low as 1:12);
and the high velocity and small VT breaths do not penetrate injured areas of lung with
high resistance, allowing for maturation and/or healing.
Table 2-A Differences between HFOV and HFJV.
Device tI:tE ratio Inspiratory time
Waveform Exhalation phase
CMV needed
Endotracheal tube adaptor
HFOV 1:3 to 1:1 0.02-0.1 s Squared or
sinusoidal
Active No No
HFJV 1:12 to 1:1.8
0.02-0.034 s Peaked Passive Yes Yes
The passive exhalation phase during HFJV gives this modality a distinct advantage in
certain scenarios. Gas trapping is less likely if the absolute expiratory time is
lengthened: the longer expiration period facilitates more complete passive exhalation.
Preterm infants with increased airway resistance and long time constants, as might
occur with pulmonary interstitial emphysema, need a longer expiration period to
accommodate exhalation (compared to patients with RDS that have shorter time
constant, less compliant lungs). Otherwise they may develop lung over-inflation and
enter a vicious cycle of progressive gas trapping that can’t be broken unless the
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absolute expiratory time is shortened. Time in expiration is not the only factor that
impacts upon gas trapping. If exhalation is active, airway collapse may ensue and gas
trapping is exacerbated. Friedlich et al (2003) presented the results of a cross-over
study from HFOV to HFJV in patients with refractory hypoxaemia. Ten patients crossed
over to HFJV and the survival rate was 90 % (2). They attributed this high survival rate
to their exploitation of the low tI:tE ratio and the passive exhalation phase possible
during HFJV.
2.2 Using a High-frequency Jet Ventilator
High-frequency jet ventilators deliver small tidal volume breaths at a very rapid rate.
Breaths may be as small as 0.5-1 mL kg-1 and can be delivered at a frequency of 4-12 Hz
(240-720 breaths/min). The Life Pulse high-frequency jet ventilator (Bunnell
Incorporated, Salt Lake City, U.S.A.) is a pressure limited and time cycled ventilator
with adjustable peak inspiratory pressure (PIP) and inspiratory time (tI). The jet pulse is
delivered via a special jet tracheal tube adaptor (replacing the normal adaptor) which
has a pressure monitoring port and jet port (Figure 2-C). Inspired gases pass through
the jet port from the jet ventilator and expired gases move out passively to the
breathing system. Opposite the jet port a pressure monitoring port feeds back
information to the ventilator.
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Figure 2-C Jet Port Adaptor (Bunnell Inc., Salt Lake City, Utah, U.S.A.)
The Life Pulse high-frequency jet ventilator displays set parameters and monitored
variables. The central control panel details PIP, respiratory rate, tI and calculates the
tI:tE ratio. The monitored variables are PIP, ∆P, PEEP, servo pressure and Paw (Figure
2-D). ∆P and Paw are calculated values.
Figure 2-D The Life Pulse high-frequency jet ventilator. The central control panel displays the current (now) and to be altered (new) PIP, respiratory rate and tI along
63
with the tI:tE ratio. The uppermost panel displays the monitored variables: PIP, ∆P, PEEP, servo pressure and Paw.
Humidified gas travels to the patient through a patient box. This patient box is a flow
interrupter with a pinch valve which controls the breaths delivered to the patient. The
patient box is located in close proximity to the patient to minimise dampening of the
breaths between the flow interrupter and the endotracheal tube. The specialised jet
adaptor which replaces the endotracheal tube adaptor fits a pressure monitoring line
which provides feedback to the ventilator to control the pressure of subsequent
breaths. It also measures PEEP, providing a monitor of PEEP set on the conventional
ventilator (Figure 2-E).
Figure 2-E Configuration of the jet ventilator and conventional ventilator in the preterm lamb model used in the studies presented in this thesis. Gas from the jet ventilator passes through the patient box and a pinch valve acts as a flow interrupter to create the jet breaths. Gas enters the specialised jet adaptor through the green tube and a pressure monitoring line provides feedback to the jet ventilator. Expired gases pass through the expiratory limb of the CMV circuit.
The parameters to set on the jet ventilator are PIP, respiratory rate and tI. PIP is
discussed under ‘The role of the high-frequency jet ventilator’. The respiratory rate can
64
be set between 240 and 720 breath/min (4-12 Hz). Slower rates are chosen if there is
resistance to expiration (e.g. pulmonary hyperinflation, pulmonary interstitial
emphysema). There are no data regarding the optimal respiratory rate during HFJV and
it will vary according to the disease process. An initial rate of 420 breaths/min is most
often chosen (172). The default tI setting is 0.02 s and works best in most situations.
This very short tI provides very small tidal volumes and keeps alveolar pressures low.
Such a short tI prevents the PIP set on the jet ventilator from being completely
transmitted to the alveoli. If the respiratory rate is decreased, and the tI remains the
same, the time in expiration is increased. Since exhalation is passive this facilitates the
movement of gas and secretions along the airways and out of the lungs.
2.2.1 The Role of the Conventional Ventilator
The jet ventilator is set up in tandem with a conventional mechanical ventilator which
provides the bias flow and PEEP, and channels expired gases away from the patient.
The conventional ventilator may be set to continuous positive airway pressure (CPAP)
mode or a ventilation mode where occasional CMV breaths are delivered. The role of
the conventional ventilator during HFJV in providing PEEP, and therefore controlling
Paw, is a major determinant of oxygenation in patients requiring HFJV. The role of
PEEP and Paw in optimising oxygenation is well documented (173-175).
2.2.1.1 Alveolar Recruitment and Positive End-expiratory Pressure
Positive end-expiratory pressure (PEEP) is the positive airway pressure maintained at
the end of expiration. It is used during mechanical ventilation to recruit alveoli and to
prevent alveolar collapse. PEEP is provided by the conventional mechanical ventilator
65
and the jet ventilator monitors PEEP to alert the clinician to a discrepancy between set
and measured values and the development of auto, or inadvertent, PEEP. Choosing the
most appropriate level of PEEP will depend on the need for alveolar recruitment, the
haemodynamic state of the patient and the pulmonary pathophysiology. Excessive
PEEP will compromise cardiac output as the positive intra-thoracic pressure at the end
of expiration will, to some degree, impact upon the low pressure part of the
circulation, the venous return and the right atrium and ventricle. If venous return is
compromised, cardiac output will fall and this may have implications for the systemic
circulation. An algorithm (Figure 2-F) details the decision making process for altering
PEEP and incorporating CMV breaths. This algorithm has been developed from
experience in a clinical setting, however there is no documented evidence to support
the size, shape, frequency and duration of CMV breaths delivered during HFJV.
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Figure 2-F Algorithm for optimising PEEP when changing from CMV to HFJV with or without CMV breaths (176)
The relationship between PEEP and pulmonary blood flow (PBF) is complex and the
clearance of lung liquid during the first few minutes of ventilation will also impact
upon the haemodynamic effects of PEEP. The massive increase in PBF after birth (8
fold) results from the rapid increase in PaO2 at this time, the release of nitric oxide
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from the pulmonary vascular bed and aeration of the lung. Crossley et al (2007) found
that increasing PEEP (to a critical point) improved oxygenation at the same time as
reducing PBF. As increased oxygenation should promote pulmonary vascular
vasodilation it may be that increasing PEEP led to a gradual decrease in the number of
atelectatic regions and therefore improved oxygenation (177). Blood flow through
those recruited regions may have been reduced due to the compression of the
capillaries. A reduction in PBF is reported with increasing levels of PEEP and it is
postulated it may result from an increase in the alveolar capillary transmural pressure
causing capillary compression (177-179). Likewise, too little PEEP may also
compromise PBF, as extra-alveolar vessels lose support. In the preterm lung, however,
increased airway pressures caused by increasing PEEP have less compressive effects on
capillaries than in the mature lung as preterm lungs are less compliant. Increased lung
compliance following antenatal steroids or surfactant administration at delivery may,
however, increase the sensitivity of pulmonary vessels to changes in airway pressure
(177).
Optimising PEEP is essential during mechanical ventilation. Not only can PEEP prevent
alveolar de-recruitment and interfere with cardiac output, it will affect the pressure
cost of ventilation. Changing PEEP settings over a range of just a few cmH2O can
reduce the required PIP by as much as 50 % (171). These authors concluded that
optimising PEEP is especially important given that ventilator parameters (frequency,
PEEP, PIP and VT) that yield adequate ventilation with safe distension of recruited
alveoli are severely limited if lungs have collapsed because of inadequate PEEP (171).
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The algorithm for finding optimal PEEP during HFJV suggests that this is most easily
achieved when HFJV and CMV breaths are combined (Figure 2-F). The PEEP
optimisation process is based upon findings from HFOV and at the present time is not
substantiated by systematic research using HFJV in a controlled environment. The aim
of the algorithm is to assess the response to removing CMV breaths by changing to
CPAP mode on the conventional ventilator. The assessment variable is SpO2 as it is a
continuous non-invasive measurement but arterial blood gas analyses should also
guide decision making. If the SpO2 is stable and in the target range without CMV
breaths then PEEP is deemed to be adequate to maintain end-expiratory lung volume.
However, if SpO2 decreases after CMV breaths are turned off then the current PEEP
level is assumed to be below the closing pressure. In this scenario, CMV breaths should
be re-introduced and PEEP should be increased to improve alveolar stabilisation.
Optimal lung volume may be defined as having achieved satisfactory SpO2 when FiO2 is
approaching 0.21. To achieve this goal in acute atelectatic lung disease, PEEP may be
substantially higher than traditionally used in conventional ventilation. PEEP should be
decreased if cardiac output is compromised or if oxygenation is adequate and SpO2
doesn’t fall with a decrease in PEEP (176).
If PEEP is too low, alveoli will collapse and atelectasis will develop. If PEEP is too high
there is a risk of alveolar overdistension, impedance of pulmonary perfusion,
cardiovascular depression, poor right heart filling and impedance of venous return
(Figure 2-G).
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Figure 2-G During mechanical ventilation PEEP must be optimised to prevent ventilator induced lung injury and haemodynamic compromise (55).
The importance of optimising PEEP and the impact it has on oxygenation was
demonstrated in a study comparing the arterial blood gas and PBF parameters of
preterm lambs delivered by caesarean section at 126 d gestation. The effect of PEEP
was compared in groups of lambs that had received antenatal steroids, exogenous
surfactant, both or neither and ventilated in volume guarantee mode with CMV. When
VT was constant and PEEP was maintained at 8 cmH2O the effect of PEEP on
oxygenation was larger than the effect of antenatal steroid treatment or exogenous
surfactant administration (177). The improvement in oxygenation was not at the
expense of increased PaCO2 or decreased arterial blood pressure. Surprisingly
antenatal steroid treatment and postnatal surfactant had little impact on the effect of
PEEP on PBF.
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During HFJV, PEEP is set on the conventional ventilator and the jet ventilator displays
the measured value. This information is particularly useful if measured PEEP is higher
than set PEEP. This discrepancy indicates inadvertent intrinsic PEEP and most often
reflects an inappropriately high HFJV rate promoting gas trapping. This information is
also useful if a patient is changed from one ventilator to another. The measured PEEP
can be used to ensure that PEEP is maintained in the changeover (180).
2.2.1.2 Oxygenation and Mean Airway Pressure
Mean airway pressure is a measured value that is displayed on the jet ventilator. It is
determined by the relationship between frequency, tI, PIP and PEEP:
Equation 2-C
time cycle
time) exp x PEEP(time) insp x PIP (HFJVaw
P
where Paw = mean airway pressure, HFJV = high-frequency jet ventilation, PIP = peak
inspiratory pressure, PEEP = positive end-expiratory pressure.
Mean airway pressure is considered the primary determinant of oxygenation and is in
the most part manipulated by changing PEEP (174). During HFJV, Paw is closer to PEEP
than PIP as the tI:tE ratio range can be changed (by altering respiratory frequency) from
1:12 up to 1:1.8. Mean airway pressure is attenuated significantly during HFJV so the
pressure in the alveoli is considerably lower than at the endotracheal tube connector
(157). If Paw is too high, airways and alveoli may over distend and rupture. Conversely,
if Paw is too low airways and alveoli may collapse.
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In the clinical setting, the oxygenation index is calculated to determine the impact of
Paw on oxygenation. The oxygenation index describes the relationship between FiO2,
Paw and PaO2 and a lower value implies better arterial oxygenation (175):
Equation 2-D
OI = 2
2
aO
100 x aw xFiO
P
P
where OI = oxygenation index, FiO2 = fractional inspired oxygen concentration, Paw =
mean airway pressure, PaO2 = partial pressure of oxygen in arterial blood.
Calculation of the OI is a useful bedside tool for comparison of oxygenation between
different ventilation strategies that use different mean airway pressures. It is also used
for prognostication (181).
2.2.2 The Role of the High-frequency Jet Ventilator
The parameters that can be set on the Life Pulse high-frequency jet ventilator are PIP,
respiratory rate and tI. Given that carbon dioxide elimination is proportional to
respiratory rate and VT2 (Equation 2-A) and that VT is determined by lung compliance
and ∆P (PIP-PEEP) it is the high-frequency jet ventilator that provides the most control
over PaCO2 (compared to the conventional ventilator).
2.2.2.1 Carbon dioxide Elimination, HFJV Peak Inspiratory Pressure and ∆P
The PIP during HFJV is altered to achieve a target range for PaCO2 according to serial
arterial blood gas analysis. Increasing PIP will increase ∆P at the same PEEP, and
therefore decrease PaCO2. Conversely, decreasing PIP will decrease ∆P at the same
PEEP and increase PaCO2. Non-invasive methods for assessing CO2 elimination include
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capnography and transcutaneous monitoring but neither technique has a fast enough
response time for the frequency of breaths during HFJV. Transcutaneous monitoring is,
however, more commonly used as a trending tool.
While alterations in PIP will primarily affect PaCO2 they may also affect oxygenation if
Paw changes in parallel with PIP, as is the case if PEEP remains the same. At higher
respiratory frequencies during HFJV, PIP has a greater impact on Paw. Thus to maintain
Paw, and oxygenation, but alter PaCO2, PEEP must be changed at the same time as PIP.
The change in PEEP will be in the opposite direction to the change in PIP to maintain
Paw.
When compared to HFOV in an animal model, the PIP required to achieve comparable
pH and PaCO2 was significantly lower during HFJV (123). This difference is probably
greater than the actual values suggest as the jet ventilator measures PIP at the
proximal airway and this pressure will be significantly attenuated at the level of the
alveolus (55). If lung compliance is poor however, pressure attenuation is less marked
(182). High-frequency oscillatory ventilators do not display PIP or PEEP; rather
management of PaCO2 is by altering ∆P (amplitude). Despite the nomenclature, ∆P will
determine PIP, which will be higher during HFOV.
The difference between PIP (set and measured on the jet ventilator) and PEEP
(determined by the conventional ventilator settings) is equivalent to the amplitude of
the airway pressure waveform. ∆P is therefore a calculated value displayed on the
ventilator and will change as PIP and PEEP are altered. This will determine the VT of
each breath and control PaCO2. During HFJV the VT is as low as 0.1-1.0 mL kg-1 but will
change according to lung and chest wall compliance (183). Regardless of compliance
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the VT during HFJV remains considerably smaller than anatomical and equipment dead
space. The high velocity, central inspiratory flow spike generated during HFJV will
penetrate the dead space and ventilate alveoli (183). Concurrent measurement of VT
will provide information on compliance. For a given ∆P increasing VT indicates relatively
high compliance and decreasing VT indicates relatively poor compliance.
2.2.3 Monitoring during High-frequency Jet Ventilation
The Life Pulse high-frequency jet ventilator monitors airway pressures to provide
continuous feedback information to drive PIP. It also measures servo pressure and
PEEP. The former is the pressure generated within the ventilator that is required to
meet the set PIP and the latter monitors PEEP set on the conventional ventilator.
2.2.3.1 Servo Pressure
The concept of servo pressure is important during HFJV. It refers to the automatically
generated driving pressure that the ventilator itself creates to deliver a breath that
meets the set PIP. Servo pressure will change as lung volume and compliance change
and a decrease in servo pressure may occur as a result of worsening compliance,
endotracheal tube obstruction, the need for airway suctioning, tension pneumothorax,
right maintstem bronchial intubation or a deliberate decrease in ∆P. Servo pressure
will increase with any increase in ∆P, or as compliance improves, the circuit
disconnects or there is an airleak that does not put the lung under tension (184).
Monitoring servo pressure will provide information on lung compliance and contribute
to an understanding of the pathophysiology of lung disease and the progression of this
disease during mechanical ventilation. Servo pressure is measured in pounds per
74
square inch (psi) by the Life Pulse high-frequency jet ventilator and will change from
breath to breath giving an early indication of changes within the lungs.
2.2.3.2 Positive End-Expiratory Pressure
The difference between PEEP set on the conventional ventilator and PEEP measured by
the jet ventilator should be negligible. If measured PEEP is higher than set PEEP it is an
indication of auto PEEP. This term refers to the development of end-expiratory
pressure within the patient, most likely due to gas trapping, creating resistance to
expiration. In this instance the VT can’t escape the lungs, servo pressure decreases and
PaCO2 increases. Decreasing the respiratory rate will help minimise the effects of this
inadvertent PEEP. Remaining at a lower respiratory rate is indicated until the set and
monitored PEEP correlate again. If CMV breaths are being delivered at this time,
decrease the frequency of these first, then the frequency of jet breaths.
2.3 High-Frequency Jet Ventilation in the Clinical Environment
Homogeneous restrictive lung disease caused by uniform restriction from extra
pulmonary pathology is difficult to manage with CMV. As a result HFV strategies have
been employed but there are no randomised controlled trials to support the use of
one strategy over another (157). While there are various reports of the use of HFJV in
infants, children and adults, the decision making process during patient management
with HFJV is not evidence based.
High-frequency jet ventilation is theoretically superior for the management of patients
with gas-trapping as the passive exhalation phase and relatively short tI:tE ratio
75
facilitates the passage of gas along the periphery of the airways for exhalation. Early
studies report more rapid and more frequent resolution of pulmonary interstitial
emphysema (PIE) with no difference in the incidence of adverse side effects (157). A
rabbit study found that gas trapping was significantly greater during HFJV when
compared to HFOV and attributed the difference to the passive exhalation phase and
relatively short tI:tE ratio (185). This finding contradicts the clinical experience and
highlights the need to compare these ventilation strategies in a controlled setting.
In the 1980s, HFJV was employed in situations where other ventilation modalities had
failed. Early studies document the safety and effectiveness of HFJV in rescue situations
(100, 142, 186, 187). The outcomes of these studies reflected the severity of the
condition of the patients at the time of changeover to HFJV. More recently, however,
attention is shifting to the earlier use of HFJV in infants with RDS that is not yet
complicated by airleak (138).
To date there are 4 randomized controlled trials of HFJV (100, 138, 139, 188). Differing
entry criteria, treatment strategies and the definition of primary outcomes have
complicated interpretation of the results of these trials. The main features of each trial
are summarised in Table 2-B and are evaluated below with respect to their potential
benefits and adverse consequences as both early and rescue therapies in preterm
babies.
2.3.1 High-frequency Jet Ventilation as a Rescue Therapy
A Cochrane review published in 2006 eliminated all but one publication from their
examination of the use of HFJV as a rescue ventilator strategy (141). The study
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reviewed infants who had developed PIE within the first 7 days of life while receiving
CMV (100). Cross-over to HFJV occurred if the patient deteriorated. The results
demonstrated no statistically significant difference in the overall mortality between
the HFJV and CMV groups, no difference in the incidence of BPD in survivors, of new
air leaks after change-over, of total intraventricular haemorrhage, of necrotising
tracheobronchitis at autopsy or of airway obstruction (100). This study only included
144 infants and surfactant administration was not standard practice. Furthermore, the
long term neurological outcomes were not examined. These limitations made it
difficult to conclude that HFJV was a useful rescue therapy in preterm infants.
2.3.2 HFJV used Early in the Management of Respiratory Distress Syndrome
Despite the limited data generated from randomised controlled trials examining the
use of HFJV as a rescue therapy, neonatologists considered HFJV a worthy alternative
therapy when others had failed (100, 142, 186, 187). This acknowledgement led to
investigation of HFJV early in the time-course of patient management (138, 139, 188).
There are only 3 small randomised controlled trials of early HFJV. Carlo et al (1990)
studied 45 infants and found no difference in the incidence of adverse outcomes when
HFJV was compared to CMV in preterm infants. This study used a non-commercially
available high-frequency jet ventilator and none of the infants received surfactant.
Wiswell et al (1996) concluded that the incidence of adverse neurological outcomes
was higher in infants managed with HFJV compared to those managed with CMV (139).
A major factor in this study was the incidence of hypocapnia in the HFJV group of
infants. Despite comparable arterial blood gas values prior to randomisation these
infants had a significantly lower PaCO2 during the study. This complicates
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interpretation of the negative finding for HFJV in this study and is characteristic of the
era of hyperventilation with HFJV. The use of higher PEEP has since decreased the
incidence of hypocapnia during HFJV as higher PEEP allows for a smaller ∆P (at the
same PIP). Lastly, Keszler et al (1997) compared HFJV with CMV in preterm infants.
Despite guidelines for both protocols, during analysis they subdivided the HFJV group
into a low pressure (lung volume) strategy and an optimal volume strategy as some
centres violated the trial protocol. Concern stemming from Wiswell’s earlier study
inadvertently created a unique opportunity to compare 2 different HFJV strategies.
The optimal volume strategy was designed to provide alveolar recruitment, optimise
lung volume and improve V/Q matching, while minimising FiO2 and ∆P. This strategy
also included the delivery of 2 to 5 CMV breaths/min and maintenance of adequate
Paw with relatively high PEEP. They showed that the optimal volume strategy
improved oxygenation, decreased exposure to hypocapnia and reduced the incidence
of high grade IVH and/or PVL (138). This study is the only one in which surfactant was
administered to each infant before entry. They concluded that HFJV reduced the
incidence of BPD at 36 w and the need for home oxygen in preterm infants with
uncomplicated RDS. Further, they surmised that there was no increase in adverse
outcomes compared to CMV. While the results of this study created promise for HFJV
as an early management tool for preterm babies, larger studies have not been
performed.
Table 2-B Summary of randomised controlled trials of high-frequency jet ventilation in preterm babies.
Study n Description Eligibility Surfactant Antenatal Steroids
Ventilators Outcome Measures
Carlo et al 1990
45 Single centre, crossover if failing the assigned therapy, mean randomisation age 14 h (CMV) vs. 15.5 h (HFJV)
Preterm, < 24 h of age, 1000-2000 g, RDS
0 % NR Locally developed, non commercially available jet ventilator with a time-cycled, pressure-limited infant ventilator (Bear Cub) for CMV
Mortality at 28 d, CLD at 28 d, ALS, progression of IVH, success after crossover, days on mechanical ventilation, days on supplemental oxygen, mechanical ventilation at 28 d
Keszler et al 1991
144 (CMV n=70, HFJV n-74)
Multicentre (15), crossover if failing the assigned therapy, mean randomisation age 29.3 w post conceptional age
Preterm, < 7 d of age, < 750 g at birth, PIE
0 % NR Life Pulse (Bunnell) with standard time-cycled, pressure-limited infant ventilators for CMV
Mortality at 28-30 d, success in the original assignment, CLD at 28-30 d, IVH, new airleak, necrotising tracheobronchitis, airway obstruction, CLD in survivors
Wiswell et al 1996
73 (CMV n=36, HFJV n=37)
Single centre, crossover if failing the assigned therapy, mean randomisation age 7.1 h (CMV) vs. 7.3 h (HFJV)
Preterm (< 33 w), 500-2000 g at birth, < 24 h of age at randomisation, chest roentgenographic findings consistent with RDS, requirement for ventilator support with FiO2 > 0.3 and PIP > 16 cmH2O
CMV 97 % HFJV 92 %
CMV 19 % HFJV 22 %
Life Pulse (Bunnell) with a time-cycled, pressure-limited infant ventilator (Bear Cub) for CMV
IVH, periventricular echodensities, cystic PVL, supplemental oxygen at 28 d and 36 w, mortality at 28 d and 36 w, days on mechanical ventilation, days in hospital
Keszler 130 Multicentre (8), Preterm (<36 w), 700- 100 % NR Life Pulse BPD at 28 d and 36 w post-
CMV = conventional mechanical ventilation, HFJV = high-frequency jet ventilation, RDS = respiratory distress syndrome, NR = not recorded, CLD = chronic lung disease, ALS
= airleaks, IVH = intraventricular haemorrhage, PIE = pulmonary interstitial emphysema, BPD = bronchopulmonary dysplasia, PVL = periventricular leukomalacia, PDA =
patent ductus arteriosus, NEC = necrotising enterocolitis, ROP = retinopathy of prematurity.
et al 1997
(CMV n=65, HFJV n=65)
crossover if failing the assigned therapy, mean randomisation age 8.3 h (CMV) vs. 8.1 h (HFJV) HFJV groups subdivided (following analysis) to HF-OPT (optimal volume strategy) and HF-LO (low pressure strategy)
1500 g, requirement for mechanical ventilation with FiO2 > 0.3 at 2-12 h after surfactant administration, received surfactant by 8 h of age, < 20 h of age, and mechanically ventilated for < 12 h
(before entry to study)
(Bunnell) with standard time-cycled, pressure-limited infant ventilators for CMV
conceptional age, survival, gas exchange, airway pressures, airleak, IVH, PVL, pulmonary haemorrhage, PDA, NEC, ROP
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2.3.3 Clinical Strategies
The ventilation strategy employed by the clinician has important physiological
implications for infants treated with HFJV. Optimising lung volume has been
recognised as an important goal during HFJV (138) but as this requires the use of a
relatively high PEEP it may cause a decrease in cardiac output. This decrease in cardiac
output may be less profound in a preterm infant’s non-compliant lung but is
nevertheless a potential adverse side effect. The initial settings chosen for HFJV are
PIP, respiratory frequency, tI, PEEP (on the conventional ventilator). The latter
parameters will determine the tI:tE ratio. Furthermore, a decision must also be made
about whether or not CMV breaths should be delivered.
2.3.3.1 PIP
The optimal PIP during HFJV is determined by PaCO2. The initial setting however has
been made in accordance with the CMV PIP prior to cross-over. Keszler et al (1997)
started the HFJV PIP at the same level as CMV PIP (138) and found that it needed to be
decreased rapidly thereafter to avoid hypocapnia. This comparison to CMV PIP may be
valid, but it is likely that a higher PIP can be used during HFJV for less haemodynamic
expense given that the HFJV airway pressure waveform is attenuated (55). While
changes during patient management are more intuitive (based upon PaCO2) the
starting point is arguably more difficult to determine and there is little evidence in the
literature to support a particular decision making process.
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2.3.3.2 tI:tE ratio
The respiratory frequency and tI determine the tI:tE ratio during HFJV. As the
exhalation phase is passive during HFJV, the potential for gas trapping is decreased at
lower respiratory frequencies. The initial respiratory frequency and tI is rarely
documented in the literature. Wiswell et al (1996) commenced at 420 breaths/min and
0.02 s respectively, giving an tI:tE ratio of 1 : 6.1. Changes to these settings are made if
there is evidence of gas-trapping, in which case respiratory frequency is decreased,
increasing the tI:tE ratio (up to a maximum of 1:12). The advantages of the passive
exhalation phase are greater when respiratory frequency is decreased.
2.3.3.3 PEEP
The initial PEEP has been set at 6-8 cmH2O (138) and 4-5 cmH2O (139) but the key to
maintaining oxygenation is maintaining Paw. PEEP is therefore altered to optimise
oxygenation but the maximum PEEP tolerated has not been investigated. Paranoia
about the expense of high PEEP during CMV has been transferred to HFJV. The use of
low PEEP during HFJV necessitated delivery of CMV breaths and high jet PIP to achieve
adequate Paw, alveolar recruitment and satisfactory oxygenation. A direct
consequence of this approach was the generation of large ∆P, which promoted
hypocapnia. As the attenuation of the airway pressure waveform during HFJV is better
understood, higher PEEPs are being used, but once again, there is little evidence in the
literature to support or refute a suitable PEEP range during HFJV.
Transitioning from HFOV to HFJV may be indicated in some patients, especially in the
instance of airleak or PIE. To maintain open alveoli when changing from one modality
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to the other, maintenance of Paw is essential. Bass et al (2007) investigated the
accuracy of the Life Pulse high-frequency jet ventilator for measuring Paw during HFOV
and then determined a calculation for the PEEP setting required to maintain Paw (180).
In their ex vivo study they found the Life Pulse to be an accurate monitor of airway
pressure during HFOV and predicted PEEP was related to actual PEEP by Equation 2-E:
Equation 2-E
Actual PEEP = (Predicted PEEP x 1.12) – 2.38
The greatest difference was 2 cmH2O (180). This work makes the transition from HFOV
to HFJV less likely to derecruit alveoli, but the same comparison does not exist for
CMV.
2.3.3.4 CMV Breaths during HFJV
The delivery of CMV breaths during HFJV is used to recruit alveoli but the optimal
frequency and characteristics of these breaths is not known. The literature reports
CMV breath rates delivered 2-5 times/min with an tI of 0.5-0.8 s (138) or 5-10
times/min (139). The impact of CMV breath PIP, tI and frequency on oxygenation, CO2
removal, VILI and cardiac output is unknown.
2.4 Summary
The development of HFJV as a clinical tool has been somewhat haphazard. This has led
to a period of HFJV characterised by hypocapnia (most likely due to PEEP paranoia) and
uncertainty regarding the incidence of adverse outcomes (especially neurological). The
recognition of its usefulness and the importance of an open lung ventilation strategy
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have certainly contributed to more recent success with HFJV and it is widely used in
NICUs in the U.S.A. The full potential of HFJV is yet to be realised and with a better
understanding of the impact of different pressures and HFJV and CMV combinations
the management of preterm infants will improve.
The mechanisms of gas transport during HFJV are unique and to exploit this
therapeutic tool to its full potential a thorough understanding of these mechanisms
and the consequences of particular strategies in different clinical scenarios is
necessary. Despite extensive use, and in spite of limited randomised controlled trials,
the intricacies of gas exchange, airway pressure changes along the bronchial tree are
documented, but their impact on lung tissue and the cardiovascular system are for the
most part unknown. This thesis explores HFJV in an experimental setting with
emphasis on alveolar recruitment manoeuvres and their impact on blood flow and
lung tissue.
82
83
3 General Methodology
This chapter provides a detailed description of the materials and methodology relevant
to the studies described in this thesis. Information specific to a particular protocol is
presented within the relevant chapter. All animal procedures were approved by the
University of Western Australia animal ethics committee, according to the guidelines
of the National Health and Medical Research Council of Australia code of practice for
the care and use of animals for scientific purposes.
3.1 Animal Breeding and Welfare
Merino ewes between 5 and 6 years of age were mated over a 24 hour period. Oestrus
and ovulation was synchronised by the insertion of intra-vaginal sponges for the 14
days prior to mating. Pregnancy was confirmed 55-85 days later by transabdominal
ultrasound examination.
Date mated pregnant ewes were transported from the farm of origin in Darkan,
Western Australia at no later than 100 d gestation to either the Large Animal Facility at
the University of Western Australia or the Shenton Park Biomedical Research Facility.
During the week prior to transport, ewes were inspected by a veterinary surgeon to
ensure there was no evidence of disease or injury and that their body condition score
was adequate (minimum 2.5/5). At the Large Animal Facility, ewes were housed in
either single raised pens adjacent to one another or a raised communal pen (4 m x 4.7
m maximum capacity 16 sheep). They were fed a ration of chaff, lupins and pellets
84
with free access to water through a dripper or in a bucket. At the Shenton Park
Biomedical Research Facility, ewes were run in a paddock and supplementary feed was
supplied if necessary.
3.1.1 Nutrition
3.1.1.1 Instrumented Lambs
For studies involving surgical instrumentation of the fetus prior to delivery, food was
withheld from the ewes 24 hours prior to surgery to minimise the potential for
regurgitation and aspiration of rumen contents, to decrease the incidence of bloat and
to reduce compression of the caudal vena cava when positioned in dorsal recumbency.
All ewes had free access to water up until the induction of anaesthesia.
3.1.1.2 Non-instrumented Lambs
Ewes bearing lambs that were not instrumented in utero had food withheld overnight
prior to non-recovery surgical delivery. All ewes had free access to water up until the
induction of anaesthesia.
3.1.2 General Anaesthesia and Instrumentation
3.1.2.1 Lambs Instrumented in utero
At 128-130 days of gestation, anaesthesia was induced with an intramuscular injection
of xylazine (0.2 mg kg-1; Xylazil 20 mg mL-1, Troy Laboratories, Australia) and ketamine
(15 mg kg-1; Ketamil 100 mg mL-1, Troy Laboratories, Australia). A cuffed oral
endotracheal tube (7.5 mm internal diameter, Portex Ltd, England) was positioned and
secured to facilitate maintenance of anaesthesia with isoflurane in 100 % oxygen
85
delivered through a circle breathing system. Throughout surgery the ewe was
ventilated with a volume cycled bag-in-the-bottle mechanical ventilator to maintain
normocapnia (Ohio 7000, Ohio Medical Products, Division of Airco Inc. Madison,
Wisconsin). A side stream capnograph enabled breath to breath non-invasive
assessment of end expiratory CO2 concentration.
Studies involving pulmonary blood flow and pulmonary arterial blood pressure
measurements required instrumentation of the fetus prior to caesarean delivery. The
anaesthetised ewe was positioned in dorsal recumbency and the abdomen was clipped
and prepared for surgery. The uterus was exposed through a ventral midline incision
and the fetal head was located and exposed via hysterotomy at a poorly vascularised
site of the uterine wall. Two incisions were made for twin pregnancies.
The fetus was partially exteriorised to facilitate access to the lateral thorax. An incision
was made at the fourth intercostal space on the left side. Exposure of the heart was
achieved by blunt dissection through the intercostal muscles and pleura and incision of
the pericardium. The left pulmonary artery was isolated by careful blunt dissection and
an ultrasonic flow probe (4R, Transonic Systems, Ithaca, NY) was positioned around it,
upstream of the ductus arteriosus (Figure 3-A). A tapered polyvinyl intravenous
catheter was inserted through the main trunk of the pulmonary artery by direct
puncture and secured so the tip was located in the left main pulmonary artery. A
suture in the wall of the artery secured the catheter in place. The thoracotomy incision
was closed using silk suture material in a simple continuous pattern.
86
Figure 3-A Instrumentation of the fetus: Pulmonary artery flow probe and pulmonary artery catheter in situ
3.1.2.2 Non-instrumented Lambs
An intravenous injection of medetomidine (0.02 mg kg-1; Domitor 1 mg mL-1, Pfizer
Animal Health, U.S.A.) and ketamine (10 mg kg-1; Ketamil 100 mg mL-1, Troy
Laboratories, Australia) was administered to induce anaesthesia in pregnant ewes at
128-130 days of gestation. A subarachnoid (spinal) injection of lidocaine (3 mL;
Lignocaine 20 mg mL-1, Troy Laboratories, Australia) was administered with access
through the intervertebral space between the 4th and 5th or 5th and 6th lumbar
vertebrae. A midline laparotomy incision was made, the position of the lamb was
determined by palpation, and the uterus was incised to facilitate exteriorisation of the
entire fetus.
87
3.1.3 Caesarean Delivery
Immediately prior to delivery, a cuffed oral endotracheal tube (4.5 mm internal
diameter, Portex Ltd, England) was positioned under direct vision and secured with an
umbilical tape tie around the back of the lamb’s ears. Lung fluid was suctioned with
gentle negative pressure and 100 mg kg-1 surfactant was administered [either
beractant; (Survanta 25 mg of phospholipids mL-1, Abbott Laboratories, U.S.A.) or
poractant; (Curosurf 80 mg of phospholipids mL-1, Chiesi Pharmaceuticals Ltd, Parma,
Italy)]. The umbilical vessels were clamped and the lamb was delivered, weighed and
commenced immediately on the assigned ventilation protocol. A cord blood sample
was collected from the umbilical artery as the lamb was delivered and analysed
immediately for baseline blood gas status.
After commencement of ventilation, catheters were inserted into an umbilical artery
and an umbilical vein. The arterial catheter was advanced to approximately 15 cm and
the venous catheter to 9 cm. Serial arterial blood gas samples were collected from the
umbilical artery throughout the 2 or 3 hour ventilation period. Anaesthetic and
analgesic drugs were administered through the umbilical venous catheter to ensure
the lamb was unconscious and nonresponsive to the procedures. Propofol (Repose; 10
% Norbrook Laboratories Ltd., Victoria, Australia) and remifentanil (Ultiva; 1 mg vial
requiring reconstitution, Abbott Laboratories, U.S.A.) were infused at 0.1 mg kg-1min-1
and 0.05 µg kg-1min-1 respectively, in the first instance, and the rate adjusted according
to clinical effect.
The ewe was euthanased immediately after delivery of the lamb(s) by an intravenous
injection of pentobaritone (100 mg kg-1) (Valabarb 325 mg mL-1 Jurox, Australia) into
88
the uterine vein or superficial abdominal vein. Death was confirmed by absence of a
heart beat during thoracic auscultation.
3.2 Ventilator Set-up
The high-frequency jet ventilator (Life Pulse High Frequency Ventilator, Bunnell Inc.,
Salt Lake City, U.S.A.) was used in tandem with a pressure limited, time cycled infant
ventilator with a humidifier (MR850 Humidifier, Fisher and Paykel Healthcare,
Auckland, N.Z.). Studies performed in 2007 used a Bourns ventilator (Bourns Life
Systems BP 200 Infant Pressure Ventilator, California, U.S.A.) for CMV. In subsequent
years CMV was delivered via a Drager Babylog (Babylog 8000+, Drägerwerk, Lubeck,
Germany). A LifePort™ Adaptor (Bunnel Inc, Utah, USA) replaced the usual tracheal
tube adaptor (Figure 3-B) to facilitate injection of inspired gas in high velocity spurts
and monitoring of pressure for estimation of the PIP and mean airway pressure (Paw)
at the end of the tracheal tube.
89
Figure 3-B Ventilator Set Up: Life Pulse High-frequency Jet Ventilator with Conventional Mechanical Ventilator. The High-frequency Jet Ventilator (HFJV) is used in tandem with a Conventional Mechanical Ventilator (CMV). Humidified inspiratory gases pass from the HFJV to the Patient box where a pinch valve interrupts the inspiratory gas to create the jet breaths. This is connected to the LifePort adaptor which in turn connects to a standard tracheal tube. At the LifePort Adaptor, the CMV circuit connects and humidified inspiratory gases pass along the inspiratory limb and expired gases pass along the expiratory limb.
3.3 Data Collection
Continuous information regarding the ventilator frequency, PIP, amplitude of the
pressure waveform (ΔP), positive end expiratory pressure (PEEP), servo pressure, Paw,
rectal temperature, pulse rate, oxyhaemoglobin saturation and fractional inspired
oxygen concentration (FiO2) was available. If the lamb was instrumented, pulmonary
blood flow, pulmonary arterial blood pressure and systemic arterial blood pressure
90
were also monitored continuously. Arterial blood samples were collected and data was
recorded at predetermined intervals. Any unexpected or unusual event was noted.
Arterial blood gas samples were collected as planned and analysed immediately to
provide information for decision making. HFJV PIP was adjusted to achieve permissive
hypercapnia (PaCO2 45-55 mmHg): HFJV PIP was increased (to a maximum of 40
cmH2O) if PaCO2 was above and decreased if PaCO2 was below this range. The initial
fractional inspired oxygen concentration (FiO2) was always 0.4 and adjusted according
to the oxyhaemoglobin saturation (SpO2). FiO2 was increased in increments of 0.1-0.2
(to a maximum of 1.0) if SpO2 dropped below 89 %.
3.3.1 Pulmonary Arterial Blood Pressure and Blood Flow Measurements
Left pulmonary arterial blood pressure was measured and recorded in real time using a
digital data acquisition system (Powerlab 8SP, AD Instruments, N.S.W., Australia). The
catheter was connected to a pressure transducer (Maxxim Medical, Tx, U.S.A.) and the
signal was amplified before it was recorded. The sampling frequency was 1 KHz. Mean
pressure values were calculated from the relevant pressure signals.
Left pulmonary arterial blood flow was also recorded in real time using a volume flow
meter (Transonic Systems T108, Neomedix Pty. Ltd., N.S.W., Australia). The transonic
flowmeter incorporates wide beam illumination whereby two transducers pass
ultrasonic signals back and forth, alternately intersecting the flowing blood in
upstream and downstream directions. The flowmeter derives an accurate measure of
the ‘transit time’ it takes for a sound wave to travel from one transducer to the other.
The difference between the upstream and downstream integrated transit times is a
91
measure of blood volume flow and is independent of vessel diameter (Figure 3-C). The
transducers and flowmeter were calibrated every day before recordings began. Zero
and 100 mmHg was calibrated on the transducers and 0 and 400 mL min-1 on the
flowmeter.
Figure 3-C Widebeam Illumination: Schematic view of the perivascular Transonic ultrasonic volume flow-sensor. Using wide beam illumination, two transducers pass ultrasonic signals back and forth, alternately intersecting the flowing liquid in upstream and downstream directions. The flowmeter derives an accurate measure of the ‘transit time’ it takes for the wave of ultrasound to travel from one transducer to the other. The difference between the upstream and downstream integrated transit times is a measure of volume flow rather than velocity (189)
Pulmonary waveform data were recorded continuously throughout the ventilation
period (190). Measurements of mean PBF and pulse-by-pulse minimum values at the
end of diastole and systole were computed from the PBF waveform over 5 consecutive
waveforms at each time point. Mean systolic and diastolic PBF was also calculated at
each time point by including 5 consecutive waveforms. Pulsatility Index, a measure of
downstream resistance to blood flow, was calculated:
Equation 3-A
)cycles econsecutiv 5 over flow systolic peak mean
diastole after flow minimumflow systolic Peak(yIndexPulsatilit
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All data were analysed using a computer software package (Chart v4.2 Powerlab,
ADInstruments, N.S.W., Australia).
3.4 Euthanasia and Post Mortem
The ewes were euthanased immediately following delivery. Lambs were euthanased at
delivery (unventilated controls) or the end of the ventilation protocol. Pentobarbitone
(100 mg kg-1; Valabarb 325 mg mL-1, Jurox, Australia) was administered by intravenous
injection into either the uterine vein (ewe) or umbilical vein (lamb) and death was
confirmed when thoracic auscultation of the heart was negative. Prior to euthanasia
the lungs were ventilated with 100% oxygen for 3 minutes, to promote alveolar
collapse. A third of the dose of pentobarbitone was delivered, the chest cavity was
evacuated, the tracheal tube clamped, and the remainder of the dose of
pentobarbitone delivered to euthanase the lamb.
The thoracic cavity of each lamb was opened. The trachea was isolated and a short
tracheal tube was inserted and secured with the tip positioned 3 cm above the carina.
The gas volume of the lung was measured as the volume used to inflate the lung to 40
cmH20 pressure and maintain it at that pressure for 30 s. The lung was then deflated
sequentially, and the residual volume recorded at 40, 20, 15, 10, 5 and 0 cmH2O
pressure to yield a deflation pressure-volume curve.
Bronchoalveolar lavage (BAL) was performed on the left lung three times with saline
and BAL fluid was snap frozen in liquid nitrogen. Cytospin samples were collected and
prepared immediately for cytology and protein analysis by the Lowry method (191).
93
The right upper lung lobe was fixed to 30 cmH2O in formalin for 24 hours prior to
preparation of tissue for immunohistochemistry. The tissue was washed in 0.1 molar
phosphate buffered saline (3 x 10 minutes), 30% ethanol (1 x 10 minutes), 50% ethanol
(1 x 10 minutes) and 70% ethanol (1 x 10 minutes). Lung sections were processed in
alcohol and xylene overnight then embedded in paraffin wax for cutting 5 µm slices for
histopathology.
Samples of the right lower lobe were collected and snap frozen in liquid nitrogen, then
stored at -80°C for later quantification of mRNA using Real-Time Polymerase Chain
Reaction (RT-PCR).
3.4.1 Cell Population of Bronchoalveolar Lavage Fluid
Cytospin samples of BAL fluid were prepared immediately after collection of BAL fluid.
10 mL of fluid was centrifuged for 10 minutes at 4°C, the supernatant removed to
leave a pellet of cells. This was resuspended and diluted with trypan blue for total cell
counts. Differential cell counts were performed at a later date following Diff-Quik
staining to allow identification and differentiation of inflammatory cells, epithelial cells
and other cells (including junk cells and mucoid cells). Total cell count per milliliter of
BAL fluid enabled calculation of total cell count per lamb based upon the formula:
Equation 3-B
Cell count per kg bodyweight = AWc) x BW)(AWv x (LL
RLLL
where LL = left lung weight (g), RL = right lung weight (g), BW = body weight (g), AWv =
alveolar wash volume (mL), AWc = alveolar wash cell count mL-1 BAL fluid.
94
At 40x magnification 200 cells were counted to identify inflammatory, epithelial and
other cells. Further differentiation of inflammatory cells to mononuclear cells
(including macrophages), neutrophils, lymphocytes, eosinophils and basophils was also
performed. The division of cell numbers within the 200 cell sample was used to
extrapolate to the total cell population.
3.4.2 Bronchoalveolar Lavage Protein Assay
Bronchoalveolar lavage fluid was stored at -80°C until the protein assay was
performed. A standard curve (0-1000 µg mL-1) was created for each set of samples.
Each sample was prepared according to the Lowry method and measured in triplicate
with the Versa Max Microplate Reader Spectrophotometer. Three milliliters of Reagent
1 (Na2CO3 with K+ tartrate, CuSO4 and water) was added to each sample and left to
stand for 10 minutes. Two hundred microlitres of Reagent 2 (Follin’s Reagent and
water) was then added and mixed immediately. After a 30 minute incubation period
(at room temperature) the samples were analysed with the spectrophotometer.
3.4.3 Immunohistochemistry
Immunohistochemical stains were performed for myeloperoxidase (MPO) and
inducible nitric oxide synthetase (iNOS). The dilution of the primary antibody was
1:500 for MPO and 1:250 for iNOS. Biotinylated secondary antibody was diluted 1:200
for each. Anti-rabbit secondary antibody was used for the MPO stain and anti-mouse
for the iNOS stain. A control slide provided by the manufacturer was used for the MPO
staining. Antigen retrieval was performed in NaCitrate buffer (pH 8.45) at 80°C for 30
minutes then room temperature for 60 minutes.
95
Three slices of lung were collected from each lamb, processed overnight and
embedded in paraffin wax. Five micron sections were cut from each block. Each animal
therefore had 3 areas of lung tissue for examination. From each slide 10 fields were
examined giving 30 fields (40x magnification) for each lamb. Each field was
photographed using SPOT insight 4MP, 2048 x 2048 colour mosaic camera with infra
red filter, 14 bit, 20 MHz, C Mount, Firewire, SPOT software through the C Mount
Adaptor 1.2x lens for the Olympus BX Microscope. The number of MPO or iNOS
positive cells were counted in each of the 10 fields/slide and the total cellular area of
those fields was quantified using densitometry software (Image-Pro Plus v4). An
average for each slide was calculated, and then averages for each animal given 3 slides
were examined. The results are expressed as number of positive cells per cellular area
(nm2).
3.4.4 Qualitative Polymerase Chain Reaction
Total RNA was isolated from 30 mg of homogenised lung tissue using the RNeasy Mini
kit (Qiagen, U.S.A.) according to the manufacturer’s instructions. The contaminating
genomic DNA was removed by an on-column DNaseI digestion performed using the
DNaseI digestion kit (Qiagen, U.S.A.). One microgram of RNA was then reverse
transcribed into complementary DNA in a 20 µL reaction with QuantiTect® Reverse
Transcription Kit (Qiagen, U.S.A.). The primers used for amplifying IL-1β and IL-6 (192)
and IL-8, EGR1 and CTGF (111) have been described elsewhere. Amplification and
detection of specific products were conducted on the Rotor-gene 3000 real time PCR
system (Corbett Life Science) with the published cycle profiles using Rotor-gene SYBR
Green PCR Kit (Qiagen, U.S.A.) following the manufacturer’s instructions. The
96
expression levels of genes of interest were normalised into 18S RNA (193) using the 2-
∆∆CT method (194) and presented as expression ratio relative to the unventilated
control group.
3.4.5 Myeloperoxidase Activity in Lung Tissue
Myeloperoxidase (MPO) activity was measured spectrophotometrically using methods
described by McCabe et al in 2001 and Faith et al in 2008 (195, 196). Minor
modifications were made: lung tissue samples were homogenised in 50 nmol L-1
potassium-phosphate buffer (pH 6.0), containing 5 mg mL-1
hexadecyltrimethylammonium bromide. The samples were subjected to 3 cycles of
freeze-thaw, followed by sonication. The suspensions were then centrifuged at 10 000
rpm for 10 minutes. Ten microlitres of supernatant was mixed with 290 µL of 50 mmol
L-1 phosphate buffer (pH 6.0), containing 0.167 mg o-dianisidine dihydrochloride and
0.0005 % hydrogen peroxide in a standard 96 well microtiter plate. The changes in
absorbance at 30 second intervals were recorded at 450 nm. The MPO activity was
then normalised to the total protein content of the tested samples. Activity was
expressed as units of MPO activity per mg of protein, where one unit of MPO was
defined as the amount needed to degrade 1 µmol of hydrogen peroxide per minute at
room temperature.
3.5 Statistical Analyses
Parametric data were analysed with a Student’s t test (for comparisons against one
other group) and one way analysis of variance (ANOVA) for multiple group
comparisons. Non-parametric data were compared with a Rank Sum test for
97
comparisons between two groups or ANOVA on Ranks for multiple comparisons. A
difference was considered significant if p < 0.05. Statistical analyses were performed
using SigmaStat (Version 3.5, Systat Software Incorporated, U.S.A.) and values in the
text are expressed as the mean ± the standard deviation (SD), standard error of the
mean (SEM), if multiple values were collected for each data point or Median (25, 75
centile).
98
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4 High Positive End‐Expiratory Pressure during High Frequency
Jet Ventilation Improves Oxygenation and Ventilation in Preterm
Lambs.
Gabrielle C Musk1, Graeme R Polglase1, J Bert Bunnell2, Carryn J McLean1, Ilias Nitsos1,
Yong Song1 and J Jane Pillow1.
1 School of Women’s and Infants’ Health, University of Western Australia, Perth, Western
Australia, 6009, Australia.
2 Bunnell Inc, Salt Lake City, Utah, USA and Department of Bioengineering, University of
Utah, Salt Lake City, Utah, USA.
This first study investigated the role of positive end‐expiratory pressure for alveolar
recruitment during high‐frequency jet ventilation.
122
Abstract
Increasing positive end‐expiratory pressure (PEEP) is advocated to recruit alveoli during
high‐frequency jet ventilation (HFJV) but its effect on cardiopulmonary physiology and
lung injury is poorly documented. We hypothesised that high PEEP would recruit alveoli
and reduce lung injury but compromise pulmonary blood flow (PBF). Preterm lambs of
anaesthetised ewes were instrumented, intubated and delivered by cesarean section after
instillation of surfactant. HFJV was commenced with a positive end‐expiratory pressure
(PEEP) of 5 cmH2O. Lambs were allocated randomly at delivery to remain on constant
PEEP (PEEPconst, n=6) or to recruitment via stepwise adjustments in PEEP (PEEPadj, n=6) to
12 cmH2O then back to 8 cmH2O over the initial 60 min. Pulmonary blood flow was
measured continuously while ventilatory parameters and arterial blood gases were
measured at intervals. At postmortem, in situ pressure‐volume deflation curves were
recorded, and bronchoalveolar lavage fluid and lung tissue were obtained to assess
inflammation. PEEPadj lambs had lower pressure amplitude, fractional inspired oxygen
concentration, oxygenation index and PBF, and more compliant lungs. Inflammatory
markers were lower in the PEEPadj group. Adjusted PEEP during HFJV improves
oxygenation and lung compliance and reduces ventilator requirements despite reducing
pulmonary perfusion.
123
Introduction
High‐frequency ventilation (HFV) is advocated as a lung protective ventilation strategy for
the treatment of respiratory distress syndrome (RDS) in preterm newborn infants. HFV has
proven particularly useful for optimising lung volume, reducing atelectotrauma and
volutrauma, and therefore reducing injurious lung stimuli associated with
bronchopulmonary dysplasia (BPD) (1‐3). High frequency jet ventilation (HFJV) and high‐
frequency oscillatory ventilation (HFOV) are the two main forms of HFV used in neonatal
intensive care units and while there is substantial research on the optimal approach to
lung volume optimisation in preterm RDS using HFOV, data are limited for HFJV. To date
there is one multicentre controlled trial and one single centre controlled trial comparing
the use of HFJV (with a low positive end‐expiratory pressure (PEEP) strategy) to
conventional mechanical ventilation (CMV) for treatment of preterm RDS (4, 5). The
results give conflicting information regarding the respiratory and neurological outcomes of
neonates treated with HFJV. Increased adverse neurological outcomes for HFJV group in
the Wiswell study (1996) were attributed to hypocarbia (6). Subgroup analysis in the
Keszler trial (1997) suggested that the use of low PEEP was associated with an increased
risk for grade III‐IV intraventricular hemorrhage or periventricular leukomalacia. An
important limitation of these trials is that the HFJV groups were not compared to a true
“lung protective” CMV strategy. Further research assessing the pros and cons of optimised
versus low PEEP in HFJV is therefore warranted.
124
HFJV is characterised by a (normally) fixed brief inspiratory time, passive expiration, and
coupling with a conventional ventilator for provision of conventional breaths, positive
end‐expiratory pressure (PEEP) and bias flow. The current recommended strategy for
treatment of RDS with HFJV is to commence HFJV early in the disease process with a peak
inspiratory pressure (PIP) just below that being used during conventional mechanical
ventilation (CMV) (http://www.bunl.com/7%20Steps%20NO%20QT.html). The initial PEEP
is set to achieve a mean airway pressure (Paw) equal to that used prior to commencement
of HFJV. The primary method advocated for optimising lung volume recruitment in HFJV is
by incrementing PEEP until stable peripheral oxyhemoglobin saturation (SpO2) is achieved,
with low‐rate CMV breaths added to HFJV as a supplementary method for recruiting
collapsed alveoli.
Despite the detailed guidelines provided for optimising lung volume during initiation of
HFJV, the evidence basis for this approach is limited. Whereas inadequate PEEP will
encourage airway collapse and atelectasis and initiate the lung injury sequence, excessive
PEEP promotes alveolar overdistension, impedes pulmonary perfusion, decreases venous
return, and depresses cardiovascular function (7‐9).
We hypothesised that a PEEP driven lung volume recruitment protocol would enhance
arterial oxygenation and ventilation without promoting lung injury during the initiation of
ventilation in a preterm ovine model of RDS. Furthermore, we hypothesised that these
effects would be achieved at the expense of pulmonary perfusion. We aimed to compare
125
the effect of an adjusted PEEP strategy on pulmonary blood flow (PBF), blood gases and
lung injury with a constant low PEEP strategy in an instrumented preterm lamb model.
126
Materials and Methods
All animal procedures were approved by the University of Western Australia animal ethics
committee, according to the guidelines of the National Health and Medical Research
Council of Australia code of practice for the care and use of animals for scientific purposes
(10).
Animals, Instrumentation and Delivery
Single and twin‐bearing date‐mated merino ewes were anaesthetised at 128‐130 d
gestation (term is ~ 150 d) with intramuscular xylazine (0.5 mg kg‐1, Troy Laboratories,
N.S.W., Australia) and ketamine (20 mg kg‐1, Parnell Laboratories, N.S.W., Australia), and
intubated (7.5 mm cuffed tracheal tube, Portex Ltd. England). Maternal anaesthesia was
maintained with halothane in 100% O2. The fetus was exteriorised and a right lateral
thoracotomy was performed. A flow probe (4R, Transonic Systems, Ithaca, NY) was
positioned around the left pulmonary artery and a catheter was inserted into the main
pulmonary artery (7). The fetus was intubated orally (4.5 mm cuffed tracheal tube, Portex
Ltd. England), lung fluid was suctioned and intra‐tracheal surfactant (100 mg kg‐1:
Survanta, 25 mg of phospholipids mL‐1, Abbott Laboratories, U.S.A.) was administered
prior to caesarian section delivery of the lamb. Unventilated controls (UVC; n=6) were
euthanased (pentobarbitone 100 mg kg‐1 i.v.) at delivery without instrumentation.
127
Postnatal care
Instrumented lambs were dried, weighed and randomised to one of two ventilation
groups: constant PEEP (PEEPconst; n=6) and adjusted PEEP (PEEPadj; n=6). They were
commenced on HFJV according to a predetermined protocol (Figure 1). Umbilical venous
and arterial catheters were inserted. Propofol (0.1 mg/kg/min; Repose 10%, Norbrook
Laboratories Ltd., Victoria, Australia) and remifentanil (0.05 µg/kg/min; Ultiva, Abbott
Laboratories, U.S.A.) were infused continuously through an umbilical vein for anaesthesia
and analgesia. An umbilical arterial catheter was used for continuous measurement of
systemic arterial blood pressure and intermittent sampling to assess gas exchange and
acid‐base balance. Rectal temperature was monitored continuously and maintained
between 38° and 39° C (normothermic for newborn lambs).
The Oxygenation Index (OI) was calculated as OI=2
2
PaO
100 x Paw xFiO where FiO2 is fractional
inspired oxygen concentration, Paw is mean airway pressure and PaO2 is partial pressure
of oxygen in arterial blood.
High Frequency Jet Ventilation
HFJV (Life Pulse High Frequency Ventilator, Bunnell Inc., Salt Lake City, U.S.A.) coupled to a
pressure‐limited time‐cycled infant conventional ventilator (Bourns Life Systems BP 200
Infant Pressure Ventilator, California, U.S.A.) was commenced immediately following
delivery. HFJV was commenced using a respiratory rate of 420 breaths per minute (7 Hz)
128
and peak inspiratory pressure (PIP) of 40 cmH2O. HFJV PIP was adjusted to a maximum of
40 cmH2O to target moderate permissive hypercapnia (Partial pressure of carbon dioxide
in arterial blood (PaCO2) 45‐55 mmHg). The initial FiO2 of 0.4 was adjusted to maintain
peripheral oxyhemoglobin saturation (SpO2) of 90‐95%. Inspiratory time (tI) was fixed at
0.02 s. No conventional ventilator breaths were applied during the 2 hour study period.
PEEP was maintained at 5 cmH2O in the PEEPconst group for the duration of the study.
Lambs in the PEEPadj group were stabilised on a PEEP of 5 cmH2O for 10 min then PEEP
was incremented at 10 min (8 cmH2O), 15 min (10 cmH2O) and 20 min (12 cmH2O). PEEP
was decreased by 2 cmH2O at 35 min and 60 min and then maintained at 8 cmH2O from
60 min until euthanasia at 120 min.
Continuous measurements of PBF, pulmonary artery blood pressure (PAP) and systemic
arterial blood pressure (ABP) were processed via calibrated pressure transducers (Maxxim
Medical, Tx, U.S.A.). Data were amplified and stored on a digital data acquisition system
(Powerlab 8SP, ADInstruments, N.S.W., Australia). Pulmonary waveform analysis was
performed at regular time points as described previously (11). Pulsatility Index, a measure
of downstream resistance to blood flow, was calculated as (peak systolic flow – minimum
flow after diastolic flow)/mean peak systolic flow over five consecutive cardiac cycles).
Tidal volume (VT) was measured continuously using an electronic flowmeter (Florian,
Acutronics, CH). Ventilator settings (respiratory rate, PIP, PEEP; Paw, Delta P (∆P), Servo
Pressure and ti) were recorded at intervals. After final measurements were obtained, the
129
FiO2 was increased to 1.0 for 2 minutes, the tracheal tube was occluded for 3 min, and the
lamb was euthanased (100 mg kg‐1 pentobarbitone i.v.).
Post‐mortem
The lung was exposed by thoracotomy, and an in situ deflation pressure volume curve was
obtained (12). The right upper lung lobe was inflation fixed (30 cmH2O) in formalin and
samples of the right lower lobe were snap frozen for molecular analyses. Bronchoalveolar
lavage (BAL) was performed on the left lung for cytology and protein analysis by the Lowry
method. Differential cell counts were performed on cytospin samples of the BAL fluid
stained with Diff‐Quik (Fronine Lab Supplies, N.S.W., Australia).
Total RNA was isolated from 30 mg of homogenised lung tissue using the RNeasy Mini kit
(Qiagen, U.S.A.) according to the manufacturer’s instructions. The contaminating genomic
DNA was removed by an on‐column DNaseI digestion performed using the DNaseI
digestion kit (Qiagen, U.S.A.). One microgram of RNA was then reverse transcribed into
complementary DNA in a 20 µL reaction with QuantiTect® Reverse Transcription Kit
(Qiagen, U.S.A.). The primers used for amplifying interleukin (IL) 1β and IL‐6 (13) and IL‐8,
early growth response (EGR) 1, connective tissue growth factor (CTGF) and cysteine rich
61 (CYR 61) have been described elsewhere (14). Amplification and detection of specific
products were conducted on the Rotor‐gene 3000 real time PCR system (Corbett Life
Science) with the published cycle profiles using Rotor‐gene SYBR Green PCR Kit (Qiagen,
U.S.A.) following the manufacturer’s instructions. The expression levels of genes of
130
interest were normalized into 18S RNA (15) using the 2‐∆∆CT method (16) and presented as
expression ratio relative to the unventilated control group (UVC).
Myeloperoxidase (MPO) activity was measured spectrophotometrically using methods
described by McCabe et al in 2001 and Faith et al in 2008 (17, 18). The MPO activity was
normalized to the total protein content of the tested samples. Activity was expressed as
units of MPO activity per mg of protein, where one unit of MPO was defined as the
amount needed to degrade 1 µmol of hydrogen peroxide per minute at room
temperature.
Statistical Analyses
For comparison of 2 groups of ventilated animals at specific time points, a Mann‐Whitney
Rank Sum test (non‐parametric data) or a Student’s t‐test (parametric data) was used.
Comparisons of two ventilated groups against the unventilated controls used one‐way
analysis of variance (ANOVA). A two‐way repeated measure ANOVA was used to
determine the effect of PEEP on PBF and Pulsatility Index and the effect of time on PIP,
Delta P, Paw and Servo Pressure. Data in the text and legends are expressed as mean
(SEM) or median (25,75 centile) unless otherwise stated. Analyses were performed using
SigmaStat (Version 3.5, Systat Software Incorporated, U.S.A.) with p<0.05 considered
statistically significant.
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Results
Baseline characteristics of lambs in each group were not different (Table 1).
Ventilator Settings and Lung Mechanics
Changes in Paw reflected the different PEEP protocols (Figure 2A) but as PIP decreased in
both groups, the amplitude of the airway pressure waveform (∆P) also decreased (Figure
2B). The VT was higher in PEEPadj group (pooled time points: p <0.01) despite a significantly
lower ∆P (Figure 2C). Servo pressure was lower (p = 0.026) in the PEEPadj group at 120 min
(Table 2). Pressure‐volume curves showed a higher volume achieved per unit of pressure
for the PEEPadj lambs compared to the PEEPconst lambs (p = 0.003 at 40 cmH2O) and for
both of the ventilated groups compared to the UVCs (Figure 2D).
Oxygenation
The target SpO2 of 90% to 95% was achieved in both groups within the first 10 minutes
and was maintained throughout the ventilation period. FiO2 and OI were lower in the
PEEPadj group compared to the PEEPconst lambs from 45 minutes until the end of the
ventilation period (Figure 3A and 3B).
Blood gases
The target PaCO2 was achieved within 10 minutes in both groups and was maintained
between 45‐55 mmHg throughout the ventilation procedure (Table 2). The pH, arterial
132
lactate concentration and base excess (BE) were comparable between the two groups for
the duration of the ventilation period (Table 2).
Hemodynamic consequences of different PEEP protocols
PBF increased over the first 10 min of ventilation in both groups and there were no
significant differences between the groups at any time point between 10 and 120 min.
PBF decreased by approximately 8% in the PEEPconst group and by approximately 48% in
the PEEPadj group between 10 and 120 min (p = 0.507 and p = 0.026 respectively) (Figure
4A). After an initial decrease during the transition from fetal to neonatal circulation, the
Pulsatility Index increased in both groups over time and was significantly higher in the
PEEPadj group over much of the first 60 min (Figure 4B). The pulmonary and systemic
arterial blood pressures were no different between the groups at any time point (Table 2).
End diastolic and end systolic pulmonary blood flow were significantly decreased at 15
min in the PEEPadj group (p<0.001 for each parameter). For all pulmonary artery variables,
the difference between PEEPconst and PEEPadj was temporary (Figure 4C and 4D).
Lung Injury
Bronchoalveolar Lavage Fluid
BAL fluid protein concentration was higher in ventilated groups than in the unventilated
controls, but no difference between PEEPadj and PEEPconst was observed. The cell
populations of BAL fluid did not differ between the groups (Table 3).
133
Lung Tissue
Compared to unventilated controls, IL‐1β, IL‐6, IL‐8, CTGF and EGR1 mRNA was elevated in
the PEEPconst group compared to UVCs, whereas CYR 61 expression was significantly
elevated in the PEEPadj group compared to UVCs. The expression of IL‐1β, IL‐6, EGR1 and
CTGF was greater in the PEEPconst group compared to PEEPadj. There was no difference in
MPO activity (Table 3) between each of the three groups.
134
Discussion
Alveolar recruitment during HFJV can be achieved by delivering CMV breaths or adjusting
PEEP, or both. To examine the role of PEEP for alveolar recruitment during HFJV, this study
aimed to compare the effect of an adjusted versus a constant PEEP protocol on
oxygenation, ventilation, pulmonary haemodynamics and lung injury during HFJV. We
showed that lambs in the PEEPadj group had better oxygenation and lung compliance but
decreased PBF compared to PEEPconst lambs. Markers of lung injury were higher in the
PEEPconst group in this short‐term study.
The correlation between Paw and oxygenation during HFJV is well understood. As
expected by study design, the Paw of the PEEPadj group was significantly higher than the
PEEPconst group throughout the ventilation period. The benefit of higher Paw was
evidenced by lower FiO2 requirements from as early as 45 min, supporting the concept
that a high Paw strategy enhances arterial oxygenation, decreasing the FiO2 required
during ventilation (19). In a clinical setting, PEEP is increased and FiO2 is decreased until
the SpO2 or PaO2 plateaus or falls. When FiO2 is stable at 0.21 optimal PEEP has been
achieved (20). The OI describes the relationship between FiO2, Paw and PaO2: a lower
value implies better arterial oxygenation (21). After the initial decrease in OI within the
PEEPconst group, the OI increased over the remainder of the study. In the PEEPadj group it
remained relatively low and stable despite the higher Paw, suggesting more efficient gas
exchange due to either airway stenting, or increased gas‐exchange surface due to
effective volume recruitment.
135
Servo pressure is the automatically controlled driving pressure of the ventilator and
changes in response to altered monitored airway pressure, to ensure that the ventilator
will continue to deliver inspiratory gas to meet the set PIP (22). Servo pressure increases
as PIP increases, with increased lung and/or chest wall compliance, a decrease in airway
resistance, or if there is an air‐leak or circuit disconnection. A decrease in servo pressure
however, indicates either a reduction in set PIP, or worsening compliance and increased
resistance to gas flow, obstruction of the tracheal tube, tension pneumothorax, the
requirement for suctioning or right mainstem bronchus intubation
(www.bunl.com/ServoSlidesNEW.html). Our finding of a lower Servo pressure throughout
most of the study period in PEEPadj lambs despite evidence of increased lung compliance,
likely reflects the reductions in PIP required to maintain moderate permissive
hypercapnea and avoid overventilation.
The haemodynamic consequences of the ventilation protocol were assessed by
measurement of PBF, PAP, systemic ABP, pulse rate and calculation of Pulsatility Index.
Pulsatility Index is directly related to resistance to blood flow. An increase in intrathoracic
pressure during any kind of positive pressure ventilation impacts on venous return,
cardiac output, right ventricular end diastolic volume, PBF and pulmonary vascular
resistance (23). Increased PEEP reduces PBF during conventional ventilation of very
premature lambs by increasing pulmonary vascular resistance (PVR) (8). A similar decrease
in PBF in response to Paw driven alveolar recruitment maneuvers during HFOV of up to
69.3% is also reported (7). Importantly, during HFOV, the fall in PBF persisted after Paw
136
was decreased following recruitment (7). The mechanism causing a fall in PBF as PEEP is
increased may include an increase in the alveolar capillary transmural pressure causing
capillary compression (8, 24). The non‐compliant immature lung may be less susceptible
to capillary compression than a mature lung (8, 24). Factors that increase lung compliance
(e.g. antenatal corticosteroids, exogenous surfactant, lung volume recruitment) may
increase the sensitivity of PBF to changes in airway pressure (24). We observed improved
oxygenation at the expense of PBF when PEEP was increased up to 12 cmH2O, as
previously described during HFOV (8). However, this decrease in PBF was relatively small
and of shorter duration relative to the change associated with increasing Paw during
HFOV (7).
The fall in PBF in the PEEPadj group reversed rapidly as PEEP was initially decreased,
suggesting the impact of a PEEP recruitment strategy on PBF during HFJV was not
sustained. During HFOV however, PBF does not recover after a temporary increase in Paw.
(7). Furthermore, the impact of increased PEEP on end diastolic and end systolic blood
flow coincided with the initial increase in PEEP from 5 cmH2O to 8 cmH2O at 10 min. The
difference between the 2 groups was temporary and despite further increases in PEEP in
the PEEPadj group, the blood flow variables did not remain significantly different. The
maintenance of a higher Paw in the PEEPadj group likely contributed to the continued slow
decline in PBF in this group and returning to PEEP of 5 cmH2O (instead of 8 cmH2O) may
have been prudent. Nevertheless, a direct comparison of HFJV and HFOV in a controlled
137
setting is warranted to determine the impact of each of these ventilation strategies on the
extent and duration of effect on pulmonary haemodynamics.
Lung inflammation is a prelude to lung injury. We examined inflammatory markers that
we anticipated would be increased at 120 min in response to lung injury (14, 25‐27). There
was a clear increase in lung injury across the range of inflammatory markers for the
PEEPconst group, compared to the unventilated controls and for the PEEPconst group
compared to PEEPadj. These findings support our hypothesis regarding reduced lung injury
with PEEP recruitment of the lung during HFJV.
Myeloperoxidase activity has been shown to correlate with IL‐6 expression (17) but our
results did not show a clear relationship between these variables. The MPO activity was
not different between the groups, despite a trend for an increase in the ventilated groups.
It is possible that this is a result of maternal anaesthesia, and this finding in itself warrants
further investigation. It is also possible that a Type II statistical error as a result of the
small group sizes prohibited a demonstrable difference between the groups.
There are a number of limitations to our study. We wanted to examine the physiological
changes associated with each PEEP alteration and required at least 10 minutes to
accommodate and document any changes. Consequently, the adjusted PEEP protocol we
studied does not reflect standard clinical practice given that the 60 min period for
increasing and decreasing PEEP is considerably longer than optimally used in a clinical
setting. and potentially masked significant differences between the ventilated groups.
138
Secondly, surfactant was administered to the lambs prior to the commencement of
ventilation. This practice may not be achieved routinely in a clinical setting. However, our
goal was to isolate the effect of PEEP and standardise and optimise all other aspects of
care. Thirdly, the lambs in our study were anaesthetised and underwent an invasive
surgical procedure. The haemodynamic effects of anaesthesia combined with the physical
impact of instrumentation on lung inflation are likely to impact physiological outcomes.
Lastly, the flowmeter used for measuring VT slightly overestimates tidal volume at 7 Hz
(28). However, as HFJV frequency was constant throughout the study, we would not
expect this to affect comparisons between the ventilatory groups.
In conclusion, adjusted PEEP during HFJV improves oxygenation and lung compliance and
reduces ventilator requirements despite reducing pulmonary perfusion. The majority of
markers of injury were higher when PEEP was constant during HFJV. Evaluation of PEEP
recruitment manoeuvers in human patients is indicated to explore the efficacy of the
technique in the target patient population.
139
Acknowledgements
Surfactant was donated by Abbott Australia. The Life Pulse High Frequency Ventilators
were supplied on long‐term loan by Bunnell Incorporated. We would like to express our
sincere appreciation to the members of the Ovine Research Group for technical assistance
and JRL Hall and Co. for provision and early antenatal care of the ewes.
140
Tables
Table 1: Baseline characteristics
UVC PEEPconst PEEPadj
n (male) 6 (3) 6 (4) 6 (4)
Twin (singleton) 4 (2) 6 (0) 6 (0)
Birthweight (kg) 2.9 (0.4) 3 (0.3) 2.8 (0.2)
Gestational Age (d) 127.2 (0.4) 128 (0.8) 128 (0.8)
Cord pH 7.15 (0.1) 7.15 (0.05) 7.09 (0.1)
Cord PaCO2 (mmHg) 81 (13) 73.8 (7.4) 89.7 (20.8)
UVC = Unventilated Control, PEEPconst = Constant Positive End Expiratory Pressure,
PEEPadj = Adjusted Positive End Expiratory Pressure. Values are mean (SEM).
141
Table 2: Cardiovascular and Respiratory Physiological Measurements
PEEPconst PEEPadj
Time 10 min 60 min 120 min 10 min 60 min 120 min
PIP (cmH2O) 40 (0) 34.5 (2.5) 30.2 (2.3) 40 (0) 32.8 (1.9) 27.9 (1)
Paw (cmH2O) 14.1 (0.9) 13.5 (0.6) 12 (0.6) 14.5 (0.2) 16.7 (0.4)† 13.5 (0.3)†
Servo Pressure (psi) 6.6 (0.8) 8.3 (0.3) 7 (0.3) 7.8 (0.4) 6.8 (0.7) 5.9 (0.3)†
PaCO2 (mmHg) 52.0 (4.6) 41.5 (2) 43.3 (2.6) 39.8 (3.4) 38.2 (3.5) 41.4 (3.1)
pH 7.1 (0.1) 7.4 (0.02) 7.4 (0.04) 7.3 (0.1) 7.3 (0.04) 7.4 (0.03)
Lactate (mg dL‐1) 83.9 (13.4) 67.4 (10.7) 48.1 (10.9) 87.2 (22.6) 67.2 (19) 50.4 (20.1)
Base Excess ‐8.8 (2.1) ‐2.2 (1.4) ‐0.9 (2.1) ‐6.8 (3.2) ‐5.4 (1.9) ‐1.5 (2.2)
Left PAP (mmHg) 43.6 (4.3) 35.6 (3.8) 43.2 (6.8) 48.5 (1.6) 37.4 (3.4) 43.8 (5.6)
Systemic ABP (mmHg) 53.5 (6.2) 50.1 (4.4) 60.3 (6.7) 53 (3.2) 50.2 (3.1) 53.3 (3.4)
Pulse rate (bpm) 140 (7) 170 (15) 194 (8)* 145 (8) 181 (15) 210 (8)*
PIP = Peak Inspiratory Pressure, ∆P = Pressure differential (PIP – PEEP), Paw = Mean
Airway Pressure, psi = pounds per square inch, PaCO2 = partial pressure of carbon dioxide
in arterial blood, PAP = pulmonary arterial blood pressure, Systemic ABP = systemic
arterial blood pressure, bpm = beat per minute. * p<0.001 time 120 compared to time 10
in the same group, † p<0.05 PEEPadj compared to PEEPconst at the same time. Values are
mean (SEM).
142
Table 3: Post mortem inflammatory markers
UVC PEEPconst PEEPadj
BAL fluid
Protein concentration (mg mL‐1) 14.7 (7.4) 120.2 (28)* 89.2 (18.5)*
Inflammatory cells (x 103 kg‐1) 0.3 (0.02) 7.5 (3.9)* 2.0 (1.0)
Neutrophils (x 103 kg‐1) 0 4.4 (2.4) 0.4 (0.2)
Mononuclear cells (x 103 kg‐1) 0.3 (0.02) 2.7 (1.4)* 1.5 (0.8)
Lymphocytes (x 103 kg‐1) 0 0.4 (0.3) 0.1 (0.1)
Lung Tissue
IL‐1β fold change 0.6 (0.4, 3.2) 262.9 (159.3, 350)** 37.2 (18.2, 53.7)‡
IL‐6 fold change 1.0 (0.6, 1.6) 95.5 (57.2, 155.4)* 13.4 (3.2, 23.8)‡
IL‐8 fold change 0.7 (0.6, 2.0) 77.6 (26.1, 101.1)** 28.0 (6.3, 53.9)
EGR1 fold change 1.1 (0.3, 2.9) 48.9 (33.5, 94.7)* 15.6 (5.4, 27.1)‡
CTGF fold change 0.8 (0.6, 2.4) 31.2 (13.3, 44.3)* 4.0 (1.97, 5.9)‡
CYR61 fold change 0.9 (0.7, 1.3) 9.7 (5.4, 20.1) 11.0 (5.9, 12.2)*
MPO ac vity † 18.2 (1.4) 29.3 (3.8) 26.6 (6.1)
IL = interleukin, EGR = early growth response, CTGF = connective tissue growth factor, CYR
61 = cysteine rich 61, MPO = myeloperoxidase. † units per mg of protein where 1 unit of
MPO is defined as the amount needed to degrade 1 µmol of hydrogen peroxide per
minute at room temperature. *p<0.05 compared to UVC, **p<0.001 compared to UVC, ‡
p<0.05 compared to PEEPconst. Values are mean (SEM) or median (25,75 centile).
143
Figure 1
144
Figure 2
145
Figure 3
146
Figure 4
147
Figure Legends
Figure 1 Constant and Adjusted PEEP protocols: Lambs were ventilated at either a
constant PEEP of 5 cmH2O (PEEPconst, solid line) or with stepwise increments and
decrements to PEEP (PEEPadj, grey). Values are Mean (SEM).
Figure 2 Ventilator Settings and Lung Mechanics: A: Mean Airway Pressure (Paw), B:
Delta P (ΔP), C: Tidal Volume, D: Deflation limb of the post‐mortem in situ pressure‐
volume curves. ◊ = PEEPconst, ● = PEEPadj, = Unventilated control. * p<0.05 compared to
PEEPconst ** p<0.05 compared to unventilated control. Values are Mean (SEM).
Figure 3 Oxygenation: A: Fractional inspired oxygen concentration (FiO2), B: Oxygenation
index (OI) over the 120 min ventilation period. ◊ = PEEPconst, ● = PEEPadj. * p<0.05
compared to PEEPconst. Values are Mean (SEM).
Figure 4 Pulmonary Perfusion: A: Pulmonary Blood Flow, B: Pulsatility Index over the 120
min ventilation period, C: End Diastolic Blood Flow, D: End Systolic Blood Flow. ◊ =
PEEPconst, ● = PEEPadj , * p<0.05 compared to PEEPadj ** p<0.05 time 120 min compared to
time 10 min. Values are Mean (SEM).
148
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153
5 The Impact of Conventional Breath Inspiratory Time during
High‐frequency Jet Ventilation in Preterm Lambs
Gabrielle C Musk,1* Graeme R Polglase,1 Yong Song,1 and J Jane Pillow1
1School of Women’s and Infants’ Health, the University of Western Australia, M550, 35
Stirling Highway, Crawley, 6009, Western Australia, Australia.
This is the first study investigating the role of conventional mechanical ventilator breaths
for alveolar recruitment during high‐frequency jet ventilation. We isolated the effect of
the inspiratory time of the conventional mechanical ventilation breath in this study.
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Abstract
Background: The delivery of conventional mechanical ventilator (CMV) breaths during
high‐frequency jet ventilation (HFJV) is advocated to recruit and stabilise alveoli.
Objectives: To establish if CMV breath duration delivered during HFJV influences gas
exchange, lung mechanics and lung injury.
Methods: Sedated newborn preterm lambs at 128 d gestational age were studied. HFJV (7
Hz, PEEP 8 cmH2O, PIPHFJV 40 cmH2O, FiO2 0.4) with superimposed CMV breaths (PIPCMV 25
cmH2O, rate 5 breaths/min) was commenced after delivery and continued for 2 h. CMV
breath inspiratory time (tI) was either 0.5 s (HFJV+CMV0.5: n=8) or 2.0 s (HFJV+CMV2.0:
n=8). Age matched unventilated controls (UVC) were included for comparison.
Results: Serial arterial blood gas analyses were performed. PIPHFJV was adjusted to target a
PaCO2 of 45‐55 mmHg. FiO2 was adjusted to target SpO2 90‐95 %. Static deflation
pressure‐volume curves, bronchoalveolar lavage (BAL) and lung tissue samples were
obtained post‐mortem. Gas exchange, ventilation parameters, static lung compliance and
BAL inflammatory markers were not different between HFJV+CMV0.5 and HFJV+CMV2.0.
Both ventilation groups had higher BAL inflammatory markers and increased iNOS positive
cells on histology compared to UVC, whilst lung tissue IL‐1β and IL‐6 mRNA expression was
higher in the HFJV+CMV2.0 group compared to the UVC group.
Conclusions: Preterm lambs were ventilated effectively with HFJV and 5 CMV
breaths/min. CMV breath duration did not alter blood gas exchange, ventilation
parameters ex vivo static lung mechanics or markers of lung injury over a 2 h study,
155
although consistent trends towards increased inflammatory markers with the longer tI
suggest greater risk of injury.
156
Introduction
High‐frequency jet ventilation (HFJV) is a lung protective ventilation strategy for patients
with respiratory distress syndrome (RDS) (1, 2). As with any ventilation strategy, the aim
during HFJV is to recruit, stabilise and maintain open alveoli to facilitate efficient gas
exchange and to prevent lung injury (3, 4). The primary method advocated for optimising
lung volume recruitment during HFJV includes recruiting collapsed units with low‐rate
conventional mechanical ventilator (CMV) breaths superimposed on the HFJV waveform
(1, 5). The peak inspiratory pressure (PIP) of the CMV breath (CMVPIP) is optimally set
above the opening pressure (lower inflection point) on the inflation pressure‐volume
curve. Alveoli are stabilised in expiration by incrementing positive end‐expiratory pressure
(PEEP) until the target oxyhemoglobin saturation (SpO2) is achieved and maintained
without a background CMV rate (6, 7)
While the delivery of CMV breaths during HFJV is advocated, there is little information to
guide the selection of the inspiratory time (tI) of the CMV breath (8). Extending the
duration of the CMV breath beyond that required to complete tidal volume delivery may
promote aeration of alveolar units with long time constants, but will expose aerated lung
regions to unnecessarily high peak inspiratory pressures for extended times.
The duration of the inspiratory phase of the CMV breath potentially affects the delivered
CMV tidal volume (depending on the time constant of the lung), and the duration of
exposure of alveoli to the set PIPCMV. When HFJV is applied to poorly compliant, atelectatic
157
lungs, the lungs will inflate completely with a short tI. Whereas a longer tI may enhance
recruitment by facilitating aeration of long time constant acini, this approach may expose
existing aerated units to barotrauma.
We aimed to compare the effect of different duration CMV breaths during HFJV on HFJV
ventilator requirements, gas exchange and markers of lung injury in a preterm lamb model
of respiratory distress syndrome. We hypothesised that in the setting of preterm
respiratory distress syndrome, a CMV breath with a long tI would promote lung injury,
without improvement in arterial oxygenation.
158
Materials and Methods
All animal procedures were approved by the University of Western Australia animal ethics
committee, according to the guidelines of the National Health and Medical Research
Council of Australia code of practice for the care and use of animals for scientific purposes
(9).
Animals, Instrumentation, and Delivery
An intravenous injection of medetomidine (0.02 mg kg‐1, Pfizer Animal Health, U.S.A.) and
ketamine (10 mg kg‐1, Troy Laboratories, Australia) was administered to induce
anaesthesia in pregnant ewes at a mean (SD) of 128 (0.8) d gestation, prior to spinal
anaesthesia (3 mL lidocaine, 20 mg mL‐1, Troy Laboratories, Australia) and surgical
delivery. Lambs were randomised to either unventilated controls (UVC, n=5) that were
euthanased at delivery (pentobarbitone 100 mg kg‐1 i.v., Valabarb, Jurox, Australia)
immediately prior to postmortem, or to one of two HFJV strategies. For ventilation groups,
the fetus was exteriorised through a uterine incision, intubated orally (4.5 mm ID cuffed
tracheal tube, Portex Ltd. England), suctioned and intratracheal surfactant was instilled
(100 mg kg‐1: Survanta, Abbott Laboratories, U.S.A.) prior to delivery of the lamb.
Postnatal care
Lambs were dried, weighed and commenced on their randomised ventilation strategy:
HFJV with CMV breaths delivered over 0.5 s (HFJV+CMV0.5; n=8); and HFJV with CMV
breaths delivered over 2.0 s (HFJV+CMV2.0; n=8). Propofol (0.1 mg kg‐1 min‐1; Norbrook
159
Laboratories Ltd., Victoria, Australia) and remifentanil (0.05 µg kg‐1min‐1; Abbott
Laboratories, U.S.A.) were infused continuously via an umbilical venous catheter for
anaesthesia and analgesia. An umbilical arterial catheter was sampled intermittently to
assess gas exchange and acid‐base status. Core body temperature was monitored
continuously and maintained between 38° C and 39° C. Oxygenation Index (OI) was
calculated as:
OI=1.36 x aO
100 x aw xFiO
2
2
P
P
where FiO2 is fractional inspired oxygen concentration, Paw is mean airway pressure
(cmH2O) and PaO2 is partial pressure of oxygen in arterial blood (mmHg). The constant
1.36 is for conversion of mmHg to cmH2O.
Mechanical Ventilation
HFJV (Life Pulse High Frequency Ventilator, Bunnell Inc., Salt Lake City, U.S.A.) coupled to
pressure‐limited time‐cycled infant conventional ventilator (Bourns Life Systems BP200
Infant Pressure Ventilator, California, U.S.A.) was commenced immediately following
administration of exogenous surfactant. Initial HFJV settings included a respiratory rate of
420 breaths/min (7 Hz), FiO2 0.4 and a PIPHFJV of 40 cmH2O. PIPHFJV was adjusted to achieve
permissive hypercapnia (partial pressure of carbon dioxide in arterial blood (PaCO2) 45‐55
mmHg) to a maximum of 40 cmH2O. FiO2 was altered to maintain SpO2 at 90‐95 %. HFJV tI
was fixed at 0.02 s. Intermittent CMV breaths using the assigned tI were superimposed on
160
the HFJV waveform to a PIPCMV of 25 cmH2O and a rate of 5 breaths/min. The PEEP was
constant throughout at 8 cmH2O.
High‐frequency jet ventilation measured variables (PIPHFJV, PEEP; Paw,HFJV, pressure
differential (∆PHFJV) and servo pressure) were recorded at regular intervals. After obtaining
final measurements at 2 h, the FiO2 was increased to 1.0 for 2 min followed by a 3 min
tracheal tube occlusion to facilitate lung collapse for post‐mortem pressure‐volume curve
and then euthanasia (pentobarbitone 100 mg kg‐1 i.v. Jurox, Australia).
Post‐mortem
The collapsed lung was exposed by thoracotomy, inflated slowly to 40 cmH2O, and an in
situ deflation pressure volume curve was recorded (10). Bronchoalveolar lavage (BAL) fluid
was collected from the left lung for protein analysis (11, 12) and cytology. Differential cell
counts were performed on cytospin samples of the BAL fluid stained with Diff‐Quik
(Fronine Lab Supplies, N.S.W., Australia). The right upper lung lobe was inflation fixed (30
cmH2O) in 10 % formalin. Immunohistochemical staining for inducible nitric oxide
synthase (iNOS) was performed on 5 µm sections of lung tissue (13). Positive cells were
identified and quantified per field as number of cells per total cellular area (nm2) using
densitometry (Image‐Pro Plus version 4.5, Media Cybernetics, U.S.A.).
RNA was extracted from the left lung and reverse transcribed to cDNA (QuantiTect®
Reverse Transcription Kit, Qiagen, U.S.A.). Expression of IL‐1β and IL‐6 was measured by
qRT‐PCR (14) and normalized to 18S RNA (15) using the 2‐∆∆CT method (16).
161
Statistical Analyses
Kruskal‐Wallis one way analysis of variance was used to compare groups at specific time
points while the effect of CMV strategy on ventilator requirements and physiological
changes over the duration of the study was determined using two‐way repeated measure
analysis of variance. A Holm Sidak post‐hoc test was used to determine significance
(p<0.05). Analyses were performed using SigmaStat (Version 3.5, Systat Software
Incorporated, U.S.A.). Data are expressed as mean (SEM) unless otherwise stated.
162
Results
Baseline characteristics of the lambs were not different between groups (Table 1).
Physiological Measurements
HFJV and CMV monitored pressures (PIPHFJV, Paw,HFJV, ∆PHFJV or servo pressure, Fig. 1),
oxygen requirements (FiO2, Fig. 2A) or indices of arterial oxygenation (PaO2 and OI, Fig. 2B
and 2C) were not different between the two ventilation groups over the 2 h study. PaCO2
was similar for each group throughout the study (Fig. 2D). PaCO2 initially fell rapidly, but
subsequently increased with ventilator adjustment and stabilised within the target range
by 60 min.
Post‐mortem static lung compliance was comparable for HFJV+CMV0.5 and HFJV+CMV2.0.
Lung volume was higher in both ventilated groups compared to the UVC group at 15
cmH2O, 20 cmH2O and 40 cmH2O (Fig. 3).
Lung Injury
The total protein concentration of BAL fluid was higher in both ventilated groups
compared to UVCs, but did not differ between HFJV+CMV0.5 and HFJV+CMV2.0 (p=0.38).
Similarly, there was no difference in the total inflammatory cell population of the BAL fluid
between the ventilated groups (p=0.16), although total inflammatory cell counts were
increased compared to UVCs (Table 2).
163
The number of iNOS positive cells was not different between the ventilated groups. Lung
IL‐1β and IL‐6 mRNA expression was significantly greater in the HFJV+CMV2.0 group (Table
2) compared to UVCs.
164
Discussion
Although low rate CMV breaths are often used to assist alveolar recruitment during HFJV,
few data exist to guide clinicians on the selection of rate, size or duration of such CMV
breaths. We investigated the effect of two different inspiratory times for low rate CMV
breaths during HFJV and found no significant differences in gas exchange, ventilation
parameters, static lung compliance or inflammatory markers (as determined by BAL
protein, tissue iNOS or pro‐inflammatory cytokine mRNA expression) between a CMV tI of
0.5 s and 2.0 s.
The primary purpose of CMV breaths during HFJV is to promote alveolar recruitment via
alveolar exposure to a distending pressure above the opening pressure threshold. In
previous studies using similar gestation naïve lambs, we have consistently observed a
pressure of 25 cmH2O to be above the lower inflection point (17). In the current study we
expected the opening pressure to be lower than our previous experience given that the
ventilated lambs received prophylactic surfactant, thus an initial PIPCMV of 25 cmH2O
should achieve recruitment. Although PIPCMV was weaned alongside PIPHFJV to remain 5
cmH2O lower than PIPHFJV throughout the study period, the PIPCMV did not fall below 20
cmH2O or differ between the two ventilation groups at any time during the study.
Nonetheless, it is possible that the PIPCMV was lower than the opening pressure and that
this may account for the failure to demonstrate differences between the short and long
duration PIPCMV.
165
To isolate the effect of CMV breath tI we kept other CMV variables constant between the
two ventilated groups: a CMV breath rate of 5 breaths/min reflects the recommended
upper limit for common clinical practice (1). Likewise, a constant PEEPCMV of 8 cmH2O was
maintained throughout the study. We showed previously that a constant PEEP of 5 cmH2O
was insufficient to stabilise alveoli (18) during HFJV.
Our selection of 0.5 s for the short tI was guided by our previous experience using CMV in
preterm lambs: in the 128 d preterm lamb, 0.5 s is sufficient to complete inspiration with a
constant ventilator flow of 8 L/min with a waveform pattern comparable to that achieved
using 0.3 s ‐ 0.4 s inspiratory times commonly used in small babies (19). We rationalised
the selection of 2.0 s as the long tI for comparison as it would provide at least 1.5 s of
pressure plateau. If an extended tI for the CMV breath enhanced lung volume recruitment
during HFJV, we would have expected improved oxygenation in the group receiving the
2.0 s CMV breath. The absence of any difference in FiO2, PaO2 or OI between the two
ventilator groups suggests a 2.0 s CMV breath has no volume recruitment advantage over
a 0.5 s CMV breath during HFJV in the preterm lamb over 2 hours in the postnatal
transition period.
Whilst no specific advantage of inspiratory time was demonstrated, there was evidence of
improved compliance for both CMV tI strategies: servo pressure initially stayed high
despite a rapid decrease in PIPHFJV and ∆PHFJV, indicative of increased lung compliance.
Servo pressure is the automatically controlled driving pressure of the jet ventilator which
changes in response to monitored airway pressure to ensure that the ventilator continues
166
to deliver sufficient inspiratory gas flow to meet the set PIPHFJV (20). There were no
differences between the ventilator groups in the servo pressure at any time point,
suggesting that similar improvements in compliance were achieved for both groups during
the initial phases. Interpretation of servo pressure in the latter half of the study is more
complicated: decreases in servo pressure in the latter half of the study paralleled further
reductions in PIPHFJV and ΔPHFJV suggesting that the previously achieved level of
compliance was at least maintained. Reduced servo pressure in the absence of
corresponding reductions in ΔPHFJV would have implied decreased compliance and loss of
lung volume. There was a slow increase in OI towards the end of the study in both
ventilated groups likely indicative of alveolar instability and collapse due to the use of
Paw,HFJV or PEEPHFJV that was too low to maintain end expiratory lung volume.
We deliberately used a constant PEEP strategy throughout the study to isolate the effect
of different CMV tI strategies during HFJV on oxygenation from the potentially
confounding effects arising from altered PEEP. In clinical practice, however, adequacy of
PIPCMV and PEEPCMV settings would be important considerations in assessing response to
changes in the OI. Increasing PEEPCMV rather than decreasing PIPHFJV may have effectively
decreased ΔPHFJV and controlled PaCO2 whilst maintaining Paw,HFJV and optimal lung
volume. Regardless, Paw,HFJV was not different between the two groups over the study
duration despite our rigid protocol.
Low rate CMV has a negligible contribution to ventilation during HFJV, during which CO2
removal is primarily influenced by HFJV frequency and HFJV tidal volume. The absence of
167
difference in HFJV parameters between the two ventilator groups is therefore not
surprising given the same HFJV ventilator strategy was used to target the desired PaCO2
range. The initial rapid fall in PaCO2 values to a level below the target range is most likely
related to the initial PIPHFJV selection.
In the absence of benefits of a specific CMV tI strategy on oxygenation or ventilation, the
question remains whether a long CMV breath tI is more injurious than a shorter breath.
Lung injury is preceded by lung inflammation, evidenced by increased inflammatory cells
in the airspaces and lung tissue that release pro‐inflammatory mediators (21). We
hypothesised that lung injury would be increased in the HFJV+CMV2.0 group compared to a
shorter CMV breath tI, and chose to measure inflammatory markers that we anticipated
would increase within 120 min (22‐25). Whereas HFJV+CMV was injurious in both groups
compared to UVC, there were no statistical differences in mRNA expression or static lung
compliance between the two ventilated groups. Nonetheless, all markers of injury (BAL
protein concentration, BAL inflammatory cells, lung tissue iNOS positive cells and lung
mRNA pro‐inflammatory cytokine expression), showed a consistent pattern of higher
levels in the 2.0 s group compared to the 0.5 s group. The lack of statistically significant
difference is likely related to a type II error. Regardless, any potential increase in injury of
a 2.0 s CMV breath tI compared to 0.5 s, is likely to be subtle at most over a 2 h time
frame. Based on our observed differences in the oxygenation index and IL‐6 and IL‐1β a
group size of 21 would have been required to detect a statistically significant difference
with 80% power.
168
Potential mechanisms promoting injury with a long CMV breath tI likely relate to extended
exposure of distal airways and alveoli to high continuous inflating pressures. Using a
PIPCMV less than the PIPHFJV, we deliberately prevented the CMV breath from interrupting
the HFJV breaths. Thus, theoretically at least, longer exposure to ‘jet‐stacking’ may have
been a consequence of this strategy for the HFJV+CMV2.0 group. However, the amplitude
of the HFJV pulses is markedly damped by the time the HFJV pressure waves are
transmitted to the periphery, and jet‐stacking does not result in PIPHFJV more than a few
cmH2O greater than the preset PIPHFJV. It is more likely that any consequence of longer
CMV breath inspiratory times are related to the PIPCMV (which is fully transmitted to the
periphery) rather than any stacking of HFJV breaths resulting from this approach.
Conclusions
We found no significant differences in gas exchange or markers of injury and inflammation
between HFJV with CMV breaths delivered with an tI of 0.5 s or 2.0 s over 2 h. However, a
consistent tendency for increased inflammatory markers when CMV breaths were
delivered with an tI of 2.0 s compared to a tI of 0.5 s suggests the use of longer CMV
breath inspiratory times may be more injurious if continued over a longer time frame, and
warrants further investigation. Further studies to provide evidence to support strategies
aiming to establish optimal lung volume recruitment during HFJV are recommended.
169
Acknowledgements
We would like to express our sincere appreciation to the members of the Ovine Research
Group, Ilias Nitsos and Carryn McLean, for technical assistance, JRL Hall and Co. for
provision and early antenatal care of the ewes and Professor Karen Simmer for support
and encouragement.
170
Tables
Table 1: Baseline characteristics
UVC HFJV+CMV0.5 HFJV+CMV2.0
n (male) 6 (3) 8 (6) 8 (4)
Twin (singleton) 4 (2) 8 (0) 8 (0)
Birthweight (kg) 2.9 (0.4) 2.9 (0.2) 3.0 (0.2)
Cord pH 7.15 (0.04) 7.15 (0.05) 7.16 (0.05)
Cord PaCO2 (mmHg) 81.0 (5.8) 73.8 (9.2) 80.6 (10.1)
UVC = Unventilated Control, HFJV+CMV0.5 = HFJV with CMV breaths delivered with
an inspiratory time of 0.5 s, HFJV+CMV2.0 = HFJV with CMV breaths delivered with an
inspiratory time of 2.0 s. Values are mean (SEM).
171
Table 2: Post‐mortem inflammatory markers
UVC HFJV+CMV0.5 HFJV+CMV2.0
BAL fluid
Protein concentration (mg mL‐1)
14.7 (7.4) 164.1 (27.2)* 197.8 (32.5)*
Total Inflammatory Cells (cells x 106 kg‐1)
1.9 (0.8) 44.3 (14.8)* 83.5 (22.6)*
Neutrophils (cells x 106 kg‐1)
0.5 (0.5) 31.3 (15.2)* 66.0 (20.9)*
Mononuclear cells (cells x 106 kg‐1)
1.4 (0.4) 12.2 (4.0)* 16.6 (3.2)*
Lymphocytes (cells x 106 kg‐1)
0.03 (0.01) 0.2 (0.2) 0.2 (0.1)
Lung Tissue
iNOS positive cells (cells (nm2)‐1)
0 154.9 (62)* 219.3 (83)*
IL‐1β (fold change)# 1.0 (0.6, 1.6) 64.1 (42.2, 110.9) 82.1 (57.2, 120.8)†
IL‐6 (fold change)# 0.6 (0.4, 3.2) 238.2 (66.3, 295.6) 266.1 (179.9, 490.5)†
UVC = Unventilated Control, HFJV+CMV0.5 = HFJV with CMV breaths delivered with
an inspiratory time of 0.5 s, HFJV+CMV2.0 = HFJV with CMV breaths delivered with an
inspiratory time of 2.0 s. * p<0.01 compared to UVC, † p=0.04 compared to UVC, #
n=7 for analyses. Values are mean (SEM) or median (25th, 75th centile) for parametric
and non‐parametric data respectively.
172
Figure 1
173
Figure 2
174
Figure 3
175
Figure Legends
Figure 1 Ventilator variables: A: Peak Inspiratory Pressure (PIP); B: Mean Airway Pressure
(Paw) calculated by the high‐frequency jet ventilator; C: ∆P (Pressure differential); D: Servo
Pressure (psi = pounds per square inch). ∆ HFJV with CMV breaths delivered over 0.5 s
(HFJV+CMV0.5), о HFJV with CMV breaths delivered over 2.0 s (HFJV+CMV2.0). Solid fill =
PIPHFJV, gray fill = PIPCMV.
Figure 2 Ventilation and Oxygenation: A: Fractional inspired oxygen concentration (FiO2);
B: Partial pressure of oxygen in arterial blood (PaO2); C: Oxygenation Index; D: Partial
pressure of carbon dioxide in arterial blood (PaCO2); E: pH. ∆ HFJV with CMV breaths
delivered over 0.5 s (HFJV+CMV0.5), о HFJV with CMV breaths delivered over 2.0 s
(HFJV+CMV2.0).
Figure 3 Pressure Volume Curve: ∆ HFJV with CMV breaths delivered over 0.5 s
(HFJV+CMV0.5), о HFJV with CMV breaths delivered over 2.0 s (HFJV+CMV2.0),
Unventilated control (UVC). * p<0.05 HFJV+CMV0.5 compared to UVC, ** p<0.01
HFJV+CMV2.0 compared to UVC.
176
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preterm infants with uncomplicated respiratory distress syndrome. Pediatrics
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2. Carlo W, Siner, B, Chatburn, RL, Robertson, S, Martin, RJ. 1990 Early randomised
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3. Courtney SE, Asselin JM 2006 High‐frequency jet and oscillatory ventilation for
neonates: Which strategy and when? Respir Care Clin N Am 12:453‐467
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Kramer BW 2007 Pulmonary and systemic endotoxin tolerance in preterm fetal
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17. Pillow JJ, Sly PD, Hantos Z 2004 Monitoring of lung volume recruitment and
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18. Musk GC, Polglase GR, Bunnell JB, McLean CJ, Nitsos I, Song Y, Pillow JJ 2010 High
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180
181
6 The Effect of Conventional Breath Peak Inspiratory Pressure
during High‐frequency Jet Ventilation in Preterm Lambs
Gabrielle C Musk 1, Graeme R Polglase2 and J Jane Pillow 1.
1School of Women’s and Infants’ Health, University of Western Australia, Perth, Australia.
2The Ritchie Centre, Monash Institute of Medical Research, Monash University,
Melbourne, Australia.
This is the second study investigating the role of conventional mechanical ventilator
breaths for alveolar recruitment during high‐frequency jet ventilation. We isolated the
effect of the peak inspiratory pressure of the conventional mechanical ventilation breath
in this study.
182
Abstract
Alveoli are recruited with conventional mechanical ventilator (CMV) breaths during high‐
frequency jet ventilation (HFJV). We assessed the effect of CMV breath peak inspiratory
pressure (PIP) on gas exchange, ventilator requirements and lung injury during HFJV.
Preterm lambs of anaesthetised ewes were delivered surgically at 128 d gestation
(term=150 d) and randomised to an unventilated control group (UVC) or one of 3
ventilated groups: HFJV; or HFJV with 5 CMV breaths/min to a PIPCMV either 5 cmH2O
below or above PIPHFJV (HFJV+CMVlow and HFJV+CMVhigh). Set PEEP was maintained at 8
cmH2O. PIPHFJV and FiO2 were adjusted to maintain PaCO2 45‐55 mmHg and SpO2 88‐95 %.
Lambs were euthanased after 2 h and a post mortem performed. FiO2 was lowest in the
HFJV+CMVhigh group from 60 min. Oxygenation index increased over time in the HFJV
group. In situ lung volume at 40 cmH2O and bronchoalveolar lavage (BAL) protein content
and inflammatory cell count was higher in all the ventilated groups compared to UVC. BAL
neutrophil count was higher in the HFJV+CMVlow group compared to UVC. Lung tissue IL‐6
mRNA was higher in HFJV+CMVlow compared to UVC. The most physiological benefit with
the least evidence of harm was apparent in the HFJV+CMVhigh group of preterm lambs.
183
Introduction
High‐frequency jet ventilation (HFJV) is a novel mode of high‐frequency ventilation
offering the potential for lung protective low tidal volume ventilation during the
management of respiratory distress syndrome (1, 2). In neonates, HFJV is delivered in
tandem with a conventional ventilator that provides positive end‐expiratory pressure
(PEEP), bias flow for spontaneous breaths, a passage for exhaled gases and a means of
delivering sigh breaths. Alveolar recruitment during HFJV is achieved by altering PEEP or
by delivering conventional mechanical ventilator (CMV) breaths (3). Neonatal clinical
protocols suggest that during HFJV CMV breaths should be delivered at very low rates (0‐3
breaths per minute), with more frequent CMV breaths (5‐10 breaths per minute) used to
recruit collapsed alveoli (4). Despite the widespread use of such protocols in neonatal
units over the last few decades, studies that explore how these breaths should be
delivered are limited.
The peak inspiratory pressure (PIP) of CMV breaths delivered during HFJV will determine
whether or not HFJV breaths are interrupted during delivery of CMV breaths.
Conventional mechanical ventilator breaths delivered to a PIPCMV higher than the HFJV
breaths will interrupt the HFJV breaths while those delivered to a PIPCMV lower than the
HFJV breaths will allow the HFJV breaths to stack on top of the CMV breath, delivering
peak pressures in excess of those set on the ventilator and potential barotrauma.
However, at any given PIPHFJV, a higher PIPCMV will deliver a higher tidal volume, with the
potential of increased volutrauma compared to that resulting from a PIPCMV below the
184
PIPHFJV. Whether a series of CMV breaths that interrupt the HFJV breaths are more
harmful than those that don’t is unknown.
We aimed to investigate the effect of 2 different CMV breath PIPCMV settings during HFJV.
We hypothesised that during HFJV in a preterm lamb model of RDS, CMV breaths
delivered to a PIPCMV greater than the PIPHFJV would promote lung injury.
185
Materials and Methods
All animal procedures were approved by the University of Western Australia animal ethics
committee, according to the guidelines of the National Health and Medical Research
Council of Australia code of practice for the care and use of animals for scientific purposes
(5).
Animals, Instrumentation and Delivery
Anaesthesia was induced in ewes at 128‐130 days of gestation with an intravenous
injection of medetomidine (0.02 mg/kg, Pfizer Animal Health, U.S.A.) and ketamine (10 mg
kg‐1, Troy Laboratories, Australia) followed by a subarachnoid (spinal) injection of lidocaine
(3 mL, 20 mg mL‐1, Troy Laboratories, Australia). The fetus was exteriorised surgically and
intubated orally (4.5 mm cuffed tracheal tube, Portex Ltd. England). Lung fluid was
suctioned and intra‐tracheal surfactant (100 mg kg‐1: 25 mg of phospholipids mL‐1, Abbott
Laboratories, U.S.A.) instilled prior to delivery of the lamb. Unventilated controls (UVC,
negative controls: n=8) were euthanased (pentobarbitone 100 mg kg‐1 i.v. Jurox, Australia)
at delivery.
Postnatal care
Lambs were dried, weighed and randomised to one of three ventilation groups: HFJV only
(HFJV; n=8); HFJV with 5 CMV breaths delivered to a PIP 5 cmH2O below the HFJV PIP
(HFJV+CMVlow, n=8); or HFJV with 5 CMV breaths delivered to a PIP 5 cmH2O above the
HFJV PIP (HFJV+CMVhigh, n=8).
186
Propofol (0.1 mg kg‐1 min‐1, Norbrook Laboratories Ltd., Victoria, Australia) and
remifentanil (0.05 µg kg‐1 min‐1, Abbott Laboratories, U.S.A.) were infused continuously
through an umbilical vein for anaesthesia and analgesia. An umbilical arterial catheter was
sampled intermittently to assess gas exchange and acid‐base status. Rectal temperature
was monitored continuously and maintained at 38 – 39 °C (normothermic for newborn
lambs). Oxygenation Index (OI) was calculated as OI = 2
2
aO
100 xaw xFiO
P
P where FiO2 is
fractional inspired oxygen concentration, Paw is mean airway pressure measured by the
HFJV ventilator and PaO2 is partial pressure of oxygen in arterial blood.
High Frequency Jet Ventilation
HFJV (Life Pulse High Frequency Ventilator, Bunnell Inc., Salt Lake City, U.S.A.) coupled to a
pressure‐limited time‐cycled conventional ventilator (Dräger, Babylog 8000+, Drägerwerk
AG, Lübeck, Germany) was commenced immediately following delivery. Initial HFJV
settings were respiratory rate 420 breaths/min, PIPHFJV 30 cmH2O and inspiratory time (tI)
0.02 s. PIPHFJV was adjusted to target permissive hypercapnia (PaCO2 45‐55 mmHg) to a
maximum of 40 cmH2O. The initial FiO2 (0.4) was adjusted to maintain SpO2 88‐95 %.
CMV breaths were delivered to a PIP (PIPCMV) 5 cmH2O above or below PIPHFJV, and with a
rate of 0 or 5 breaths/min as per randomisation. PIPCMV was adjusted in parallel with
PIPHFJV to maintain the predefined low and high PIP strategy relative to the PIPHFJV. A PEEP
of 8 cmH2O and tI of 0.5 s, were maintained throughout the study.
187
After obtaining final measurements, the FiO2 was increased to 1.0 for 2 minutes. The
tracheal tube was occluded for 3 min to facilitate lung collapse before the lamb was
euthanased (pentobarbitone 100 mg kg‐1 i.v. Jurox, Australia).
Post‐mortem
The lung was exposed by thoracotomy, and an in situ deflation pressure volume curve was
obtained (6). The right upper lung lobe was inflation fixed (30 cmH2O) in formalin and
samples of the right lower lobe were snap frozen for molecular analyses. Bronchoalveolar
lavage (BAL) was performed on the left lung for cytology and protein analysis (7).
Differential cell counts were performed on cytospin samples of the BAL fluid stained with
Diff‐Quik (Fronine Lab Supplies, N.S.W., Australia). Immunohistochemical staining for
myeloperoxidase (MPO) was performed on 5 µm sections of lung tissue (8). Positive cells
were identified and quantified per field as number of cells per total cellular area (nm2)
using densitometry (Image‐Pro Plus version 4.5, Media Cybernetics, U.S.A.) by a blinded
observer (GCM). RNA was extracted from lung tissue and reverse transcribed to cDNA
(Bioscript, Bioline, N.S.W., Australia) for measurement of IL‐1β and IL‐6 mRNA expression
by qRT‐PCR (9).
Statistical Analyses
Kruskal‐Wallis one way analysis of variance was used to compare groups at specific time
points. The effect of time on ventilator requirements and physiological changes was
determined using two‐way repeated measure analysis of variance. Analyses were
188
performed using SigmaStat (Version 3.5, Systat Software Incorporated, U.S.A.) with p <
0.05 considered statistically significant. Data are expressed as mean (SEM) unless stated
otherwise.
189
Results
Baseline characteristics of lambs in each group were not different (Table 1).
Ventilator Settings
The PIPCMV was 5.3 (0.6) cmH2O higher than PIPHFJV throughout the study in the
HFJV+CMVhigh group and 3.8 (1.5) cmH2O lower than PIPHFJV in the HFJV+CMVlow group.
There were no differences in PIPHFJV, Paw,HFJV, ΔPHFJV and servo pressure between the
groups (Figure 1A, 1B, 1C and 1D).
Gas Exchange
The target SpO2 was achieved in all groups within the first 10 min and maintained for the
duration of the ventilation period. FiO2 was lower in the HFJV+CMVhigh group from 60 min
compared to the HFJV+CMVlow group (p<0.03) and from 90 min compared to the HFJV
group (p<0.03) (Figure 2A). The oxygenation index increased over time in the HFJV group
(p=0.02) but did not change over time in either of the HFJV+CMV groups (Figure 2B).
The target PaCO2 was achieved within 30 min and maintained in all groups (Figure 2C). The
HFJV+CMVhigh group tended to have PaCO2 at the higher end of the target range but it was
not different to the other ventilated groups. There were no differences in arterial pH
(Figure 2D).
Static lung compliance
190
HFJV+CMVlow (p<0.02) and HFJV+CMVhigh (p<0.01) were more compliant than the HFJV
group as determined by a post‐mortem deflation static pressure‐volume curve. All
ventilated groups had a higher static compliance compared to the UVC group. (Figure 3).
Bronchoalveolar Lavage Fluid
The total protein concentration of BAL fluid was higher in each of the ventilated groups
compared to the UVC group, however there were no significant differences between the
ventilatory strategies. There were more inflammatory cells in the BAL fluid from each of
the ventilated groups compared to the UVC group (p<0.01). There were more neutrophils
in the HFJV+CMVlow group compared to UVC (p<0.05) (Table 2).
Lung Tissue
The number of MPO positive cells in the lung tissue of ventilated groups was comparable
between all groups (Table 2). The mRNA expression of IL‐1β was similar between all the
ventilated groups. The mRNA expression of IL‐6 was higher in HFJV+CMVlow compared to
the UVC group (Table 2).
191
Discussion
While the delivery of CMV breaths during HFJV is advocated to recruit alveoli, there are
few data to justify the selection of CMV breath parameters. CMV breaths delivered during
HFJV improved static lung compliance and in the HFJV+CMVhigh group this was associated
with decreased oxygen requirements and a consistent trend towards lower expression of
pro‐inflammatory markers. The use of a PIPCMV higher than the set PIPHFJV appeared more
protective than a PIPCMV lower than the set PIPHFJV. These findings suggest that in the
setting of atelectatic lungs at the initiation of ventilation in preterm lambs with a constant
PEEP, the inclusion of CMV breaths may improve lung volume recruitment. Of the two
strategies including CMV breaths, the strategy using PIPCMV above PIPHFJV appeared to
have the greatest physiological benefit with the least evidence of harm, contrary to our
hypothesis that this approach may invoke barotrauma.
Alveolar recruitment during HFJV can be achieved by adjusting PEEP, delivering occasional
CMV breaths, or both approaches in parallel. PEEP facilitates alveolar stabilisation and
avoidance of collapse. PEEP may also contribute to recruitment after achievement of
partial aeration if PEEP exceeds the opening pressure of atelectatic lung units, or by
assisting airway patency through the action of alveolar‐airway attachments in areas of
recruited lung. The CMV breaths open the lung primarily by exposing the distal alveoli to a
pressure greater than the opening pressure and by the delivery of recruiting volumes. The
PIPCMV determines the delivered volume of inflation above the opening pressure for a
given lung impedance. If the delivered PIPCMV and consequently the delivered tidal volume
192
are excessive, the cost of alveolar recruitment from CMV breaths may outweigh any
benefit of more rapid alveolar recruitment.
We compared HFJV+CMVlow and HFJV+CMVhigh with HFJV alone to investigate the
cost:benefit ratio for inclusion of CMV breaths, and size of those conventional breaths for
effective alveolar recruitment during HFJV. If PIPCMV is above PIPHFJV, the HFJV breaths are
interrupted for the duration of the CMV breath. Provided the PIPCMV is above the opening
pressure of the lung, this breath will act to recruit lung volume. Although this approach
(PIPCMV > PIPHFJV) has been the preferred clinical strategy, CMV breaths may be injurious if
they induce excessive parenchymal stretch (10, 11). As PIPHFJV is primarily determined by
the need to achieve satisfactory clearance of CO2, the situation often arises whereby
PIPHFJV is increased significantly above PIPCMV. In this scenario, (PIPCMV < PIPHFJV), the
phenomenon of ‘HFJV‐stacking’ is observed such that the peak pressure in the central
airways may be higher than either the preset PIPCMV or PIPHFJV. However, as the HFJV
waveform is delivered at high velocity, the PIPHFJV is substantially attenuated such that the
absolute PIP during an HFJV stacked CMV breath is usually only marginally above the
preset PIPHFJV (unpublished observations). Thus it is unlikely that the distal alveoli are
exposed to excessive peak pressures. Therefore, the most important proviso is that the
PIPCMV remains higher than the opening pressure of the lung, such that it recruits lung
volume, without being so high that it promotes cyclic overdistension. As volutrauma is a
more important determinant of lung injury than barotrauma (12), we hypothesised that
the HFJV+CMVhigh strategy would be more injurious than a HFJV+CMVlow approach.
193
Whereas we observed differences in oxygenation outcome variables, there were no
intergroup differences in the pro‐inflammatory markers between the ventilated groups.
There was however a consistent trend towards less injury in the HFJV+CMVhigh group in all
pro‐inflammatory markers studied compared to the HFJV and HFJV+CMVlow group.
Without accurate measurements of lung volume or CMV breath tidal volume (inaccurately
reported by the conventional ventilator during HFJV), we were unable to determine
whether this trend towards less injury in the HFJV+CMVhigh group is a consequence of
interrupted HFJV breaths, or more effective CMV breath related volume recruitment.
Some variability in delivered tidal volumes has beneficial effects in preterm lambs as
lambs managed with a variable ventilation strategy had improved in vivo lung compliance
without increased lung injury (J Pillow, unpublished data). Intermittent delivery of
relatively large breaths to a relatively high pressure is less likely to facilitate alveolar
recruitment more so than uniform breaths delivered repeatedly. The HFJV+CMVhigh group
in this study is likely to have received the most variable set of breaths over 2 h. This
variation may have contributed to the favourable results for this group.
We deliberately kept PEEP constant for the duration of this study to isolate the effect of
including CMV breaths and the magnitude of those breaths on oxygenation, ventilation
and lung injury. PEEP during HFJV is used primarily to stabilise alveoli and prevent alveolar
derecruitment (13‐16) although it can contribute to lung volume recruitment (17). We
selected a PEEP of 8 cmH2O as previous studies (18) found a PEEP of 5 cmH2O insufficient
to stabilise alveoli (18). Nonetheless, the increased OI in the HFJV group and the
comparable trend in the HFJV+CMV groups over the 2 h suggests progressive atelectasis
194
and inadequate PEEP stabilisation of the lung at end expiration. In clinical practice, an
increment in PEEP as PIPHFJV and ∆PHFJV are weaned may be essential to avoid alveolar
collapse.
In earlier studies we had observed the lower inflection point of similar gestation preterm
lambs as 20‐25 cmH2O (19) and hence expected that an initial PIPCMV of 25 cmH2O (in the
HFJV+CMVlow group) would be above the critical opening pressure. Nonetheless, it seems
likely that the HFJV+CMVlow group generated insufficient pressures and tidal volumes to
achieve effective recruitment and hence to provide the same benefit as the HFJV+CMVhigh
group.
We chose inflammatory markers of ventilation induced lung injury that typically increase
within 120 min (20‐23), and confirmed previous observations using CMV strategies that
any ventilation, even using HFJV alone, increases inflammatory markers in the premature
lung (17, 24). With only 8 lambs per group we cannot exclude a type II statistical error for
detection of significant differences between the ventilated groups. Even so, the lack of
significant differences between the different ventilated groups suggests that such
differences, if present, are likely to have minimal significance over a 2 h ventilation period.
Nonetheless, the trends towards increased BAL protein and inflammatory cells, and lung
tissue MPO and interleukin expression in the HFJV+CMVlow group compared to other
ventilated groups and the UVC group are suggestive of increased lung injury in this group.
A possible explanation may be the trend to lower mean airway pressures in the
HFJV+CMVlow group compared to the other two ventilated groups, resulting from the fall
195
in ΔP over the study period. In clinical practice, a drop in mean airway pressure with
weaning of the PIPHFJV could be offset by increases in PEEP, however by design, PEEP was
maintained at a constant level throughout the study duration, in all groups.
A higher static compliance in the HFJV+CMVhigh group was expected given the improved
oxygenation and likely increased lung volume in vivo protecting the lung from injury.
Whereas the OI increased over time in the HFJV group, it is unlikely that there was
significant alveolar recruitment in either the HFJV only or the HFJV+CMVlow group as with
the exception of the 15 min time point, their oxygenation indexes did not differ
subsequently and FiO2 remained elevated throughout the study. It is intriguing therefore,
that the static lung compliance was lower in the HFJV group compared to the
HFJV+CMVlow group given the trends towards increased lung injury in the latter group.
One explanation for this finding may be that the intermittent CMV breaths in the
HFJV+CMVlow group stimulated surfactant production and release. Further studies to
quantify surfactant protein mRNA may help to elucidate the answer to this question.
In summary, CMV breaths delivered to a PIPCMV 5 cmH2O above the PIPHFJV provided the
most physiological benefit with the least evidence of harm over a 2 h period following
initiation of ventilation in preterm lambs. Our results support the judicious use of
infrequent, but appropriately targeted, CMV breaths for alveolar recruitment during
initiation of HFJV in acute neonatal respiratory distress syndrome. Further studies
investigating CMV breath parameters during HFJV in the target patient population are
indicated.
196
Acknowledgements
Surfactant was donated by Abbott Australia. The Life Pulse High Frequency Ventilators
were supplied on long‐term loan by Bunnell Incorporated. We would like to express our
sincere appreciation to the members of the Ovine Research Group, Ilias Nitsos, Carryn
McLean, Richard Dalton and Yong Song for technical assistance and JRL Hall and Co. for
provision and early antenatal care of the ewes.
197
Tables
Table 1: Baseline lamb data
UVC HFJV HFJV+CMVlow HFJV+CMVhigh
n (male) 5 (3) 8 (5) 8 (3) 8 (4)
Birth weight (kg) 2.6 (0.3) 2.6 (0.6) 2.8 (0.6) 3.1 (0.3)
Gestational age (d) 129.6 (0.5) 129.1 (0.8) 129.1 (0.6) 129.5 (0.5)
Cord pH 7.11 (0.18) 7.15 (0.09) 7.09 (0.04) 7.11 (0.13)
Cord PaCO2 (mmHg) 88.9 (28) 77.1 (14.3) 99.3 (24.1) 84.0 (37.0)
Term = 150 d. UVC = unventilated control; HFJV = high‐frequency jet ventilation
(HFJV) alone; HFJV+CMVlow = HFJV with 5 conventional mechanical ventilation
(CMV) breaths/min to a peak inspiratory pressure 5 cmH2O below HFJV peak
inspiratory pressure; HFJV+CMVhigh = HFJV with 5 CMV breaths/min to a peak
inspiratory pressure 5 cmH2O above HFJV peak inspiratory pressure. Values are
mean (SD). Cord samples were collected from the umbilical artery.
198
Table 2: Post‐mortem inflammatory markers
UVC HFJV HFJV+CMVlow HFJV+CMVhigh
BAL fluid
Protein (mg mL‐1) 153.0 (85.9) 209.8 (90.5)* 353.3(212.1)* 273.(169.7)*
Total Inflammatory Cells
(x 106 kg‐1) 1.9 (0.8) 43.0 (13.9)* 90.2 (28.3)* 36.8 (15.0)*
Neutrophils (x 106 kg‐1) 0.5 (0.5) 14.2 (5.9) 34.7 (18.1)* 12.1 (6.4)
Mononuclear cells (x 106 kg‐1) 1.4 (0.4) 25.5 (11)* 53 (33.2)* 24.5 (10.9)*
Lung Tissue
MPO positive cells (cells (nm2)‐1) 0 (0, 1.5) 0.8 (0, 14.2) 1.1 (0, 4.4) 2.5 (0, 6.8)
IL‐1β (fold change) 1.0 (0.5, 1.4)
4.6 (1.6, 8.8)
5.6 (2.1, 9.4)
2.4 (1.7, 8.1)
IL‐6 (fold change) 1.0(0.7, 1.5)
19.4(1.0, 29.8)
21.5*(4.6, 38.1)
10.7 (6.9, 22.0)
*p<0.05 compared to UVC. Values are Mean (SD) or median (25, 75 centile).
199
Figure 1
200
Figure 2
201
Figure 3
202
Figure Legends
Figure 1 Ventilator Parameters: A: High‐frequency jet ventilation inspiratory pressure
(PIPHFJV) (cmH2O)was adjusted to target PaCO2 45‐55 mmHg; B: HFJV Mean Airway
Pressure (cmH2O); C: HFJV Delta P (HFJVPIP ‐ HFJVPEEP) (cmH2O); D: HFJV Servo Pressure
(psi). Grey diamond = HFJV, open circle = HFJV+CMVlow, solid circle = HFJV+CMVhigh.
Figure 2 Gas Exchange: A: FiO2 * p<0.03 HFJV+CMVhigh compared to HFJV+CMVlow, # p
<0.03 HFJV+CMVhigh compared to HFJV B: Oxygenation Index * p = 0.02 time 120
compared to time 5 for HFJV C: PaCO2 (mmHg); D: pH. Grey diamond = HFJV, open circle =
HFJV+CMVlow, solid circle = HFJV+CMVhigh.
Figure 3 Pressure‐Volume Curve: # p<0.02 HFJV+CMVhigh compared to HFJV, ## p<0.02
HFJV+CMVlow compared to HFJV, * p<0.01 compared to unventilated controls (UVC). Grey
diamond = HFJV, open circle = HFJV+CMVlow, solid circle = HFJV+CMVhigh, open triangle =
UVCs.
203
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Cell Mol Physiol 296:L510‐L518
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207
7 Alveolar Recruitment with Five or Twenty Conventional
Mechanical Ventilator Breaths per minute during High‐frequency
Jet Ventilation in Preterm Lambs.
Gabrielle C Musk1, Graeme R Polglase2 and J Jane Pillow 1.
1School of Women’s and Infants’ Health, University of Western Australia, Perth, Australia.
2The Ritchie Centre, Monash Institute of Medical Research, Monash University,
Melbourne, Australia.
3Centre for Neonatal Research and Education, University of Western Australia, Perth,
Australia.
This is the third study investigating the role of conventional mechanical ventilator breaths
for alveolar recruitment during high‐frequency jet ventilation. We isolated the effect of
conventional ventilator breath frequency in this study.
208
Abstract
Alveoli are recruited with conventional mechanical ventilator (CMV) breaths during high‐
frequency jet ventilation (HFJV). We assessed the impact of CMV breath frequency on gas
exchange, ventilator requirements and lung injury during HFJV.
Preterm lambs of anaesthetised ewes were delivered surgically at 128 d gestation
(term=150 d) and randomised to an unventilated control group (UVC) or one of 3
ventilated groups including HFJV alone or HFJV with either 5 or 20 CMV breaths/min to a
PIPCMV 5 cmH2O below PIPHFJV (HFJV+CMV5 or HFJV+CMV20). Set PEEP was maintained at 8
cmH2O. PIP and FiO2 were adjusted to maintain PaCO2 45‐55 mmHg and SpO2 88‐95 %.
Lambs were euthanased after 2 hours and a post mortem performed.
Physiological variables and ventilator requirements did not differ between groups.
Compared to other ventilated groups the HFJV+CMV20 group had higher measured PEEP
(auto PEEP) from 15 min and higher FiO2 from 30 min. In situ lung volume at 40 cmH2O
was higher in the HFJV+CMV5 group compared to all other groups. Bronchoalveolar lavage
(BAL) fluid inflammatory cell count was higher in HFJV+CMV groups and BAL protein
concentration was higher in all ventilated groups compared to UVCs. Surprisingly, lung
tissue IL‐1β and IL‐6 mRNA was lowest in HFJV+CMV20 compared to other ventilated
groups. Whereas the improved static compliance in the HFJV+CMV5 group suggests
potential benefit of low‐rate CMV breaths during HFJV during the initiation of HFJV in
acute neonatal respiratory distress syndrome, the mixed inflammatory outcomes in the
209
HFJV+CMV groups suggests further studies to clarify the optimal frequency of CMV
breaths are required.
210
Introduction
High‐frequency jet ventilation (HFJV) is considered a lung protection ventilation strategy
suitable for ventilatory support of preterm infants with respiratory distress syndrome
(RDS) (1, 2). A high‐frequency jet ventilator is set up in tandem with a conventional
ventilator that provides positive end‐expiratory pressure (PEEP), bias flow for
spontaneous breaths, a passage for exhaled gases and a means of delivering sigh breaths.
This tandem arrangement facilitates alveolar recruitment manoeuvers during HFJV which
may be achieved by altering PEEP or by delivering conventional mechanical ventilator
(CMV) breaths (3). Neonatal clinical protocols suggest that during routine HFJV, CMV
breaths should be delivered at very low rates (0‐3 breaths per minute), with more
frequent CMV breaths (5‐10 breaths per minute) used to recruit collapsed alveoli (4).
Whereas protocols are based on the presumption that excessive CMV breaths are
injurious, there are no experimental data that define the optimal frequency for CMV
breaths during HFJV in the setting of acute neonatal RDS.
Compared to the small volume HFJV breath, CMV breaths are more likely to cause cyclic
stretch of the lung parenchyma and contribute to volutrauma (5). The frequency of CMV
breaths during HFJV may therefore influence volume recruitment. However, whereas
occasional sigh breaths may promote recruitment, repetitive cyclic stretch of the lung
parenchyma may also promote injury. The CMV breath parameters that must be selected
include the inspiratory time (tI), the peak inspiratory pressure (PIP) and the respiratory
frequency.
211
We aimed to investigate the effect of 2 different CMV breath frequencies during HFJV. We
hypothesised that during HFJV in a preterm model of RDS, 20 CMV breaths/min would be
more injurious to the lung than 5 CMV breaths/min.
212
Materials and Methods
All animal procedures were approved by the University of Western Australia animal ethics
committee, according to the guidelines issued by the National Health and Medical
Research Council of Australia (6).
Animals, Instrumentation and Delivery
Pregnant ewes at 128‐130 days of gestation were anaesthetised with an intravenous
injection of medetomidine (0.02 mg/kg, Pfizer Animal Health, U.S.A.) and ketamine (10 mg
kg‐1, Troy Laboratories, Australia) followed by a subarachnoid (spinal) injection of lidocaine
(3 mL, 20 mg mL‐1, Troy Laboratories, Australia). The fetus was exteriorized surgically and
intubated orally (4.5 mm cuffed tracheal tube, Portex Ltd. England). Lung fluid was
suctioned and intra‐tracheal surfactant (100 mg kg‐1: 25 mg of phospholipids mL‐1, Abbott
Laboratories, U.S.A.) instilled prior to delivery of the lamb. Unventilated controls (UVC,
negative controls: n=8) were euthanased at delivery with pentobarbitone (100 mg kg‐1 i.v.
Jurox, Australia).
Postnatal care
Lambs were dried, weighed and randomised to one of three ventilation groups: HFJV only
(HFJV; n=8); HFJV with 5 or 20 CMV breaths delivered to a PIP 5 cmH2O below the PIPHFJV
(HFJV+CMV5, n=8; HFJV+CMV20, n=8).
213
Propofol (0.1 mg kg‐1 min‐1, Norbrook Laboratories Ltd., Victoria, Australia) and
remifentanil (0.05 µg kg‐1 min‐1, Abbott Laboratories, U.S.A.) were infused continuously
through an umbilical vein for anaesthesia and analgesia. An umbilical arterial catheter was
sampled intermittently to assess gas exchange and acid‐base status. Rectal temperature
was monitored continuously and maintained between 38° and 39° C (normothermic for
newborn lambs). Oxygenation Index (OI) was calculated as OI = 2
2
aO
100 x aw x FiO
P
P where FiO2
is fractional inspired oxygen concentration, Paw is mean airway pressure measured by the
jet ventilator and PaO2 is partial pressure of oxygen in arterial blood.
High Frequency Jet Ventilation
HFJV (Life Pulse High Frequency Ventilator, Bunnell Inc., Salt Lake City, U.S.A.) coupled to a
pressure‐limited time‐cycled conventional ventilator (Dräger, Babylog 8000+, Drägerwerk
AG, Lübeck, Germany) was commenced immediately following delivery. Initial HFJV
settings were: respiratory rate 420 breaths/min; PIPHFJV 30 cmH2O; and inspiratory time (tI)
0.02 s. PIPHFJV was adjusted to achieve permissive hypercapnia (PaCO2 45‐55 mmHg) to a
maximum of 40 cmH2O. The initial FiO2 (0.4) was adjusted to maintain SpO2 88‐95 %.
CMV breaths were delivered to a PIPCMV 5 cmH2O below the PIPHFJV, and with a rate of 0, 5
or 20 breaths/min as per randomisation. PIPCMV was adjusted in parallel with PIPHFJV to
maintain PIPCMV 5 cmH2O below the PIPHFJV. A PEEP of 8 cmH2O and tI of 0.5 s for CMV
breaths, were maintained throughout the study.
214
After final measurements were obtained, the FiO2 was increased to 1.0 for 2 minutes,
after which the tracheal tube was occluded for 3 min to facilitate lung collapse before the
lamb was euthanased (100 mg kg‐1 i.v. Jurox, Australia).
Post‐mortem
The lung was exposed by thoracotomy, and an in situ deflation pressure volume curve was
obtained (7). The right upper lung lobe was inflation fixed (30 cmH2O) in formalin and
samples of the right lower lobe were snap frozen for molecular analyses of IL‐1β and IL‐6
mRNA expression by qRT‐PCR (8). Bronchoalveolar lavage (BAL) was performed on the left
lung for cytology and protein analysis (9). Differential cell counts were performed on
cytospin samples of the BAL fluid stained with Diff‐Quik (Fronine Lab Supplies, N.S.W.,
Australia). Immunohistochemical staining for myeloperoxidase (MPO) was performed on 5
µm sections of lung tissue (10). Positive cells were identified and quantified per field as
number of cells per total cellular area (nm2) using densitometry (Image‐Pro Plus version
4.5, Media Cybernetics, U.S.A.) by a blinded observer (GCM).
Statistical Analyses
Kruskal‐Wallis one way analysis of variance was used to compare groups at specific time
points. The effect of time on ventilator requirements and physiological changes was
determined using two‐way repeated measure analysis of variance. Analyses were
performed using SigmaStat (Version 3.5, Systat Software Incorporated, U.S.A.) with p<0.05
215
considered statistically significant. Data are expressed as mean (SEM) unless stated
otherwise.
216
Results
Baseline characteristics of lambs in each group, including weight and cord blood gas
status, were not different (Table 1).
Ventilator Settings
The PIPHFJV, Paw,HFJV, ΔPHFJV and servo pressure were similar between the 3 ventilated
groups during the 2 h ventilation period (Figure 1A, 1B, 1C and 1D). The measured PEEP
was higher than set PEEP in HFJV+CMV20 animals at all time points (p=0.03; Figure 1E).
Gas Exchange
The target SpO2 was achieved in all groups within the first 10 minutes and maintained for
the duration of the ventilation period (data not shown). PaO2 was not different between
groups for the duration of the study (data not shown). From 90 minutes, FiO2 and OI were
significantly higher in the HFJV+CMV20 group compared to HFJV+CMV5 and HFJV (Figure
2A and 2B). The target PaCO2 was achieved within 15 minutes and maintained in all
groups (Figure 2C). There were no differences in arterial pH (Figure 2D).
Static lung compliance
The lungs of animals in the HFJV+CMV5 group were more compliant compared to HFJV
and HFJV+CMV20 (p<0.02) as determined by a post‐mortem static pressure‐volume curve.
As expected, unventilated control lambs had the lowest compliance compared to all
ventilated groups (p<0.01; Figure 3).
Bronchoalveolar Lavage Fluid
217
The total protein concentration of BAL fluid was higher in all the ventilated groups
compared to the UVC group. The total inflammatory cell counts, and the differential
neutrophil counts, were higher in the BAL fluid from HFJV+CMV5 and HFJV+CMV20 groups
compared to the UVC group (p<0.01; Table 2) but were not different to HFJV alone. The
differential mononuclear cell counts were higher in each of the ventilated groups
compared to the UVC group.
Lung Tissue
The number of MPO positive cells in the lung tissue of ventilated groups was low and
comparable between all groups (Table 2). The expression of IL‐1β mRNA was higher in all
the ventilated groups compared to HFJV+CMV20 (p<0.05). The expression of IL‐6 mRNA
was higher in the HFJV+CMV5 group compared to the UVC group and higher in both HFJV
alone and HFJV+CMV5 compared to HFJV+CMV20 (p<0.05) (Table 2).
218
Discussion
While the delivery of CMV breaths during HFJV is advocated to recruit alveoli, there are
few studies justifying the selection of CMV breath parameters, including breath frequency.
In the current study, we showed that 20 CMV breaths/min increased oxygen requirements
and decreased static lung compliance compared to a strategy using 5 CMV breaths/min,
but resulted in less up‐regulation of pro‐inflammatory markers in the lung.
PIPHFJV was comparable between groups and between the commencement and the end of
the study within each group. The PIPCMV was maintained 5 cmH2O below the PIPHFJV in this
study. Recent work from this group compared PIPCMV in relation to PIPHFJV and found that
the most physiological benefit, with the least evidence of harm, was apparent if the PIPCMV
was 5 cmH2O above the PIPHFJV when 5 CMV breaths/min were delivered during HFJV
(unpublished data – see Chapter 6). These results were not available at the time the
current study was performed, and it may be that we have hindered our ability to
demonstrate a difference between our HFJV+CMV groups by using insufficient CMVPIP to
recruit the lung effectively in either HFJV alone, or HFJV+CMV5.
ΔPHFJV was not different during the study but there was a trend towards a higher ΔPHFJV in
the HFJV only group for the second hour of the study. Given that during high‐frequency
ventilation, ventilation is proportional to the product of respiratory frequency and tidal
volume (VT)2 (11), this finding demonstrates how little the CMV breaths contribute to
overall minute ventilation during HFJV. Even when 20 CMV breaths/min were delivered,
219
the contribution to overall minute volume did not translate to a lower ΔPHFJV. This trend
may be due to a low VT of the CMV breaths and despite delivering 20 breaths/min, the
impact on PaCO2 was negligible.
PEEP during HFJV is provided by the conventional ventilator and used primarily to stabilise
alveoli and prevent alveolar derecruitment (11‐14). To avoid confounding the
interpretation of CMV breath strategy on outcomes, PEEP was kept constant for the
duration of this study. We selected a PEEP of 8 cmH2O as a previous study found a PEEP of
5 cmH2O insufficient to stabilise alveoli (15). Despite our attempts to maintain a constant
PEEP between and within groups there was a negative difference between PEEP set on the
conventional ventilator and PEEP measured by the jet ventilator for the duration of the
study in the HFJV+CMV20 group. This discrepancy is indicative of auto PEEP, or gas
trapping, and might be expected at higher CMV breath frequencies. This auto PEEP should
translate to a higher Paw but there was no difference in Paw between the groups. Gas
trapping, however, will compromise oxygenation and may explain the higher FiO2
requirements and OI in the HFJV+CMV20 group. The value for Paw on the high‐frequency
jet ventilator is an estimate of the mean pressure at the tip of the tracheal tube and may
not reflect intrapulmonary pressure. It is therefore possible that mean pressure at the
parenchymal level was higher and compromised pulmonary blood flow and oxygenation.
We assessed oxygenation by recording the FiO2 required to achieve a target SpO2 and by
calculating OI. Paw is a determinant of oxygenation (16), but was similar for all groups,
leaving differences in oxygenation parameters attributable to variables other than Paw.
220
There was a higher inspired oxygen requirement and OI in the HFJV+CMV20 group which
suggests either inferior alveolar recruitment, or shunting due to impaired pulmonary
blood flow and increased pulmonary vascular resistance. Although Paw was not different
between groups, it is estimated by the high‐frequency jet ventilator at the tip of the
tracheal tube and it is conceivable that inadvertent PEEP translated to a higher mean
pressure at the parenchyma level that may have impaired pulmonary blood flow and
oxygenation.
In the absence of a clear advantage of higher CMV breath frequency for lung volume
recruitment and arterial oxygenation, we expected the HFJV+CMV20 approach to be
associated with increased potential for lung injury due to increased frequency of distal
alveolar exposure to the PIPCMV (which is transmitted along the airways) and volutrauma
from the associated cyclic tidal volume. Despite choosing inflammatory markers of
ventilation injury that typically increase within 120 min (17‐20), the results provided
conflicting evidence with a trend towards higher BAL protein and inflammatory cell counts
in the HFJV+CMV groups compared to the HFJV only group, but a higher compliance in the
HFJV+CMV5 group compared to either the HFJV+CMV20 and the HFJV only group on the
post‐mortem static pressure‐volume curve. The finding of reduced pro‐inflammatory
markers in the lung tissue for the HFJV+CMV20 group was similarly unexpected.
Inadvertent PEEP may have protected the HFJV+CMV20 lungs from atelectatrauma at the
expense of impaired oxygenation.
221
The limitations of this study may contribute to the lack of significant differences in lung
injury markers. With 8 animals per group we cannot exclude a type II statistical error for
detection of small significant differences between groups. It is also possible that the
strategies were not injurious enough to elicit a strong inflammatory response. The
combination of endogenous surfactant administered immediately after delivery, the short
duration of the study and a modestly high level of PEEP throughout the study may have
limited injury in all three ventilated groups. Furthermore, we have a single snapshot of
inflammation in the lungs and captured a higher BAL neutrophil population in the
HFJV+CMV groups along without an increase in IL‐1β and IL‐6 mRNA in the HFJV+CMV20
group. It is possible that a longer ventilation period may have resulted in an increase in
these cytokines in all groups.
In summary the use of a high CMV breath rate during HFJV strategy increased oxygen
requirements and decreased static lung compliance over a 2 h study compared to a low
CMV breath frequency strategy. The increased compliance of the post‐mortem static
deflation curve in the absence of a significant increase in pro‐inflammatory markers in the
low‐rate CMV breath HFJV strategy over HFJV alone supports the judicious use of
infrequent CMV breaths during initiation of HFJV in acute neonatal respiratory distress
syndrome. The lack of significantly enhanced oxygenation in the HFJV+CMV5 group
compared to HFJV alone suggests that specific targeting of the PIPCMV to achieve
recruitment may be essential to derive further benefit from inclusion of low‐rate CMV
breaths during HFJV. Further studies investigating the use of CMV breaths during HFJV
222
that measure the effect of these breaths on lung volume, gas exchange and lung injury
during neonatal RDS are indicated.
223
Acknowledgements
Surfactant was donated by Abbott Australia. The Life Pulse High Frequency Ventilators
were supplied on long‐term loan by Bunnell Incorporated. We would like to express our
sincere appreciation to the members of the Ovine Research Group, Ilias Nitsos, Carryn
McLean, Richard Dalton and Yong Song for technical assistance and JRL Hall and Co. for
provision and early antenatal care of the ewes.
224
Tables
Table 1: Baseline lamb data
UVC HFJV HFJV+CMV5 HFJV+CMV20
n (male) 5 (3) 8 (5) 8 (3) 8 (4)
Birth weight (kg) 2.6 (0.3) 2.6 (0.6) 2.8 (0.6) 3.0 (0.5)
Gestational age (d) 129.6 (0.5) 129.1 (0.8) 129.1 (0.6) 129.2 (0.7)
Cord pH 7.11 (0.18) 7.15 (0.09) 7.09 (0.04) 7.14 (0.12)
Cord PaCO2 (mmHg) 88.9 (28) 77.1 (14.3) 99.3 (24.1) 86.0 (22.3)
Term = 150 d. UVC = unventilated control; HFJV = high‐frequency jet
ventilation (HFJV) alone; HFJV+CMV5 = HFJV with 5 conventional
mechanical ventilation (CMV) breaths/min to a peak inspiratory pressure
5 cmH2O below HFJV peak inspiratory pressure; HFJV+CMV20 = HFJV with
20 CMV breaths to a peak inspiratory pressure 5 cmH2O below HFJV peak
inspiratory pressure. Values are mean (SD). Cord samples were collected
from the umbilical artery.
225
Table 2: Post‐mortem inflammatory markers
UVC HFJV HFJV+CMV5 HFJV+CMV20
BAL fluid
Protein (mg mL‐1) 83.2 (23.) 306.5 (27)* 437.4 (105)* 411.4 (37)*
Total Inflammatory Cells
(x 106 kg‐1) 1.93 (0.8) 43.0 (13.9) 90.2 (28.3)* 81.3 (30.6)*
Neutrophils (x 106 kg‐1) 0.5 (0.5) 14.2 (5.9) 34.7 (18.1)* 39.3 (18.1)*
Mononuclear cells (x 106 kg‐1) 1.4 (0.4) 25.5 (11)* 53 (33.2)* 41.8 (15.2)*
Lung Tissue
MPO positive cells (cells (nm2)‐1) 0 (0, 1.5)
0.8 (0, 14.2)
1.1 (0, 4.4)
3.0 (0, 7.0)
IL‐1β (fold change) 1.0 (0.5, 1.4)
4.6† (1.6, 8.8)
5.6† (2.1, 9.4)
0.7 (0.2, 1.4)
IL‐6 (fold change) 1.0 (0.7, 1.5)
19.4† (1.0, 29.8)
21.5*† (4.6, 38.1)
2.5 (0.6, 5.7)
*p<0.05 compared to UVC, †p<0.05 compared to HFJV+CMV20. Values are Mean (SEM) or
median (25, 75 centile).
226
Figure 1
227
Figure 2
228
Figure 3
229
Figure Legends
Figure 1 Ventilator Parameters: A: High‐frequency jet ventilation peak inspiratory
pressure (PIPHFJV) (cmH2O)was adjusted to target PaCO2 45‐55 mmHg; B: HFJV Mean
Airway Pressure (cmH2O); C: HFJV Delta P (HFJVPIP ‐ HFJVPEEP) (cmH2O); D: HFJV Servo
Pressure (psi); E: The difference between set PEEP and measured PEEP (cmH2O). *p<0.05
HFJV+CMV20 compared to HFJV+CMV5 and HFJV. Grey diamond = HFJV, open circle =
HFJV+CMV5, solid circle = HFJV+CMV20.
Figure 2 Gas Exchange: A: FiO2 * p<0.03 HFJV+CMV20 compared to HFJV+CMV5, # p<0.03
HFJV+CMV20 compared to HFJV; B: Oxygenation Index * p=0.04 HFJV+CMV20 compared to
HFJV+CMV5, # p=0.05 HFJV+CMV20 compared to HFJV; C: PaCO2 (mmHg); D: pH. Grey
diamond = HFJV, open circle = HFJV+CMV5, solid circle = HFJV+CMV20.
Figure 3 Pressure‐Volume Curve: * p<0.02 HFJV+CMV5 compared to HFJV, # p<0.02
HFJV+CMV5 compared to HFJV+CMV20, ** p<0.01 unventilated controls (UVC) compared to
all ventilated groups. Grey diamond = HFJV, open circle = HFJV+CMV5, solid circle =
HFJV+CMV20, open triangle = UVCs.
230
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frequency oscillatory ventilation. Journal of Maternal‐Fetal and Neonatal Medicine
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Kramer BW 2007 Pulmonary and systemic endotoxin tolerance in preterm fetal
sheep exposed to chorioamnionitis. J Immunol 179:8491‐8499
11. Courtney SE, Asselin JM 2006 High‐frequency jet and oscillatory ventilation for
neonates: Which strategy and when? Respir Care Clin N Am 12:453‐467
12. Venegas J, Fredberg J 1994 Understanding the pressure cost of ventilation: Why
dose high‐frequency ventilation work? Critical Care Medicine 22:S49‐S57
13. Bass A, Gentile M, Heinz J, Craig D, Hamel D, Cheifetz I 2007 Setting positive end‐
expiratory pressure during jet ventilation to replicate the mean airway pressure of
oscillatory ventilation. Respiratory Care 52:50‐55
14. Crossley KJ, Morley CJ, Allison BJ, Polglase GR, Dargaville PA, Harding R, Hooper SB
2007 Blood gases and pulmonary blood flow during resuscitation of very preterm
lambs treated with antenatal betamethasone and/or curosurf: Effect of positive
end‐expiratory pressure. Pediatr Res 62:37‐42
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positive end‐expiratory pressure during high‐frequency jet ventilation improves
oxygenation and ventilation in preterm lambs. Pediatr Res 69:319‐324
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of neonatal respiratory care. Mosby Elsevier, Philadelphia, pp 232‐233
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233
8 A Comparison of High‐frequency Jet Ventilation and High‐
frequency Oscillatory Ventilation with Conventional Mechanical
Ventilation in Preterm Lambs
Gabrielle C Musk 1, Graeme R Polglase 1,2, J Bert Bunnell 3, Ilias Nitsos 1, David Tingay
4, J Jane Pillow 1,5
1 School of Women’s and Infants’ Health, The University of Western Australia, Perth,
Australia.
2 The Ritchie Centre, Monash Institute of Medical Research, Monash University,
Clayton, Australia.
3 Bunnell Inc, Salt Lake City, Utah, USA and Department of Bioengineering, University of
Utah, Salt Lake City, Utah, USA.
4 Murdoch Children’s Research Institute, Melbourne, Australia
5 Centre for Neonatal Research and Education, The University of Western Australia,
Perth, Australia
This is the final study in this thesis. We compared an optimal high‐frequency jet
ventilation strategy against a high‐frequency oscillatory ventilation and a gentle
conventional mechanical ventilation strategy in our preterm lamb model of respiratory
distress syndrome. The high‐frequency jet ventilation strategy was developed based
upon the results of the previous study chapters.
234
Abstract
Conventional mechanical ventilation (CMV), high‐frequency oscillatory ventilation
(HFOV) and high‐frequency jet ventilation (HFJV) are accepted ventilatory strategies
for respiratory distress syndrome (RDS) in preterm babies. We hypothesised these
strategies would successfully manage oxygenation and ventilation, but that HFJV and
HFOV would cause the least lung injury. Furthermore, we hypothesised that HFJV
would have the least impact on pulmonary blood flow. Preterm lambs of anaesthetised
ewes were instrumented, intubated and delivered by caesarean section after
intratracheal suction and instillation of surfactant. Each lamb was managed for 3 hours
according to a predetermined algorithm for ventilator support consistent with the
open lung approach. Pulmonary blood flow was measured continuously and pulsatility
index was calculated, while ventilatory parameters and arterial blood gases were
measured at intervals. At postmortem, in situ pressure‐volume deflation curves were
recorded, and bronchoalveolar lavage fluid and lung tissue were obtained to assess
inflammation. There was no difference in arterial oxygenation despite lower mean
airway pressure during CMV for most of the study. HFOV animals had a higher PaCO2
at multiple time points. The lack of significant differences in end systolic and end
diastolic PBF for the majority of the study, lung injury data and static lung compliance
demonstrate that in the absence of airleaks each of these strategies can be employed
in the clinical setting with a comparable pressure cost of ventilation.
235
Introduction
The lungs of preterm infants are vulnerable to ventilation induced lung injury (1).
Preterm newborns often require invasive ventilatory support which exposes them to
positive intrathoracic pressures, the risk of lung injury and compromised cardiac
output. The three contemporary ventilatory strategies most frequently used for
preterm babies with respiratory distress syndrome (RDS) are synchronised, volume‐
targeted/guaranteed conventional mechanical ventilation (CMV), high‐frequency jet
ventilation (HFJV) and high‐frequency oscillatory ventilation (HFOV).
A number of clinical trials have compared HFOV and CMV for the management of
preterm infants with acute RDS but the results are conflicting (2, 3). Cools et al (2010)
found in their systematic review and meta‐analysis HFOV and CMV equally effective (2)
while Bhuta and Henderson‐Smart (1998) found little difference other than a decrease
in new pulmonary airleak following treatment with HFOV (3). Variation in study design,
ventilator management and outcome measures between studies make it difficult to
substantiate a clear benefit of one strategy over another. Perhaps the greatest
limitation to these studies was failure to pursue a protective lung ventilation approach
which relies upon opening the lung and keeping it open to optimise ventilation and
perfusion (4, 5). HFJV and CMV have also been compared in a number of studies but
similar issues exist in that an open lung strategy was not always employed, or achieved
(6‐8).
A direct comparison of all three of these ventilatory strategies for neonatal RDS has
not been performed in a controlled setting. Each modality has unique features which
236
provide justification for its use in certain scenarios. A CMV strategy delivers breaths of
a similar size and at a similar frequency to spontaneous ventilation while high‐
frequency strategies deliver breaths smaller than dead space volume at frequencies
varying from 3‐20 Hz depending on the specific ventilator used. High‐frequency
ventilation offers potential lung protective ventilation (9‐14) as the low tidal volumes
reduce the risk of cyclic volutrauma. The airway pressure waveform is attenuated
along the airways during HFJV and HFOV which may decrease the pressure and flow
cost of ventilation induced lung injury (15).
There are important differences between HFJV and HFOV which give rise to potential
physiological advantages of one strategy over another: optimal mean airway pressure
(Paw) is lower during HFJV compared to HFOV because less of the respiratory cycle is
spent in inspiration; HFJV enhances mucociliary clearance by combining fast
inspirations with relatively slow, passive exhalations (I:E ratio may be as low as 1:12);
and the use of small tidal volume (VT) breaths during HFJV at low frequencies, in
combination with low inspiratory to expiratory ratios, make HFJV especially useful in
patients with gas trapping. Both strategies deliver high velocity and small VT breaths
that do not penetrate injured areas of lung with high resistance, allowing for
maturation and healing (15). HFOV, however, has an active expiratory phase which
makes it useful at higher respiratory frequencies. This feature is advantageous for
lungs that are poorly compliant and that have a higher corner frequency, as it
facilitates gas exchange but avoids unnecessary barotrauma (16). HFOV has been
investigated in and ex vivo and gas flow and gas exchange is more extensively
documented (16, 17). This extensive data from HFOV has facilitated its widespread use
237
in neonatal intensive care units. Furthermore, during HFOV, a conventional mechanical
ventilator is not required and a standard endotracheal tube adaptor is suitable.
Early recruitment of the functional residual capacity immediately after birth may
facilitate the commencement of ventilation on the deflation limb of the pressure‐
volume curve. The delivery of a sustained inflation before commencing a particular
ventilation strategy has consistently decreased the requirement for subsequent
aggressive alveolar recruitment manoeuvres in both animal and human studies (18‐
20).
The aim of this study was to compare HFJV and HFOV with a moderate lung protective
CMV strategy in a preterm lamb model of RDS. We hypothesised that each strategy
would facilitate acceptable oxygenation and ventilation, but that the low delivered
tidal volumes of HFJV and HFOV would result in less evidence of ventilation induced
lung injury than a CMV strategy. Furthermore, based upon the findings of Polglase et al
(2008) (21) we hypothesised that compared to HFOV, HFJV would cause less reduction
of pulmonary blood flow due to a proportionately greater duration of the cycle spent
at the positive end expiratory pressure with less impairment of venous return.
238
Materials and Methods
All animal procedures were approved by the University of Western Australia animal
ethics committee, in accordance with guidelines of the National Health and Medical
Research Council of Australia (22).
Animals, Instrumentation and Delivery
Twin‐bearing date‐mated merino ewes were anaesthetised at 128‐130 d gestation
(term ≈ 150 d) with intramuscular xylazine (0.5 mg kg‐1, Troy Laboratories, N.S.W.,
Australia) and ketamine (20 mg kg‐1, Parnell Laboratories, N.S.W., Australia) and
intubated (7.5 mm cuffed tracheal tube, Portex Ltd. England). Maternal anaesthesia
was maintained with isoflurane in 100 % O2. The fetus was exteriorized via
hysterotomy and a right lateral thoracotomy was performed. A flow probe (4R,
Transonic Systems, Ithaca, NY) was positioned around the left pulmonary artery and a
catheter was inserted into the main pulmonary artery (21). The fetus was intubated
orally (4.5 mm cuffed tracheal tube, Portex Ltd. England), lung fluid was suctioned and
intra‐tracheal surfactant (100 mg kg‐1, Abbott Laboratories, U.S.A.) was administered
prior to delivery of the lamb. Unventilated controls (UVC; n=6) were euthanised
(pentobarbitone 100 mg kg‐1 i.v. Jurox, Australia) at delivery without instrumentation,
suctioning or surfactant. Remaining lambs were randomized to one of three ventilation
groups including conventional mechanical ventilation (CMV: n=6), high‐frequency jet
ventilation (HFJV; n=8) and high‐frequency oscillatory ventilation (HFOV; n=8) as
outlined below.
239
Ventilation
Instrumented lambs were dried, weighed. Functional residual capacity was established
by delivering 2 sustained inflations to 30 cmH2O (20 s and 10 s duration respectively)
with an infant T‐piece resuscitator (NeopuffTM, Fisher & Paykel Healthcare, Auckland,
New Zealand), immediately before ventilation was commenced using the assigned
ventilation strategy as detailed below. The initial FiO2 was 0.4 for all groups.
Conventional Mechanical Ventilation
Initial settings for positive pressure ventilation (PPV) with volume guarantee (VG)
included: respiratory rate 50 breaths/min; tI 0.5 s; VG 5 mL/kg; positive end‐expiratory
pressure (PEEP) 7 cmH2O; and a peak inspiratory pressure limit (PIPlimit) of 30 cmH2O
(Babylog 8000+, Drägerwerk, Lubeck, Germany). After 5 min, the VG was increased to 7
mL/kg and PIPlimit was increased to 40 cmH2O. FiO2 was altered to target SpO2 88‐94 %,
VT was altered to target PaCO2 45‐55 mmHg and PEEP was altered according to the
response of SpO2 to changes in FiO2 (Figure 1A).
High‐frequency Jet Ventilation
High‐frequency jet ventilation (Life Pulse™, Bunnell Inc., Salt Lake City, U.S.A.) coupled
to a pressure‐limited time‐cycled infant conventional ventilator (Babylog 8000+,
Drägerwerk, Lubeck, Germany) was commenced with the following initial settings:
respiratory rate 420 breaths/min (7 Hz); peak inspiratory pressure (PIPHFJV) 40 cmH2O;
PEEP 8 cmH2O; FiO2 0.4 and tI was fixed at 0.02 s. PIPHFJV was adjusted to achieve
permissive hypercapnia (PaCO2 45‐55 mmHg) to a maximum of 40 cmH2O. FiO2 was
240
altered to target SpO2 88‐94 % and CMV breaths were delivered according to the
protocol algorithm (Figure 1 B) to target lung volume recruitment.
High‐frequency Oscillatory Ventilation
High‐frequency oscillatory ventilation (3100A, Care Fusion, CA, U.S.A.) was
commenced as follows: frequency 12 Hz (720 breaths/min); Paw 16 cmH2O; tI 33 %
(tI:tE 1:2) and an amplitude (ΔP) of 30 cmH2O. The amplitude was adjusted to achieve
permissive hypercapnia (PaCO2 45‐55 mmHg) to a maximum of 50 cmH2O. The Paw
was adjusted to optimise oxygenation, preceding adjustments to FiO2 to target SpO2
88‐94 % (Figure 1 C).
Postnatal care
Propofol (0.1 mg/kg/min; Norbrook Laboratories Ltd., Victoria, Australia) and
remifentanil (0.05 µg/kg/min; Abbott Laboratories, U.S.A.) were infused continuously
through an umbilical vein for anaesthesia and analgesia. An umbilical arterial catheter
was used for intermittent sampling to assess gas exchange and acid‐base balance.
Rectal temperature was monitored continuously and maintained between 38° and 39°
C (normothermic for newborn lambs). Ventilator settings and physiological data were
recorded at intervals. After final measurements were obtained, the FiO2 was increased
to 1.0 for 2 min after which the tracheal tube was occluded for 3 min to facilitate lung
collapse prior to euthanasing the lamb (pentobarbitone 100 mg kg‐1 i.v. Jurox,
Australia).
241
Physiological Analyses
Continuous measurements of pulmonary blood flow (PBF) and pulmonary artery blood
pressure (PABP) were processed via calibrated pressure transducers (Maxxim Medical,
Tx, U.S.A.). Data were amplified and digitally recorded (Powerlab 8SP, ADInstruments,
N.S.W., Australia). Pulmonary waveform analysis was performed at regular time points
as described previously (23) to quantifiy changes in pulmonary blood flow throughout
the cardiac cycle. Pulsatility Index (PI), a measure of downstream resistance to blood
flow, was calculated as (peak systolic flow – minimum flow after systolic peak)/mean
peak systolic flow and averaged over five consecutive cardiac cycles.
The Oxygenation Index (OI) was calculated as OI=2
2
aO
100 x aw xFiO
P
P where FiO2 is fractional
inspired oxygen concentration, Paw is mean airway pressure and PaO2 is partial
pressure of oxygen in arterial blood. An increase in the OI suggests deterioration in
arterial oxygenation.
Post‐mortem
The lung was exposed by thoracotomy, and an in situ deflation pressure volume curve
was obtained (24). The right upper lung lobe was inflation fixed (30 cmH2O) in formalin
and samples of the right lower lobe were snap frozen for molecular analyses.
Bronchoalveolar lavage (BAL) was performed on the left lung for cytology and protein
analysis by the Lowry method (25, 26). Differential cell counts were performed on
cytospin samples of the BAL fluid stained with Diff‐Quik (Fronine Lab Supplies, N.S.W.,
Australia).
242
RNA was extracted from the left lung and reverse transcribed to cDNA (QuantiTect®
Reverse Transcription Kit, Qiagen, U.S.A.). Expression of IL‐1β and IL‐6 was measured
by qRT‐PCR (27) and normalized to 18S RNA (28) using the 2‐∆∆CT method (29).
Statistical Analyses
Kruskal‐Wallis one way analysis of variance on ranks was used to compare groups at
specific time points while the effect of ventilator strategy on ventilator requirements
and physiological changes over the duration of the study were determined using two‐
way repeated measure analysis of variance. Posthoc comparisons were performed
using the Holm‐Sidak method. Analyses were performed using SigmaStat (Version 3.5,
Systat Software Incorporated, U.S.A.) with p<0.05 considered statistically significant.
Data are expressed as mean (SEM) unless otherwise stated.
243
Results
Baseline characteristics of lambs in each group were not different (Table 1).
Ventilation and Oxygenation
PaCO2 was higher (and above the target range) in the HFOV group compared to both
HFJV and CMV at 45, 60, 120 and 180 min (Figure 2A). The ∆P was highest for HFOV
from 20 min (Figure 2B).
FiO2 commenced at 0.4 in all ventilated groups. At 60 and 75 min, FiO2 was higher in
the HFOV group compared to HFJV and CMV groups. At 75 min FiO2 was higher in the
HFJV group compared to the CMV group (Figure 2C). The alveolar‐arterial difference in
partial pressure of oxygen (AaDO2) was higher in the HFOV group at 60 and 75 min
(Figure 2D). Paw was lowest in the CMV group for most of the study while HFJV Paw
was lower than HFOV Paw at 45, 120, 150 and 180 min (Figure 2E).
Pulmonary Blood Flow
End systolic and end diastolic pulmonary blood flows were comparable until 150 min
from which time HFJV lambs had lower flows (Figure 3A and 3B respectively).
Pulsatility Index was comparable between all groups for the duration of the study
(Figure 3C).
Post‐mortem
BAL fluid protein concentration and cell populations were similar between all groups.
The expression of IL‐1β and IL‐6 were not different between groups (Table 2). There
244
was no difference in the static lung compliance, as assessed by the deflation limb of
the post‐mortem pressure‐volume curve, between the ventilated groups. The UVC
group had lower static lung compliance (Figure 4A).
245
Discussion
This study aimed to compare a CMV strategy that included moderate PEEP (7 cmH2O)
with both HFJV and HFOV over a 3 hour ventilation study using a preterm lamb model
of RDS. Gas exchange was comparable for both CMV and HFJV lambs, however, HFOV
did not achieve oxygenation and ventilation targets within the algorithm utilised.
Pulmonary vascular resistance was similar for all groups though HFJV caused shunting
of blood at the end of the ventilation period. Despite these differences in gas exchange
and pulmonary blood flow there were no differences in the markers of lung injury.
Each lamb was managed identically immediately after delivery and then according to a
predetermined ventilation strategy specific algorithm that aimed to open the lungs,
keep the lungs open and maintain optimal oxygenation as described by the open lung
approach (4, 5). The initial sustained inflations were performed as a specific lung
volume recruitment manoeuvre for all animals so the subsequent ventilation strategy
followed a lung volume recruitment process which was less aggressive than it would
have been otherwise. In the CMV group, the initial PEEP was chosen on the basis of
previous observations in the preterm lamb model to maintain end‐expiratory lung
volume above the closing volume as PEEP below the inflection point will increase the
potential for atelectotrauma (30). During HFOV, we used a stepwise recruitment
strategy as standardly performed in clinical practice (31, 32). The maximum Paw in this
group was limited to, but did not reach 26 cmH2O as we expected that the prophylactic
administration of surfactant and the delivery of sustained inflations immediately prior
to HFOV would establish ventilation above the critical closing pressure of the lungs
246
(33). For the HFJV group we chose to use a combination of intermittent CMV breaths
and incrementing PEEP to recruit and stabilise alveoli. The maximum PIPHFJV was set at
40 cmH2O as we have previously successfully ventilated lambs without needing to
exceed this limit (34). The number of CMV breaths was limited to 5 breaths/min in line
with clinical recommendations and our previous experience that this CMV breath rate
was sufficient to provide physiological benefit at initiation of ventilation (unpublished
data ‐ see Chapter 7). Despite our previous experience that PIPCMV 5 cmH2O above
PIPHFJV provides greater physiological benefit with the least evidence of harm
(unpublished data – see Chapter 6), the PIPCMV was kept 5 cmH2O below PIPHFJV as
results of the previous study were not fully available at the time of the study. All
ventilation protocols aimed to minimise cyclic stretch within the lung using a
permissive hypercapnia approach.
The comparison of 3 different ventilation strategies is a challenge as ventilator settings
and displayed measurements vary between modalities. During CMV and HFJV, Paw is
determined by PIP, PEEP, respiratory frequency, and the tI:tE ratio (35). During CMV,
PIP, ∆P and Paw are fully transmitted to the distal airways so pressure measurements
at the airway opening closely approximate alveolar pressures (15). During high‐
frequency ventilation (both HFJV and HFOV), however, the ∆P is attenuated in the
distal airways and alveoli. During HFJV, pressure monitored at the airway opening via
the custom designed tracheal tube adaptor closely approximates the mean pressure at
the distal tip of the tracheal tube (36). In contrast, during HFOV the mean pressure is
measured at the airway opening and hence does not reflect mean pressure drop
across the tracheal tube.
247
One of the aims of this study was to compare the physiological responses to different
ventilation strategies in a standardised model of RDS. Oxygenation and ventilation
variables were recorded to determine whether or not the different strategies achieved
similar physiological outcomes. In the HFOV group we were unable, without exceeding
the preset limits of Paw and ∆P, to approach optimal oxygenation and ventilation and
there were times when FiO2 and PaCO2 were higher in this group. Oxygen
requirements followed a similar trend between the groups with a gradual increase in
FiO2 and AaDO2 over the 3 h study indicating progressive ventilation‐perfusion
mismatching. Nonetheless, FiO2 requirements and AaDO2 were increased from 60‐75
min ventilation in the HFOV group compared to both HFJV and CMV lambs. The Paw
during HFOV, however, was highest throughout the study but the indices of
oxygenation were worst for this group. Given the correlation of Paw and oxygenation it
is surprising that this higher Paw did not translate to a lower FiO2 in this group. FiO2
tended to be higher in the HFOV group from soon after initiation of ventilation hence it
is conceivable that the limit of 26 cmH2O placed on Paw may have limited achievement
of an open lung. Secondly, while an tI:tE ratio of 1:2 during HFOV may minimise gas
trapping, it may also create a mean pressure drop across the tracheal tube that
becomes clinically significant (37). If alveolar pressure is lower than Paw displayed on
the ventilator, the benefits to oxygenation may not manifest. Conversely, it is also
feasible that Paw remained too high for too long during HFOV and the associated
barotrauma caused the deterioration in oxygenation in this group. Another
explanation is that the sustained inflations may have been more effective at volume
recruitment than we appreciated and our subsequent ventilation strategy caused
overinflation of the lungs in the HFOV lambs.
248
There were differences in ventilation which may be attributed to the upper limits set
on the parameter primarily responsible for carbon dioxide elimination (VT/kg for CMV,
∆P for HFJV and ∆P for HFOV). The ∆P measurements during HFOV reflect measured
pressure at the proximal end of the tracheal tube, while ∆P for HFJV are calculated (by
the ventilator algorithm) to estimate the airway pressure at the distal end of the
tracheal tube. The pressure transmitted along the airway will be affected by lung
compliance where reduced lung compliance will increase transmission of ∆P from the
airway opening to the alveoli (38) The ∆P recorded during HFOV may be attenuated by
between 40 and 80 % when measured just distal to the tracheal tube (32). This means
the highest ∆P of 50 cmH2O during HFOV is likely to have created between 20 and 40
cmH2O of pressure in the trachea, which creates a range which includes the highest ∆P
during HFJV of 27.5 cmH2O. Nevertheless we did not achieve the target PaCO2 for the
HFOV animals and in hindsight a higher limit on ∆P may have been appropriate.
There was a predictable initial increase in PBF in all groups with the transition from
fetal to neonatal circulation as the pulmonary vasculature dilates. Pulmonary blood
flow subsequently steadily became more negative over the remainder of the study in
all groups. Given the fall in PBF was mirrored by a steady increase in pulsatility index,
particularly in the first 90 min after birth, these findings are most likely explained by an
increase in pulmonary vascular resistance and right to left shunting of blood through
fetal vascular channels (39).
The significant fall in end systolic and end diastolic pulmonary blood flow in the HFJV
animals over the last 30 min of the study compared to both CMV and HFOV groups was
unexpected and is difficult to explain. We had expected the magnitude of change in
249
pulmonary blood flow to mirror changes in Paw in all the ventilated groups as it has
been reported that increasing Paw decreases PBF and increases pulmonary vascular
resistance during CMV (40), HFJV (34) and HFOV (21). With a Paw consistently lower in
HFJV than that observed in the HFOV group, we would have expected that HFJV would
be less likely to negatively impact pulmonary blood flow and to have a lower pulsatility
index compared to HFOV. Although the pulsatility index increased in the HFJV group
over the last 30 min of the study, it was not significantly higher than the pulsatility
index in either the CMV or HFOV groups throughout the study and hence does not
explain the late fall in PBF in the HFJV animals.
Given that we followed an algorithm for optimal ventilation with each strategy, and
there were not marked differences in ventilator parameters it is not surprising that we
did not find a difference in lung injury markers. A comparison such as ours has not
been performed before but there are a multitude of studies assessing lung injury in
response to mechanical ventilation in neonates and animals (41‐44). The results of
studies comparing lung injury following HFOV and CMV vary and demonstrate either
little difference in the alveolar leakage and systemic inflammation in neonates (45),
probable attenuation of early activation of inflammation and clotting in preterm lambs
during HFOV when compared to CMV (46) and reduced pro‐inflammatory cytokines in
HFOV treated neonates compared to CMV (47). There are fewer studies comparing
lung injury during HFJV but a comparison of HFJV and HFOV in rabbits showed a clear
reduction in lung injury following HFJV (48). The lack of difference in the markers of
lung injury we chose to study suggests that the strategies were equivalent in their
ability to produce inflammation in the lung.
250
There are a number of limitations to this study. We aimed to compare three different
ventilation strategies utilising an open lung approach but failed to achieve our target
arterial blood gas parameters in all the groups. The basis for the higher PaCO2 in the
HFOV group is unknown and the higher FiO2 required to achieve equivalent SpO2 is
difficult to interpret given that the limit of Paw was reached. Furthermore, the
ventilation period was short which may have prevented a difference in lung injury
markers. The changes in end systolic and end diastolic blood flow are also difficult to
interpret in light of the short duration of the study. It is also important to acknowledge
that surfactant was administered to the lambs immediately after delivery, which may
not always be achieved in clinical practice. The lambs in our study were anaesthetised
and underwent an invasive surgical procedure. The hemodynamic effects of
anaesthesia combined with the physical impact of instrumentation on lung inflation
are likely to impact physiological outcomes. Nevertheless, these limitations were for
the most part standard across each group and this model provides the first direct
comparison of these three lung protective ventilation strategies in an animal model.
In conclusion, the open lung approach to CMV, HFJV and HFOV were all suitable for the
respiratory management in the context of our model of RDS in preterm lambs. The lack
of significant differences in end systolic and end diastolic PBF for the majority of the
study, lung injury data and static lung compliance demonstrate that in the absence of
airleaks each of these strategies can be employed in the clinical setting with a
comparable pressure cost of ventilation.
251
Acknowledgements
We would like to express our sincere appreciation to the members of the Ovine
Research Group, Ilias Nitsos and Carryn McLean, for technical assistance and JRL Hall
and Co. for provision and early antenatal care of the ewes.
252
Tables
Table 1: Baseline lamb data
UVC CMV HFJV HFOV
n (male) 6 (3) 6 (3) 8 (4) 8 (6)
Birth weight (kg) 3.9 (0.2) 3.5 (0.2) 3.6 (0.2) 3.5 (0.2)
Gestational age (d) 129.0 (0.5) 128.8 (0.3) 129.2 (0.3) 129.0 (0.3)
Cord pH 7.16 (0.03) 7.21 (0.03) 7.19 (0.03) 7.20 (0.04)
Cord PaCO2 (mmHg) 75.7 (3.6) 75.4 (6.7) 75.6 (5.2) 81.9 (10.5)
UVC = Unventilated Control, CMV = Conventional Mechanical Ventilation, HFJV = High‐
frequency Jet Ventilation, HFOV = High‐frequency Oscillatory Ventilation. Values are
mean (SEM).
253
Table 2: Post mortem inflammatory markers
UVC CMV HFJV HFOV
BAL fluid
Protein (mg mL‐1) 183.2 (37.9) 363.3 (71.4) 272.3 (37.9) 275.9 (28.1)
Total Inflammatory Cells
(x 106 kg‐1) 1 (0.3) 17.6 (18.9) 6.7 (1.7) 6.9 (2.6)
Neutrophils (x 106 kg‐1) 0.6 (0.3) 14.4 (18) 4.9 (1.4) 4.9 (2.1)
Mononuclear cells (x 106 kg‐1) 0.3 (0.1) 3.2 (3) 1.5 (0.4) 1.7 (0.8)
Lung Tissue
IL‐1β (fold change) 1.4
(0.7, 2.2)
4.8
(3.2, 18.0)
7.9
(3.1, 15.4)
10.9
(3.1, 53.1)
IL‐6 (fold change) 1.2
(0.5, 2.4)
3.0
(2.2, 33.5)
6.8
(2.2, 19.8)
11.8
(2.4, 40.4)
UVC = Unventilated Control, CMV = Conventional Mechanical Ventilation, HFJV = High‐
frequency Jet Ventilation, HFOV = High‐frequency Oscillatory Ventilation, IL‐1β =
interleukin 1 beta, IL‐6 = interleukin 6. Values are mean (SEM) or median (25th, 75th
centile) for parametric and non‐parametric data respectively.
254
Figure 1A
255
Figure 1B
256
Figure 1C
257
Figure 2
258
Figure 3
259
Figure 4
260
Figure Legends
Figure 1 Ventilation strategies:
A: Conventional mechanical ventilation. PPV = Positive pressure ventilation, VG =
volume guarantee, tI = inspiratory time, VT = tidal volume, PIP = peak inspiratory
pressure, PEEP = positive end‐expiratory pressure SpO2 = oxyhaemoglobin saturation
measured by the pulse oximeter, FiO2 = fractional inspired oxygen concentration,
PaCO2 = partial pressure of carbon dioxide in arterial blood.
B: High‐frequency jet ventilation strategy. HFJV = high‐frequency jet ventilation, tI =
inspiratory time, PIP = peak inspiratory pressure, CMV = conventional mechanical
ventilation, PEEP = positive end‐expiratory pressure, CPAP = continuous positive
airway pressure, SpO2 = oxyhaemoglobin saturation measured by the pulse oximeter,
FiO2 = fractional inspired oxygen concentration, PaCO2 = partial pressure of carbon
dioxide in arterial blood.
C: High‐frequency oscillatory ventilation strategy. Paw = mean airway pressure, tI =
inspiratory time, I:E = inspiratory to expiratory time ratio, SpO2 = oxyhaemoglobin
saturation measured by the pulse oximeter, FiO2 = fractional inspired oxygen
concentration, PaCO2 = partial pressure of carbon dioxide in arterial blood.
Figure 2 Ventilation and Oxygenation: A: Partial pressure of carbon dioxide in arterial
blood (PaCO2), B: Airway pressure differential (∆P) (cmH2O), C: Fractional inspired
oxygen concentration (FiO2), D: Alveolar ‐ arterial difference in the partial pressure of
oxygen (AaDO2) (mmHg), E: Mean airway pressure (Paw). *p<0.05 HFJV compared to
261
CMV, #p<0.05 HFOV compared to CMV, ^ p<0.05 HFJV compared to HFOV. Closed
circle = CMV, closed square = HFJV, closed triangle = HFOV.
Figure 3 Pulmonary Blood Flow: A: End Systolic Pulmonary Blood Flow (PBF), B: End
Diastolic Pulmonary Blood Flow, C: Pulsatility Index. *p<0.05 HFJV compared to CMV, ^
p<0.05 HFJV compared to HFOV. Closed circle = CMV, closed square = HFJV, closed
triangle = HFOV.
Figure 4 Static Lung Compliance: Deflation limb of the post‐mortem in situ pressure‐
volume curves, * p<0.05 HFJV compared to CMV, ^ p<0.05 HFJV compared to HFOV, **
p<0.05 UVC compared to HFJV, HFOV and CMV. Closed circle = CMV, closed square =
HFJV, closed triangle = HFOV, open diamond = UVC.
262
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9 Discussion
The overall aim of this collection of studies was to systematically investigate high‐frequency
jet ventilation (HFJV), in a preterm lamb model of respiratory distress syndrome (RDS), to
establish an evidence base to support clinical HFJV strategies in neonatal intensive care
units. Although HFJV has been available to clinicians for over 20 years, extensive in vivo
studies exploring HFJV for RDS have not been performed. This has led to the development of
clinical strategies that lack an evidence based structure. Furthermore, the results of
randomised controlled trials have not demonstrated convincingly a clear benefit of HFJV
over other ventilatory strategies in preterm infants. In order to elucidate optimal ventilator
settings during HFJV the impact of positive end‐expiratory pressure (PEEP) and the size,
duration and frequency of conventional mechanical ventilator (CMV) breaths delivered
during HFJV were examined. The results of these controlled studies led to the composition
of an algorithm which was used to compare HFJV with high‐frequency oscillatory ventilation
(HFOV) and a gentle CMV strategy in the preterm lamb model.
We hypothesised that increasing PEEP would recruit alveoli and improve oxygenation at the
expense of pulmonary blood flow and lung injury. Further, we hypothesised that CMV
breaths delivered at a longer inspiratory time, to a higher peak inspiratory pressure and
more frequently would recruit alveoli and improve oxygenation at the expense of lung
injury. To this end we investigated and documented the effect of HFJV on ventilation
(elimination of CO2), oxygenation, pulmonary blood flow, static lung compliance and lung
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injury. These investigations were undertaken by categorising the ventilator parameters
responsible for alveolar recruitment and maintenance of an open lung during HFJV into
PEEP and CMV breaths. The CMV breaths delivered during HFJV were then compared by
isolating the effect of 2 different CMV breath inspiratory times, peak inspiratory pressure
settings and frequencies. We chose a preterm lamb model for these studies as this non‐
primate animal model has been extensively utilised in the past. This widespread application
of the preterm lamb model has enabled correlation of lung developmental stages between
Ovis aries and Homo sapiens. Furthermore, ventilatory equipment used for human babies
can be used in the laboratory without modifications as the size of a preterm lamb is close to
a term baby.
The high‐frequency jet ventilator we used must be set up in tandem with a conventional
ventilator. The conventional ventilator, amongst other functions, delivers PEEP. PEEP is
utilised during mechanical ventilation to prevent alveolar collapse at the end of expiration,
but also to recruit and stabilise alveoli. PEEP is usually employed during mechanical
ventilation of preterm babies but the consequences of PEEP may outweigh the benefit. The
first study in this thesis: “"High Positive End‐Expiratory Pressure during High‐Frequency Jet
Ventilation Improves Ventilation in Preterm Lambs" investigated PEEP recruitment
manoeuvres and found that incrementing PEEP up to 12 cmH2O facilitated lung volume
recruitment without significant adverse effects on pulmonary blood flow, ex vivo lung
compliance and lung injury. The premise of this study challenged common clinical practices
which rarely increased PEEP to this level. The results were presented the following year
(2008) and clinical strategies have been altered to increase PEEP to a higher level for
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alveolar recruitment manoeuvres based upon the results of that first study (personal
communication: Rob Graham, R.R.T./N.R.C.P., N.I.C.U., Sunnybrook Health Sciences Centre
Toronto,Ontario,Canada).
The subsequent studies examined the parameters of the CMV breaths that can be delivered
during HFJV using a ventilated control group and an unventilated control group for
comparison. A comparison of CMV breaths delivered over 0.5 s and 2 s was made over a 2 h
study: “The Impact of Conventional Breath Inspiratory Time during High‐frequency Jet
Ventilation in Preterm Lambs”. Consistent with our hypothesis for this study we concluded
that a longer CMV breath inspiratory time was more injurious to the lung given the trend for
increased inflammatory markers in this group. To follow this study a comparison of CMV
breaths delivered to a peak inspiratory pressure (PIP) 5 cmH2O above and below the HFJV
PIP was made: “The Effect of Conventional Breath Peak Inspiratory Pressure during High‐
frequency Jet Ventilation in Preterm Lambs”. The results of this study were contrary to our
hypothesis that CMV breaths delivered to a PIP 5 cmH2O above the HFJV PIP would be more
injurious. In both of the aforementioned CMV breath studies we delivered 5 CMV
breaths/min so our final study compared CMV breaths delivered 5 or 20 times/min:
“Alveolar Recruitment with Five or Twenty Conventional Mechanical Ventilator Breaths per
minute during High‐frequency Jet Ventilation in Preterm Lambs”. The CMV breaths in this
study were delivered to a PIP 5 cmH2O below the HFJV PIP and while more frequent CMV
breaths increased oxygenation requirements and compromised static lung compliance there
was no clear evidence of greater lung injury in this group. An issue with this study was the
lack of a third treatment group. There was an unventilated control group, a HFJV only
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ventilated control group and 2 HFJV+CMV groups. In hindsight the addition of another group
in which 20 CMV breaths were delivered to PIP 5 cmH2O above the HFJV PIP would have
been interesting, especially in light of the results of the CMV breath PIP comparison study.
We attributed the differences in the CMV breath PIP comparison study to less effective lung
volume recruitment when 5 CMV breaths/min were delivered to a PIP 5 cmH2O below HFJV
PIP. This finding left us comparing a strategy of sub optimal lung volume recruitment (5 CMV
breaths/min to a low PIP) with a strategy that we hypothesised would cause greater lung
injury (20 CMV breaths/min) Whether optimal lung volume recruitment can be achieved
when CMV breaths are delivered to PIP higher than HFJV breaths or when CMV breaths are
delivered at a faster rate (>5 CMV breaths/min) is difficult to say without comparing a high
CMV PIP, high CMV breath rate group to the groups we already have. However, the high
CMV breath rate group did demonstrate convincing inadvertent PEEP and this should
increase Paw which, to a point, may improve oxygenation, at the expense of lung injury. The
combination of inadvertent PEEP when CMV breaths are delivered more often with a CMV
breath PIP higher than HFJV PIP may cause greater lung injury. The data we have suggest
CMV breaths delivered to a PIP 5 cmH2O above HFJV PIP is better, but the optimal frequency
for these breaths is unknown and warrants further investigation.
The final study: “A Comparison of High‐frequency Jet Ventilation and High‐frequency
Oscillatory Ventilation with Conventional Mechanical Ventilation in Preterm Lambs”
provided an opportunity to compare the 3 strategies most commonly utilised in the
neonatal intensive care unit. This direct comparison has not been performed previously and
gave us a unique opportunity to investigate the ventilatory efficacy of these 3 strategies in a
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controlled environment. Our hypothesis was not supported by our results insofar as the
cardiovascular side effects of HFJV were not substantially different to the other strategies.
At best, we demonstrated that when an open lung approach drives the decision making
process there was no clear benefit of one strategy over another in our preterm lamb model
of RDS.
There are a number of limitations to the studies in this thesis. The short duration of
ventilation prevented extensive temporal data collection, which limits the relevance of the
information to the clinical setting as babies requiring HFJV will be ventilated for days,
potentially weeks. Nevertheless, the acute effect of changes in airway pressures is still
relevant to the neonatal intensive care unit environment. An animal model will always
introduce species specific characteristics which may or may not be recognized. Furthermore,
the use of animals in research is bound by the Australian code of practice for the care and
use of animals for scientific purposes. Working within the code requires anaesthesia of the
pregnant ewe for caesarean delivery, and therefore transplacental transfer of anaesthetic
drugs to the fetus, and the administration of additional anaesthesia and analgesia to the
lamb following delivery. The influence of anaesthetic drugs was controlled as it was
standard throughout the studies, however, it remains that these drugs will affect the
cardiovascular system and the target population is unlikely to be subject to the same
pharmacological restraint.
A major limitation of these studies was the sample size. Our largest study group had 8
animals which, in light of the variability of some data, decreased the power of these studies.
Larger groups would have increased study power, but the financial and animal welfare
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expense was too great to accommodate this. This limitation is particularly applicable to the
final study: “A Comparison of High‐frequency Jet Ventilation and High‐frequency Oscillatory
Ventilation with Conventional Mechanical Ventilation in Preterm Lambs” where the decision
making process was governed by the physiological status of the lamb at preset time points.
We wanted to manage this group as though they were individual patients where survival
was imperative. This was in contrast to the preceding studies where a predetermined
ventilatory strategy was set without aiming for survival, but rather, aiming to document the
effects of individual ventilator settings. It is also noteworthy that comparing the efficacy of
ventilation of one strategy compared to another was difficult as we targeted a tight PaCO2
and SpO2 range. This meant that our primary outcome variables were other indicators of
physiology (not including PaCO2 and SpO2) and injury.
It is also important to note that our preterm lamb model of RDS represents just one of the
many clinical presentations that may require ventilatory support. Preterm babies are at risk
of a number of different lung diseases and may not be commenced on a ventilator protocol
immediately after delivery. With this in mind, the results of these studies cannot be directly
translated to babies with heterogeneous lung disease or those requiring ‘rescue’ from
another ventilatory strategy. Nonetheless these data are still useful for more evidence
based decision making in the context of an individual patient.
The results of these studies contribute to the sparse data on HFJV and have provided
fundamental information that will enable a more evidence based approach to clinical
decision making in the neonatal intensive care unit. Future work in this area should focus on
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the target population and incorporate randomised controlled trials comparing HFJV to other
ventilatory strategies.
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Appendix
Musk GC, Polglase GR, Bunnell JB, McLean CJ, Nitsos I, Song Y and Pillow JJ 2011 High
Positive End‐Expiratory Pressure during High‐Frequency Jet Ventilation Improves
Oxygenation and Ventilation in Preterm Lambs. Pediatric Research 69(4):319‐324
See attached pdf