Changes in Amyloid-beta Correlate with Neurological Status after … · Changes in Amyloid-beta...

Changes in Amyloid-beta Correlate with Neurological Status after TBI

David L. Brody, MD PhD

Assistant Professor of Neurology Washington University, St. Louis, MO

Keystone Symposium

February 29, 2012

Disclosures • Support:

– NIH K08 to D. Brody – Burroughs Wellcome Career Award in the

Biomedical Sciences to D. Brody – NIH R01 and P01 grants to D. Holtzman – Eli Lilly and Co. provided antibodies.

• Conflicts of interest: none.

Outline • Introduction • Human Studies

– A-beta dynamics correlate with neurological status – A-beta is inversely related to markers of axonal injury

• Experimental Animal Studies – Soluble extracellular A-beta is reduced after TBI in

PDAPP and Tg2576 mice – Insoluble intraaxonal A-beta is increased after TBI in

3xTg-AD and APP/PS1 mice • Conclusions and Future Directions

TBI is the best documented environmental risk factor for Alzheimer’s disease (AD),

• In a metaanalysis of 9 studies, TBI increased the risk of AD ~ 1.8 fold.

(Mortimer et. al Int. Journal of Epidemiology 20 Suppl 2 S28-35 1991)

• Removing the possibility of recall bias, well-documented moderate to severe TBI in WWII veterans was found to be a strong risk factor for AD, with hazard ratios of 2.3 to 4.5.

(Plassman et. al. Neurology 55 1158-66 2000).

• The victim of a single, severe TBI at age 22.

• Partial recovery of cognitive function, but then developed a progressive dementia starting at age 32.

• AD-like changes at the time of his death at 38

Clinton et. al. Neuropath Appl Neurobiol 17 69-71 1991).

• Diffuse Aβ plaques in 46 of 152 cases of fatal TBI, as young as 10 years old without Down syndrome or familial AD.

Roberts et. al. J. Neurol, Neurosurg. & Psychiatry 57 419-425 1994.

Shared Pathology of TBI and AD

Aβ plaques appear in areas of Diffuse Axonal Injury in Humans

• Smith et. al., J. Neurosurgery 98 1072 (2003)

Aβ

Aβ

Diffuse Axonal Injury: APP → NF ↓

colocalization

colocalization

Amyloid-beta deposition in Alzheimer’s disease is largely extracellular

Yamaguchi et al. Am J Path 1979

Early, diffuse plaques (arrows) from frontal cortex in an 81 year old with AD.

Brendza et al. J Comp Neurol 2003

Mature, neuritic plaque (arrowhead)

Amyloid-beta dynamics in the extracellular space of the

human brain

Amyloid-beta deposition related to default activity in humans

Buckner et al J Neurosci 2005

Microdialysis involves exchange of extracellular fluid and solutes across a semi-permeable membrane.

Cirrito et al J. Neurosci 2003

Regulation of extracellular amyloid-beta levels in animal models.

Kamenetz et al., Neuron 2003 (slice cultures)

Cirrito et al., Neuron 2005 (in vivo microdialysis)

Microdialysis in the Human Brain Microdialysis

Catheter

External Ventricular

Drain

MicrodialysisCatheter


Drain

Courtesy of CMA

Poca et al, J Neurotrauma 2006 Brody, Magnoni et al Science 2008


– A-beta dynamics correlate with neurological status

– A-beta is inversely related to markers of axonal injury • Experimental Animal Studies

– Soluble extracellular A-beta is reduced after TBI in PDAPP and Tg2576 mice

– Insoluble intraaxonal A-beta is increased after TBI in 3xTg-AD and APP/PS1 mice

• Conclusions and Future Directions

Amyloid-beta Dynamics Correlate with Neurological Status in the

Injured Human Brain David L. Brody*, Sandra Magnoni* Kate E. Schwetye, Michael L. Spinner, Thomas J. Esparza, Nino Stocchetti, Gregory J. Zipfel,

David M. Holtzman Washington University, St. Louis, MO and Ospedale Maggiore Policlinico, Milan, Italy

Methods • 18 patients participated in this study: 17 with acute brain injury and 1 undergoing

craniotomy for unruptured aneurysm clipping. • All protocols were approved by the Human Research Protection Offices at Washington

University, St. Louis and the Ospedale Maggiore Policlico, Milan. • Written informed consent was provided by next of kin. • All microdialysis catheters (CMA70, 20kDa nominal MW cutoff or CMA71, 100 kDa

nominal MW cutoff) were placed by experienced neurosurgeons in conjunction with another interventional procedure, typically placement of an intracranial pressure monitoring device.

• Sterile human albumin was added to sterile CMA perfusion fluid to a final concentration of 0.15% (CMA70 catheters) or 1.5% (CMA71 catheters, for oncotic balance)

• Flow rate was 0.3 µl/min (18 µl/hour) • Samples were acquired every 1-2 hours in CMA microdialysis tubes. • Samples were immediately refrigerated on ice and frozen at -80°C within 12 hours of

acquisition. • 96-well plate format ELISAs were used to measure amyloid-beta.

– Aβ1-x: m266 (recognizes aa 13-28) used to capture, 3D6 (recognizes aa 1-5) used to detect

– Aβ1-42: 21F12 (specific for Aβ42) used to capture, 3D6 used to detect

– Aβ1-40: 2G3 (specific for Aβ40) used to capture, 3D6 used to detect

0 24 48 720

500

1000

1500

2000

0

5

10

15

20

Aβ1-xUrea

time (h)

ISF

Aβ

1-x

(pg/

ml) U

rea (mg/dl)

Aneurysmal Subarachnoid HemorrhageaGold tip of microdialysis catheter in right frontal lobe white matter

Aβ dynamics in a human patient

Urea was measured in the same samples as a control for stable catheter function (Ronne-Engstrom et al J Neurosurg 2001).

Brody, Magnoni et al, Science 2008

0 24 48 72 96 120 1440

500

1000

1500

0

5

10

15

time (h)

ISF

Aβ

1-x

(pg/

ml) U

rea (mg/dl)

Traumatic Brain Injury: Contusionc


Median Aβ increase was 58% over 3 days

*** p=.0002 Wilcoxon signed rank test

N=9 TBI patients with catheters in apparently normal brain regions (triangles)

N=3 TBI patients with catheters in pericontusional regions (x-symbols)

N=6 SAH patients (open circles)

Hours 0-1

2

Hours 60

-72

0

500

1000

1500

20005000

10000 ***B

rain

ISF

Aβ

1-x

(pg/

ml)

Hours 0-1

2

Hours 60

-72

Hours 0-1

2

Hours 60

-720

5

10

15

20

Bra

in I

SF U

rea

(mg/

dl)

Hours 0-1

2

Hours 60

-72


What about cerebrospinal fluid?

0 24 48 720

1000

2000

3000

4000Brain ISF (Microdialysis)Ventricular CSF

Approx time since injury (h) Aβ 1

-x (p

g/m

l)

Aβ levels appear overall lower in microdialysis samples than in ventricular CSF

ISF dynamics not reflected in ventricular CSF.



Drain



Drain


Aβ recovery by microdialysis is incomplete

0.0 0.5 1.00

500

1000

1500

2000 Zero-Flow Extrapolation

flow rate (µ l/min)

ISF

Aβ

40 (p

g/m

l)

in vivo

in vitro

010203040

% re

cove

ryat

0.3

µl /

min

Recovery appears to be ~30% in vivo and in vitro.

In contrast, recovery of small molecules like glutamate, lactate, pyruvate, glucose, urea etc. has been reported to be ~70-90% (Hillared et al J Neurotrauma 2005).


Unruptured aneurysm patient(no neurological injury)

0 12 240

500

1000

1500

2000

2500

3000

time (hours)

Estim

ated

true

ISF

A β1-

40 (p

g/m

l)

Estimated true ISF levels =

measured levels / fractional recovery

Fractional recovery calculated from zero-flow extrapolation.

Brain IS

F

Brain IS

F, corr.

for r

ecov.

Ventri

cular

CSF

10

100

1000

10000Aβ

1-x

(pg/

ml)

After correction for partial recovery, brain ISF levels appear similar on average to ventricular CSF levels.

Box: median and interquartile range, Whiskers: 5-95% confidence interval, Circles: outliers.


0.125 0.2

5 0.5 1 2 4 80.25

0.5

1

2

4

8

N.S.

Ventricular CSF Aβ1-x (fold change)

Brai

n IS

F Aβ 1

-x(fo

ld c

hang

e)

No correlation of dynamics between brain ISF and ventricular CSF


Aβ1-42

0 2000 40000

25

50

75200250300

Spearman r = 0.84

Brain ISF Aβ1-x (pg/ml)

Brai

n IS

F Aβ

1-42

(pg/

ml)

0 10000 200000

100

200

300

400

500Spearman r = 0.62

Ventricular CSF Aβ1-x (pg/ml)

Vent

ricul

ar C

SF A

β1-

42 (p

g/m

l)


What underlies these dynamcs in the injured brain?

• Correlations with other microdialysis parameters.

• Correlations with other aspects of cerebral physiology.

• Correlations with neurological status.

N.S.

0 5 100

500

1000

1500

2000

3000

5000

Lactate (mM)

ISF

Aβ

1-x

(pg/

ml)

Spearman r = .18, p=.0003

0.0 0.1 0.2 0.3 0.40

500

1000

1500

2000

3000

5000

Pyruvate (mM)

ISF

Aβ

1-x

(pg/

ml)

Spearman r = -.40, p<.0001

10 20 30 400

500

1000

1500

2000

3000

5000

Lactate / Pyruvate

ISF

Aβ

1-x

(pg/

ml)

a b c

Correlations with Redox State

Elevated lactate/pyruvate ratio reflects impaired oxidative metabolism. (Hillered et al J Neurotrauma 2005)

Elevated lactate can be a marker of synaptic activity (Bero et al Nature Neurosci 2011)

Spearman r = .45, p<.0001

0 1 2 3 40

500

1000

1500

2000

3000

5000

Glucose (mM)

ISF

Aβ

1-x

(pg/

ml)

N.S.

0 10 20 30 40 500

500

1000

1500

2000

3000

5000

PtiO2 (mm Hg)

ISF

Aβ

1-x

(pg/

ml)

Correlations with Oxygen and Glucose Delivery


N.S.

0.1 1 10 1000

500

1000

1500

2000

3000

5000

Glutamate (µM)

ISF

Aβ

1-x

(pg/

ml)

Glutamate Toxicity?


0 10 20 300

500

1000

1500

2000

50001000015000

Spearmanr = -.56,

p<.0001 forICP>20

N.S. overall

Intracranial Pressure (mm Hg)

ISF

Aβ

1-x

(pg/

ml)

N.S.

40 60 80 100 1200

500

1000

1500

2000

50001000015000

Cerebral Perfusion Pressure (mm Hg)IS

F Aβ

1-x

(pg/

ml)

Intracranial Pressure


N.S.

35 36 37 38 39 40 410

500

1000

1500

2000

3000

5000

Cerebral Temperature ( °C)

ISF

Aβ

1-x

(pg/

ml)

Cerebral Temperature Spearman r = -.26, p<.0001

0 1 2 3 40

500

1000

1500

2000

3000

5000

|Cerebral Temperature (°C)- 37.2|

ISF

Aβ

1-x

(pg/

ml)

Normal cortical temperature 37.2 ± 0.6 °C estimated by MR spectroscopy (Corbett et al. JCBFM 1997)


Overall Neurological Status

The Glasgow Coma Score (Teasdale and Jennett, Lancet 1974)

Eye Opening: 1-4

Best Verbal Response: 1-5

Best Motor Response: 1-6

Total: 3-15

3579111315

Rapid Recovery

Gla

sgow

Com

a Sc

ore

0 12 24 36 48 60

0.5

1

2

4

time (hours)

ISF

Aβ

1-x

(fold

cha

nge

from

bas

elin

e)

0 24 48 72 96 120 144 168

0.5

1

2

4

Prolonged Coma:Glasgow Coma Score = 4

time (hours)

ISF

Aβ

1-x

(fold

cha

nge

from

bas

elin

e)

567891011Clinical Fluctuations

Gla

sgow

Com

a Sc

ore

345678

Secondary Insult (ischemia)

Gla

sgow

Com

a Sc

ore

0 24 48 72 96

0.5

1

2

4

time (hours)

ISF

Aβ

1-x

(fold

cha

nge

from

bas

elin

e)

36 48 60 72 84 96

0.25

0.5

1

2

time (hours)

ISF

Aβ

1-x

(fold

cha

nge

from

bas

elin

e)

Global Neurological Status


Correlation with Change in Global Neurological Status

-5 -4 -3 -2 -1 0 1 2 3 4 5 6 7 8 9 10

0.25

0.5

1

2

4

8

Spearman r = 0.82, P<0.0001for |Change in GCS| ≥ 2

change in Glasgow Coma Score from baseline

Brai

n IS

F Aβ

1-x

(fold

cha

nge

from

bas

elin

e)


-5 -4 -3 -2 -1 0 1 2 3 4 5 6 7 8 9 10

0.25

0.5

1

2

4

8

Overall: Spearman r = .20, p=.009|Change in GCS| ≥ 2: Spearman r = 0.22, p=.05

change in GCS from baseline

Vent

ricu

lar

CSF

Aβ

1-x

(fol

d ch

ange

from

bas

elin

e)

-5 -4 -3 -2 -1 0 1 2 3 4 5 6 7 8 9 10

0.25

0.5

1

2

4

8

Spearman r = .65, p<.0001

change in GCS from baseline

ISF

Aβ

1-x

(fol

d ch

ange

from

bas

elin

e)

a b

Ventricular CSF vs Brain ISF

These dynamics correlating with changes in global neurological status are are not as clearly reflected in ventricular CSF amyloid-beta.


AcuteBrainInjury:[ISF Aβ ]unknown

Stabilization:[ISF Aβ ] stable

Recovery:[ISF Aβ ] increases

Secondary Insults:[ISF Aβ ] decreases

Model


What does this mean? • Amyloid-beta can be measured by

microdialysis in the human brain. – Selected scientific questions about amyloid-

beta physiology can be addressed using this technique.

– In principle, assessment of the effects of candidate therapeutics on amyloid-beta levels in the most relevant compartment- the human brain- could be made.

– However, it is not feasible or ethical to perform microdialysis studies in many patients, which may limit widespread use of this approach.

• Microdialysate amyloid-beta changes correlate with global neurological status changes. – We hypothesize that amyloid-beta levels

correlate with neurological status because both are related to synaptic activity.

– In mice, extracellular amyloid-beta levels are regulated by local synaptic activity.

– It is likely that synaptic activity is reduced following brain injury, and increases with recovery.

Synaptic Activity

?

?


– A-beta dynamics correlate with neurological status – A-beta is inversely related to markers of axonal

injury • Experimental Animal Studies

– Soluble extracellular A-beta is reduced after TBI in PDAPP and Tg2576 mice

– Insoluble intraaxonal A-beta is increased after TBI in 3xTg-AD and APP/PS1 mice

• Conclusions and Future Directions

Tau Elevations in the Brain Extracellular Space Correlate with

Reduced Amyloid-β Levels and Predict Adverse Clinical Outcomes after Severe Traumatic Brain Injury.

Tau and Amyloid-beta dynamics

Magnoni et al, Brain 2011

Tau and Amyloid-beta dynamics

Diffuse

Injury

Contusion

0

10000

20000

30000 **

Initi

al 2

4h M

ean

Tau

(pg/

ml)

Diffuse

Injury

Contusion

0

1000

2000

3000

4000 **

Initi

al 2

4h M

ean

Aβ

1-x (

pg/m

l)

Diffuse

Injury:

1-12

h

Diffuse

Injury:

61-72

h0

10000

20000

30000

40000 *

Mea

n Ta

u (p

g/m

l)

Contusion: 1

-12h

Contusion: 6

1-72h

*

A B

C D

Contusion: 1

-12h

Contusion: 6

1-72h

*

Diffuse

Injury:

1-12

h

Diffuse

Injury:

61-72

h0

1000

2000

3000 N.S.

Mea

n Aβ

1-x (

pg/m

l)


Neurofilament Light Chain- Another marker of Axonal Injury

A

B

0 10000 20000 300000

1000

2000

3000

4000 Spearman r = 0.60, p=0.013

ContusionDiffuse Injury

Initial 24h Mean Tau (pg/ml)

Initi

al 2

4h M

ean

NF-

L (p

g/m

l)

0 24 48 72 96 120 144 1680

10000

20000

30000

0

1000

2000

3000

4000Tau (pg/ml)NF-L

Time (h)

Tau

(pg/

ml)

NF-L (pg/m

l)


1 2 3 4 5 6 7 81

10

100

Spearman r = -0.40, N.S.

Clinical Outcome at 6 months(Glasgow Outcome Score-Extended)

Initi

al 2

4h M

ean

Glu

tam

ate

(mM

)

A B

1 2 3 4 5 6 7 80

10000

20000

30000

40000 Spearman r = -0.61, p=0.015


Initi

al 1

2h M

ean

Tau

(pg/

ml)

1 2 3 4 5 6 7 80

1000

2000

3000

4000

Diffuse InjuryContusion

Spearman r = 0.45, p=0.11


Initi

al 1

2h M

ean

Aβ

(pg/

ml)

C D E

1 2 3 4 5 6 7 80

1

2

3 Spearman r = 0.23, N.S.


Initi

al 2

4h M

ean

Glu

cose

(mM

)

1 2 3 4 5 6 7 810

20

30

40100

150 Spearman r = -0.45, N.S.


Initi

al 2

4h M

ean

Lact

ate/

Pyru

vate

Tau levels (but not Amyloid-beta levels) correlate with Clinical Outcomes


Unresolved Questions Regarding Amyloid-beta Dynamics after TBI: Rationale for

Development of an Animal Model

Schwetye et al, Neurobiology of Disease 2010

Normal, risingHigh, rising further

Low, recovering

TBIHuman catheters placed ?

?

?

time (h; TBI at t=0)

% b

asel

ine

Aβ 1

-x

A B

Immunohistochemistryreveals Aß deposits

Microdialysis samplesextracellular (ISF) Aß

Extracellularinsoluble

Extracellularsoluble

Intracellularinsoluble

Intracellularsoluble

microdialysis

Tissue homogenates measurea mixture of Aß in all 4 pools







microdialysis


Traumatic Brain Injury Reduces Soluble Extracellular Amyloid-β in Mice: A Combined Microdialysis- Controlled Cortical Impact Study

Katherine E. Schwetye, John R. Cirrito, Thomas J. Esparza, Christine L. Mac Donald, David M. Holtzman, and David L. Brody

Controlled Cortical Impact TBI in Mice

Brody et al., J Neurotrauma 2007

Combined Microdialysis and Controlled Cortical Impact TBI in Mice

38°

0.5 mm

Bregma -2.80 mm

2.0 mm

probe

1.3 mm canula

38°

0.5 mm

Bregma -2.80 mm

2.0 mm

probe

1.3 mm canula

A B

C D

tether inflow

outflow

probe probe tract

guide canula


Controlled Cortical Impact

TBI Acutely Reduces

Extracellular Amyloid-beta in

Mice


-18 -12 -6 0 6 12 18 24 30

50

100

150

200 SHAM (n = 6)2.0 mm (n = 6)

*

time (h; SHAM or TBI at t=0)

% b

asel

ine

Aβ 1

-x

-18 -12 -6 0 6 12 18 24 30

50

100

150

200 SHAM (n = 5)2.0 mm (n = 5)

*


% b

asel

ine

Aβ x

-40

-18 -12 -6 0 6 12 18 24 30

50

100

150

200

1.5 mm (n = 5)1.0 mm (n = 5)

*

SHAM (steady state)2.0 mm (steady state)

ns


% b

asel

ine

Aβ 1

-x

-18 -12 -6 0 6 12 18 24 30 36 42

50

100

150 SHAM (n = 4)2.0 mm (n = 4)

*


% b

asel

ine

Aβ x

-40

-12 0 12 24 36 48 60 72 84 96 108 120

50

100

150

1.5 mm (n = 5)


% b

asel

ine

Aβ 1

-xA B

C

E

D

PDAPP+/- PDAPP+/-

Tg2576+/- C57Bl6 WT

Tg2576+/- / hApoE2

Fractional Amyloid-beta Recovery is Unchanged

0.0 0.5 1.0 1.50.0

0.5

1.0

1.5

2.0

flow rate (µL/min)

[Aβ ]

/pre

-TB

I [Aβ ]

, 0.1

uL/

min

0.0 0.5 1.0 1.50.0

0.1

0.2

0.3

0.4

flow rate (µL/min)

-18 -12 -6 0 6 12 18 24 30

50

100

150


% b

asel

ine

urea

Pre Post0

25

50

75

100

% b

asel

ine

urea

-18 -12 -6 0 6 12 18 24 30

25

50

75

100

125


Mea

n %

[Aβ

] IN

C

A

Pre Post0

25

50

75

100

% [A

β] IN

K0, pre-TBI = 9.5 ± 2.2 mL/(min*mm2) K0, post-TBI = 10.2 ± 1.6 mL/(min*mm2)

B


Only PBS-soluble Amyloid-beta is affected in PDAPP mice


SHAM2.0 mm

0

10

20

30

40

SHAM2.0 mm

0

1

2

3

4*

A β(p

g/m

L) /

prot

ein

(ug/

mL)

SHAM2.0 mm

0

1

2

3 *

SHAM2.0 mm

0

5

10

15

SHAM2.0 mm

0

5

10

15

PBS

CO

RTE

X

Carbonate pH 11

SHAM2.0 mm

0

5

10

15

20

GuanidineA C E

B D F

HIP

POC

AM

PUS

Reductions in Amyloid-beta Correlate with Reduced Neuronal Activity





PDAPP and Tg2576 mice – Insoluble intra-axonal A-beta is increased after

TBI in 3xTg-AD and APP/PS1 mice • Conclusions and Future Directions

Controlled Cortical Impact Traumatic Brain Injury in 3xTg-AD Mice Causes Acute Intra-axonal

Amyloid-beta Accumulation Hien T. Tran, Frank M LaFerla, David M. Holtzman, and

David L. Brody

Amyloid-beta Pathology in White Matter

Tran et al, J Neurosci 2011

Amyloid-beta Increased only in Guan-Soluble Fraction


Amyloid-beta Colocalized With Intracellular Markers of Axonal Injury


Conclusions 1 • Abeta can be measured by microdialysis in the

human brain. • Microdialysate Abeta levels do not mimic

ventricular CSF Abeta levels. – Microdialysate reflects local extracellular fluid around the

catheter, whereas CSF drains from extracellular fluid throughout the brain.

• Abeta levels change substantially (up to 8-fold) over hours to days. – This is in contrast to the traditional view of Abeta as a

peptide that slowly accumulates over many years. – However, it is consistent with recent studies in mouse

brain (Cirrito et al) and human lumbar CSF (Bateman et al) which have demonstrated rapid dynamics.

Conclusions 2 • Surprisingly, Abeta levels increase over time after injury.

– Previous studies have suggested that total brain homogenate Abeta levels may be acutely increased after TBI.

– It is possible that there is a dissociation between soluble, extracellular Abeta levels measured by microdialysis and total brain Abeta levels, which include intracellular and extracellular, soluble and insoluble Abeta.

• Microdialysate Abeta changes may correlate with global neurological status changes. – These changes may be consistent with recent findings that

extracellular Abeta levels are governed by local synaptic activity. – It is likely that synaptic activity is reduced following TBI, and

increases with recovery. – Other correlates of low amyloid-beta (high lactate/pyruvate ratio,

low glucose, high ICP, extremes of temperature) all would be expected to impair synaptic activity.

Conclusions 3 • Amyloid-beta levels are decreased in the extracellular

fluid in PDAPP, Wild-type, and Tg-2576 mice. – This result is concordant human microdialysis studies.

• Amyloid-beta levels are increased in the insoluble form within injured axons in 3xTg-AD and APP/PS1 mice, – This pathology is consistent with what is seen in human TBI

patients and pigs subjected to experimental TBI. – This result suggests that presenilin function may drive

intraaxonal amyloid-beta production or aggregation, as both of these mice have human presenilin mutations.

• Amyloid-beta levels are not uniform, but have distinct dynamics in different compartments.

Distinct dynamics in different compartments


Normal, risingHigh, rising further

Low, recovering

TBIHuman catheters placed ?

?

?


% b

asel

ine

Aβ 1

-x

A B







microdialysis








microdialysis


How Does It All Fit Together?

SYNAPTIC ACTIVITY

mouse studies

human studies

?

? ISF Aβ


Axonal Injury (Increased tau, NF-L) Reduced Neuronal and Synaptic Activity Reduced Extracellular Soluble Amyloid-beta

Increase Insoluble Intraaxonal Amyloid-beta

Future Directions • Detection of Abeta oligomers in patients with AD:

– Question: under what circumstances could microdialysis be performed in AD patients?

• Amyloid plaque imaging (PIB)-guided catheter placement. – Coregistration of PET with CT marking catheter location

• Pharmacodynamic studies of Abeta- modifying therapeutics in the human brain. – Candidates: gamma secretase inhibitors, beta secretase inhibitors,

statins… • Diffusion Tensor Imaging- guided probe placement

– Axonal injury vs. normal white matter. – Additional microdialysis markers of axonal injury: Tau, NF-L.

• Correlations with EEG • Comparisons across ApoE genotypes.

– ApoE4 is associated with worse outcomes following TBI, and increased risk of AD. [Talk and Poster from Rachel Bennett]

Acknowledgements: • We would like to especially thank the participants and their families

for their invaluable contributions. • We appreciate the assistance of our neurosurgical collegues for

referring patients, Dr. M. Diringer for serving as the independent safety monitor, Dr. M. Pluderi for placing several microdialysis catheters, and P. Bianchi, G. Brandi, M. Carbonara, U. Deledda, L. Magni, E. Milner, J. Sagar and T. Zoerle for help collecting samples.

• Valuable discussions with Drs. R. Bateman, R. Bullock, J. Cirrito, R. Dacey, M. Diringer, L. Hillered, P. Hutchinson, J. Ladensen, W. Powers and N. Temkin are acknowledged.

• We are grateful to Eli Lilly and Co. for providing the antibodies used in the Aβ ELISAs.

• This work was supported by NIH NS049237 (DLB), a Burroughs Wellcome Career Award in the Biomedical Sciences (DLB), NIH AG13956 (DMH), and Cure Alzheimer’s Fund (DMH)

0.25 0.5 1 2 40.25

0.5

1

2

4

N.S.

Lactate (fold change)

ISF

Aβ

1-x

(fold

cha

nge)

0.25 0.5 1 2 40.25

0.5

1

2

4

N.S.

Pyruvate (fold change)

ISF

Aβ

1-x

(fold

cha

nge)

0.25 0.5 1 20.25

0.5

1

2

4

N.S.

Lactate / Pyruvate (fold change)

ISF

Aβ

1-x

(fold

cha

nge)

0.25 0.5 1 2 4 80.25

0.5

1

2

4

N.S.

Glucose (fold change)

ISF

Aβ

1-x

(fold

cha

nge)

Spearman r = -.34, P<0.0001

0.01 0.1 10.25

0.5

1

2

4

Glutamate (fold change)

ISF

Aβ

1-x

(fold

cha

nge)

0.25 0.5 1 2 40.25

0.5

1

2

4

N.S.

PtiO2 (fold change)

ISF

Aβ

1-x

(fold

cha

nge)

0.25 0.5 1 2 40.25

0.5

1

2

4

8

N.S.

Intracranial Pressure (fold change)

ISF

Aβ

1-x

(fold

cha

nge)

Spearman r = .27, P<0.0001

0.5 1 20.25

0.5

1

2

4

8

Cerebral Perfusion Pressure(fold change)

ISF

Aβ

1-x

(fold

cha

nge)

Spearman r = .34, P<0.0001

0.90 0.95 1.00 1.05 1.100.25

0.5

1

2

4

Cerebral Temperature (fold change)

ISF

Aβ

1-x

(fold

cha

nge)

A B C

D

G H

F

I

E

A B

Sedati

on Incre

ase

Sedati

on Dec

rease

0.0

0.5

1.0

1.5

2.0

2.5N.S.

Bra

in IS

F Aβ

1-x

fold

cha

nge

Propofol

Barbitu

ates

Benzo

diazep

ines

Opiates

None0

2000

4000

6000

8000

**

**

Bra

in IS

F Aβ

1-x

(pg/

ml)

Overall, brain ISF Aβ levels were highest when patients were not sedated (**P <0.005, ANOVA followed Bonferroni pairwise post-hoc testing). Sedation with propofol was associated with lower Aβ levels than sedation with benzodiazepines (** P =.00025). However, sedation was not varied systematically; less severely injured patients typically received less intensive sedation, and the choice of agents was left to the discretion of the treating physicians. B. There was no difference in the fold changes in Aβ associated with times when sedation was increased vs. times when sedation was decreased (P=0.66). Ratios are slightly greater than 1 because brain ISF Aβ in general was rising over time. When this analysis was restricted to changes in short-acting agents (primarily propofol, midazolam, and fentanyl), the results were essentially unchanged (P=0.42).

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