Kisten Et Al 2009

VOLUME 40 NUMBER 3 MARCH 2009 195

QUINTESSENCE INTERNATIONAL

Since its introduction by Haywood and

Heymann,1 home dental bleaching has been

the most commonly used method for tooth

whitening. The original technique, published

in 1989, used 10% carbamide peroxide in a

vacuum-formed custom tray. Nightguard vital

bleaching is an effective and simple method

of lightening extrinsically stained or discol-

ored teeth.2 One of the changes in the origi-

nal technique has been the use of carbamide

peroxide in concentrations greater than 10%,

claiming greater effectiveness and less time.3

However, several studies have reported

important adverse effects, such as tooth sen-

sitivity,48 soft tissue changes,9,10 hard tissue

changes,11,12 genotoxic effect in bacteria and

cultured cells,1315 cytotoxic effects,16,17 and

gingival irritation,3,79,1821 due to multiple and

prolonged exposures of bleaching agents.

According to Haywood et al,2 approximately

two-thirds of patients who undergo bleaching

treatment with carbamide peroxide experi-

ence tooth and/or gingival sensitivity. Gingival

Effect of reservoirs on gingival inflammation after home dental bleachingGiovanna A. Kirsten, DDS1/Andrea Freire, DDS2/

Antonio Adilson S. de Lima, DDS, MDS, PhD3/

Sergio Aparecido Igncio, DDS, MDS, PhD4/

Evelise M. Souza, DDS, MDS, PhD4

Objective: To evaluate the influence of reservoirs on the gingival mucosa of patients sub-

mitted to at-home bleaching with 16% carbamide peroxide. Method and Materials:

Nineteen nonsmoking male patients, 18 to 25 years of age, were submitted to home

bleaching with a 16% carbamide peroxide gel for 2 consecutive hours for 21 days. The

custom-made mouth trays were made with a reservoir on only the left side and cut anatom-

ically 1 mm beyond the gingival margin. Smears of the gingival mucosa were obtained by

the exfoliation cytology in liquid media technique before (control), immediately after, and

30 and 45 days after treatment. The samples were processed in the laboratory and evalu-

ated according to Papanicolaous criteria of malignity. Statistical analysis was carried out

by McNemar test, 2 proportions test, and Wilcoxon test with a level of significance of 1%.

Results: The presence of a reservoir in the custom tray resulted in an increase of inflam-

mation only immediately after the bleaching procedure. After 30 and 45 days, the differ-

ence between inflammation on the sides with and without a reservoir was not statistically

significant. Significant differences were found in the degree of inflammation, classified as

predominantly mild on the nonreservoir side and moderate on the reservoir side (P < .01).

Conclusions: A 16% carbamide peroxide bleaching gel caused gingival inflammation

immediately after the procedure and persisted until 45 days after the bleaching treatment.

The use of a reservoir in the custom tray for home bleaching resulted in higher rates and

higher intensity of gingival inflammation. (Quintessence Int 2009;40:195202)

Key words: carbamide peroxide, dental bleaching, exfoliative cytology, gingival

inflammation, reservoir, tray

1MDS student, School of Dentistry, Pontifical Catholic University

of Paran, Curitiba, Paran, Brazil.

2PhD student, School of Dentistry, Pontifical Catholic University

of Paran, Curitiba, Paran, Brazil.

3Professor, School of Dentistry, Pontifical Catholic University of

Paran, Curitiba, Paran, Brazil.

4Adjunct Professor, School of Dentistry, Pontifical Catholic

University of Paran, Curitiba, Paran, Brazil.

Correspondence: Dr Evelise M. Souza, Programa de Pos-gradu-

ao em Odontologia, School of Dentistry, Pontifical Catholic

University of Paran, R. Imaculada Conceio, 1155, Curitiba

PRBrazil 80215-901. Fax: 55 41 3271-1405. Email: evesouza@

yahoo.com

Kirsten.qxd 4/9/09 11:29 AM Page 195

196 VOLUME 40 NUMBER 3 MARCH 2009


Kir s ten et a l

irritation can be associated with the chem-

istry of the bleaching agent5 or the design of

the tray.22,23 However, Haywood and Heyman24

claim that most irritations appear to involve

the nightguard itself and are rarely due to

chemical irritation.

Clinical trials have evaluated gingival

health by measuring the Bleeding Index,

Gingival Index, and Plaque Index.3,21

However, these methods are visual and

demand an adequate calibration of the

examiners involved in the study. Numerical

scales2527 and questionnaires3,9,28 are also

used for the assessment of gingival irrita-

tion, but these methods rely on the

patients perception of discomfort and are

dependent on the pain threshold of each

individual. Histopathologic studies29,30 are

more accurate but too invasive, because a

biopsy of the tissue must be obtained for

cell evaluation. However, exfoliative cytology

has also been used for evaluation of

changes in the cells of oral epithelium.3034

This method is noninvasive and painless

and can be more accurate because of the

standardization of the technique and cali-

bration of the examiner.

The liquid-based cytology has been con-

sidered more advantageous than the con-

ventional technique because it produces

more homogeneous samples, due to the

reduction in the amount of mucus and blood

in the preparation, and less clumping of

epithelial cells.35 Thereby, the liquid-based

cytology technique reduces the proportion

of specimens classified as technically unsat-

isfactory for evaluation,36 decreasing false-

negative or false-positive results.33 Also, the

liquid-based cytology method allows for the

preparations of more than 1 slide per sample

collected, and because of the long storage

life of the liquid-fixative solution, the material

can be preserved and is therefore available

for additional analyses.30

The aim of this in vivo study was to evalu-

ate the effect of using reservoirs on the gin-

gival inflammation of patients undergoing

at-home dental bleaching treatment with

16% peroxide carbamide. The null hypothesis

of this study was that there would be no

difference in the gingival inflammation with

or without the use of reservoirs.

METHOD AND MATERIALS

The experimental protocol of the present

study was approved by the Ethics Committee

of the Pontifical Catholic University of Parana.

SubjectsMale university dental students 18 to 25

years old were invited to participate in the

study. For inclusion, subjects had to be non-

smokers, not submitted to dental bleaching

before the study, and without extensive

restorations in anterior teeth. For exclusion,

the subjects were submitted to a cytologic

evaluation by the liquid-based cytology to

assure that the selected subjects did not

have gingival inflammation prior to the study.

A total of 19 subjects were selected for the

dental bleaching procedure.

Custom tray fabricationAlginate impressions of the maxillary arches

of the subjects were made with Jeltrate Plus

(Dentsply), and casts were fabricated from

dental stone (Herodent, Vigodent). The

Fig 1 Stone cast with reservoirs made on only the left sideof the maxillary arch with a light-activated gingival barrier.

Fig 2 A silicone tray trimmed anatomically on the labialsurface 1 mm beyond the gingival margin.




Kir s ten et a l

reservoirs were made on only the left side of

each subjects cast with a light-activated

gingival barrier (Top Dam, FGM Dental

Products). The material was applied so that

the labial surface was covered with the

exception of 1 mm mesially, distally, cervical-

ly, and incisally (Fig 1). The trays were made

by a vacuum-formed process with a silicone

soft sheet (FGM Dental Products). The

excess was trimmed anatomically on the labial

surface 1 mm beyond the gingival margin

(Fig 2).

Bleaching procedureThe fit of the tray was carefully inspected,

and adjustments were made to ensure that

the tray did not abrade the tissue and was

well-adapted. Subjects were instructed ver-

bally and via hands-on practical demonstra-

tion in the use of the bleaching gel.27 The

16% carbamide peroxide gel (Whiteness

Perfect 16%, FGM Dental Products) was

used 2 hours per day for 21 days. The sub-

jects were asked to return the trays at the

completion of treatment.

Cells collectionExfoliated cells of the gingival margin

between the maxillary canine and premolar

were obtained by liquid-based exfoliative

cytology (Fig 3). Initially, the mouth was

rinsed with water to remove excess debris

and bacteria. The squamous epithelial cells

were collected using a cytobrush and uni-

versal collection medium kit (UCM)

(Universal Collection Medium of DNA-Citoliq

System, Digene Brasil). The cells were col-

lected before, immediately after, and 30 and

45 days after the bleaching treatment. To

compare the degree of inflammation after

dental bleaching, the cells were collected

bilaterally at the sides with and without

reservoirs.

Cytologic preparationsAn aliquot of 200 L UCM was filtered

through Filtrogene polycarbonate membrane

filters (Digene Brasil) with a pore size of 5 m

and diameter of 25 mm. The filter was placed

in prepgene press (Digene Brasil), which was

attached to glass slides. The former were

immediately fixed in absolute alcohol for 20

minutes, and smears were then stained with

routine Papanicolaou stain.

Morphologic analysisEach slide was assessed using a binocular

light microscope Olympus BX50 (Olympus)

at 400 magnification. All cellular features

were coded according to Papanicolaous

classification37: (0) insufficient smear, (1) nor-

mal smear, (2) normal smear with inflamma-

tory changes, (3) dysplastic smear, (4) smear

suggesting neoplastic changes, and (5) neo-

plastic smear. Inflammatory cytologic results

were categorized as absent (0 cells/field),

mild (1 to 5 cells/field), moderate (6 to 20

cells/field), or severe (> 20 cells/field). The

type of predominant cells (cellularity) in each

smear was also analyzed.

Statistical analysisAll data were tabulated, and statistical tests

were performed with SPSS for Windows

13.0 (SPSS). The McNemar test, two propor-

tions test, and Wilcoxon test were used in

this study with a level of significance of 1%.

Fig 3 A brush collecting cells onthe gingival margin between themaxillary canine and premolar forexfoliative cytology.




Kir s ten et a l

RESULTS

The McNemar test found statistically signif-

icant differences in the type of epithelial

cells among the control group (before) and

other periods of evaluation after the bleaching

procedure (immediately after and 30 and

45 days after), characterized by an increase

in the amount of inflammation cells (Table 1).

The presence of a reservoir in the custom

tray resulted in an increase of inflammation

only immediately after the bleaching procedure

(P = .00758), as detected by the two propor-

tions test. At 30 and 45 days after bleaching,

the difference in the amount of inflammatory

cells on the sides with and without reservoirs

was not statistically significant.

The Wilcoxon test detected statistically

significant differences in the inflammation

rates on the reservoir side immediately after

bleaching when compared to 30 and 45

days. However, these were not statistically dif-

ferent from each other. On the other side of

the tray, without a reservoir, the 3 observation

periods showed no statistical differences in

the inflammation prevalence (P > .01).

Figure 4 demonstrates the amount of

inflammatory cells before bleaching (control)

and immediately after bleaching on the sides

without and with a reservoir of the tray.

Statistically significant differences were

found in the inflammation intensity between

both sides of the tray immediately following

and 45 days after bleaching (P < .01). At

Table 1 No. and percentage of patients submittedto home bleaching with 16% carbamide peroxide with inflammation during differentperiods of observation (n = 19)

Period of observation Reservoir Inflammation (%)

Before (control) 0 (0)a

Immediately Without 13 (68.0)b

following With 19 (100.0)c

30 days Without 5 (26.3)b

With 3 (15.8)b

45 days Without 8 (42.1)b

With 4 (21.1)b

Groups connected by the same letter are not statistically different (P > .01).Fig 4 Liquid-based cytology (Papanicolaou, origi-nal magnification 400). (a) Normal aspect of thecells before bleaching (control). (b) Epithelial andinflammatory cells immediately after bleaching onthe nonreservoir side of the tray. (c) Epithelial andinflammatory cells immediately after bleaching onthe reservoir side of the tray.

a b

c




Kir s ten et a l

these periods of evaluation, there was a

prevalence of mild inflammation at the side

of the tray without a reservoir (Fig 5) and

moderate inflammation at the side with a

reservoir (Fig 6).

DISCUSSION

The inflammatory response is closely related

to the process of repair. Inflammation serves

to destroy, dilute, or isolate the injurious

agent, but in turn, it sets into motion a series

of events that, as far as possible, heal and

reconstitute the damaged tissue. On the

other hand, inflammation and repair may be

potentially harmful. Chronic inflammation is

considered to be inflammation of prolonged

duration (weeks or months) in which active

inflammation, tissue destruction, and attempts

at healing are proceeding simultaneously.

This kind of inflammation frequently begins

insidiously, as a low-grade and often asymp-

tomatic response.38

The signs of gingival inflammation after

dental bleaching treatments may not be

observed by visual inspection. Therefore, an

accurate method, such as exfoliative cytology,

must be used to confirm the presence or

absence of inflammatory cells, as well as the

intensity of the inflammation.3436

Although some authors19,25 claimed that

patients experience mild gingival sensitivity

that lasts for a few days after dental bleach-

ing, the present study demonstrated that

16% carbamide peroxide gel, used 2 hours

per day for 3 weeks, was capable of causing

morphologic changes in the human gingival

epithelium not only immediately following

the treatment but also up to 45 days after.

Immediately after treatment, the reservoir

side exhibited a prevalence of moderate

inflammation, whereas on the nonreservoir

side, a greater incidence of mild inflammation

was found. At the 30-day observation, the

intensity of inflammation was similar for both

sides of the tray. By contrast, at the 45-day fol-

low-up, inflammation was largely absent on

the nonreservoir side, while a prevalence of

70

60

50

40

30

20

10

0

% o

f in

flam

ed c

ells

Immediatelyafter

30 days 45 days

Absent

Mild

Moderate

Severe

Immediatelyafter

70

60

50

40

30

20

10

0

% o

f in

flam

ed c

ells

30 days 45 days

Absent

Mild

Moderate

Severe

Fig 5 Inflammation intensity on the nonreservoirside after different periods of observation.

Fig 6 Inflammation intensity on the reservoir sideafter different periods of observation.




Kir s ten et a l

moderate inflammation was seen on the

reservoir side. Additionally, the reservoir in

the tray resulted in even more inflammation in

45 days than the nonreservoir side immedi-

ately after the bleaching procedure.

One possible explanation for these find-

ings could be related to custom-made trays.

The anatomic design, 1 mm beyond the gin-

gival margin, and the flexibility of the silicone

tray could have allowed extrusion of the

bleaching gel. Perhaps a more rigid material

and/or a trimming parallel to the incisal or

occlusal plane could have avoided or

reduced the gingival irritation. Although tray

designs seek to avoid covering the attached

gingiva, the interdental papillae are still

exposed to the bleaching gel.7 Therefore,

total avoidance of soft tissue contact is

impossible.

According to Haywood et al,8 the pres-

ence of reservoirs decreases the retention of

the tray, allowing more room for the gel but

also reducing the adaptation of the tray. This

could explain why the presence of a reservoir

caused more gingival inflammation in this

study. Accordingly, the examiners of the pres-

ent study observed clinical signs of inflamma-

tion in the subjects after 1 week of bleaching

on only the reservoir side of the tray.

In a clinical investigation by Matis et al26 no

difference was found in gingival sensitivity

between areas bleached with and without

reservoirs. However, this parameter was evalu-

ated based on a daily record and classification

of tooth and gingival sensitivity into categories

attributed by the patients.

The presence of severe inflammation 45

days after bleaching was observed only in 2

patients, 1 in the reservoir side and the other

in the side without reservoir. This fact could

be explained by the possible residual effect

of the bleaching agent or the plaque accu-

mulation due to patients negligent hygiene.

Additionally, there is an individual factor

involved in the gingival tissue response to

toxic agents, since the concentration of sali-

vary modulators varies among patients. The

role of saliva and its modulators must be

taken into account: They act as protective

agents of gingival tissue. Tipton et al39

demonstrated that whole saliva, lactoperoxi-

dase, and catalase, at sufficient concentra-

tions, could provide complete or nearly com-

plete protection from the toxic effects of car-

bamide peroxide, removing the hydrogen

peroxide generated during its degradation.

Bleaching agents are cytotoxic to human

gingival fibroblasts, increasing the effects on

cell viability and morphology, and on the pro-

liferation and production of fibronectin and

collagen.39 In an in vitro study, Koulaouzidou

et al17 investigated the cytotoxic effect of a

bleaching agent on 2 fibroblast cell lines and

found that both were sensitive to urea perox-

ide. These authors17 suggested that the

potential damage to oral tissues in vivo may

be considerable because of the direct and

long-term exposure of the tissues to the

bleaching agents.

Therefore, indiscriminate, frequent, or

prolonged treatments, even under profes-

sional supervision, might increase the potential

damage of bleaching agents to the

periodontal tissues, leading to chronic

inflammation. Because the presence of

inflammatory cells was observed 45 days

after the bleaching procedure, this treatment

must not be given frequently, so as to allow

healing of the injured tissues. The dental cli-

nician must be careful to indicate and super-

vise bleaching procedures and be conscious

about the damage that they could cause to

their patients.

CONCLUSIONS

The 16% carbamide peroxide home bleach-

ing caused gingival inflammation not only

immediately after the procedure but also until

45 days following the bleaching treatment.

The use of a reservoir in the custom tray for

home bleaching resulted in higher rates and

higher intensity of gingival inflammation.

ACKNOWLEDGMENTS

The authors thank FGM Dental Products for the supply

of bleaching agents and trays.




Kir s ten et a l

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