ORIGINAL Branco ARTICLE et al
Early Supra- and Subgingival Plaque Formation in Experimental Gingivitis in Smokers and Never-Smokers Paula Brancoa/Patricia Weidlichb/Rui Vicente Oppermannc/ Cassiano Kuchenbecker Rösingc Purpose: To evaluate supragingival and subgingival plaque formation on the dentogingival area in smokers and neversmokers using the experimental gingivitis model and a plaque scoring system that considers the presence of an area free of plaque between plaque and the gingival sulcus called the plaque free zone (PFZ). Materials and Methods: Male volunteers, 9 current smokers and 10 never-smokers, refrained from oral hygiene procedures in the maxillary incisors and canines (test teeth) for 25 days. Under conditions of clinically healthy gingiva (phase 1) and gingival inflammation (phase 2), the supragingival plaque formation pattern was observed for 4 days in the dentogingival area. Gingival crevicular fluid was also measured. Plaque was dyed with fucsine and its presence was recorded by a calibrated examiner based on a 3-criteria scoring system: 0 – absence of stained plaque; 1 – presence of stained plaque and supragingival PFZ; 2 – presence of stained plaque and absence of PFZ, indicating that subgingival plaque formation has taken place. Results: In both phases, smokers presented a significantly lower relative frequency of sites with subgingival plaque compared to never-smokers (P < 0.001). Mean gingival crevicular fluid was significantly higher in the presence of gingival inflammation for both groups (P = 0.001), whereas smokers demonstrated a significantly lower frequency of gingival bleeding than did non-smokers (23.6% vs 66.1%; P < 0.001). Conclusion: Smokers presented significantly lower percentages of sites with subgingival plaque in all experimental periods and presented less gingival inflammation as shown by GBI and gingival crevicular fluid quantification. Key words: dental plaque, gingival crevicular fluid, gingivitis, smoking Oral Health Prev Dent 2015;13:13-20 doi: 10.3290/j.ohpd.a32669
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t has been demonstrated that dental plaque initially develops on the tooth surface adjacent to the gingival margin (Quirynen and van Steenberghe, 1989). Supragingival plaque accumulation may lead to gingivitis development within 2 to 3 weeks (Löe et al, 1965; Theilade et al, 1966), subject to interindividual variation or even to specific microbiota (Page et al, 1997; Haffajee and Socransky, 2001).
a
Master’s Student, Graduate Programme in Dentistry, Lutheran University of Brazil, Canoas, RS, Brazil.
b
Associate Professor, Section of Periodontology, Faculty of Dentistry, Federal University of Rio Grande do Sul, Porto Alegre, RS, Brazil.
c
Professor, Section of Periodontology, Faculty of Dentistry, Federal University of Rio Grande do Sul, Porto Alegre, RS, Brazil.
Correspondence: Patricia Weidlich, Rua Ramiro Barcelos, 2492, Porto Alegre, RS, Brazil 90035-003. Tel: +55-51-3308-5318, Fax: +55-51-3308-5318. Email: [email protected]
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Submitted for publication: 11.07.12; accepted for publication: 26.05.13
Bass (1946) observed the presence of a plaquefree area in extracted teeth, which was limited coronally by the gingival margin and on the root surface by the coronal portion of the junctional epithelium. Brady (1973) called this the ‘plaque free zone’ (PFZ), delimited apically by the periodontium and cervically by the microbial deposit. Some studies have demonstrated that the PFZ is not stained by disclosing agents and can be identified clinically as a thin line between the gingival margin and the stained plaque. It has been observed in extracted teeth and in vivo in analyses of both supragingival and subgingival plaque formation processes (Brady, 1973; Saglie et al, 1975; Friedman et al, 1992; Weidlich et al, 2001). Studies concerning supragingival plaque are mostly performed on extracted teeth, making it impossible to evaluate the role of the PFZ in the development of microbial deposits and its relation-
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ship to the gingival margin. It is clear that some external factors such as smoking, which has been associated with a higher prevalence and greater severity of periodontitis, may interfere in this process. However, the biological mechanisms involved in tobacco consumption are not completely understood. Pioneer studies demonstrated that smokers presented higher amounts of plaque and supragingival calculus (Sheiham, 1971; Preber and Kant, 1973). The majority of authors, however, corroborate that there are no significant differences in the plaque accumulation pattern between smokers and nonsmokers (Rosa et al, 2000; Shiloah et al, 2000;
day 0
days 1, 2, 3, and 4
Gingival crevicular fluid quantification Professional tooth cleaning Plaque dye VPI, GBI
Gingival crevicular fluid quantification Plaque dye PFZ assessment
Calsina et al, 2002). Additionally, local effects of nicotine are related to the masking of clinical signs of gingival inflammation due to vasoconstriction, which slows the blood flow, leading to less marginal bleeding (Bergström, 1990; Gomes et al, 2007). Therefore, better understanding of the process of dental plaque formation is still necessary, including an analysis of variables that may interfere with its development. In this sense, the PFZ may have some clinical implications for the transition from supra- to subgingival plaque as well as the inflammatory process. Intervention studies that evaluate the possible influence of smoking on plaque development and gingival inflammation may provide additional information in the role of tobacco in the pathogenesis of periodontal diseases. The aim of the present study was to evaluate supragingival and subgingival plaque formation on the dentogingival area by means of the experimental gingivitis model and a plaque scoring system that considers the presence of a plaque free zone (PFZ) in smokers and never-smokers.
MATERIALS AND METHODS Study design
days 7 and 14
Gingival crevicular fluid quantification
day 21
Gingival crevicular fluid quantification Professional tooth cleaning Plaque dye VPI, GBI
days 22, 23 and 24
Gingival crevicular fluid quantification Plaque dye PFZ assessment VPI, GBI Professional tooth cleaning
day 25
Gingival crevicular fluid quantification Plaque dye PFZ assessment
Fig 1 Study flowchart. VPI: visible plaque index; GBI: gingival bleeding index; PFZ: plaque-free zone.
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The present study was performed by means of a parallel experimental gingivitis design in maxillary incisors and canines (test teeth). Nineteen male volunteers, selected after an interview and periodontal examination, were included in the study: 9 current smokers (mean of 15 cigarettes per day) aged 18–42 years (mean 28.6 ± 6.7 years) and 10 never-smokers aged 19–28 years (mean 23.8 ± 3.1 years). Individuals with caries, misaligned and/or restored/decayed teeth in the cervical or middle third of the buccal area of the test teeth, attachment loss, antibiotic treatment in the last 6 months and need of prophylactic dental treatment were not included. The study protocol was approved by the Institutional Review Board of the Lutheran University of Brazil, and all participants signed an informed consent form. Participants were instructed not to modify eating habits, to use a standard dentifrice without any antimicrobial agents (Close-Up, Unilever; São Paulo, Brazil) and to refrain from oral hygiene procedures in the maxillary dentition during the whole 25-day experimental period (Fig 1). In phase 1, starting with clinical gingival health, the supragingival plaque formation pattern in the dentogingival area
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was observed daily for 4 days. After day 4, participants continued refraining from oral hygiene up to day 21, when professional prophylaxis was performed to allow the observation of supragingival plaque formation under conditions of gingival inflammation (phase 2). During the experimental period, gingival fluid secretion was utilised as an inflammatory indicator. Test teeth were submitted to gingival fluid quantifi cation according to Egelberg (1966). After gentle rinsing with water, a Periopaper (Oraflow; New York, NY, USA) strip was inserted in the entrance of the buccal crevice at each test tooth for 3 min. Subsequently, the strips were transferred to petri dishes, wet with one drop of 0.2% nihidrine solution and dried at 75°C in a microbiological oven for 15 min. The area of the strip dyed by nihidrine was measured digitally using Adobe Photoshop. Gingival crevicular fluid quantification took place on days 0 to 4, 7, 14 and 21 to 25. The clinical evaluation of the presence of plaque and gingivitis was assessed by the Visible Plaque Index (VPI) and Gingival Bleeding Index (GBI) (Ainamo and Bay, 1975) at days 0, 21 and 25. At the beginning of each phase (days 0 and 21), professional prophylaxis was performed on the test teeth with rubber cups (KG Sorensen; Viking, Brazil), prophy paste (Pert, SS White; Rio de Janeiro, Brazil) and dental floss (Sanifill; São Paulo, Brazil), in order to obtain completely plaque free surfaces as disclosed by fucsine. To stain the plaque, teeth were rinsed with water, dried and isolated with cotton rolls. Fucsine solution (Replanic T, Iodontec; Porto Alegre, Brazil) was applied with a syringe and disposable needles (Becton Dickinson; Franklin Lakes, New Jersey, USA) in order to better access the sulcus area without touching the surface. After 30 s, teeth were rinsed, dried and evaluated with the scoring system proposed by Maliska et al (2006). In brief, the scoring system evaluates the initial plaque formation in the dentogingival area and is based on 3 criteria: 0 – absence of stained plaque; 1 – presence of stained plaque and a supragingival PFZ; 2 – presence of stained plaque and absence of PFZ, indicating that subgingival plaque formation has taken place. The assessment was performed on the buccal surface of the test teeth. A marking pen (Pilot Pen do Brasil; São Paulo, Brazil) was used to indicate 2 reference points on the buccal gingiva, 1–2 mm apical to the gingival margin, longitudinally dividing the buccal surface in three equal thirds: mesio-buccal, buccal and disto-buccal. The same
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trained and calibrated examiner (PB) who was supervised by the gold-standard examiner (PW) in a previous study (Maliska et al, 2006) performed all assessments. To test for reproducibility, 40 patients from the University dental clinic were assessed twice by the gold-standard examiner and the examiner of the present study. Kappa statistics demonstrated values of 0.79 and 0.76 for interand intraexaminer agreement, respectively. After the end of the experiment, professional prophylaxis was performed in order to facilitate the re-establishment of gingival health. All participants were followed-up until basal amounts of gingival crevicular fluid were present.
Statistical analysis The mean volume of gingival crevicular fluid was obtained by the mean linear measurement of the paper strip for smokers and never-smokers on each test day of phases 1 and 2. Intragroup comparisons were tested using the dependent t-test and intergroup comparisons using repeated-measures ANOVA. The frequency distribution of each PFZ score was obtained and compared over time using the Kruskall-Wallis test and intergroup comparisons were performed with the Wilcoxon signedrank test. Statistical significance was set at 5%.
RESULTS Table 1 demonstrates the data on VPI, GBI and probing depth (PD) on days 0, 21 and 25 for both groups. No statistically significant differences were observed in the percentage of sites without visible plaque (VPI = 0) and in mean PD between smokers and never-smokers at baseline. The percentage of gingival bleeding varied at the beginning of phase 2, with never-smokers presenting higher levels of gingival bleeding after 21 and 25 days of not performing oral hygiene as compared to smokers. Tables 2 and 3 present the mean volumes of gingival crevicular fluid in smokers and never-smokers throughout the study. When plaque accumulation occurred in the absence of gingival inflammation (phase 1), a gradual and significant increase was observed in the amount of gingival crevicular fluid during the 4-day period within each group. Neversmokers and smokers did not present significant differences in gingival crevicular fluid volume, except on day 3, when smokers presented a lower
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Table 1 Visible plaque index (VPI), gingival bleeding index (GBI) and pocket depth (PD) for smokers and never-smokers
Never-smokers
Smokers
P*
Beginning of phase 1 (day 0)
97
98.8
0.29
Beginning of phase 2 (day 21)
30.4
22.2
0.07
End of phase 2 (day 25)
54.8
53.5
0.45
Beginning of phase 1 (day 0)
100
100
Beginning of phase 2 (day 21)
33.9
76.4
< 0.01
End of phase 2 (day 25)
65.5
93.8
< 0.01
1.12 ± 0.36
1.15 ± 0.35
0.51
% sites with VPI = 0
% sites with GBI=0
PD day 0 (mean ± SD) * Independent samples t-test.
Table 2 Gingival crevicular fluid volume (mean ± SD) in the absence of gingival inflammation in never-smokers and smokers Absence of gingival inflammation
Never-smokers
day 0
day 1
day 2
day 3
day 4
0.02±0.04
0.06±0.13
0.10±0.13
0.15±0.17
0.20±0.20
0.001
0.001
0.001
0.001
0.04±0.07
0.05±0.09
0.12±0.15
0.24±0.18
0.001
0.001
0.001
0.001
0.32
0.03
0.37
0.31
P* Smokers
0.05±0.13
P* P#
0.12
* Intragroup comparison, dependent samples t-test. # Intergroup comparisons, repeated measures ANOVA.
Table 3 Gingival crevicular fluid volume (mean ± SD) in the presence of gingival inflammation in never-smokers and smokers Presence of gingival inflammation
Never-smokers
day 0
day 1
day 2
day 3
day 4
0.68±0.31
0.63±0.41
0.58±0.25
0.60±0.23
0.65±0.27
0.001
0.001
0.001
0.001
0.48±0.28
0.51±0.28
0.51±0.29
0.61±0.31
0.001
0.001
0.001
0.001
0.02
0.17
0.08
0.52
P* Smokers
0.64±0.30
P* P#
0.49
* Intragroup comparison, dependent samples t-test. # Intergroup comparisons, repeated measures ANOVA.
mean volume of gingival crevicular fluid. Considering gingival crevicular fluid in the presence of gingival inflammation (phase 2), intragroup comparisons demonstrate a significant increase in its volume during the 4-day period, with intergroup differences observed on day 1 between smokers and neversmokers. The mean gingival crevicular fluid volume was significantly higher in the presence of gingival
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inflammation for both groups. However, no statistically significant differences were observed related to smoking status. Figure 2 demonstrates the formation of subgingival plaque in never-smokers and smokers in the absence of gingival inflammation (phase 1). A significant increase was found in percent of sites with subgingival plaque (PFZ score = 2) during the 4
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never-smokers 100
*
*
*
smokers *
100
0
9
14.1 80
80
26.2
26.4 33.3
42.9 % sites
60
% sites
60
40
40 68.1 67.9
20
71.4 2.4
17.9
0
day 1
day 2
14.9
56.5 0.6 day 3
77.8
20
85.1
70.8
0
0
day 4
66.7 day 1
22.9 day 2
2.8 day 3
22 0 day 4
Fig 2 Early subgingival plaque formation in the absence of gingival inflammation in smokers and never-smokers. PFZ = 0: absence of stained plaque (dark gray); PFZ = 1: presence of stained plaque and supragingival PFZ (light gray); PFZ = 2: presence of stained plaque and absence of PFZ (presence of subgingival plaque) (black). *Intergroup comparisons, Friedman and Wilcoxon tests (P < 0.001).
never-smokers 100
*
*
*
smokers *
100
6.3
14.1
60
60 % sites
80
% sites
80
51.8 40
19.4
54.2
50 40
71.5 20
0
84.5
64.9
19.6 day 1
92.9
57.6 20
45.2 3 day 2
15.5 0 day 3
7.1 0 day 4
41 0
43.8 day 1
22.9 day 2
4.9 day 3
24.3 4.2 day 4
Fig 3 Early subgingival plaque formation in the presence of gingival inflammation in smokers and never-smokers. PFZ = 0: absence of stained plaque (dark gray); PFZ = 1: presence of stained plaque and supragingival PFZ (light gray); PFZ = 2: presence of stained plaque and absence of PFZ (presence of subgingival plaque) (black). *Intergroup comparisons, Friedman and Wilcoxon tests (P < 0.001).
days of plaque accumulation for both groups. Throughout the experiment, smokers presented a significantly lower relative frequency of sites with subgingival plaque compared to never-smokers. Similar results can be observed in Fig 3, when the formation of subgingival plaque occurred in the presence of gingival inflammation (phase 2). It was observed that the presence of gingival inflamma-
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tion accelerated the formation of subgingival plaque both in smokers and never-smokers. However, smokers presented significantly lower percentages of sites with subgingival plaque (PFZ score = 2) in all experimental evaluations as compared to never-smokers.
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DISCUSSION The present study analysed the plaque formation pattern in smokers and never-smokers under different gingival inflammatory conditions after an experimental gingivitis model and demonstrated that the presence of gingival inflammation accelerates subgingival plaque formation both in smokers and never-smokers. Additionally, significantly lower frequencies of sites with subgingival plaque were found in smokers independent of the inflammatory condition. Exclusively male volunteers were included in order to avoid hormonal fluctuations as confounders of the results (Lindhe and Attsfröm, 1967). Also, it is important to highlight that the examiner was calibrated with good levels of reproducibility, both intra-examiner and compared to the gold-standard examiner from the original study that developed the scoring system (Landis and Koch, 1977; Maliska et al, 2006). Plaque was assessed using the scoring system developed by Maliska (2006) and considers the presence of a PFZ. Knowledge about the transformation of supragingival deposits into subgingival plaque is incomplete; however, some studies report the presence of a plaque-free zone located apical to the supragingival plaque and coronal to the gingival margin (Quyrinen and van Steenberghe, 1985; Bergström, 1990; Weidlich et al, 2001). This supragingival PFZ is present during initial plaque formation, but it vanishes after 72–96 h of accumulation. The disappearance of the PFZ demonstrates that subgingival plaque has already developed. Scanning electron microscopic analyses demonstrate that subgingival plaque formation evolves from the supragingival plaque. Concomitant with these observations, an increase in gingival crevicular fluid volume was observed, together with a coronal migration of the gingival margin due to oedema, resulting in the establishment of subgingival plaque and a subgingival PFZ (Weidlich et al, 2001). The PFZ scoring system developed by Maliska et al (2006) has been shown to be reproducible and allows evaluation of the transformation of supragingival into subgingival plaque. The distribution of scores 0, 1 and 2 of the system used here demonstrated that the PFZ was pronounced at the beginning of the study, but gradually became less apparent during 4 days of plaque accumulation. It should be emphasised that a score of 2 means that subgingival plaque is already present. The moment at which a site no longer has a score
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of 1 should be considered the initiation of subgingival plaque formation. In this context, smokers presented lower frequencies of subgingival plaque than did never-smokers. This could be explained by the local action of cigarette components on periodontal tissues that may affect both microbial challenge and host response. The majority of studies that observe lower degrees of gingivitis or marginal bleeding in smokers suppose that this is related to nicotine-induced peripheral vasoconstriction, resulting in a decreased inflammatory pattern as well as a decreased host defense (Bergström, 1990; Giannopoulou et al, 2003; Gomes et al, 2007). Differences between smokers and non-smokers in relation to clinical parameters are important. Our results demonstrated an increase in VPI during the study which did not differ between smokers and never-smokers; this was similar to the results of several other studies that could not show differences in the plaque formation pattern (Shiloah et al, 2000; Calsina et al, 2002; Salvi et al, 2005; Gomes et al, 2007). However, some reports found higher amounts of plaque in smokers than in nonsmokers (Ainamo, 1971; Preber and Bergström, 1985; Machuca et al, 2000). In relation to GBI, the present study found that smokers exhibited lower levels of clinically detectable gingivitis than neversmokers. These differences were only observed on days 21 and 25, since GBI was not assessed at other time points in order to not interfere in the plaque analysis by disrupting the formed plaque with the probe. These findings corroborate those of Ainamo (1971), in which the gingival index was lower in individuals that smoked more than 20 cigarettes per day, and the results of Preber and Bergström (1985), which demonstrated that gingival redness and bleeding were lower in individuals who smoked at least 20 cigarettes per day. On the other hand, Salvi et al (2005) demonstrated that smokers and never-smokers had similar patterns of gingival inflammation during 21 days of experimental plaque accumulation. Different lifetime smoking exposure can be observed among studies and this may partially explain the differences related to gingival bleeding in smokers. In phase 2, from day 22, a new occurrence of scores 0 and 1 was observed, with a lower frequency of score 2 both in smokers and in never-smokers. From day 22 to the end of the experiment, a marked decrease in scores 0 and 1 was observed, with an increase in the presence of subgingival plaque (score 2). The conversion of scores 0 and 1 into score 2 in never-smokers was faster that in
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smokers. This observation is of extreme interest in the context of the present investigation. Although an explanation for this phenomenon is not a direct objective of this study, some suppositions can be made. First, the utilised variable – PFZ – has a different concept from the majority of published studies and may be an interesting tool for studies that aim at investigating plaque formation patterns. Secondly, the role of gingival inflammation as a modifier of plaque formation should be highlighted. It is well-established that de novo plaque formation is faster in inflamed that in non-inflamed areas (Hillam and Hull, 1977; Ramberg et al, 1994; Ramberg et al, 1995). This fact was clearly observed in the present study, especially in never-smokers. It would be unwarranted to suppose that smoking inhibits plaque formation. The present study demonstrated that smokers showed less gingival inflammation both in terms of GBI and gingival crevicular fluid volume. Therefore, it may be postulated that the lower level of inflammation observed contributed to less plaque accumulation, as it is well established that gingival inflammation favours plaque accumulation. This is confirmed by the differences observed between smokers and neversmokers in the present study. On the other hand, it is important to point out that smoking modifies the host response, leading to higher periodontal breakdown, probably by different mechanisms of tissue destruction (Salvi et al, 1997; Bergström et al, 2000; Johnson and Hill, 2004; Moimaz et al, 2009). A study that evaluated the microbiological and pro-inflammatory characteristics during early plaque formation demonstrated that smokers present an initial plaque colonisation that is diverse, unstable and contains more periodontopathogenic microorganisms than in non-smokers. Additionally, smokers presented higher levels of pro-inflammatory mediators as compared to non-smokers since the first day of plaque accumulation (Kumar et al, 2011). In this sense, smoking seems to play a central role both in microbiota composition and in host response starting with early plaque formation.
CONCLUSION The present study analysed supragingival and subgingival plaque formation in smokers and neversmokers under different gingival inflammatory conditions and concluded that smokers presented significantly lower percentages of sites with subgingival plaque during the entire experimental period
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and less gingival inflammation as shown by both GBI and gingival crevicular fluid volume.
REFERENCES 1. Ainamo J, Bay I. Problems and proposals for recording gingivitis and plaque. Int Dent J 1975;25:229–235. 2. Ainamo J. The seeming effect of tobacco consumption on the occurrence of periodontal disease and dental caries. Suom Hammaslaak Toim 1971;67:87–94. 3. Bass CC. A demonstrable line on extracted teeth indicating the location of the outer border of the epithelial attachment. J Dent Res 1946;25:401–415. 4. Bergström J. Oral hygiene compliance and gingivitis expression in cigarette smokers. Scand J Dent Res 1990;98:497–503. 5. Bergström J. Photogrammetric registration of dental plaque accumulation in vivo. Acta Odontol Scand 1981;39:275–284. 6. Bergström J, Eliasson S, Dock J. A 10-year prospective study of tobacco smoking and periodontal health. J Periodontol 2000;71:1338–1347. 7. Brady JM. A plaque-free zone on human teeth--scanning and transmission electron microscopy. J Periodontol 1973;44:416–428. 8. Calsina G, Ramón JM, Echeverría JJ. Effects of smoking on periodontal tissues. J Clin Periodontol 2002;29:771–776. 9. Egelberg J. Permeability of the dento-gingival blood vessels. 1. Application of the vascular labelling method and gingival fluid measurements. J Periodontal Res 1966;1: 180–191. 10. Friedman MT, Barber PM, Mordan NJ, Newman HN. The „plaque-free zone“ in health and disease: a scanning electron microscope study. J Periodontol 1992;63:890–896. 11. Giannopoulou C, Cappuyns I, Mombelli A. Effect of smoking on gingival crevicular fluid cytokine profile during experimental gingivitis. J Clin Periodontol 2003;30:996–1002. 12. Gomes SC, Piccinin FB, Susin C, Oppermann RV, Marcantonio RA. Effect of supragingival plaque control in smokers and never-smokers: 6-month evaluation of patients with periodontitis. J Periodontol 2007;78:1515–1521. 13. Haffajee AD, Socransky SS. Relationship of cigarette smoking to the subgingival microbiota. J Clin Periodontol 2001;28:377–388. 14. Hillam DG, Hull PS. The influence of experimental gingivitis on plaque formation. J Clin Periodontol 1977;4:56–61. 15. Johnson GK, Hill M. Cigarette smoking and the periodontal patient. J Periodontol 2004;75:196–209. 16. Kornman KS, Loesche WJ. The subgingival microbial flora during pregnancy. J Periodontal Res 1980;15:111–122. 17. Landis JR, Koch GG. The measurement of observer agreement for categorical data. Biometrics 1977;33:159–174. 18. Lindhe J, Attsfröm R. Gingival exudation during the menstrual cycle. J Periodontal Res 1967;2:194–198. 19. Löe H, Theilade E, Jensen SB. Experimental gingivitis in man. J Periodontol 1965;36:177–187. 20. Machuca G, Rosales I, Lacalle JR, Machuca C, Bullón P. Effect of cigarette smoking on periodontal status of healthy young adults. J Periodontol 2000;71:73–78.
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Branco et al 21. Maliska AN, Weidlich P, Gomes SC, Oppermann RV. Measuring early plaque formation clinically. Oral Health Prev Dent 2006;4:273–278. 22. Moimaz SA, Zina LG, Saliba O, Garbin CA. Smoking and periodontal disease: clinical evidence for an association. Oral Health Prev Dent 2009;7:369–376. 23. Page RC, Offenbacher S, Schroeder HE, Seymour GJ, Kornman KS. Advances in the pathogenesis of periodontitis: summary of developments, clinical implications and future directions. Periodontol 2000 1997;14:216–248. 24. Preber H, Bergström J. Occurrence of gingival bleeding in smoker and non-smoker patients. Acta Odontol Scand 1985;43:315–320. 25. Preber H, Kant T. Effect of tobacco-smoking on periodontal tissue of 15-year-old schoolchildren. J Periodontal Res 1973;8:278–283. 26. Quirynen M, van Steenberghe D, Vuylsteke M. The possibility of measuring plaque growth in vivo within 24 hours. J Periodontal Res 1985;20:321–328. 27. Quirynen M, van Steenberghe D. Is early plaque growth rate constant with time? J Clin Periodontol 1989;16:278– 283. 28. Ramberg P, Axelsson P, Lindhe J. Plaque formation at healthy and inflamed gingival sites in young individuals. J Clin Periodontol 1995;22:85–88. 29. Ramberg P, Lindhe J, Dahlén G, Volpe AR. The influence of gingival inflammation on de novo plaque formation. J Clin Periodontol 1994;21:51–56. 30. Rosa GM, Lucas GQ, Lucas ON. Study of the crevicular fluid flow rate in smokers. Acta Odontol Latinoam 2000;13:51–60.
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31. Saglie R, Johansen JR, Tollefsen T. Plaque-free zones on human teeth in periodontitis. J Clin Periodontol 1975;2:190–197. 32. Salvi GE, Lawrence HP, Offenbacher S, Beck JD. Influence of risk factors on the pathogenesis of periodontitis. Periodontol 2000 1997;14:173–201. 33. Salvi GE, Ramseier CA, Kandylaki M, Sigrist L, Awedowa E, Lang NP. Experimental gingivitis in cigarette smokers: a clinical and microbiological study. J Clin Periodontol 2005; 32:441–447. 34. Sheiham A. Periodontal disease and oral cleanliness in tobacco smokers. J Periodontol 1971;42:259–263. 35. Shiloah J, Patters MR, Waring MB. The prevalence of pathogenic periodontal microflora in healthy young adult smokers. Periodontol 2000;71:562–567. 36. Theilade E, Wright WH, Jensen SB, Löe H. Experimental gingivitis in man. II. A longitudinal clinical and bacteriological investigation. J Periodontal Res 1966;1:1–13. 37. Trombelli L, Tatakis DN, Scapoli C, Bottega S, Orlandini E, Tosi M. Modulation of clinical expression of plaque-induced gingivitis. II. Identification of ‘high-responder’ and ‘low-responder’ subjects. J Clin Periodontol 2004;31:239– 252. 38. Van der Weijden GA, Timmerman MF, Danser MM, Nijboer A, Saxton CA, Van der Velden U. Effect of pre-experimental maintenance care duration on the development of gingivitis in a partial mouth experimental gingivitis model. J Periodontal Res 1994;29:168–173. 39. Weidlich P, Lopes de Souza MA, Oppermann RV. Evaluation of the dentogingival area during early plaque formation. J Periodontol 2001;72:901–910.
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Early Supra- and Subgingival Plaque Formation in Experimental Gingivitis in Smokers and Never-Smokers Paula Brancoa/Patricia Weidlichb/Rui Vicente Oppermannc/ Cassiano Kuchenbecker Rösingc Purpose: To evaluate supragingival and subgingival plaque formation on the dentogingival area in smokers and neversmokers using the experimental gingivitis model and a plaque scoring system that considers the presence of an area free of plaque between plaque and the gingival sulcus called the plaque free zone (PFZ). Materials and Methods: Male volunteers, 9 current smokers and 10 never-smokers, refrained from oral hygiene procedures in the maxillary incisors and canines (test teeth) for 25 days. Under conditions of clinically healthy gingiva (phase 1) and gingival inflammation (phase 2), the supragingival plaque formation pattern was observed for 4 days in the dentogingival area. Gingival crevicular fluid was also measured. Plaque was dyed with fucsine and its presence was recorded by a calibrated examiner based on a 3-criteria scoring system: 0 – absence of stained plaque; 1 – presence of stained plaque and supragingival PFZ; 2 – presence of stained plaque and absence of PFZ, indicating that subgingival plaque formation has taken place. Results: In both phases, smokers presented a significantly lower relative frequency of sites with subgingival plaque compared to never-smokers (P < 0.001). Mean gingival crevicular fluid was significantly higher in the presence of gingival inflammation for both groups (P = 0.001), whereas smokers demonstrated a significantly lower frequency of gingival bleeding than did non-smokers (23.6% vs 66.1%; P < 0.001). Conclusion: Smokers presented significantly lower percentages of sites with subgingival plaque in all experimental periods and presented less gingival inflammation as shown by GBI and gingival crevicular fluid quantification. Key words: dental plaque, gingival crevicular fluid, gingivitis, smoking Oral Health Prev Dent 2015;13:13-20 doi: 10.3290/j.ohpd.a32669
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t has been demonstrated that dental plaque initially develops on the tooth surface adjacent to the gingival margin (Quirynen and van Steenberghe, 1989). Supragingival plaque accumulation may lead to gingivitis development within 2 to 3 weeks (Löe et al, 1965; Theilade et al, 1966), subject to interindividual variation or even to specific microbiota (Page et al, 1997; Haffajee and Socransky, 2001).
a
Master’s Student, Graduate Programme in Dentistry, Lutheran University of Brazil, Canoas, RS, Brazil.
b
Associate Professor, Section of Periodontology, Faculty of Dentistry, Federal University of Rio Grande do Sul, Porto Alegre, RS, Brazil.
c
Professor, Section of Periodontology, Faculty of Dentistry, Federal University of Rio Grande do Sul, Porto Alegre, RS, Brazil.
Correspondence: Patricia Weidlich, Rua Ramiro Barcelos, 2492, Porto Alegre, RS, Brazil 90035-003. Tel: +55-51-3308-5318, Fax: +55-51-3308-5318. Email: [email protected]
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Submitted for publication: 11.07.12; accepted for publication: 26.05.13
Bass (1946) observed the presence of a plaquefree area in extracted teeth, which was limited coronally by the gingival margin and on the root surface by the coronal portion of the junctional epithelium. Brady (1973) called this the ‘plaque free zone’ (PFZ), delimited apically by the periodontium and cervically by the microbial deposit. Some studies have demonstrated that the PFZ is not stained by disclosing agents and can be identified clinically as a thin line between the gingival margin and the stained plaque. It has been observed in extracted teeth and in vivo in analyses of both supragingival and subgingival plaque formation processes (Brady, 1973; Saglie et al, 1975; Friedman et al, 1992; Weidlich et al, 2001). Studies concerning supragingival plaque are mostly performed on extracted teeth, making it impossible to evaluate the role of the PFZ in the development of microbial deposits and its relation-
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ship to the gingival margin. It is clear that some external factors such as smoking, which has been associated with a higher prevalence and greater severity of periodontitis, may interfere in this process. However, the biological mechanisms involved in tobacco consumption are not completely understood. Pioneer studies demonstrated that smokers presented higher amounts of plaque and supragingival calculus (Sheiham, 1971; Preber and Kant, 1973). The majority of authors, however, corroborate that there are no significant differences in the plaque accumulation pattern between smokers and nonsmokers (Rosa et al, 2000; Shiloah et al, 2000;
day 0
days 1, 2, 3, and 4
Gingival crevicular fluid quantification Professional tooth cleaning Plaque dye VPI, GBI
Gingival crevicular fluid quantification Plaque dye PFZ assessment
Calsina et al, 2002). Additionally, local effects of nicotine are related to the masking of clinical signs of gingival inflammation due to vasoconstriction, which slows the blood flow, leading to less marginal bleeding (Bergström, 1990; Gomes et al, 2007). Therefore, better understanding of the process of dental plaque formation is still necessary, including an analysis of variables that may interfere with its development. In this sense, the PFZ may have some clinical implications for the transition from supra- to subgingival plaque as well as the inflammatory process. Intervention studies that evaluate the possible influence of smoking on plaque development and gingival inflammation may provide additional information in the role of tobacco in the pathogenesis of periodontal diseases. The aim of the present study was to evaluate supragingival and subgingival plaque formation on the dentogingival area by means of the experimental gingivitis model and a plaque scoring system that considers the presence of a plaque free zone (PFZ) in smokers and never-smokers.
MATERIALS AND METHODS Study design
days 7 and 14
Gingival crevicular fluid quantification
day 21
Gingival crevicular fluid quantification Professional tooth cleaning Plaque dye VPI, GBI
days 22, 23 and 24
Gingival crevicular fluid quantification Plaque dye PFZ assessment VPI, GBI Professional tooth cleaning
day 25
Gingival crevicular fluid quantification Plaque dye PFZ assessment
Fig 1 Study flowchart. VPI: visible plaque index; GBI: gingival bleeding index; PFZ: plaque-free zone.
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The present study was performed by means of a parallel experimental gingivitis design in maxillary incisors and canines (test teeth). Nineteen male volunteers, selected after an interview and periodontal examination, were included in the study: 9 current smokers (mean of 15 cigarettes per day) aged 18–42 years (mean 28.6 ± 6.7 years) and 10 never-smokers aged 19–28 years (mean 23.8 ± 3.1 years). Individuals with caries, misaligned and/or restored/decayed teeth in the cervical or middle third of the buccal area of the test teeth, attachment loss, antibiotic treatment in the last 6 months and need of prophylactic dental treatment were not included. The study protocol was approved by the Institutional Review Board of the Lutheran University of Brazil, and all participants signed an informed consent form. Participants were instructed not to modify eating habits, to use a standard dentifrice without any antimicrobial agents (Close-Up, Unilever; São Paulo, Brazil) and to refrain from oral hygiene procedures in the maxillary dentition during the whole 25-day experimental period (Fig 1). In phase 1, starting with clinical gingival health, the supragingival plaque formation pattern in the dentogingival area
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was observed daily for 4 days. After day 4, participants continued refraining from oral hygiene up to day 21, when professional prophylaxis was performed to allow the observation of supragingival plaque formation under conditions of gingival inflammation (phase 2). During the experimental period, gingival fluid secretion was utilised as an inflammatory indicator. Test teeth were submitted to gingival fluid quantifi cation according to Egelberg (1966). After gentle rinsing with water, a Periopaper (Oraflow; New York, NY, USA) strip was inserted in the entrance of the buccal crevice at each test tooth for 3 min. Subsequently, the strips were transferred to petri dishes, wet with one drop of 0.2% nihidrine solution and dried at 75°C in a microbiological oven for 15 min. The area of the strip dyed by nihidrine was measured digitally using Adobe Photoshop. Gingival crevicular fluid quantification took place on days 0 to 4, 7, 14 and 21 to 25. The clinical evaluation of the presence of plaque and gingivitis was assessed by the Visible Plaque Index (VPI) and Gingival Bleeding Index (GBI) (Ainamo and Bay, 1975) at days 0, 21 and 25. At the beginning of each phase (days 0 and 21), professional prophylaxis was performed on the test teeth with rubber cups (KG Sorensen; Viking, Brazil), prophy paste (Pert, SS White; Rio de Janeiro, Brazil) and dental floss (Sanifill; São Paulo, Brazil), in order to obtain completely plaque free surfaces as disclosed by fucsine. To stain the plaque, teeth were rinsed with water, dried and isolated with cotton rolls. Fucsine solution (Replanic T, Iodontec; Porto Alegre, Brazil) was applied with a syringe and disposable needles (Becton Dickinson; Franklin Lakes, New Jersey, USA) in order to better access the sulcus area without touching the surface. After 30 s, teeth were rinsed, dried and evaluated with the scoring system proposed by Maliska et al (2006). In brief, the scoring system evaluates the initial plaque formation in the dentogingival area and is based on 3 criteria: 0 – absence of stained plaque; 1 – presence of stained plaque and a supragingival PFZ; 2 – presence of stained plaque and absence of PFZ, indicating that subgingival plaque formation has taken place. The assessment was performed on the buccal surface of the test teeth. A marking pen (Pilot Pen do Brasil; São Paulo, Brazil) was used to indicate 2 reference points on the buccal gingiva, 1–2 mm apical to the gingival margin, longitudinally dividing the buccal surface in three equal thirds: mesio-buccal, buccal and disto-buccal. The same
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trained and calibrated examiner (PB) who was supervised by the gold-standard examiner (PW) in a previous study (Maliska et al, 2006) performed all assessments. To test for reproducibility, 40 patients from the University dental clinic were assessed twice by the gold-standard examiner and the examiner of the present study. Kappa statistics demonstrated values of 0.79 and 0.76 for interand intraexaminer agreement, respectively. After the end of the experiment, professional prophylaxis was performed in order to facilitate the re-establishment of gingival health. All participants were followed-up until basal amounts of gingival crevicular fluid were present.
Statistical analysis The mean volume of gingival crevicular fluid was obtained by the mean linear measurement of the paper strip for smokers and never-smokers on each test day of phases 1 and 2. Intragroup comparisons were tested using the dependent t-test and intergroup comparisons using repeated-measures ANOVA. The frequency distribution of each PFZ score was obtained and compared over time using the Kruskall-Wallis test and intergroup comparisons were performed with the Wilcoxon signedrank test. Statistical significance was set at 5%.
RESULTS Table 1 demonstrates the data on VPI, GBI and probing depth (PD) on days 0, 21 and 25 for both groups. No statistically significant differences were observed in the percentage of sites without visible plaque (VPI = 0) and in mean PD between smokers and never-smokers at baseline. The percentage of gingival bleeding varied at the beginning of phase 2, with never-smokers presenting higher levels of gingival bleeding after 21 and 25 days of not performing oral hygiene as compared to smokers. Tables 2 and 3 present the mean volumes of gingival crevicular fluid in smokers and never-smokers throughout the study. When plaque accumulation occurred in the absence of gingival inflammation (phase 1), a gradual and significant increase was observed in the amount of gingival crevicular fluid during the 4-day period within each group. Neversmokers and smokers did not present significant differences in gingival crevicular fluid volume, except on day 3, when smokers presented a lower
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Table 1 Visible plaque index (VPI), gingival bleeding index (GBI) and pocket depth (PD) for smokers and never-smokers
Never-smokers
Smokers
P*
Beginning of phase 1 (day 0)
97
98.8
0.29
Beginning of phase 2 (day 21)
30.4
22.2
0.07
End of phase 2 (day 25)
54.8
53.5
0.45
Beginning of phase 1 (day 0)
100
100
Beginning of phase 2 (day 21)
33.9
76.4
< 0.01
End of phase 2 (day 25)
65.5
93.8
< 0.01
1.12 ± 0.36
1.15 ± 0.35
0.51
% sites with VPI = 0
% sites with GBI=0
PD day 0 (mean ± SD) * Independent samples t-test.
Table 2 Gingival crevicular fluid volume (mean ± SD) in the absence of gingival inflammation in never-smokers and smokers Absence of gingival inflammation
Never-smokers
day 0
day 1
day 2
day 3
day 4
0.02±0.04
0.06±0.13
0.10±0.13
0.15±0.17
0.20±0.20
0.001
0.001
0.001
0.001
0.04±0.07
0.05±0.09
0.12±0.15
0.24±0.18
0.001
0.001
0.001
0.001
0.32
0.03
0.37
0.31
P* Smokers
0.05±0.13
P* P#
0.12
* Intragroup comparison, dependent samples t-test. # Intergroup comparisons, repeated measures ANOVA.
Table 3 Gingival crevicular fluid volume (mean ± SD) in the presence of gingival inflammation in never-smokers and smokers Presence of gingival inflammation
Never-smokers
day 0
day 1
day 2
day 3
day 4
0.68±0.31
0.63±0.41
0.58±0.25
0.60±0.23
0.65±0.27
0.001
0.001
0.001
0.001
0.48±0.28
0.51±0.28
0.51±0.29
0.61±0.31
0.001
0.001
0.001
0.001
0.02
0.17
0.08
0.52
P* Smokers
0.64±0.30
P* P#
0.49
* Intragroup comparison, dependent samples t-test. # Intergroup comparisons, repeated measures ANOVA.
mean volume of gingival crevicular fluid. Considering gingival crevicular fluid in the presence of gingival inflammation (phase 2), intragroup comparisons demonstrate a significant increase in its volume during the 4-day period, with intergroup differences observed on day 1 between smokers and neversmokers. The mean gingival crevicular fluid volume was significantly higher in the presence of gingival
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inflammation for both groups. However, no statistically significant differences were observed related to smoking status. Figure 2 demonstrates the formation of subgingival plaque in never-smokers and smokers in the absence of gingival inflammation (phase 1). A significant increase was found in percent of sites with subgingival plaque (PFZ score = 2) during the 4
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never-smokers 100
*
*
*
smokers *
100
0
9
14.1 80
80
26.2
26.4 33.3
42.9 % sites
60
% sites
60
40
40 68.1 67.9
20
71.4 2.4
17.9
0
day 1
day 2
14.9
56.5 0.6 day 3
77.8
20
85.1
70.8
0
0
day 4
66.7 day 1
22.9 day 2
2.8 day 3
22 0 day 4
Fig 2 Early subgingival plaque formation in the absence of gingival inflammation in smokers and never-smokers. PFZ = 0: absence of stained plaque (dark gray); PFZ = 1: presence of stained plaque and supragingival PFZ (light gray); PFZ = 2: presence of stained plaque and absence of PFZ (presence of subgingival plaque) (black). *Intergroup comparisons, Friedman and Wilcoxon tests (P < 0.001).
never-smokers 100
*
*
*
smokers *
100
6.3
14.1
60
60 % sites
80
% sites
80
51.8 40
19.4
54.2
50 40
71.5 20
0
84.5
64.9
19.6 day 1
92.9
57.6 20
45.2 3 day 2
15.5 0 day 3
7.1 0 day 4
41 0
43.8 day 1
22.9 day 2
4.9 day 3
24.3 4.2 day 4
Fig 3 Early subgingival plaque formation in the presence of gingival inflammation in smokers and never-smokers. PFZ = 0: absence of stained plaque (dark gray); PFZ = 1: presence of stained plaque and supragingival PFZ (light gray); PFZ = 2: presence of stained plaque and absence of PFZ (presence of subgingival plaque) (black). *Intergroup comparisons, Friedman and Wilcoxon tests (P < 0.001).
days of plaque accumulation for both groups. Throughout the experiment, smokers presented a significantly lower relative frequency of sites with subgingival plaque compared to never-smokers. Similar results can be observed in Fig 3, when the formation of subgingival plaque occurred in the presence of gingival inflammation (phase 2). It was observed that the presence of gingival inflamma-
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tion accelerated the formation of subgingival plaque both in smokers and never-smokers. However, smokers presented significantly lower percentages of sites with subgingival plaque (PFZ score = 2) in all experimental evaluations as compared to never-smokers.
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DISCUSSION The present study analysed the plaque formation pattern in smokers and never-smokers under different gingival inflammatory conditions after an experimental gingivitis model and demonstrated that the presence of gingival inflammation accelerates subgingival plaque formation both in smokers and never-smokers. Additionally, significantly lower frequencies of sites with subgingival plaque were found in smokers independent of the inflammatory condition. Exclusively male volunteers were included in order to avoid hormonal fluctuations as confounders of the results (Lindhe and Attsfröm, 1967). Also, it is important to highlight that the examiner was calibrated with good levels of reproducibility, both intra-examiner and compared to the gold-standard examiner from the original study that developed the scoring system (Landis and Koch, 1977; Maliska et al, 2006). Plaque was assessed using the scoring system developed by Maliska (2006) and considers the presence of a PFZ. Knowledge about the transformation of supragingival deposits into subgingival plaque is incomplete; however, some studies report the presence of a plaque-free zone located apical to the supragingival plaque and coronal to the gingival margin (Quyrinen and van Steenberghe, 1985; Bergström, 1990; Weidlich et al, 2001). This supragingival PFZ is present during initial plaque formation, but it vanishes after 72–96 h of accumulation. The disappearance of the PFZ demonstrates that subgingival plaque has already developed. Scanning electron microscopic analyses demonstrate that subgingival plaque formation evolves from the supragingival plaque. Concomitant with these observations, an increase in gingival crevicular fluid volume was observed, together with a coronal migration of the gingival margin due to oedema, resulting in the establishment of subgingival plaque and a subgingival PFZ (Weidlich et al, 2001). The PFZ scoring system developed by Maliska et al (2006) has been shown to be reproducible and allows evaluation of the transformation of supragingival into subgingival plaque. The distribution of scores 0, 1 and 2 of the system used here demonstrated that the PFZ was pronounced at the beginning of the study, but gradually became less apparent during 4 days of plaque accumulation. It should be emphasised that a score of 2 means that subgingival plaque is already present. The moment at which a site no longer has a score
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of 1 should be considered the initiation of subgingival plaque formation. In this context, smokers presented lower frequencies of subgingival plaque than did never-smokers. This could be explained by the local action of cigarette components on periodontal tissues that may affect both microbial challenge and host response. The majority of studies that observe lower degrees of gingivitis or marginal bleeding in smokers suppose that this is related to nicotine-induced peripheral vasoconstriction, resulting in a decreased inflammatory pattern as well as a decreased host defense (Bergström, 1990; Giannopoulou et al, 2003; Gomes et al, 2007). Differences between smokers and non-smokers in relation to clinical parameters are important. Our results demonstrated an increase in VPI during the study which did not differ between smokers and never-smokers; this was similar to the results of several other studies that could not show differences in the plaque formation pattern (Shiloah et al, 2000; Calsina et al, 2002; Salvi et al, 2005; Gomes et al, 2007). However, some reports found higher amounts of plaque in smokers than in nonsmokers (Ainamo, 1971; Preber and Bergström, 1985; Machuca et al, 2000). In relation to GBI, the present study found that smokers exhibited lower levels of clinically detectable gingivitis than neversmokers. These differences were only observed on days 21 and 25, since GBI was not assessed at other time points in order to not interfere in the plaque analysis by disrupting the formed plaque with the probe. These findings corroborate those of Ainamo (1971), in which the gingival index was lower in individuals that smoked more than 20 cigarettes per day, and the results of Preber and Bergström (1985), which demonstrated that gingival redness and bleeding were lower in individuals who smoked at least 20 cigarettes per day. On the other hand, Salvi et al (2005) demonstrated that smokers and never-smokers had similar patterns of gingival inflammation during 21 days of experimental plaque accumulation. Different lifetime smoking exposure can be observed among studies and this may partially explain the differences related to gingival bleeding in smokers. In phase 2, from day 22, a new occurrence of scores 0 and 1 was observed, with a lower frequency of score 2 both in smokers and in never-smokers. From day 22 to the end of the experiment, a marked decrease in scores 0 and 1 was observed, with an increase in the presence of subgingival plaque (score 2). The conversion of scores 0 and 1 into score 2 in never-smokers was faster that in
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smokers. This observation is of extreme interest in the context of the present investigation. Although an explanation for this phenomenon is not a direct objective of this study, some suppositions can be made. First, the utilised variable – PFZ – has a different concept from the majority of published studies and may be an interesting tool for studies that aim at investigating plaque formation patterns. Secondly, the role of gingival inflammation as a modifier of plaque formation should be highlighted. It is well-established that de novo plaque formation is faster in inflamed that in non-inflamed areas (Hillam and Hull, 1977; Ramberg et al, 1994; Ramberg et al, 1995). This fact was clearly observed in the present study, especially in never-smokers. It would be unwarranted to suppose that smoking inhibits plaque formation. The present study demonstrated that smokers showed less gingival inflammation both in terms of GBI and gingival crevicular fluid volume. Therefore, it may be postulated that the lower level of inflammation observed contributed to less plaque accumulation, as it is well established that gingival inflammation favours plaque accumulation. This is confirmed by the differences observed between smokers and neversmokers in the present study. On the other hand, it is important to point out that smoking modifies the host response, leading to higher periodontal breakdown, probably by different mechanisms of tissue destruction (Salvi et al, 1997; Bergström et al, 2000; Johnson and Hill, 2004; Moimaz et al, 2009). A study that evaluated the microbiological and pro-inflammatory characteristics during early plaque formation demonstrated that smokers present an initial plaque colonisation that is diverse, unstable and contains more periodontopathogenic microorganisms than in non-smokers. Additionally, smokers presented higher levels of pro-inflammatory mediators as compared to non-smokers since the first day of plaque accumulation (Kumar et al, 2011). In this sense, smoking seems to play a central role both in microbiota composition and in host response starting with early plaque formation.
CONCLUSION The present study analysed supragingival and subgingival plaque formation in smokers and neversmokers under different gingival inflammatory conditions and concluded that smokers presented significantly lower percentages of sites with subgingival plaque during the entire experimental period
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and less gingival inflammation as shown by both GBI and gingival crevicular fluid volume.
REFERENCES 1. Ainamo J, Bay I. Problems and proposals for recording gingivitis and plaque. Int Dent J 1975;25:229–235. 2. Ainamo J. The seeming effect of tobacco consumption on the occurrence of periodontal disease and dental caries. Suom Hammaslaak Toim 1971;67:87–94. 3. Bass CC. A demonstrable line on extracted teeth indicating the location of the outer border of the epithelial attachment. J Dent Res 1946;25:401–415. 4. Bergström J. Oral hygiene compliance and gingivitis expression in cigarette smokers. Scand J Dent Res 1990;98:497–503. 5. Bergström J. Photogrammetric registration of dental plaque accumulation in vivo. Acta Odontol Scand 1981;39:275–284. 6. Bergström J, Eliasson S, Dock J. A 10-year prospective study of tobacco smoking and periodontal health. J Periodontol 2000;71:1338–1347. 7. Brady JM. A plaque-free zone on human teeth--scanning and transmission electron microscopy. J Periodontol 1973;44:416–428. 8. Calsina G, Ramón JM, Echeverría JJ. Effects of smoking on periodontal tissues. J Clin Periodontol 2002;29:771–776. 9. Egelberg J. Permeability of the dento-gingival blood vessels. 1. Application of the vascular labelling method and gingival fluid measurements. J Periodontal Res 1966;1: 180–191. 10. Friedman MT, Barber PM, Mordan NJ, Newman HN. The „plaque-free zone“ in health and disease: a scanning electron microscope study. J Periodontol 1992;63:890–896. 11. Giannopoulou C, Cappuyns I, Mombelli A. Effect of smoking on gingival crevicular fluid cytokine profile during experimental gingivitis. J Clin Periodontol 2003;30:996–1002. 12. Gomes SC, Piccinin FB, Susin C, Oppermann RV, Marcantonio RA. Effect of supragingival plaque control in smokers and never-smokers: 6-month evaluation of patients with periodontitis. J Periodontol 2007;78:1515–1521. 13. Haffajee AD, Socransky SS. Relationship of cigarette smoking to the subgingival microbiota. J Clin Periodontol 2001;28:377–388. 14. Hillam DG, Hull PS. The influence of experimental gingivitis on plaque formation. J Clin Periodontol 1977;4:56–61. 15. Johnson GK, Hill M. Cigarette smoking and the periodontal patient. J Periodontol 2004;75:196–209. 16. Kornman KS, Loesche WJ. The subgingival microbial flora during pregnancy. J Periodontal Res 1980;15:111–122. 17. Landis JR, Koch GG. The measurement of observer agreement for categorical data. Biometrics 1977;33:159–174. 18. Lindhe J, Attsfröm R. Gingival exudation during the menstrual cycle. J Periodontal Res 1967;2:194–198. 19. Löe H, Theilade E, Jensen SB. Experimental gingivitis in man. J Periodontol 1965;36:177–187. 20. Machuca G, Rosales I, Lacalle JR, Machuca C, Bullón P. Effect of cigarette smoking on periodontal status of healthy young adults. J Periodontol 2000;71:73–78.
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Branco et al 21. Maliska AN, Weidlich P, Gomes SC, Oppermann RV. Measuring early plaque formation clinically. Oral Health Prev Dent 2006;4:273–278. 22. Moimaz SA, Zina LG, Saliba O, Garbin CA. Smoking and periodontal disease: clinical evidence for an association. Oral Health Prev Dent 2009;7:369–376. 23. Page RC, Offenbacher S, Schroeder HE, Seymour GJ, Kornman KS. Advances in the pathogenesis of periodontitis: summary of developments, clinical implications and future directions. Periodontol 2000 1997;14:216–248. 24. Preber H, Bergström J. Occurrence of gingival bleeding in smoker and non-smoker patients. Acta Odontol Scand 1985;43:315–320. 25. Preber H, Kant T. Effect of tobacco-smoking on periodontal tissue of 15-year-old schoolchildren. J Periodontal Res 1973;8:278–283. 26. Quirynen M, van Steenberghe D, Vuylsteke M. The possibility of measuring plaque growth in vivo within 24 hours. J Periodontal Res 1985;20:321–328. 27. Quirynen M, van Steenberghe D. Is early plaque growth rate constant with time? J Clin Periodontol 1989;16:278– 283. 28. Ramberg P, Axelsson P, Lindhe J. Plaque formation at healthy and inflamed gingival sites in young individuals. J Clin Periodontol 1995;22:85–88. 29. Ramberg P, Lindhe J, Dahlén G, Volpe AR. The influence of gingival inflammation on de novo plaque formation. J Clin Periodontol 1994;21:51–56. 30. Rosa GM, Lucas GQ, Lucas ON. Study of the crevicular fluid flow rate in smokers. Acta Odontol Latinoam 2000;13:51–60.
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31. Saglie R, Johansen JR, Tollefsen T. Plaque-free zones on human teeth in periodontitis. J Clin Periodontol 1975;2:190–197. 32. Salvi GE, Lawrence HP, Offenbacher S, Beck JD. Influence of risk factors on the pathogenesis of periodontitis. Periodontol 2000 1997;14:173–201. 33. Salvi GE, Ramseier CA, Kandylaki M, Sigrist L, Awedowa E, Lang NP. Experimental gingivitis in cigarette smokers: a clinical and microbiological study. J Clin Periodontol 2005; 32:441–447. 34. Sheiham A. Periodontal disease and oral cleanliness in tobacco smokers. J Periodontol 1971;42:259–263. 35. Shiloah J, Patters MR, Waring MB. The prevalence of pathogenic periodontal microflora in healthy young adult smokers. Periodontol 2000;71:562–567. 36. Theilade E, Wright WH, Jensen SB, Löe H. Experimental gingivitis in man. II. A longitudinal clinical and bacteriological investigation. J Periodontal Res 1966;1:1–13. 37. Trombelli L, Tatakis DN, Scapoli C, Bottega S, Orlandini E, Tosi M. Modulation of clinical expression of plaque-induced gingivitis. II. Identification of ‘high-responder’ and ‘low-responder’ subjects. J Clin Periodontol 2004;31:239– 252. 38. Van der Weijden GA, Timmerman MF, Danser MM, Nijboer A, Saxton CA, Van der Velden U. Effect of pre-experimental maintenance care duration on the development of gingivitis in a partial mouth experimental gingivitis model. J Periodontal Res 1994;29:168–173. 39. Weidlich P, Lopes de Souza MA, Oppermann RV. Evaluation of the dentogingival area during early plaque formation. J Periodontol 2001;72:901–910.
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