Tobacco smoking and neutrophil activity in patients with periodontal disease

BACKGROUND: Tobacco smoking has considerable negative effects on periodontal health. The mechanisms behind these effects are incompletely understood but may be related to the host response. The aim of the present study was to investigate the influence of tobacco smoking on the gingival crevicular fluid (GCF) levels of elastase, lactoferrin (LF), alpha-1-antitrypsin (alpha-1-AT), and alpha-2-macroglobulin (alpha-2-MG) under periodontally diseased conditions. METHODS: The study population included 15 smokers (5 women and 10 men) aged 34 to 69 years and 17 non-smokers (5 women and 12 men) aged 31 to 81 years. Clinical registration of gingival index (GI), plaque index (PI), probing depth, as well as sampling of GCF were made at 3 sites with severe lesions and 3 sites with moderate lesions in each individual. The elastase activity was measured with a chromogenic low molecular substrate and the LF, alpha-1-AT, and alpha-2-MG concentrations with ELISA. RESULTS: The results showed that, with regard to severe lesions, smokers had a significantly lower concentration of alpha-2-MG as well as significantly lower total amounts of alpha-2-MG and alpha-1-AT than non-smokers. With regard to moderate lesions, smokers tended to exhibit a lower concentration of alpha-2-MG, but the difference was not statistically significant. Comparing moderate and severe lesions, smokers exhibited no gradual increase with disease severity in contrast to non-smokers, who showed significantly or almost significantly increased levels of LF and alpha-2-MG in severe as compared to moderate lesions. CONCLUSIONS: The present results indicate that the levels of alpha-2-MG and alpha-1-AT are suppressed in smokers with periodontitis, suggesting that smoking interferes with these protease inhibitors. This may be one mechanism by which smoking affects the inflammatory response.     

Effect of tobacco smoking on neutrophil activity following periodontal surgery

BACKGROUND: Tobacco smoking has considerable negative effects on the outcome of periodontal treatment. The reason for the inferior therapeutical effect might be related to an altered neutrophil activity in terms of elastase and/or matrix metalloproteinase-8 (MMP-8), as well as in the protease inhibitor alpha-1-antitrypsin (alpha-AT) and alpha-2-macroglobulin (alpha-2-MG) activities. The aim of the present study, therefore, was to elucidate the effect of tobacco smoking on gingival crevicular fluid (GCF) levels of these substances following surgical treatment. METHODS: The study population included 15 smoking and 15 non-smoking patients with moderate to severe periodontitis receiving surgical treatment. Clinical examinations and collection of GCF were done prior to surgery and 1 and 5 weeks following treatment. The elastase activity was measured with a chromogenic low-molecular substrate and the levels of alpha-1-AT, alpha-2-MG, and MMP-8 with enzyme-linked immunosorbent assay. RESULTS: The results showed unaltered levels of alpha-1-AT, alpha-2-MG, and MMP-8 in smokers following surgery. In non-smokers, the levels of alpha-1-AT and alpha-2-MG increased, whereas MMP-8 levels decreased. The levels of elastase remained unaltered in both smokers and non-smokers. CONCLUSIONS: These results indicate that in the presence of smoking, the levels of alpha-1-AT, alpha-2-MG, and MMP-8 remained unaltered during the recovery period following surgical treatment. This is interpreted as a possible interference of smoking with the treatment response and may, in part, explain the clinical evidence of an inferior treatment outcome in smokers.     

Elastase in gingival crevicular fluid from smokers and non-smokers with chronic inflammatory periodontal disease

OBJECTIVE: To compare elastase concentrations in gingival crevicular fluid (GCF) from individual sites of smokers and non-smokers.
MATERIALS AND METHODS: Twelve pairs of smokers and non-smokers with untreated, moderate to advanced chronic inflammatory periodontal disease were matched for gender, age, ethnicity and the clinical and radio-graphic extent of disease. Durapore filter strip samples were collected over 30 s from two mesiopalatal sites on upper left posterior teeth. Samples were analysed for: I) polymorphonuclear neutrophil leucocyte (PMNL) cell counts; 2) PMNL elastase-αI-antitrypsin complex in the GCF supernatant by ELISA; and 3) functional elastase, free or bound to α2-macroglobulin, estimated from activity against N-tert-butoxycarbonyI-alanyl-prolyl-nor-valylg-chlorothiobenzyl ester in supernatant and lysates of GCF PMNLs.
RESULTS: There were no differences in disease parameters between groups except that bleeding on probing was less extensive in smokers (P< 0.001). Cell counts and elastase content of crevicular PMNLs showed no differences between groups. Lower concentrations of elastase were found in GCF supernatants from smokers than non-smokers. This difference was observed for functional elastase (mean [s.d.] = 30.21 [17.60] against 73.77 [75.26] ng μI^-1, P <0.05) and elastase complexed with αl-antitrypsin (8.97 [6.54] ng μl^-1 against 25.71 [22.07] ng μI^-1, P < 0.001).
CONCLUSIONS: Smokers have lower elastase concentrations in GCF than non-smokers. Further investigation is required to elucidate the underlying cause and its relationship with periodontal disease.     

Acute-phase proteins in gingival crevicular fluid during experimentally induced gingivitis

The dynamics of four acute-phase proteins were investigated in gingival crevicular fluid (GCF) during the course of a 21 day experimental gingivitis study. These acute-phase proteins were the protease inhibitors α2-macroglobulin (α2-M) and α1-antitrypsin (α1-AT) and the iron-binding proteins transferrin (TF) and lactoferrin (LF). 6 healthy volunteers ceased all oral hygiene procedures for 3 weeks. GCF was sampled at seven day intervals from two sites per subject by paper strips for 30 s during the experimental gingivitis period and for two additional weeks after the reinstitution of oral hygiene. The mainly serum derived α2-M, αl-AT and TF exhibited very similar dynamics which reflects their common origin in GCF. Their levels increased significantly from baseline and remained high for at least one week after the reinstitution of oral hygiene measures (repeated measures MANOVA; α2-M: p=0.015; α1-AT: p=0.012; TF: p=0.02). This probably reflects increased vascular permeability in the gingivae and, to a lesser degree, local production by gingival inflammatory cells. In contrast to the serum derived acute-phase proteins, the neutrophil derived LF rose significantly from baseline (repeated measures MANOVA; p=0.001) but dropped rapidly after the reinstitution of oral hygiene measures. This could be because dental plaque was removed and thus neutrophil chemotactic agents in the crevice were decreased.     

Neutrophil elastase activity, levels of prostaglandin E2, and matrix metalloproteinase-8 in refractory periodontitis sites in smokers and non-smokers.

The study was aimed to determine elastase activity, levels of prostaglandin E2 (PGE2), and matrix metalloproteinase-8 (MMP-8) in gingival crevicular fluid (GCF) in 20 smokers and 20 non-smokers, mean age 47.4 (+/-2.9 SD) years with refractory periodontal diseases. GCF was collected with intracrevicular washing from four sites in each subject. Clinical assessments, included gingival index, probing depth, clinical attachment level, bleeding on probing, bone height, and plaque accumulation. Smokers had a significantly higher percentage of the gingival margin covered by plaque (P%Im), higher number of sites with probing pocket depth > or = 5 mm, higher mean values of probing pocket depth and probing attachment level (P< 0.01). Smokers had significantly higher mean levels of neutrophil elastase activity (P< 0.01) in the supernatants than non-smokers did. In sites with matching pocket depths, neutrophil elastase activity was significantly higher in smokers (P< 0.001) than in non-smokers. In sites with high levels of MMP-8 the PGE2 levels were significantly (P< 0.001) higher compared to sites with low levels in smokers as well as in non-smokers. A significant correlation was found between probing pocket depth and levels of MMP-8 (P< 0.001) and in non-smokers between probing pocket depth and levels of PGE2 (P< 0.05).     

吸烟对牙周基础治疗前后龈沟液量和弹性蛋白酶水平的影响

目的　评价吸烟是否影响牙周炎基础治疗前、后龈沟液 (gingivalcrevicularfluid ,GCF)量和龈沟液中弹性蛋白酶 (elastase ,EA)的水平。方法　将 37例男性慢性牙周炎患者分为吸烟组 (2 2例 ,12 2个牙位点 ,每日吸烟≥ 2 0支 )和非吸烟组 (15例 ,90个牙位点 )。牙周炎基础治疗前、后用滤纸条法收集GCF ,用Periotron 6 0 0 0龈沟液测量仪测定GCF量。对吸烟组 92个位点和非吸烟组 6 0个位点GCF样本 ,用底物分解法检测EA水平。结果　治疗前吸烟组GCF量 (139 2± 33 4 )U和EA水平(0 6 34± 0 5 87)明显低于非吸烟组 [GCF量 :(15 5 4± 39 7)U ,EA水平 :0 835± 0 5 72 ],P <0 0 1。治疗后 ,两组GCF量和EA水平均显著降低 (P <0 0 0 1)。但吸烟组 91个位点 (74 6 % )GCF和 70个位点(76 1% )的EA水平治疗后有改善 ;而非吸烟组高达 88个位点 (97 8% )GCF和 5 6个位点 (93 3% )的EA水平有改善 (P <0 0 1)。结论　治疗前探诊深度相同的情况下 ,吸烟组GCF量和EA水平均低于非吸烟组 ,治疗后吸烟组的GCF和EA的减少程度不如非吸烟组明显。     

Neutrophil elastase and its inhibitors in human gingival crevicular fluid during experimental gingivitis.

The relative concentrations and absolute amounts of neutrophil elastase and its two inhibitors, alpha 2-macroglobulin (alpha 2-M) and alpha 1-antitrypsin (alpha 1-AT), were determined in gingival crevicular fluid (GCF) collected from six dental students who refrained from brushing the upper left or right quadrant during three weeks. Plaque and gingival indices and flow of GCF were measured before, during, and after the three weeks of no brushing. Functional elastase, representing the enzyme complexed with alpha 2-M, was measured by use of a low-molecular-weight fluorogenic substrate. Elastolytic activity in GCF was also assayed by use of elastin as substrate. Antigenic elastase, representing the enzyme complexed with alpha 1-AT, as well as the inhibitors alpha 2-M and alpha 1-AT were measured by ELISA. After three weeks of plaque accumulation, the concentrations of both functional and antigenic elastase increased by a factor of about 3, whereas the concentrations of the inhibitors increased in a much higher proportion. No free elastase could be detected in GCF, as evidenced by the Sephadex G-75 elution profile of GCF, by the negative results obtained when elastin was used as substrate, and by the demonstration that pure enzyme kept its activity against the low-molecular-weight substrate after being saturated by alpha 2-M.     

Altered relation between granulocyte elastase and α-2-macroglobulin in gingival crevicular fluid from sites with periodontal destruction

Abstract Granulocyte elastase activity and α-2-macroglobulin (α-2-MG) were studied in the gingival crevicular fluid (GCF) from 3 categories of sites in 6 patients with gingivitis and 6 patients with periodontitis. 6 inflamed sites in each gingivitis patient were sampled on paper strips and 12 sites, 6 with and 6 without attachment loss and periodontal pockets, were selected in each periodontitis patient. To avoid the influence of increase GCF volume from deep pockets, the elastase activity and the α-2-MG were calculated per μl of GCF. The proteolytic activity of elastase was measured with a low molecular weight substrate and the antiprotease, α-2-MG, with ELISA. The measured activity could be ascribed to elastase that had been released into the gingival tissues and into the GCF prior to sampling. In the periodontitis patients, the sites with tissue destruction had a significantly higher elastase activity per site and per μl GCF and a significantly lower α-2-MG per μl than the 2 other categories of sites without tissue destruction. The destructive inflammation seems to be associated with increased release of elastase, either from more numerous or from more active granulocytes and with an increased proteolytic consumption of the inhibitor accompanied by the fast elimination of the protease-inhibitor-complex. In conclusion, the study shows a strong relationship between elastase activity and tissue destruction, a finding that supports the pathogenic theory of an involvement of granulocytes and their proteolytic enzymes in the mechanism of periodontal destruction.     

The association between neutrophil numbers and interleukin-1alpha concentrations in gingival crevicular fluid of smokers and non-smokers with periodontal disease

OBJECTIVES: To test whether neutrophil numbers are directly correlated with interleukin-1alpha (IL-1alpha) concentrations in gingival crevicular fluid (GCF) of patients with periodontitis, and to investigate the effects of smoking on these parameters. MATERIALS AND METHODS: A total of 99 GCF samples from 33 patients (14 smokers) suffering from severe chronic periodontitis were collected using Durapore filter strips. Polymorphonuclear leucocyte (PMN) numbers were counted using a Coulter cell counter and IL-1alpha levels were determined by ELISA. Total GCF protein was measured by Bio-Rad assay as a surrogate measure of GCF volume. RESULTS: Mean IL-1alpha concentrations were significantly reduced in smokers compared with non-smokers (non-smokers: 3.29+/-2.02 pg/microg protein, smokers 1.59+/-1.13 pg/microg protein). There was no association between PMN numbers and IL-1alpha concentrations found when analysed either by site or by patient. PMN numbers were not significantly different between the two groups (non-smokers: 1.16 x 10(6)+/-1.04 x 10(6); smokers: 7.30 x 10(5)+/-8.07 x 10(5)). Smoking did not affect mean total protein concentration of samples. CONCLUSIONS: Smoking significantly decreased IL-1alpha concentrations in GCF without affecting GCF volume sampled. The lack of association between IL-1alpha concentration and neutrophil numbers suggests that the reduced IL-1alpha concentrations seen in smokers is independent of any possible effect of smoking on neutrophil chemotaxis, and further suggests that smoking may directly inhibit IL-1alpha production.     

Granulocyte elastase,matrix metalloproteinase-8 and prostaglandin E2 in gingival crevicular fluid in matched clinical sites in smokers and non-smokers with persistent periodontitis

BACKGROUND/AIMS: Smokers with persistent periodontitis may have granulocytes with impaired function. This study aimed to determine the levels of granulocyte elastase, matrix metalloproteinase-8 (MMP-8) and prostaglandin E2 (PGE2) in gingival crevicular fluid (GCF) in smokers and non-smokers with persistent periodontitis. METHODS: We analyzed GCF from 70 matched sites in 29 periodontitis and 6 gingivitis sites in 34 subjects, 17 smokers, and 17 non-smokers. We also analyzed separately GCF from 28 of these subjects, 14 smokers and 14 non-smokers in 14 matched periodontitis sites. The following measurements were made: elastase complexed to alpha1-antitrypsin (EA-alpha1AT) and MMP-8 with ELISA, functional elastase with a chromogenic substrate, and PGE2 with radioimmunoassay (125I RIA). The significance of the findings was determined with Mann-Whitney test. RESULTS: In the 29 matched periodontitis sites, smokers had significantly more functional elastase (p<0.005) and more EA-alpha1AT (p<0.05) than non-smokers. In the 14 matched periodontitis sites in 14 smokers and 14 non-smokers, the former had significantly more functional elastase than the latter (p<0.001). A significant correlation was found between EA-alpha1AT and MMP-8 in smokers (p<0.05) and non-smokers (p<0.001) and a positive correlation between levels of functional elastase and MMP-8 in non-smokers (r2=0.98; p<0.001). CONCLUSIONS: Granulocyte function seems to be impaired in smokers with persistent periodontitis. The cells react to the bacterial challenge by releasing serine proteases, which reflect the degradation of connective tissue. The risk of progression of the disease is therefore higher in smokers with persistent periodontitis than in non-smokers.     

快速进展性牙周炎患者龈沟液中的弹性蛋白酶活性

目的探讨中性多形核白细胞与快速进展性牙周炎的关系。方法检测２例快速进展性牙周炎患者共１０２个位点（其中２８个位点进行治疗前后的对比）的龈沟液弹性蛋白酶活性,将人多形核白细胞弹性蛋白酶特异底物———Ｓ２４８４与龈沟液反应,测吸光度值,以反映龈沟液弹性蛋白酶活性。结果快速进展性牙周炎龈沟液中的弹性蛋白酶活性［（０．６３±０．３８）Ａｂｓ／位点］明显高于健康对照组［（０．０７±０．０５）Ａｂｓ／位点］,差异有显著性;弹性蛋白酶活性的高低与龈沟液体积、探诊深度、附着丧失和出血指数呈正相关关系;治疗后龈沟液体积和各临床指数显著降低,弹性蛋白酶活性也从治疗前的（０．７３±０．３６）Ａｂｓ／位点下降为（０．１±０．１７）Ａｂｓ／位点,差异有显著性。结论快速进展性牙周炎患者的中性多形核白细胞并不是趋化反应不足,而是过度浸润与释放溶酶体酶,起协同致炎作用。     

Elevated systemic concentrations of soluble ICAM-1 (sICAM) are not reflected in the gingival crevicular fluid of smokers with periodontitis

Raised serum levels of soluble intercellular adhesion molecule-1 (sICAM-1) in smokers could have immunomodulatory effects in periodontitis. The aim of this study was to compare serum and gingival crevicular fluid (GCF) concentrations of sICAM-1 in smokers and non-smokers with periodontal disease. sICAM-1 in serum and GCF collected from age- and gender-matched smokers (n = 14) and non-smokers (n = 14) with periodontitis were measured by ELISA. Mean serum sICAM-1 concentrations were significantly elevated in smokers (331 ng mL(-1)), compared with non-smokers (238 ng mL(-1), p = 0.008). However, the concentration of sICAM-1 in the GCF was significantly lower in the smokers (83 ng mL(-1)), compared with non-smokers (212 ng mL(-1), p = 0.013). The difference between concentrations of sICAM-1 in GCF and serum was significant only in smokers (p < 0.001). Since GCF is a serum-derived tissue exudate, these results suggest that, in smokers, circulating sICAM-1 molecules are affected either in their passage from the periodontal microvasculature or within the periodontal tissues.     

The effect of different cigarette smoking levels on gingival crevicular fluid volume and periodontal clinical parameters in Saudi Arabia

IntroductionPeriodontal disease is a chronic inflammatory condition of the periodontium. It is the main cause of tooth loss and is considered one of the biggest threats to the oral cavity. Tobacco smoking has long been associated with increased risk for periodontal, peri-implant, and other medical diseases.ObjectiveTo evaluate the effect of smoking and its level on periodontal clinical parameters (probing depth (PD), plaque index (PI), gingival index (GI), clinical attachment level (CAL), bleeding on probing (BOP), and the volume of gingival crevicular fluid (GCF)) in healthy and chronic periodontitis individuals.Material and MethodA total of 160 participants were recruited in the present study, who were equally divided into the following five groups: healthy controls (C), healthy smokers (HS), nonsmokers with periodontitis (PNS), light smokers with periodontitis (PLS), and heavy smokers with periodontitis (PHS). GCF volume and periodontal clinical parameters (PD, PI, GI, CAL, and BOP) were assessed for each participant and compared between the study groups.ResultThere was a statistically significant difference in PD, PI, GI, CAL, and BOP between healthy and periodontitis patients (p < 0.001). The mean PI, PD, and CAL were considerably higher in heavy smokers than light smokers and non-smokers (P < 0.001). In contrast, the mean GI and BOP were significantly lower in heavy smokers than in light smokers and non-smokers. There was a statistically significant difference in GCF between healthy and periodontitis patients (p < 0.001). The mean GCF readings were higher in heavy smokers than light smokers or non-smokers (P < 0.001).ConclusionThe present study confirms the influence of smoking on periodontal clinical parameters. Smoking was associated with increased PD, PI, CAL, and GCF readings; however, GI and BOP were decreased in smokers. The number of cigarettes played a key role in the volume of GCF and periodontal clinical parameters.     

Detection of stable and active periodontitis sites by clinical assessment and gingival crevicular acute-phase protein levels

The aim of the present study was to investigate whether incipient periodontal disease breakdown could be associated with changes in gingival crevicular fluid (GCF) acute-phase protein levels. In addition, the potential of clinical indices to act as predictors of significant attachment level (AL) change was investigated. AL measurements were taken at baseline and 3 months using the Florida Probe stent handpiece from a total of 384 sites in 38 patients. The average standard deviation of duplicate AL measurements was 0.423. When the tolerance method was used to detect significant AL change, 3.9% of the sites lost attachment. When a less stringent criterion of AL change of ≥1 mm was used 9.9% of the sites lost attachment during the 3-month period. With the exception of probing depth, baseline clinical parameters failed to predict AL change. Fourteen active periodontitis sites that demonstrated significant attachment loss were paired to stable periodontitis sites within the same patient. The levels of four acute-phase proteins, namely α2-macroglobulin (α2-M), α1-antitrypsin (α1-AT), transferrin (TF) and lactoferrin (LF), and also albumin (Alb) were assessed in the same gingival crevicular fluid sample using sandwich ELISAs. Results were expressed either as ng/30 s and ng/μg Alb. Acute-phase protein levels in GCF failed to differentiate between active and stable periodontitis sites at baseline. In conclusion, the degree of gingival inflammation of the tissues adjacent to the crevice/pocket seems to influence the levels of protease inhibitors and iron-binding proteins in GCF to a greater extent than probing attachment loss.     

Clinical expression of TNF-α in smoking-associated periodontal disease

Abstract. The level of TNF-α in gingival crevicular fluid (GCF) was analyzed with respect to smoking in patients with untreated moderate to severe periodontal disease including 30 current smokers, 19 former smokers and 29 non-smokers, in the age range 31–79 years, Concomitantly the occurrence of the periopathogens Actinobacillus actinomycetemcomitans (Aa), Porphyromonas gingivalis (Pg) and Prevotella intermedia (Pi) and the GCF levels of albumin, IgA and IgG were analyzed. With regard to clinical characteristics, there were no statistically significant differences between smoking groups. The occurrence of patients positive for the periopathogens Aa, Pg and Pi was 28.2%, 41.0% and 91.0%, respectively. There were no statistically significant differences between smoking groups with regard to occurrence or relative frequency of these periopathogens. An exception was a significantly lower occurrence of Aa in former smokers as compared to non-smokers. The chief novelty of the study was the observation of a clearly increased level of TNF-α in GCF associated with smoking. Both current and former smokers exhibited significantly higher levels of TNF-α in comparison to non-smokers, whereas the levels of albumin, IgA and IgG were the same irrespective of smoking. In conclusion, the present observations in patients with moderate to severe periodontal disease suggest that smoking is associated with elevated GCF levels of the cytokine TNF-α.     

Plasminogen activator system in smokers and non-smokers with and without periodontal disease

BACKGROUND: The present study assessed levels of plasminogen activator (PA) system proteins in gingival crevicular fluid (GCF) and serum of chronic gingivitis, chronic periodontitis patients and periodontally healthy subjects and evaluated how smoking influenced these levels. METHODS: Twenty chronic gingivitis; 20 chronic periodontitis patients and 20 periodontally healthy volunteers were consecutively recruited according to the inclusion criteria so that exactly half of the subjects in each category were smokers. GCF samples from four sites together with serum samples were obtained from each subject. GCF levels of tissue type PA (t-PA), urokinase type PA (u-PA), PA inhibitor-1 (PAI-1) and PA inhibitor-2 (PAI-2) and serum concentrations of cotinine, u-PA and PAI-1 were analysed by enzyme-linked immunosorbent assay. RESULTS: The only statistically significant difference between smokers and non-smokers was a lower GCF PAI-2 concentrations in healthy smokers compared with healthy non-smokers (p<0.01). Gingivitis and periodontitis patients had higher GCF concentrations of PAI-2 than healthy subjects (p<0.002 and p<0.02 respectively). The ratio of u-PA:PAI-1 and t-PA:PAI-1 were significantly higher in GCF of smokers with periodontitis compared with "healthy" smokers, whereas the ratio of t-PA:PAI-2 was significantly lower in smokers with periodontal disease (p<0.05). CONCLUSIONS: GCF levels of the PA system proteins are increased in chronic gingivitis and periodontitis compared with healthy gingiva. Smoking had only subtle effects on the GCF PA system proteins with the exception of PAI-2, and the balance of activators and inhibitors. These findings suggest one mechanism whereby smoking may exert detrimental effects on the periodontal tissues.     

Five parameters of gingival crevicular fluid from eight surfaces in periodontal health and disease

Volume and amounts of myeloperoxidase (MPO), lactoferrin (LF), aryl sulfatase (AS) and lactate dehydrogenase (LDH) were measured in gingival crevicular fluid (GCF) collected from the mesial and distal proximal surfaces of the premolars and first and second molars of 3 subject groups. Group assignment was based on subject mean gingival index (GI) and probing depth (PD) of sampled sites as follows: healthy, GI less than or equal to 0.5, PD less than or equal to 3.0; disease 1, GI greater than or equal to 1.0, PD greater than or equal to 3.0 mm; disease 2, PD greater than or equal to 4.0 mm. Attachment loss (ATL) of most sites in the 3 groups was: healthy, 0-1 mm; disease 1, 1-2 mm; and disease 2, 4-9 mm. GCF volume differed among surfaces and teeth in each of the 3 groups. The greater amount of GCF collected from posterior locations was not related to the GI and PD. Differences with sampling location in amounts of GCF constituents were restricted to MPO and LF. Most of these differences (greater amounts at posterior sites) were associated with more severe disease. Variability in amount and composition of GCF collected from different sites, therefore, should be considered in experiments which include quantitation of GCF parameters. The ratio of MPO in disease group 2 to disease group 1 was greater than similar ratios for GCF volume and LF, AS and LDH. The quantity of MPO was the only measure which differed between the 2 disease groups at all surfaces. MPO thus appears to have the greatest potential, among the measured parameters, to serve as a marker for advanced periodontal disease.     

Microassay for the detection of elastase activity in the gingival crevice

BACKGROUND, AIMS: A new microassay for the detection of elastase activity (EA) in gingival crevicular fluid (GCF) has been established. GCF was collected with Periopaper strips and quantified in a Periotron. METHODS: Enzyme activity was measured in a microtiter plate reader, using a fluorometric assay. To ensure quality and precision of the assay, recovery rates were determined at different activities with a recovery of >90%. In a 2nd step, stability of the enzyme was investigated during storage at room temperature, +4 degrees C, -22 degrees C, -88 degrees C. GCF samples retained elastase activity of almost 100% after a storage of 3 days at -22 degrees C. In a group of 12 healthy volunteers, elastase activity was assayed throughout an 18 day experimental gingivitis protocol. RESULTS: Median activity increased from 481 microU/microl at baseline to 1444 microU/microl at day 18, which was accompanied by the development of the signs of gingivitis. The increase of EA during the experimental phase of the study was highly significant (p<0.001) and correlated well with the increasing severity of gingivitis. CONCLUSION: The data suggest that elastase activity in GCF is an excellent quantitative measure of gingival inflammation.     

Influence of smoking on the outcome of periodontal surgery

Abstract. The 5-year outcome following periodontal surgery was evaluated in 57 patients that had received regular maintenance care throughout the follow-up period. The study population included 20 smokers, 20 former smokers and 17 non-smokers in the age range 37–77 years. The clinical characteristics evaluated were supragingival plaque, gingival bleeding and pocket probing depth. The region assigned for surgery was, in addition, radiographically evaluated in terms of periodontal bone height. Furthermore, the occurrence of the periopathogens Actinobacillus actinomycetemcomitans (Aa), Porphyromonas gingivalis (Pg) and Prevotella intermedia (Pi) and the gingival crevicular fluid (GCF) levels of tumor necrosis factor alpha (TNF-α) were assessed at follow-up. Plaque index was 28.5% at baseline and 32.9% at follow-up, indicating a good standard of oral hygiene, and gingival bleeding 31.7% and 24.9%, respectively, suggesting a low to moderate level of gingival inflammation. In regions assigned for surgery, pocket probing depth decreased significantly from on average 5.6 mm to 4.3 mm ( p <0.0001) and periodontal bone height increased significantly from on average 62.5% to 67.5% ( p <0.0001). In terms of bone height, the outcome was less favorable among smokers compared with non-smokers. There was a predominance of smokers among patients exhibiting loss of bone height after the 5 years of maintenance. No significant associations were found between the therapeutical outcome and supragingival plaque or subgingival occurrence of periopathogens. The associations between GCF levels of TNF-α and probing depth and bone height were unclear, whereas the level of TNF-α was significantly elevated in smokers.     

Interleukin 1 and receptor antagonist levels in gingival crevicular fluid in heavy smokers versus non-smokers

BACKGROUND/AIMS: This study aimed to investigate the concentration of the cytokine interleukin (IL)-1beta and its receptor antagonist IL-1ra in gingival crevicular fluid (GCF) in patients with adult periodontitis who were heavy smokers compared with non-smokers. METHOD: GCF samples were collected from two groups of subjects: smokers and non-smokers. Thirty-nine GCF samples were harvested from 13 subjects with moderate to severe adult periodontitis who were heavy smokers. A further 30 GCF samples were harvested from 10 subjects with moderate to severe adult periodontitis who were non-smokers. Subjects were selected from both genders and none had any relevant systemic illness, were pregnant, had recent medication or had received any periodontal therapy in the preceding 3 months. One deep bleeding site, one deep non-bleeding site and one healthy site were investigated in each subject. Clinical measurements were recorded for each site, after obtaining a GCF sample using a Periopaper strip. IL-1beta and IL-1ra were quantified using new commercially available ELISA kits (Quantikine), and could be detected in all samples. RESULTS: For smokers, the mean concentrations for IL-1beta were 2714.5 (SD 4416.2) pg/ micro L for healthy sites, 37.0 (SD 57.2) pg/ micro L for non-bleeding periodontitis sites and 24.5 (SD 29.2) pg/ micro L for bleeding periodontitis sites. The concentrations of IL-1beta for non-smokers for the same category of sites were 393.8 (SD 867.1), 74.2 (SD 107.0) and 73.1 (SD 61.0) pg/ micro L, respectively. The mean concentrations of IL-1ra for smokers were 5.8 x 10(5) (SD 9.7) pg/ micro L for healthy sites, 2.2 x 10(5) (SD 0.15) pg/ micro L for deep non-bleeding sites and 0.19 x 10(5) (SD 0.07) pg/ micro L for deep bleeding sites. The concentrations for non-smokers were: 4.1 x 10(10) (SD 3.8), 18.1 x 10(5) (SD 20.4) and 3.2 x 10(5) (SD 2.3) pg/ micro L, respectively. Significance levels of P < 0.05 were found for comparisons of healthy vs. deep bleeding and deep non-bleeding sites for IL-1beta and IL-1ra in smokers, before adjustments for multiple testing. However, none of these comparisons reached statistical significance following adjustments for multiple testing. P < 0.05 for the correlation between IL-1beta and IL-1ra at healthy sites in smokers only. Differences in GCF concentrations for IL-1beta in smokers vs. non-smokers were significant for deep bleeding sites only (P < 0.05), the mean concentration of IL-1beta being lower in GCF from smokers vs. non-smokers. All differences in GCF concentrations of IL-1ra reached statistical significance for smokers vs. non-smokers. The mean concentrations of IL-1ra in GCF were lower in smokers compared with non-smokers for all categories of sites. CONCLUSIONS: A decreased concentration of IL-1beta and also IL-1ra was found in GCF from periodontitis sites compared to healthy sites in smokers and in non-smokers, although this did not reach statistical significance following adjustments for multiple testing. For comparisons between heavy smokers and non-smokers, statistically significant differences were found in the GCF concentrations of IL-1beta from deep bleeding sites only. Statistically significant differences were found in the IL-1ra concentrations for smokers vs. non-smokers for all categories of sites.