Differential expression of CR3, FcɛRll and FcγRlll on Polymorphonuclear leukocytes in gingival crevicular fluid

Polymorphonuclear leukocytes (PMNLs) are the most numerous cell population among the cellular infiltrates in gingival crevicular fluid (GCF) and play important roles in the host-defensive system in the gingival crevices. We determined the percentage of neutrophils, cosinophils and basophils in total PMNLs by light microscopic observation using Randolph-methylene blue staining, then assessed flow cytometric differences in the expression of CR3, FcγRIII, FcɛRII. LFA-1α, and LFA-Iβ on PMNL in GCF and peripheral blood (PB) from 21 patients with adult periodontius (AP) and 13 healthy donors. Percentages of basophils and eosinophils were higher in GCF than in PB. In both AP patients and healthy subjects, expression of CR3 and FcɛRll was higher while FeγRIII was lower in GCF than in PB. The statistical analysis showed that the expressions of FcγRIII and FcɛRII on GCF PMNLs were lower in AP patients than in healthy subjects. Expressions of LFA-1α and β on GCF were similar to those on PB PMNLs. PB PMNLs stimulated in vitro with Porphyromonas gingivalts culture supernatant and fMLP displayed an expression pattern of CR3, FcγRIII and FcɛRII on GCF PMNLs. However, C5a and IL-I failed to induce changes in FcγRIII and FcɛRII. The results indicate that GCF neutrophils are activated, present enhanced adhesion and a decreased IgG-binding ability which would reflect that they are at the terminal stage of activation, and that GCF contains a larger eosinophil fraction than in PB. Moreover, these GCF eosinophils appear to be activated.     

Expression of interleukin-2 receptor and HLA-DR on lymphocyte subsets of gingival crevicular fluid in patients with periodontitis

Expression of interleukin-2 receptor (IL2R) and HLA-DR on lymphocytes of gingival crevicular fluid (GCF) was examined by two-color flow cytometric analysis. GCF from 15 patients with periodontitis was collected by crevicular washing. Mononuclear cells were isolated by Ficoll-paque gradient centrifugation from inflamed gingival tissue (GT) and peripheral blood (PB) sampled from each of the 15 patients. Lymphocyte subsets were detected by using monoclonal antibodies (mAb) of Leu 12 (CD19), Leu 4 (CD3), Leu 3a (CD4) and Leu 2a (CD8) directed to B cells, T cells, helper/inducer T cells (Th) and suppressor/cytotoxic T cells (Ts), respectively. Anti-IL2R (CD25) and anti-HLA-DR were used as lymphocyte activation markers. IL2R- or HLA-DR-positive fractions in Th, Ts and B cells were calculated. Percentage of IL2R-positive fraction in Th (IL2R+ Th) of GCF (34.0%) was significantly higher than those of GT (18.4%) and PB (13.7%). IL2R-positive fraction in B cells (IL2R+ B) of GCF was the highest among the three groups (23.9% in GCF, 12.5% in GT, 6.3% in PB). Ts did not express IL2R regardless of the origin of the samples. Compared with PB and GT, GCF showed significantly higher HLA-DR expression on Th and Ts in GCF (PB: 8.7% and 27.1%; GT: 27.9% and 50.3%; GCF: 44.7% and 65.3%). These results suggest that lymphocytes in GCF were highly activated and are related to the local host immune response in periodontitis.     

ICAM-1-expressing pocket epithelium, LFA-1-expressing T cells in gingival tissue and gingival crevicular fluid as features characterizing inflammatory cell invasion and exudation in adult periodontitis

Activated T lymphocytes constitute a major component of inflammatory cells in the early periodontal lesion, and also appear in the gingival crevicular fluid. In an attempt to clarify the relationship between the ICAM-1 (CD54) expression of pocket epithelium in gingiva and the infiltrating lymphocyte population, we carried out an analysis of CD11a⁺ (LFA-lα), CD25⁺ (IL-2Rα) and CD4⁺ (Th) cells subjacent to ICAM-1-expressing pocket epithelia and CD11a⁺ CD25⁺CD4⁺ cells in gingival crevicular fluid (GCF). GCF was collected by crevicular washing from 16 patients with periodontitis (P group) and 3 subjects with healthy gingiva (H group). Peripheral blood (PB) was collected at the same time. Mononuclear cells were isolated by Ficoll-paque gradient centrifugation from GCF and PB. Monoclonal antibodies (mAb) to CD11a, CD25, and CD4 were used for three-color flow cytometry. Gingival biopsies were obtained from 7 patients in P group and 3 subjects in H group. Serial cryostat sections (6 μm in thickness) were prepared from each biopsy, on which a double staining was performed. The number of CD11a⁺ CD25⁺ CD4⁺ cells and the fluorescence intensity of FITC conjugated anti-CD 11a were significantly higher in GCF than in PB (p≤0.001 to p≤0.01). CD11a⁺ CD25⁺CD4⁺ cells were not detected in GCF in H group. The pocket epithelia expressed CD54 in P group, but not in H group. The number of CD11a⁺, CD25⁺ and CD4⁺cells infiltrating the connective tissue subjacent to the upper, middle and lower parts of the CD54 positive pocket epithelium (n=16) was 141±26, 38±13, 144±29 (cells/0.04 mm²), respectively, whereas in the CD54 negative pocket epithelium, it was (n=5) 9±2, 3±1, 8±3. In P group, the CD11a⁺CD25⁺CD4⁺cell number in GCF correlated with CD25⁺, CD11a⁺cells in the connective tissue subjacent to the CD54⁺pocket epithelium. These results indicate that expression of ICAM-1 in pocket epithelium is relevant to the migration of CD11a, CD25, CD4 positive cells in connective tissue subjacent to the pocket epithelium into the periodontal pocket. Assessing the relationship of our findings and other adhesion molecules would offer important clues to the understanding of T cell migration in affected gingiva.     

Immunohistological analysis of memory T lymphocytes and activated B lymphocytes in tissues with periodontal disease

Memory T-cells and activated B-cells were identified in cryostat sections of adult periodontitis (AP) lesions and categorized in terms of frequency and distribution. Nineteen periodontitis biopsies were obtained at the time of periodontal surgery to remove residual periodontal pockets following the completion of initial preparation. Gingival tissues exhibited various degree of inflammation (Gl of 0-2) but probing depths of < 4 mm and < 5 mm loss of attachment. As a control, 5 gingivitis specimens (Gl of 1, probing depth and loss of attachment of ≤ 3 mm) were obtained from premolar and third molar sites requiring extraction for either orthodontic treatment or pericoronitis. Serial cryostat sections (6 μm in thickness) were prepared from each biopsy, on which a double staining avidin-biotin immunoperoxidase and avidin-biotin alkaline phosphatase technique was used to identify CD4+, CD45RO+ memory T-cells and activated CD19+ B-cells expressing CD23 or CD25. In periodontitis lesions the mean percentage of CD4+ cells expressing CD45RO was consistently high (65.9% in the crevicular (C) one-third (1/3), 61.2% in the middle (M) 1/3 and 62.5% in the oral (O) 1/3). This contrasts with the low mean percentage of CD4+, CD45RA+ naive T-cells (17.1% in the C 1/3, 14.8% in the M 1/3 and 12.4% in the O 1/3). In gingivitis specimens, the incidence of CD4+, CD45RO+ was 81.9% in the C 1/3, 81.1% in the M 1/3 and 89.0% in the O 1/3. This was higher than that of periodontitis biopsies. With CD4+, CD45RA+ the incidence was 10.0% in the C 1/3, 8.0% in the M 1/3, and 6.6% in the O 1/3 and the relationship to the periodontitis biopsies was reversed. However, the percentage of CD23+ and CD25+, CD19+ B-cells which were identified in 13 out of 19 samples from periodontitis varied significantly (0-100% for CD23, 0-36.2% for CD25) in spite of similar clinical status. The frequency of B-cells and activated B-cells in the gingivitis was much lower than that of periodontitis. These results indicate that both T-cells and B-cells were in active stage in periodontitis lesions. Differences of immunohistological features between gingivitis and periodontitis may be attributable to the heterogeneity of profiles of cytokine production by CD4+, CD45RO+“memory’ cells.     

牙周炎患者龈沟液中sICAM-1含量的研究

目的 :研究牙周炎患者龈沟液可溶性细胞间粘附分子 (sICAM - 1)的含量。方法 :牙周炎患者 15例 ,正常对照20例。用ELISA法检测牙周炎患者龈沟液sICAM - 1的含量。结果 :正常对照龈沟液sICAM - 1含量为145.014±43.745ng/ml,男女间无显著性差异 (p >0.05)。牙周炎患者龈沟液sICAM - 1的含量为23.716±8.195ng/ml,明显低于正常对照 (p <0.01)。结论 :龈沟液sICAM - 1含量的降低与牙周炎有关。     

Elevated levels of collagen cross-link residues in gingival tissues and crevicular fluid of teeth with periodontal disease

Lysylpyridinoline (LP) and hydroxylysylpyridinoline (HP) are collagen cross-link residues. Lysylpyridinoline is present in most tissues, whereas LP is present mainly in mineralized tissue. Both are elevated in tissue with increased collagen resorption. The purpose of this investigation was to assess if the concentrations of LP and HP are elevated in gingiva and gingival crevicular fluid (GCF) of teeth with advanced periodontitis (AP). We investigated human gingival biopsies of healthy teeth (n = 19) and teeth with AP (n = 43) in 49 individuals. Samples of GCF from 54 teeth with AP were collected in seven patients and compared with samples from 11 patients with experimentally induced gingivitis. Levels of LP and HP were measured by HPLC and fluorescence detection. Gingival concentrations of HP but not LP around teeth with advanced periodontitis were significantly elevated compared with teeth with healthy periodontium. While significant amounts of HP and LP were measurable in the GCF of teeth with AP, no HP and LP was identified 3 months following non-surgical periodontal therapy of the teeth or in fluid from teeth subjected to experimentally induced gingivitis. Elevated concentrations of HP and LP in GCF may serve as indicators of ongoing destruction of periodontal tissues and alveolar bone in advanced periodontitis.     

Influence of sE-selectin and L-selectin on the regulation of cell migration during chronic periodontitis

Soluble endothelial leukocyte adhesion molecule-1 (sE-selectin) levels in peripheral blood (PB) and gingival capillary blood (GCB) of both healthy donors (HD) and patients with adult periodontitis (AP) were assayed by ELISA. Binding of sE-selectin to polymorphonuclear neutrophils (PMN) from PB, GCB and crevicular fluid (GF), and expression of L-selectin and sialyl-Lewis^x (sLe^x) on these cells were analyzed by immunofluorescence and flow cytometry. No significantly enhanced serum levels of sE-selectin in patients with AP. compared to HD (28±5 ng/ml vs 19±3 ng/ml, respectively), and no differences in the concentration of sE-selectin in GCB (16±1 ng/ml vs 16±2 ng/ml, respectively) were observed. On PB-PMN no significant differences in the expression of L-selectin and sLe^x were found and binding of sE-selectin to PB-PMN was comparable between HD and patients with AP. Binding of sE-selectin to GCB-PMN was significantly higher in patients with AP compared to HD (mean channel fluorescence (MCF)=88.5± 13.2 vs MCF=24.2±5.3, respectively). The expression of sLe^x on GCB-PMN did not differ significantly between the two groups. A significant decrease in the expression of the adhesion molecule L-selectin on GCB-PMNs compared to PB-PMN was found in patients with AP but not in HD. CF-PMN showed decreased expression of both L-selectin and sLe^x compared to PMN from PB and GCB, both in HD and patients with AP. Taken together, these data suggest that PMN from patients with AP had reduced selectin-mediated adhesive capabilities to inflamed gingival endothelium.     

Secretory function of neutrophilic leukocytes of the patients with periodontal diseases

Aim of study was to examine periodontal status among 20 44 year old patients and to study the secretory function of peripheral venous blood neutrophilic leukocytes (NL) exposed to various antigens and alkaline phosphatase (AP) activity in gingival crevicular fluid (GCF) in patients suffering from gingivitis and periodontitis. Clinically were determined Russell's periodontal index (PI). Secretory function of NL affected by opsonized zymosane, non-opsonized E. coli was examined in 77 patients with gingivitis and periodontitis, and in 35 donors, free of internal diseases, by means of beta-glucuronidase (beta-GD), lysozyme (LZ). NL secreted higher levels of beta-GD in incubation medium in patients with periodontitis (p < or = 0.001) subject to degree of periodontal lesion. NL affected by various antigens secreted higher levels of LZ into non-cellular matrix in patients with gingivitis and periodontitis comparing to control environment in analogous groups. Data obtained from this study suggest that in patients with periodontitis response of NL to bacterial stimuli is specific and subject to the degree of periodontal lesion. Our study showed a significant difference of AP activity in GCF subject to pocket depth and degree of periodontal lesion. Once NL are exposed to corpuscules prone to phagocytosis, an increase in secretion of beta-GD and LZ can be explained by overall increase in secretion of NL lysosomic enzymes, thus disclosing the mechanism of inflammatory periodontal tissue damage.     

Identification and possible function of cathepsin G in gingival crevicular fluid from chronic adult periodontitis patients and from experimental gingivitis subjects

The levels of cathepsin G in gingival crevicular fluid (GCF) from chronic adult periodontitis patients and experimental gingivitis subjects were determined both by activity measurement using N-benzoyl-(DL)-phenylalanine-2-naphthyl ester as a substrate and by enzyme immunoassay using anti-human cathepsin G IgG. The activity level of cathepsin G in GCF of both periodontitis and experimental gingivitis has no significant correlation with all measured clinical parameters. Western immunoblotting using antibodies specific for cathepsin G or α1-proteinase inhibitor revealed that the difficulty in demonstrating the association of its activity with the severity of these diseases was due largely to formation of the enzyme-inhibitor complexes. By contrast, statistically significant positive correlation was found between cathepsin G content in GCF of periodontitis, which was determined by enzyme immunoassay, and such clinical parameters as the GCF volume, the gingival index and probing depth. The increased cathepsin G content with increasing severity of periodontal inflammation was markedly diminished by the initial treatment. Although no significant activity was detectable in GCF of experimental gingivitis, a rapid increase of the immunoreactive cathepsin G was found in GCF at 3–5 d after refraining from oral hygiene measures, which rapidly decreased by 10 d. The progressively increased cathepsin G between 10th and 21st d rapidly decreased by cleaning of the teeth. The results indicate that cathepsin G is involved in the host's defensive mechanism against the invasion of etiologic microbes and/or the development of either periodontitis or gingivitis.     

Estimation of soluble CD14 levels in gingival crevicular fluid and serum in diseased and healthy periodontium

ObjectiveTo estimate the levels of sCD14 in gingival crevicular fluid and serum under periodontally-healthy and diseased conditions.MethodsThe subjects were divided into three groups of 15, each as follows: healthy, gingivitis, and periodontitis. Periodontal parameters including Probing pocket depth, Clinical attachment level, Bleeding index, and Plaque index. Gingival crevicular fluid and serum samples were collected and analyzed for sCD14 levels using commercially-available ELISA kits.ResultsThe mean concentration of sCD14 in GCF was significantly lower in the gingivitis (134.5 ± 26.85 ng/mL) and periodontitis (103.23 ± 20.36 ng/mL) groups than in the healthy group (172.77 ± 46.33 ng/mL); p < 0.001. The mean serum concentration of sCD14 in the healthy group was 1528.13 ± 387.37 ng/mL, which was significantly less than that of the periodontitis group (2051.50 ± 381.10 ng/mL); p = 0.011.ConclusionsThe serum sCD14 levels in the periodontitis groups were significantly higher than those in the healthy controls. The levels of sCD14 in GCF were significantly lower in the gingivitis and periodontitis groups than in the healthy group.     

Collagenolytic activity in crevicular fluid from patients with chronic adult periodontitis, localized juvenile periodonitis and gingivitis, and from healthy control subjects

Collagenolytic activity in gingival crevicular fluid (GCF) sampled from 25 healthy control subjects, 25 gingivitis, 25 chronic adult periodontitis (CAP) and 8 LJP patients was correlated with clinical disease parameters using both the site and the patient as sampling units. Among patients collagenase activity increased with the severity of the disease in the order: healthy < gingivitis < periodontitis. Among sites , significant correlation was found between GCF collagenase activity and pocket depth in CAP and LJP, but not in gingivitis patients. Enzyme activity was also correlated with GI score in LJP, but not in CAP and gingivitis patients. In a subset of 10 patients in each of the healthy, gingivitis and CAP groups the association of enzyme activity and crevicular fluid volume (flow) was examined. Significant correlation was found between fluid volume and pocket depth in CAP patients, and between fluid volume and GI score in gingivitis patients, but no association was observed between collagenase activity and fluid volume. The Collagenolytic enzyme was shown to be a genuine vertebrate collagenase derived from unidentified host cells. The concentration of the enzyme in crevicuar fluid from CAP patients was in the order of 10 μg/ml.     

Lower protein concentration in GCF from patients with periodontitis: an indicator of host-specific inflammatory reaction

Abstract To study the local inflammatory reaction associated with gingivitis and periodontitis, the protein concentrations were measured in gingival crevicular fluid (GCF) from 3 types of site: (i) inflamed sites in patients with gingivitis alone; (ii) inflamed sites with or (iii) without tissue destruction in patients with periodontitis. The GCF was sampled with paper strips and the protein concentration was measured with the Bradford protein assay. Higher protein concentrations in GCF were found at inflamed sites in patients with gingivitis alone than at clinically similar sites without tissue destruction in patients with periodontitis. In contrast, the sites with various degrees of disease in the same periodontitis patient showed the same protein concentrations. These discrepancies indicate that the local inflammatory reaction in the gingiva is host- and not site-specific, i.e., the tissue destruction in periodontitis reflects a specific host response.     

Longitudinal evaluation of prostaglandin E2 (PGE2) and periodontal status in HIV+ patients

The study aim was to determine whether prostaglandin E(2) (PGE(2)) in gingival crevicular fluid (GCF) could serve as a risk factor for periodontitis in human immunodeficiency virus-positive (HIV(+)) patients. Clinical measurements, including gingival index (GI), plaque index, bleeding index, probing depth (PD), attachment loss (AL) and GCF samples were taken from two healthy sites (including sites with gingival recession, GI=0; PD< or =3 mm; AL< or =2 mm), three gingivitis sites (GI>0; PD< or =3 mm; AL=0) and three periodontitis sites (GI>0; PD> or =5 mm; AL> or =3 mm) of each of the 30 patients at baseline and 6-month visits. GCF samples were also taken by means of paper strips. GCF PGE(2) levels were determined by a sandwich ELISA. The progressing site was defined as a site which had 2 mm or more attachment loss during the 6-month study period. The mean amounts of PGE(2) were significantly higher in gingivitis and periodontitis sites than in healthy sites (p<0.0001). GCF levels of PGE(2) were significantly correlated with probing depth, attachment loss, CD4(+) cells, viral load, age and smoking pack-years at baseline and 6-month visits (0.0001相似文献   

Gingival crevicular fluid and serum visfatin concentration: their relationship in periodontal health and disease

Background: Visfatin is a pleiotropic mediator that acts as growth factor, cytokine, and enzyme involved in energy including nicotinamide adenine dinucleotide metabolism and was recently demonstrated to exert several proinflammatory functions. The purpose of this study is to determine the presence of visfatin in gingival crevicular fluid (GCF) and serum samples and to find out their association, if any. Methods: At the beginning of the study, 40 individuals (20 males and 20 females; age range: 23 to 53 years) were selected and divided into three groups based on the gingival index, probing depths, clinical attachment levels, and radiologic parameters (bone loss). Group 1 (10 patients with healthy periodontium; age range: 25‐36 years), group 2 (15 patients with gingivitis; age range: 25‐36 years), and group 3 (15 patients with chronic periodontitis; age range: 23‐53 years) GCF (by microcapillary pipettes) and serum (by venipuncture) samples were collected to estimate levels of visfatin using an enzyme‐linked immunosorbent assay kit. Results: Mean visfatin concentrations increased in GCF and serum with the severity of disease from healthy to gingivitis to periodontitis groups and differed significantly (P <0.05). However, it was found that GCF values were higher than serum values. The highest to lowest visfatin concentrations were found in groups 3 through 1, respectively. Visfatin in GCF and serum correlated positively with periodontal parameters in the chronic periodontitis group. Conclusions: The results suggest that GCF and serum visfatin concentrations increase with the severity of periodontal disease. Hence, visfatin values were considered an inflammatory marker in periodontal disease in GCF and serum. Visfatin also deserves further consideration as a therapeutic target.     

Gingival crevicular fluid matrix metalloproteinase-13 levels and molecular forms in various types of periodontal diseases

BACKGROUND: The purpose of this study was to evaluate the levels, molecular forms and activation degree of matrix metalloproteinase-13 (MMP-13) in the gingival crevicular fluid (GCF) of patients with periodontal diseases and to correlate these findings with periodontal clinical parameters. METHODS: Sixty one subjects participated in this study as healthy (n = 18), gingivitis (n = 17), aggressive periodontitis (AgP; n = 15) and chronic periodontitis (CP; n = 11) groups. Clinical measurements and GCF samples were obtained from each subject. The molecular forms of MMP-13 in GCF samples were analyzed by Western immunoblotting method. Differences among the groups were assessed using non-parametric statistical analysis. RESULTS: In the CP group, levels of 29-30 kDa fragment of MMP-13, total MMP-13, and activated form of MMP-13 were significantly higher than in the healthy, gingivitis and AgP groups. GCF levels of all molecular forms of MMP-13 in AgP group were similar to those of healthy and gingivitis groups. Total and activated MMP-13 levels were positively correlated with all clinical parameters. 29-30 kDa fragment levels of MMP-13 were also positively correlated with papillary bleeding index and plaque index. CONCLUSION: These results indicate that elevated GCF MMP-13 levels may play an important role in the pathogenesis of CP. These data demonstrate, for the first time, pathologically activated and elevated MMP-13 in GCF.     

Granulocyte elastase in gingival crevicular fluid

The granulocyte elastase activity and the immuno-reactive (antigenic) granulocyte elastase of gingival crevicular fluid (GCF) were studied in 16 periodontitis patients and in 10 gingivitis patients. The elastase activity was measured with a low molecular weight substrate specific for granulocyte elastase. The antigenic elastase was determined with specific antibodies against granulocyte elastase. Intracrevicular sampling of GCF with paper strips for 30 s seemed to provide representative values of elastase. The elastase activity correlated with probing depth and attachment loss and appeared to be a measure of the degree of tissue destruction. Antigenic elastase represents the number of granulocytes in GCF and should thus be related to the degree of inflammation. The periodontitis patients and the gingivitis patients both had a similar degree of inflammation as measured by antigenic elastase per microliter GCF and gingival index. The elastase activity per microliter GCF, however, was higher in the periodontitis group. Elevated granulocyte elastase activity in GCF seems to be independent of inflammation and could thus be an indicator of patients at risk for periodontitis.     

Gingival crevicular fluid collagenase-2 (MMP-8) test stick for chair-side monitoring of periodontitis

BACKGROUND: A rapid chair-side test based on the immunological detection of elevated levels of collagenase-2 (matrix metalloproteinase-8, MMP-8) in gingival crevicular fluid (GCF) was developed to identify and monitor the course and treatment of adult periodontitis. METHODS: MMP-8 was determined in GCF from periodontitis (11 patients, 90 sites), gingivitis (10 patients, 58 sites) and healthy control (8 patients, 59 sites) sites (i) by a test stick incorporating monoclonal antibodies to two epitopes on MMP-8 and (ii) by measuring MMP-8 concentration by a quantitative immunofluorometric assay. Patients with adult periodontitis were treated by scaling and root planing (SRP) and received oral hygiene instructions. GCF MMP-8 testing and clinical measurements were done before and after SRP. RESULTS: MMP-8 GCF levels and chair-side test differentiated periodontitis from gingivitis and healthy control sites. MMP-8 GCF levels > 1 mg/l and positive chair-side test identified especially severe periodontitis sites. A positive and negative test stick result, the outcome of which was rapidly detectable in 5 mins, in GCF correlated well with MMP-8 immunofluorometric assay analysis from the collected GCF samples and the severity of periodontitis. Scaling and root planing reduced the MMP-8 levels in severe periodontitis sites with positive MMP-8 test and gingival probing pocket depth (PD) > 5 mm before treatment. The test stick result and the quantitative assay were discrepant in only 18 of the 207 sites tested, thus agreement was very good (kappa = 0.81). With a threshold of 1 mg/l MMP-8 activity the chair-side test provided a sensitivity of 0.83 and specificity of 0.96 (n = 207). CONCLUSION: The MMP-8 test can be used to differentiate periodontitis from gingivitis and healthy sites as well as to monitor treatment of periodontitis. A reduction in GCF MMP-8 levels and a change in test stick result provide a means to optimize patient control during maintenance of periodontal treatment.     

牙周炎患者GCF及血清中IL-6测定及其与牙周炎的关系

目的：研究龈沟液（GCF）中白细胞介素－6（IL－6）的来源及其与牙周炎的关系。方法：采用双抗夹心酶联免疫吸附法（ELISA）测定了72例成人牙周炎，13例牙龈炎，18例健康对照组GCF IL－6含量，及17例牙周炎患者，7例健康人血清IL－6水平。结果：牙周炎患者GCF IL－6水平显著高于健康人，牙周炎患者GCF IL－6II与牙周炎患者血清中IL－6含量有显著性差异，牙龈炎患者血清IL－6含量比正常对照组含量高，牙周炎患者GCF IL－6含量与牙龈炎患者有显著性差异，重度牙周炎患者GCF中IL－6含量高于轻度牙周炎患者，牙周炎患者GCF中IL－6含量与出血指数（BI）、牙周袋深度（PD）、附着丧失（AL）有正相关关系。结论：龈沟液中IL－6主要来源于局部牙龈组织，IL－6在牙周炎的发生，发展过程中起一定作用。     

α1-Proteinase inhibitor in gingival crevicular fluid of humans with adult periodontitis: serpinolytic inhibition by doxycycline

The serum protein, α₁-proteinase inhibitor (α₁PI), defends the host against serine proteinases, e.g. PMN elastase. Using a rabbit anti-serum against human α₁-PI, this protein in GCF was quantified from a standard curve constructed from dot-blot analysis and characterized by Western blot. GCF was collected on filter paper strips from healthy (H), gingivitis (G) and adult periodontitis (AP) patients, then extracted with Tris/NaCl/CaCl₂ buffer, pH 7.6. α₁-PI concentration increased with G and was highest in AP subjects. H sites only showed intact α₁-PI (52 kDa); no degradation fragments (48 kDa) were detected. In G and AP subjects, α₁-PI degradation fragments were seen in 17% and 71% of GCF samples, respectively. Both collagenase and α₁-Pl-degrading activities in GCF increased with severity of inflammation (GCF flow). Moreover, the α₁-PI degrading (or serpinolytic) activity was characterized as a matrix metalloproteinase, probably collagenase, based on its in vitro response to a panel of different proteinase inhibitors including doxycycline. We propose: (1) that collagenase promotes periodontal breakdown not only by degrading collagen, but also by depleting α₁-PI regulation of elastase and other serine-proteinases, thereby favoring a broader attack on extracellular matrix (ECM) constituents, and (2) based on a recent longitudinal double-blind study using the techniques described above for α₁-PI analysis, that low-dose doxycycline administration to humans with adult periodontitis can inhibit this broad cascade of ECM degradation.     

Experimental gingivitis in periodontitis-susceptible subjects

Abstract The purpose of this study was to evaluate clinical, microbiological, and gingival crevicular fluid (GCF) profiles in periodontitis-resistant and periodontitis-susceptible subjects during 4 weeks of experimental gingivitis. Experimental groups of similar ages were defined as gingivitis controls (GC; n=10) and history of rapidly progressive periodontitis (RPP; n=10). respectively. Prior to baseline, all subjects achieved good plaque control (plaque index (P1I) ～0) and gingival health (gingival index (GI)=0). and had probing depths ≤4 mm on experimental teeth. For 4 weeks after baseline, oral hygiene around maxillary 2nd premolar and 1st molar teeth was inhibited by a plaque guard. The plaque guard was removed weekly for GCF sampling to determine interleukin (IL)-1β and prostaglandin (PG)E₂ amounts by ELISAs. In addition. PII. GI. probing depth, and gingival recession measurements were made. Subgingival plaque darkfield microscopy and DNA probe analysis also were performed. Results indicated that clinical signs of inflammation, microbiological patterns and GCF profiles progressed similarly in both groups. However, plaque accumulated more rapidly in the susceptible subjects. PII in RPP at 4 weeks was 2.1±0.1 compared to 1.5±0.2 in GC, with an incidence of PII>1 of 100% versus 50%, respectively (logistic regression; p<0.000l). Hence, the clinical, microbiological and host factors selected for this study were unrelated to previous susceptibility to periodontitis when evaluated in the experimental gingivitis model. However, the increased rate of plaque accumulation, following thorough plaque removal, in RPP patients suggests a potential factor in disease recurrence in these susceptible subjects.