Matrix metalloproteinase-9 and tissue inhibitor of matrix metalloproteinase-1 in blood as markers for early atherosclerosis in subjects with chronic periodontitis

Background and Objectives: An association has been found between periodontal disease and the development of atherosclerosis. We investigated the hypothesis that periodontal disease triggers the expression of matrix metalloproteinase‐9 (MMP‐9) and tissue inhibitor of matrix metalloproteinase‐1 (TIMP‐1) in blood. Increased levels of these parameters might then indicate early atherosclerosis. Material and Methods: In this cross‐sectional study, the material comprised 80 subjects with chronic periodontitis and 31 subjects with no periodontal disease. Sixteen years after diagnosis of periodontal disease ultrasonography revealed a statistically significant difference (p < 0.001) of carotid intima–media thickness between the subjects with chronic periodontitis and the periodontally healthy subjects. Matrix metalloproteinase‐9 and TIMP‐1 were analyzed from blood as periodontal and systemic inflammatory markers. The relationship between MMP‐9, TIMP‐1 and MMP‐9/TIMP‐1 as dependent variables and several independent variables (age, sex, smoking, education, body mass index, hypertension, periodontal disease and cholesterol) were analyzed in multiple logistic regression models to assess the value of the inflammatory markers in predicting carotid atherosclerosis. Results: Matrix metalloproteinase‐9 and TIMP‐1 were significantly higher in plasma from subjects with periodontal disease and atherosclerosis. Periodontal disease was identified as the principal independent predictor both for atherosclerosis (odds ratio 3.89 for increase in bilateral carotid intima–media thickness) and for increased MMP‐9, TIMP‐1 and MMP‐9/TIMP‐1 (odds ratio 2.58, 5.53 and 3.41, respectively). Classical atherosclerosis risk factors, such as increased total cholesterol, age and sex (women), were significant predictors in the model. Conclusion: Matrix metalloproteinase‐9, TIMP‐1 and MMP‐9/TIMP‐1 in blood from subjects with periodontal disease could be useful laboratory markers for increased carotid artery intima–media thickness.     

Capsaicin-induced local elevations in collagenase-2 (matrix metalloproteinase-8) levels in human gingival crevice fluid

BACKGROUND AND OBJECTIVES: Application of capsaicin on alveolar mucosa provokes pain and neurogenic vasodilatation in the adjacent gingiva. Pain-associated inflammatory reactions may initiate expression of several pro-inflammatory mediators. Collagenase-2 (matrix metalloproteinase-8: MMP-8) is the major destructive protease, especially in the periodontitis-affected gingival crevice fluid (GCF). With this background, we wished to study whether capsaicin stimulation of alveolar mucosa can induce changes in the GCF MMP-8 levels. MATERIAL AND METHODS: For 10 generally and periodontally healthy human volunteers, capsaicin (3%)-moistened filter paper was applied unilaterally to the buccal alveolar mucosa on the anterior maxilla. GCF samples were collected from the tooth at the stimulation site and from several other incisors in the upper jaw. MMP-8 levels and molecular forms in GCF samples were determined by immunofluorometric assay (IFMA) and western immunoblotting, respectively. RESULTS: Capsaicin stimulation of the alveolar mucosa induced significant local elevations in levels and activation of MMP-8 in GCF of the adjacent teeth. Western immunoblot revealed that both neutrophil- and mesenchymal-type MMP-8 isoforms were elevated and activated, together with 110 kDa high-molecular size MMP-8 species. This capsaicin-evoked MMP-8 elevation lasted several minutes after stimulation. During the experiments, no marked changes occurred in MMP-8 levels in the GCF of distantly located teeth. CONCLUSIONS: These results suggest that capsaicin-evoked neurogenic gingival inflammation can trigger the expression and activation of MMP-8 in GCF of the adjacent teeth.     

Effects of phase I periodontal treatment on gingival crevicular fluid levels of matrix metalloproteinase-3 and tissue inhibitor of metalloproteinase-1

OBJECTIVES: The aim of this study was to evaluate the effects of phase I periodontal treatment on gingival crevicular fluid (GCF) levels of matrix metalloproteinase (MMP)-3 and tissue inhibitors of metalloproteinase (TIMP)-1. METHODS: Plaque index, gingival index, pocket depth and clinical attachment loss were recorded and GCF samples were collected from 20 chronic periodontitis (CP) patients and 20 periodontally healthy controls (C) before treatment. CP patients received phase I periodontal treatment and all clinical parameters were recorded and GCF samples were collected once more after treatment. Assays were performed by an enzyme-linked immunosorbent assay. RESULTS: All of the clinical parameters improved significantly after the therapy (p<0.05). Baseline GCF levels of MMP-3 were significantly higher than C and that level was reduced significantly by treatment compared with baseline levels (p<0.05). Baseline GCF levels of TIMP-1 were lower than post-treatment levels and C (p<0.05). GCF levels of TIMP-1 increased significantly by treatment compared with baseline levels (p<0.05). CONCLUSION: This study shows that the clinical improvements after phase I periodontal therapy are accompanied by reduction in MMP-3 and increasing in TIMP-1 GCF levels.     

Experimental tooth pain elevates substance P and matrix metalloproteinase-8 levels in human gingival crevice fluid

Objective. Tooth pain can induce a neurogenic inflammatory reaction in gingiva in association with local elevations of matrix metalloproteinase (MMP)-8, which is considered the major tissue destructive protease in gingival crevice fluid (GCF). The pro-inflammatory neuropeptides released by sensory nerves coordinate the activities of the immuno-effector cells and may influence the secretion of MMP-8. With this background, we studied whether experimental tooth pain can trigger changes in GCF levels of the neuropeptide substance P (SP) and MMP-8. Material and methods. The GCF SP levels of stimulated and non-stimulated teeth were analyzed for SP using a competitive enzyme immunoassay (EIA). The GCF MMP-8 levels were determined by quantitative immunofluorometric assay (IFMA). Results. Painful stimulation of the upper central incisor caused significant elevations in GCF SP and MMP-8 levels of the stimulated tooth. At the same time, the GCF SP and MMP-8 levels of non-stimulated control teeth were unchanged. Conclusions. These data indicate that experimental tooth pain can induce local elevations of SP and MMP-8 levels in GCF simultaneously. This supports the possibility of a local neurogenic spread of inflammatory reactions from intrapulpal to surrounding periodontal tissues.     

Matrix metalloproteinase-8 levels and elastase activities in gingival crevicular fluid from chronic adult periodontitis patients

AIM: To make an initial assessment of the periodontal diagnostic potential of immunoreactive matrix metalloproteinase-8 (MMP-8) in gingival crevicular fluid (GCF) by comparison with elastase activity which has previously been associated with disease severity and progression. METHODS: GCF was collected from molar and premolar sites of 16 chronic adult periodontitis patients before treatment and 13 of this group 2 weeks after scaling and root planing. Samples were analysed for MMP-8 by immunofluorometric assay and for elastase activity with a fluorogenic substrate. RESULTS: Mean patient clinical parameters and GCF enzyme totals both decreased significantly after treatment. Total MMP-8 levels and elastase activities generally correlated significantly with gingival and bleeding indices. For GCF concentrations, only MMP-8 showed a significant fall after treatment, and some significant correlations with clinical parameters. Amounts of the 2 enzymes correlated significantly with each other. CONCLUSIONS: Similarities between MMP-8 and elastase probably reflect the fact that both enzymes are associated mainly with neutrophils: MMP-8 levels may have fallen more after treatment because the assay, unlike that for elastase, would most likely not have detected much enzyme bound to alpha-macroglobulin. The immunoassay for MMP-8 is more specific and convenient than functional collagenase assays, and might be suitable for monitoring the periodontal condition.     

Effects of sub-antimicrobial dose doxycycline therapy on crevicular fluid MMP-8, and gingival tissue MMP-9, TIMP-1 and IL-6 levels in chronic periodontitis

OBJECTIVE: To investigate whether sub-antimicrobial dose doxycycline (SDD) therapy for 120 d in chronic adult periodontitis patients had significant effects on gingival crevicular fluid (GCF) matrix metalloproteinase-8 (MMP-8) levels, and on gingival tissue MMP-9, tissue inhibitor of matrix metalloproteinases-1 (TIMP-1) and interleukin-6 (IL-6) levels. BACKGROUND: Tetracycline can significantly inhibit MMP activity in GCF and in gingival tissue, even in much lower dosage then a traditional antimicrobial dosage used in conventional therapy. Sub-antimicrobial dose doxycycline (SDD) therapy has been shown to reduce periodontal disease activity to control MMP and pro-inflammatory cytokines. METHODS: A total of 32 patients with incipient to moderate (probing pocket depth approximately 4-7 mm) chronic adult periodontitis were included in the study. Subjects were randomly assigned to two groups. After scaling and root planning (SRP), the SRP + SDD group received SDD, 20 mg bid, whereas the SRP + placebo group received placebo, 20 mg bid. In the follow-up, efficacy measures included the change in probing pocket depth (PD), clinical attachment level (CAL), bleeding on probing (BOP) and gingival crevicular fluid MMP-8 levels, gingival tissue MMP-9, TIMP-1 and IL-6 levels from baseline to 120 d. RESULTS: After 120 d, PD and CAL improved significantly in the SRP + SDD group. Initial MMP-8 levels for the SRP + SDD group and the SRP + placebo group were 407.13 +/- 114.45 ng/ml and 378.71 +/- 189.39 ng/ml, respectively, with no statistical difference between the two groups. MMP-8 levels for the SRP + SDD group and the SRP + placebo group were: 235.35 +/- 134.58 ng/ml and 364.04 +/- 219.27 ng/ml at 30 d; 157.50 +/- 95.95 ng/ml and 236.60 +/- 186.16 ng/ml at 60 d; 102.70 +/- 67.64 ng/ml and 208.56 +/- 124.54 ng/ml at 90 d; and 63.77 +/- 53.33 ng/ml and 229.13 +/- 168.09 ng/ml at 120 d, respectively. The amount of decrease in MMP-8 levels for the SRP + SDD group was statistically significant compared to that for the SRP + placebo group, especially apparent at 120 d (p < 0.05). TIMP-1 levels in both groups increased from the baseline to 120 d with statistical significance (p-value < 0.05), but there was no significant difference between the two groups. Changes in MMP-9 and IL-6 levels were not statistically significant. CONCLUSION: Adjunctive SDD therapy can improve the clinical parameters and this clinical improvement is reflected by controlled level of MMP-8 in chronic adult periodontitis after the therapy.     

Levels of tissue inhibitor of metalloproteinases-1 and matrix metalloproteinases-1 and -8 in gingival crevicular fluid following treatment with enamel matrix derivative (EMDOGAIN)

The mechanism of enamel matrix derivative (EM D) action on the periodontal wound healing process is not well understood. However, earlier in vitro studies from our laboratory demonstrated that EMD stimulated the proliferation of both periodontal ligament and gingival fibroblast cells. Therefore, the purpose of this study was to further evaluate the effect of EMD on the early wound healing process by assessing the protein levels of matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs) in gingival crevicular fluid (GCF). Sixteen patients, each of whom had one or two pairs of infrabony defects located contralaterally in the same arch, were included in this clinical trial. Thirty-six infrabony defects were randomly assigned treatment with flap surgery plus EMD or flap surgery plus placebo. At baseline and at 2, 4 and 12 week follow-up evaluation visits, GCF was sampled with paper strips. After determination of GCF volume, TIMP-1, MMP-1 and MMP-8 GCF levels were measured by an enzyme-linked immunosorbent assay. Intragroup analysis: At week 2 following surgery, when compared to baseline all parameters in each study group, except MMP-1, significantly increased (p<0.05). There were no significant differences between 4 or 12 weeks and baseline in either study group. Intergroup analysis: At 4 weeks after surgery, GCF volume and TIMP-1 levels showed a significant decrease (p<0.05) in the EMD group, when compared to the placebo group. MMP-1 levels at weeks 2, 4 and 12, and MMP-8 levels at weeks 4 and 12 were significantly lower (p < 0.05) in the EMD group compared to the placebo group. EMD compared to placebo treated sites demonstrated a more rapid return to baseline levels of TIMP-1, MMP-1 and MMP-8. These findings suggest that treatment with flap surgery and EMD, compared to flap surgery with placebo, accelerated healing at an earlier stage of wound healing following surgery.     

Matrix metalloproteinases and their inhibitors in gingival crevicular fluid and saliva of periodontitis patients

Abstract Matrix metalloproteinases (MMPs) and serine proteinases seem to be related to tissue destruction in periodontitis. The presence of MMPs in gingival crevicular fluid (GCF) and saliva, however, has not been studied comprehensively with the enzyme-linked immunosorbent assay (ELISA)-technique, We therefore examined the levels of MMP-1,-3.-8 and -9, and their endogenous inhibitor, tissue inhibitor of matrix metalloproteinases (TIMP-1). in GCF and saliva of patients with adult periodontitiss (AP) and localized juvenile periodontitis (LJP). Elevated levels of MMP-1 were detected in LJP GCF compared to AP and control GCF. Elevated levels of TIMP-1 were also detected in LJP GCF in comparison to AP and control GCF Higher MMP-8 levels were detected in AP GCF compared to LJP and control GCF. The relative low levels of MMP-3 were present in all studied GCF samples. Elevated levels of MMP-8 were further detected in saliva of AP compared to LJP and the controls. Both MMP-1 and TIMP-1 were detected in all studied saliva samples, but not significant differences were detected between the studied groups. Our ELISA-results confirm that (i) PMN MMP-8 and MMP-9 are the main collagenase and gelatinase in AP GCF, whereas GCF collagenase in LJP seems to be of the MMP-1-type; (ii) only low levels of TIMP-1. endogenous MMP-inhibitor. are present in AP GCF. which emphasises the importance of doxycycline as a possible adjunctive drug in the treatment of AP patients; (iii) tests based on specific antibodies against PMN MMPs. especially MMP-8, might serve as a reliable method of measuring and monitoring enzyme levels in GCF from different periodontitis patients.     

Risk factors for periodontitis in HIV patients

OBJECTIVE: The purpose of this study was to identify risk factors for periodontitis associated with human immunodeficiency virus (HIV) infection. METHODS: A total of 152 HIV(+) patients were recruited from the CARE clinic at the University of the Pacific School of Dentistry. Clinical measurements (gingival index, plaque index, bleeding index, probing depth, and attachment loss), gingival crevicular fluid (GCF) and subgingival plaque samples were taken from eight sites of each patient at baseline and 6-month visits. GCF neutrophil elastase was determined by measurement of p-nitroanalide resulting from hydrolysis of an elastase-specific peptide. GCF beta-glucuronidase was determined by release of 4-methylumbelliferone from hydrolysis of a specific substrate. A bacterial concentration fluorescence immunoassay was used to detect periodontopathic bacteria in subgingival plaque samples. RESULTS: Viral load, age, smoking pack-years, Fusobacterium nucleatum, Prevotella intermedia, Actinobacillus actinomycetemcomitans, Porphyromonas gingivalis, neutrophil elastase, and beta-glucuronidase were significantly correlated with clinical measurements (0.0001 < p < 0.05). Significantly higher levels of elastase, beta-glucuronidase, F. nucleatum, P. intermedia, and A. actinomycetemcomitans were found at progressing sites than in non-progressing sites (0.001 < p < 0.05). CONCLUSIONS: These data indicate that age, smoking pack-years, viral load, F. nucleatum, P. intermedia, A. actinomycetemcomitans, elastase, and beta-glucuronidase are risk factors for periodontitis in HIV(+) patients.     

Meta-analysis of the association between serum and gingival crevicular fluid matrix metalloproteinase-9 and periodontitis

Background

The authors of this meta-analysis evaluated whether there is a difference in the levels of serum and gingival crevicular fluid (GCF) matrix metalloproteinase (MMP)-9 between patients with periodontitis and periodontally healthy control participants.

The effect of topical doxycycline usage on gingival crevicular fluid MMP-8 levels of chronic and aggressive periodontitis patients: a pilot study

Abstract:  The aim of this study was to evaluate the efficacy of topical subgingival application of doxycycline hyclate (DH) gel adjunctive to non-surgical periodontal therapy on gingival crevicular fluid (GCF) matrix metalloproteinase (MMP)-8 levels in chronic and aggressive periodontitis patients. Forty teeth of 10 chronic periodontitis patients and 32 teeth of eight aggressive periodontitis patients were screened for 6 months. Scaling and root planing (SRP) was applied to the control sites and DH gel adjunctive to SRP was applied to the test sites of each patient simultaneously. GCF MMP-8 levels were analysed at baseline, 7 days; and at 1, 3 and 6 months by Sandwich Elisa Method. At 1, 3 and 6 months, probing depth ( P  < 0.0051) and plaque scores and bleeding on probing values ( P  = 0.000) significantly decreased in each group when compared with the baseline, but there was no statistically significant difference between the test and control sites. GCF MMP-8 levels reduced presenting statistically significant differences on 7 days, 1, 3 and 6 months in four of the groups ( P  < 0.05); however, intergroup differences were not statistically significant. Developing functional and immunological-based chair-side MMP tests might serve as useful adjunctive diagnostic tools when monitoring the effects of DH gel application.     

血管内皮生长因子在正常与牙周炎牙龈组织中的表达差异研究

目的 检测血管内皮生长因子(VEGF)在正常与牙周炎的牙龈组织中的表达差异。方法 采用免疫组化PV两步法,检测VEGF在20例健康成人与20例慢性中重度牙周炎患者的牙龈组织中的表达差异。结果 健康组牙龈组织VEGF表达局限于上皮的颗粒层与部分棘层,其下方的结缔组织呈弱阳性表达,实验组牙龈组织上皮全层及结缔组织均呈强阳性表达,实验组VEGF表达高于健康组,两组之间的差异具有统计学意义(上皮区P<0.01,结缔组织区P<0.05)。结论 VEGF可能参与了牙周炎牙周袋壁龈组织的病理改变过程,在牙周炎发生发展及修复机制中可能具有重要意义。     

基质金属蛋白酶组织抑制因子-1基因多态性与慢性牙周炎相关性研究

目的:探讨基质金属蛋白酶组织抑制因子-1(TIMP-1)基因多态性与慢性牙周炎易感性的相关性.方法:从广东汉族人群中收集70例慢性牙周炎(CP)患者及74 例牙周健康对照者的颊黏膜拭子,以Chelex-100法提取DNA,采用聚合酶链反应-限制性片段长度多态方法对TIMP-1 +372T/C(rs4898)、TIMP-1 +533C/T (rs1062849)多态性进行检测, χ2检验分析组间等位基因分布和基因型频率的差异.结果:TIMP-1 +372位C、T等位基因在CP组和对照组的分布差异无统计学意义(P=0.897,OR=1.062).TIMP-1 +533C/T多态性未检出.结论:TIMP-1 +372T/C多态性与中国广东汉族人慢性牙周炎的患病易感性无关,且该人群中不存在TIMP-1 +533C/T多态性.     

Longitudinal analysis of metalloproteinases, tissue inhibitors of metalloproteinases and clinical parameters in gingival crevicular fluid from periodontitis-affected patients

BACKGROUND: The aim of this work was to improve the assessment of the periodontal disease status through measurements of extracellular matrix metalloproteinases (MMPs) and their tissular inhibitors (TIMPs) in the gingival crevicular fluid from patients diagnosed with chronic periodontitis. METHODS: Gingival crevicular fluid samples from patients (n = 13) were taken from 60 sites initially, and from 51 and 41 sites, respectively, 3 and 6 months after scaling and root planing. Gingival crevicular fluid samples were also taken from healthy subjects (n = 11, 24 sites). The presence of MMP-9 and MMP-8 was assessed by zymography and immunowestern blotting, respectively. The actual MMP activity (gelatinase and collagenase) was measured using the fluorogenic substrate assay. TIMP-1 and -2 levels were measured by immunodot blot. RESULTS: The fluorogenic substrate assay determinations showed higher MMP activity in sites with probing depth > or = 4 mm, with significant reduction post-treatment. Gelatinase activity followed by zymography consisted mainly of MMP-9. A different pattern of MMP-8 in control and patient sites was found. Controls only showed species of a partially active form (69 kDa), whereas patient sites showed a high frequency of the active form (56 kDa), and in some cases the latent form (85 kDa) was also observed. The active form reduced its frequency in sites with probing depth > or = 4 mm. TIMP-1 and -2 levels in patients were significantly lower than in controls, and after treatment the recovery of TIMP-1 level similar to control was observed. CONCLUSION: Significant correlations between the severity of the periodontal disease and the actual MMP activity, the active form of MMP-8 and the low level of both TIMP-1 and TIMP-2 were found.     

实验性牙根吸收组织中基质金属蛋白酶-1及其抑制剂-1的定位表达

目的检测小型猪实验性牙根吸收组织中基质金属蛋白酶(MMP)-1及基质金属蛋白酶抑制剂(TIMP)-1的表达,探讨二者在牙根吸收过程中的作用。方法选用6头小型猪的12颗下颌乳侧切牙,随机分为4组,每组3颗下颌乳侧切牙,分别加力0、0.98、1.96、2.94 N,每2周加力1次。第1次加力后45 d切取标本,应用SABC免疫组化方法检测MMP-1及TIMP-1在根吸收区牙周组织中的定位表达。结果加力0.98、1.96、2.94 N时牙根吸收区牙周组织中MMP-1阳性染色均比不加力强,加力0.98、1.96 N时牙根吸收区牙周组织中TIMP-1阳性染色均比不加力强。结论MMP-1与TIMP-1参与细胞外基质的代谢活动;MMP-1与TIMP-1在根吸收活动中起着重要的作用。     

慢性牙周炎患者龈沟液中白细胞介素-4的检测和意义

目的检测慢性牙周炎患者牙周基础治疗前后龈沟液中白细胞介素-4(IL-4)的质量浓度,探讨IL-4与牙周炎的关系及其在牙周炎发病机制、病情进展等方面所起的作用。方法用滤纸条浸润法采集成年健康者和牙周炎患者治疗前后的龈沟液样本,用酶联免疫吸附测定检测样本中IL-4的质量浓度。结果慢性牙周炎患者龈沟液中IL-4的质量浓度低于健康对照组(P<0.05)。经牙周基础治疗1个月后,IL-4的质量浓度无明显变化,治疗前后的差异无统计意义(P>0.05);IL-4的质量浓度与探诊深度呈显著负相关,与牙龈指数和附着丧失无明显相关性。结论IL-4缺乏可能会导致牙周病的发生,IL-4可作为早期诊断牙周病和检测易患人群的敏感性指标。