Plasminogen activators and plasminogen activator inhibitors in gingival crevicular fluid of cyclosporin A-treated patients

BACKGROUND: The plasminogen activator (PA) system plays many roles in the inflammatory process and tissue remodelling and repair and is considered to play a significant role in periodontal tissue destruction and healing. The aim of this study was to evaluate the role of the PA system in cyclosporin A (CsA)-induced gingival overgrowth in renal transplant patients. METHODS: Eighteen renal transplant patients exhibiting moderate to severe CsA-induced gingival overgrowth, 10 other renal transplant patients receiving CsA therapy but showing no sign of CsA-induced gingival overgrowth (CsA-H), 16 chronic gingivitis patients (CG) and 16 systemically and periodontally healthy control subjects (H) were included in the study. Gingival crevicular fluid (GCF) samples were obtained from four randomly selected sites in each subject with the exception of the CsA-induced gingival overgrowth group, where four GCF samples were harvested from sites with severe overgrowth (CsA GO+) and from four sites without any gingival overgrowth (CsA GO-). The GCF levels of albumin, tissue-type plasminogen activator (t-PA), urokinase-type plasminogen activator (u-PA), plasminogen activator inhibitor 1 (PAI-1) and plasminogen activator inhibitor 2 (PAI-2) were analysed by enzyme-linked immunosorbent assay. The results were tested for statistical differences. RESULTS: In CsA GO+ sites t-PA levels were significantly elevated in comparison with gingivitis and healthy sites, while PAI-2 levels in these sites showed statistically significant differences only with CsA-H and gingivitis sites (p<0.05). The levels of t-PA and PAI-2 were significantly higher in CsA GO- sites compared with those of CsA-H, gingivitis and healthy sites (p<0.05). The levels of u-PA and PAI-1 failed to show significant differences between the study groups. CONCLUSIONS: The findings of the present study indicate alterations in GCF t-PA and PAI-2 levels in CsA-induced gingival overgrowth and might suggest involvement of the plasminogen activating system in the pathogenesis of this side-effect of CsA therapy. However, to what extent these molecules contribute to the pathogenesis of CsA-induced gingival overgrowth remains to be determined.     

Evaluation of t-PA, PAI-2, IL-1beta and PGE(2) in gingival crevicular fluid of rheumatoid arthritis patients with periodontal disease

AIMS: This study was undertaken to compare periodontal conditions, gingival crevicular fluid (GCF) levels of tissue-type plasminogen activator (t-PA), its inhibitor plasminogen activator inhibitor-2 (PAI-2), interleukin-1beta (IL-1beta), prostaglandin E(2) (PGE(2)) in rheumatoid arthritis (RA) patients and control groups. METHODS: Twenty-three RA patients, 17 systemically healthy patients with periodontal disease (PD), and 17 systemically and periodontally healthy subjects were recruited. GCF samples were obtained from two single-rooted teeth. Full-mouth clinical periodontal measurements were recorded at six sites/tooth. GCF samples were analysed using relevant ELISA kits. Data were tested statistically by appropriate tests. RESULTS: Total amounts of t-PA, PAI-2 and PGE(2) in GCF samples of the healthy control group were significantly lower than the other groups (p<0.05). The RA group exhibited a higher total amount of t-PA in GCF samples than the PD group (p<0.05). PAI-2, IL-1beta and PGE(2) total amounts were similar in RA and PD groups (p>0.05). CONCLUSION: The coexistence of RA and periodontitis does not seem to affect clinical periodontal findings or systemic markers of RA. Similar inflammatory mediator levels in RA and PD groups, despite the long-term usage of corticosteroids, non-steroidal anti-inflammatory drugs, suggest that RA patients may have a propensity to overproduce these inflammatory mediators.     

Detection of tissue plasminogen activator (t-PA) and plasminogen activator inhibitor 2(PAI-2) in gingival crevicular fluid from healthy, gingivitis and periodontitis patients

Background: The regulation of plasminogen activation is a key element in controlling proteolytic events in the extracellular matrix. Our previous studies had demonstrated that in inflamed gingival tissues, tissue‐type plasminogen activator (t‐PA) is significantly increased in the extracellular matrix of the connective tissue and that interleukin 1β (IL‐1β) can up regulate the level of t‐PA and plasminogen activator inhibitor‐2 (PAI‐2) synthesis by human gingival fibroblasts. Method: In the present study, the levels of t‐PA and PAI‐2 in gingival crevicular fluid (GCF) were measured from healthy, gingivitis and periodontitis sites and compared before and after periodontal treatment. Crevicular fluid from106 periodontal sites in 33 patients were collected. 24 sites from 11 periodontitis patients received periodontal treatment after the first sample collection and post‐treatment samples were collected 14 days after treatment. All samples were analyzed by enzyme‐linked immunosorbent assay (ELISA) for t‐PA and PAI‐2. Results: The results showed that significantly high levels of t‐PA and PAI‐2 in GCF were found in the gingivitis and periodontitis sites. Periodontal treatment led to significant decreases of PAI‐2, but not t‐PA, after 14 days. A significant positive linear correlation was found between t‐PA and PAI‐2 in GCF (r=0.80, p<0.01). In the healthy group, different sites from within the same subject showed little variation of t‐PA and PAI‐2 in GCF. However, the gingivitis and periodontitis sites showed large variation. These results suggest a good correlation between t‐PA and PAI‐2 with the severity of periodontal conditions. Conclusion: This study indicates that t‐PA and PAI‐2 may play a significant rôle in the periodontal tissue destruction and tissue remodeling and that t‐PA and PAI‐2 in GCF may be used as clinical markers to evaluate the periodontal diseases and assess treatment.     

Immunohistochemical demonstration of the plasminogen activator system in human gingival tissues and gingival fibroblasts

The relative distribution of urokinase-type plasminogen activator (u-PA), tissue-type plasminogen activator (t-PA), plasminogen activator inhibitor-1 (PAI-1) and plasminogen activator inhibitor-2 (PAI-2) was studied in cultured human gingival fibroblasts, healthy gingival tissues and inflamed gingival tissues by immunohistochemistry. In cultured gingival fibroblasts t-PA, u-PA and PAI-1 were expressed in cytoplasm; u-PA and PAI-1 were more intensely stained than t-PA; PAI-2 was not detectable in gingival fibroblasts. Following interleukin 1β (IL-1β) stimulation, the intensity of intracellular staining for t-PA was increased and a number of cells staining strongly for PAI-2 were seen; no difference in the intensity of immunostaining level was noted for the expression of u-PA and PAI-1 between IL-1β stimulated cells and unstimulated cells. In healthy gingival tissues, u-PA and PAI-1 displayed a wide distribution throughout all the connective tissue and epithelium; t-PA localized mainly in the connective tissue while PAI-2 showed little association with the connective tissue but did faintly stain in the epithelial layer. In inflamed gingival tissues, staining for t-PA was significantly increased in the extracellular matrix of the connective tissue, whereas staining for u-PA, PAT-I and PAI-2 was found to be slightly increased, but no significant difference was noted for staining when compared with the healthy gingival tissues. A granular distribution of t-PA, u-PA, PAI-1 and PAI-2 was noted around areas of inflammatory cell infiltration. These immunohistochemical findings indicate that the plasminogen activator system produced by fibroblasts may be influenced by the presence of the inflammatory mediator IL-1β. In addition, the significant increase of t-PA in inflamed connective tissue and the wide expression of these components around inflamed cells may contribute to connective tissue degradation and may relate to the migration and localization of monocytes/macrophages in inflamed tissue.     

Aggravation of gingival inflammatory symptoms during pregnancy associated with the concentration of plasminogen activator inhibitor type 2 (PAI-2) in gingival fluid

Gingival inflammatory symptoms are aggravated during pregnancy. In vitro studies suggest a hormonal influence on the plasminogen activator inhibitor type 2 (PAI-2), and a disturbed balance of the fibrinolytic system could help to explain pregnancy gingivitis. Gingival crevicular fluid (GCF) was sampled in 14 women in pregnant and post-pregnant states. The gingival condition was assessed by the gingival index of Løe & Silness (GI) and the amount of bacterial plaque by the plaque index of Silness & Løe (PI). The ratio of sites with gingivitis to sites with bacterial plaque was calculated (G/P-ratio). Antigen levels of tissue plasminogen activator (t-PA), urokinase-type plasminogen activator (u-PA), plasminogen activator inhibitors type 1 (PAI-1) and PAI-2 in GCF were determined with ELISAs and 17 β-oestradiol and progesterone in serum with radioimmunoassays. For each individual the differences (Δ) in hormone levels and PAs and PAIs between pregnancy and post-pregnancy were calculated. Based on differences in G/P-ratio between pregnancy and post-pregnancy, subgrouping was done into a high-reacting and a low-reacting group. For the total group, the mean G/P-ratio was 2.0 during and 1.2 after pregnancy (p=0.064). A statistically significant correlation between Δprogesterone and ΔPAI-2 was noted: the higher Δprogesterone, the lower ΔPAI-2. No other significant correlations between hormone levels and components of the fibrinolytic system were found. For the total group of women, the concentrations of PAI-2, PAI-1 and t-PA were significantly higher during than after pregnancy. The individuals in the high-reacting group, however, showed a lower or unchanged production of PAI-2 during pregnancy, while those in the low-reacting group showed a greatly increased production. The lower inhibitory capacity in terms of a low production of PAI-2 during pregnancy in women with a higher inflammatory reaction indicates that the components of the fibrinolytic system may be involved in the development of pregnancy gingivitis and implies that PAI-2 serves as an inhibitor of importance for tissue proteolysis. The present finding contributes to the explanation of pregnancy gingivitis.     

The plasminogen-activating system in gingival fluid from adults

High concentrations of tissue plasminogen activator (t-PA) and placental type plasminogen activator inhibitor (PAI-2) have previously been found in gingival crevicular fluid (GCF) of adults and children. In the present study, intra-individual comparisons were made of the concentrations of t-PA, urokinase type plasminogen activator (u-PA), PAI-1, and PAI-2 in GCF from the same sites before and after periodontal treatment in eight healthy male volunteers aged 35–46 yr. The gingival state was assessed by exudate measurement, bleeding on standardized probing, and the gingival index of Löe & Silness 3 days before the start of the trial and on the day after completing a 21-day preventive program consisting of instruction and professional cleaning once a week. Eight sites per subject were selected for enzyme analyses, all showing improvement in gingival state during the period. Sampling of GCF at the start and at the end of the trial was done with small disks of Millipore-filter. t-PA and PAI-2 were analyzed with enzyme-linked immunosorbent assays with low method errors. The mean concentrations of t-PA were 0.73 mg/1 before treatment and 0.49 mg/1 after treatment. The mean concentrations of u-PA were 84.4 ug/1 before treatment and 101.6 μg/1 after treatment. PAI-1 was found in three subjects at the detection level. The mean PAI-2 concentrations were 2.19 mg/1 before and 1.13 mg/1 after treatment. The mean molar ratio PAs/PAI-2 was 0.47 before and 0.48 after treatment. This insignificant change implies a maintained proteolytic balance and indicates that PAI-2 is an important inhibitor of tissue proteolysis.     

Gingival crevicular fluid EMAP-II, MIP-1alpha and MIP-1beta levels of patients with periodontal disease

BACKGROUND: Periodontal diseases may differ, which could be attributed to the factors that might modify the host response to microbial pathogens. The aim of this study was to examine gingival crevicular fluid (GCF) levels of EMAP-II, MIP-1alpha and MIP-1beta in patients with different periodontal diseases (EMAP-II, endothelial-monocyte activating polypeptide; MIP-1alpha, macrophage inflammatory protein-1alpha; MIP-1beta, macrophage inflammatory protein-1beta). METHODS: Eighty-two subjects were included in this study. GCF samples were collected from 26 patients with generalized aggressive periodontitis (G-AgP), 26 patients with chronic periodontitis (CP), 15 with gingivitis and 15 periodontally healthy subjects. Clinical periodontal parameters were recorded. GCF EMAP-II, MIP-1alpha and MIP-1beta levels were quantified by enzyme immunoassay. RESULTS: GCF EMAP-II levels of G-AgP group were higher than those of gingivitis and healthy groups (p<0.008). G-AgP group showed a trend for higher GCF EMAP-II levels compared with CP group (p>0.008). G-AgP, CP, gingivitis and healthy groups had comparable GCF MIP-1alpha and MIP-1beta levels. CONCLUSIONS: Our results suggest that elevated GCF EMAP-II could contribute to the pathogenesis of G-AgP. Alternatively, EMAP-II reflects the extent of the inflammatory activity in the periodontal tissues. At this point, MIP-1alpha and MIP-1beta levels in GCF do not seem to play a discriminatory role in periodontitis. Our data document for the first time the essential role of EMAP-II in the pathogenesis of different periodontal diseases.     

Gingival crevicular fluid transforming growth factor-beta1 in several forms of periodontal disease

BACKGROUND: Transforming growth factor-beta(1) (TGF-beta(1)) has significant effects on periodontal host response regulation. Limited knowledge on the role of TGF-beta(1) in various periodontal disease types and particularly in advanced periodontitis forms warranted the present study. The aim of the present study was to evaluate the gingival crevicular fluid (GCF) TGF-beta(1) levels in patients with different forms of periodontal disease. METHODS: GCF TGF-beta(1) levels were investigated in 32 chronic periodontitis (CP), 30 generalized aggressive periodontitis (G-AgP), 15 gingivitis patients and 16 periodontally healthy subjects. Periodontal status was evaluated by measuring probing depth, clinical attachment loss, plaque and bleeding on probing. TGF-beta(1) levels were analyzed by enzyme-linked immunosorbent assay. The results were expressed in terms of total amount (pg) and concentration (pg/microl). RESULTS: G-AgP and CP groups had significantly elevated GCF TGF-beta(1) total amount compared to healthy group (p<0.008). Moreover, GCF TGF-beta(1) total amount of G-AgP group was significantly higher than that of gingivitis group (p<0.008). G-AgP and CP groups had similar GCF TGF-beta(1) total amount (p>0.008). Significant correlation was found between GCF TGF-beta(1) total amount and all clinical periodontal parameters (p<0.05). CONCLUSIONS: The results of the present study suggest contribution of TGF-beta(1) to the pathogenesis of advanced chronic and aggressive periodontitis. TGF-beta(1) may thus be one of the components modulating exaggerated host response together with other major mediators of inflammation.     

Gingival crevicular fluid laminin-5 gamma2-chain levels in periodontal disease

AIM: Our study aimed to examine the molecular forms and gingival crevicular fluid (GCF) levels of laminin-5 gamma2-chain in patients with different periodontal disease, and compare the effects of P.gingivalis trypsin-like proteinase on intact laminin-5 gamma2-chain species. METHODS: Eighteen patients with generalized aggressive periodontitis (G-AgP), 29 patients with chronic periodontitis (CP), 20 with gingivitis and 20 periodontally healthy subjects were included. Probing depth, clinical attachment loss, presence of bleeding on probing and plaque were recorded. Molecular forms and GCF laminin-5 gamma2-chain levels and the effects of P. gingivalis trypsin-like proteinase on intact laminin-5 gamma2-chain were analysed by computer-quantitated Western immunoblotting. RESULTS: Laminin-5 gamma2-chain 40 and 70 kDa fragments could be detected in all groups, in varying levels.The CP group had elevated GCF laminin-5 gamma2-chain fragment levels compared with the gingivitis and healthy groups (p<0.008).The G-AgP group had GCF laminin-5 gamma2-chain fragment levels similar to the gingivitis and healthy groups (p>0.008). GCF laminin-5 gamma2-chain fragments differed clearly from the multiple lower molecular size fragments of P.gingivalis trypsin-laminin-5 gamma2-chain proteinases. CONCLUSION: Increased GCF laminin-5 gamma2-chain fragments in periodontitis sites with deep periodontal pocket suggest that these cleaved 40 and 70 kDa fragments could reflect the extent of the inflammatory reaction in CP.     

Effect of Initial Periodontal Therapy on Oxidative Stress Markers in Gingival Crevicular Fluid,Saliva, and Serum in Smokers and Non‐Smokers With Chronic Periodontitis

Background: The aim of this case‐control study with an intervention arm is to determine the effect of initial periodontal treatment on oxidative stress biomarkers in smokers and non‐smokers with chronic periodontitis (CP). Methods: The study included 47 patients with CP (24 smokers [S+P+] and 23 non‐smokers [S?P+]) and 46 periodontally healthy individuals (23 smokers [S+P?] and 23 non‐smokers [S?P?]) for a total of 93 participants. Gingival crevicular fluid (GCF), serum, and saliva samples were obtained and clinical periodontal measurements were recorded at baseline and at the first and third months after periodontal therapy. 8‐hydroxydeoxyguanosine (OHdG) and 4‐hydroxynonenal (HNE) and enzyme activity of glutathione peroxidase (GSH‐Px) were analyzed with enzyme‐linked immunosorbent assay. Results: The level of 8‐OHdD in GCF was found to be significantly higher in both periodontitis groups compared with both periodontally healthy groups. 8‐OHdG and GSH‐Px in saliva in both periodontitis groups were significantly increased compared with the S?P? group. In the S+P+ group, 4‐HNE in GCF was found to be significantly higher than in periodontally healthy participants. After initial periodontal treatment, the levels of 8‐OHdG in GCF and saliva were significantly decreased in both periodontitis groups. Conclusion: Initial periodontal therapy may be helpful for diminishing oxidative stress in periodontitis.     

慢性牙龈炎及慢性牙周炎治疗前后患牙龈沟液中t-PA及PAI活性的比较

目的 :比较慢性牙龈炎、慢性牙周炎患牙治疗前后龈沟液中t PA及PAI活性。方法 :选择慢性龈炎(n =2 8)及慢性牙周炎 (n =5 2 )患牙两组 ,在治疗前及治疗后收集龈沟液 (GCF)并记录临床指标 ,用发色底物法测定GCF中t PA及PAI的活性。结果 :t PA活性在牙龈炎组治疗前后差异无显著性 ,在牙周炎组治疗前后差异有显著性 ;PAI活性在牙龈炎组及牙周炎组治疗前后差异均有显著性 ;PAI/t PA比值在牙周炎组治疗前后差异有显著性 ,牙龈炎组治疗前后差异无显著性。结论 :PAI可以作为判断牙龈炎症程度及治疗效果较为客观的指标 ;牙周炎组中治疗后PAI/t PA显著降低的部位是否预示较差的预后 ,尚需进一步纵向研究。     

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.     

Calcitonin gene-related peptide in gingival crevicular fluid in periodontal health and disease

The aims of the present study were to investigate whether calcitonin gene-related peptide (CGRP) was present in gingival crevicular fluid in both periodontal health and disease and to study the relationship with periodontal inflammation. Gingival crevicular fluid (GCF) was collected from a healthy, a gingivitis and a periodontitis site in 18 subjects with periodontitis and from a healthy site in 19 subjects without periodontitis. The volume of GCF was measured and each sample subsequently analysed for CGRP by radioimmunoassay. In subjects with periodontitis, CGRP immunoreactivity (CGRP-IR) was not detected in any periodontitis sites, nor in 67% of gingivitis and 28% of periodontally-healthy sites. The total amount of CGRP-IR was significantly elevated in periodontally healthy (p=0.0015) and gingivitis (p=0.027) compared with periodontitis sites. CGRP-IR was present in 89% of the healthy sites sampled in control subjects at comparable levels to those in healthy sites in periodontitis subjects. It is concluded that in periodontal inflammation, particularly in deep pockets, constituents of GCF process and degrade CGRP.     

正常人、牙龈炎和牙周炎患者龈沟液内一氧化氮含量的检测

目的：检测正常人和牙周病患者龈沟液中NO含量,探讨NO在牙周病发病过程中的作用。方法：选择牙周健康组20例,牙龈炎组22例,慢性牙周炎组32例,分别采集龈沟液标本,免疫荧光法检测龈沟液内NO的含量。结果：慢性牙周炎患者和牙龈炎患者龈沟液内NO含量与牙周健康组相比均有高度显著性差异（P〈0．01）,慢性牙周炎患者龈沟液内NO含量与牙龈炎组相比有高度显著性差异（P〈0．01）。结论：牙周健康者、牙龈炎患者、慢性牙周炎患者龈沟液中能检测出NO的存在,NO参与了慢性牙周炎的发展过程,龈沟液内NO含量与慢性牙周炎炎症程度密切相关。     

不同牙周状态下龈沟液中瘦素水平的比较

目的:检测牙周健康者、慢性龈炎和慢性牙周炎患牙龈沟液中瘦素水平,为牙周炎的早期诊断提供客观指标。方法:选择牙周健康者、慢性龈炎和慢性牙周炎患者3组,治疗前记录牙周各项临床指标,并用Whatman^ 1号滤纸收集颊侧近远中牙周袋内GCF采用ELISA检测瘦素含量;同时收集血清进行瘦素水平的检测。结果:3组中慢性牙周炎组龈沟液中瘦素水平与牙周健康者龈沟液中瘦素水平比较有显著差异(P<0.05),慢性牙周炎组龈沟液中瘦素水平与慢性龈炎龈沟液中瘦素水平及慢性龈炎组龈沟液中瘦素水平与牙周健康者龈沟液中瘦素水平比较均无显著差异;牙周健康者、慢性龈炎和慢性牙周炎患者血液中瘦素水平比较均有显著差异(P<0.05)。血中的瘦素与临床指标PLI (P<0.01,r=0.593)、PD(P<0.05,r=0.920)、BI(P<0.05,r=0.862)、AL(P<0.05,r=0.846)均相关;龈沟液中的瘦素与临床指标PLI(P<0.01,r=0.813)、PD(P<0.05,r=0.962))、BI(P<0.05,r=0.720、AL(P<0.05,r=0.946)均相关。结论:龈沟液中瘦素水平可能是反映牙周组织状况的一项较为客观的指标。     

Tissue plasminogen activator (t-PA) and placental plasminogen activator inhibitor (PAI-2) in gingival fluid from 8–9-year-old children

High concentrations of tissue plasminogen activator (t-PA) and placental type plasminogen activator inhibitor (PAI-2) have previously been found in gingival crevicular fluid (GCF) of adults. In the present study, the levels were examined in 16 children aged 8–9 yr. Sampling of GCF was performed with small disks of Millipore-filter. t-PA and PAI-2 were analyzed with enzyme-linked immunosorbent assays with low method errors. The mean concentration of t-PA was slightly higher than in adults, while the mean PAI-2-concentration was slightly lower. An intraindividual study comparing healthy and inflamed sites in the children showed slightly higher concentrations in GCF from inflamed sites. No change was observed in the balance between t-PA and PAI-2.     

Gingival crevicular fluid matrix metalloproteinase-25 and -26 levels in periodontal disease

BACKGROUND: Tissue destruction associated with the progression of periodontal disease is caused by a cascade of host and microbial proteolytic enzymes. Host-derived matrix metalloproteinases (MMPs) play an important role in the degradation of the extracellular matrix. Leukolysin/membrane-type 6 (MT-6)/MMP-25, the latest member of the MT-MMP subgroup of the MMP family, is primarily expressed by neutrophils and involved in extracellular matrix turnover. Matrilysin-2/MMP-26 (endometase), a novel member of the matrilysin subgroup of the MMP family, can degrade the extracellular matrix, alpha1-antitrypsin, and activate pro-MMP-9. Our study aimed to examine the levels, molecular forms, and degrees of activation of MMP-25 and MMP-26 in gingival crevicular fluid (GCF) from patients with different periodontal diseases. METHODS: A total of 105 subjects, 35 with generalized aggressive periodontitis (GAgP), 29 with chronic periodontitis (CP), 20 with gingivitis, and 21 periodontally healthy subjects, were included in this study. Periodontal status was evaluated by measuring probing depth, clinical attachment loss, presence of bleeding on probing, and plaque. GCF MMP-25 and MMP-26 levels were analyzed by computer-quantitated Western immunoblotting using specific antibodies. RESULTS: The 57-kDa soluble pro-MMP-25 and 45- to 47-kDa active form of MMP-25 were detected by Western immunoblots in CP and GAgP GCF, and lesser levels of these soluble MMP-25 immunoreactive bands were detected in gingivitis GCF. An enhanced and similar degree of MMP-25 activation was found in GAgP, CP, and gingivitis groups. There were no detectable MMP-25 immunoreactivities in the healthy subjects' GCF. GAgP and CP groups had elevated GCF MMP-26 levels and degrees of activation compared to the gingivitis and healthy groups (P <0.008). The gingivitis group had higher GCF MMP-26 levels and degree of activation compared to the healthy group (P <0.008). CONCLUSIONS: The present study demonstrated the presence of soluble or shed forms of MMP-25 and MMP-26 in GCF of patients with different periodontal diseases. Increased levels and activation of MMP-25 and MMP-26 in GCF are associated with an enhanced severity of periodontal inflammation, suggesting that these novel MMPs can participate in the progression of periodontal diseases. They may prove to be diagnostically useful and could be targets of medication in the future.     

Granulocyte elastase activity in static and flow gingival crevicular fluid

OBJECTIVES: This study aimed to evaluate the volume of gingival crevicular fluid (GCF) and granulocyte elastase activity in static GCF (sGCF) and flow GCF (fGCF) from subjects with various periodontal conditions. METHODS: Eleven periodontally healthy, 10 gingivitis and 12 periodontitis subjects were recruited and the sites investigated consisted of healthy sites from healthy subjects (HH); healthy (HG) and gingivitis sites (GG) from gingivitis subjects; and healthy (HP), gingivitis (GP) and periodontitis sites (PP) from periodontitis subjects. fGCF samples were collected either 1 min or 5 min following sGCF collection by paper strip technique. GCF volume was determined by Periotron 6000 and granulocyte elastase activity was assayed with a specific substrate [l-pyroglutamyl-l-prolyl-l-valine-p-nitroanilide(pGluProVal-pNA)]. RESULTS: At baseline, no significant differences existed in clinical and GCF parameters between the two matched sites for subsequent collection of fGCF samples either 1 min or 5 min after sGCF sampling in all subjects. The flow exudate in HG and HP sites quickly replenished to sGCF levels, while a delayed replenishment was found in HH sites, despite the similar sGCF volumes of these sites. The GCF volume and elastase levels in the fGCF at 1 min were higher in GP sites than in GG sites (P < 0.05). Overall, depletion of elastase levels in the fGCF at 1 min was observed in all subjects, whereas elastase levels in the fGCF at 5 min had replenished to sGCF levels in HP, GP, PP sites and GG sites, but had remained at a lower level in HH and HG sites. An overall positive correlation was found between sGCF and fGCF for GCF volume and elastase activity (P < 0.001); however, this correlation varied with GCF parameters and with site conditions of the subjects concerned. CONCLUSIONS: This study shows that patterns of dynamic changes in GCF flow and elastase activity varied under different periodontal conditions. Assessment of both sGCF and fGCF may allow better insight into the dynamic change of the target components in GCF.     

Microbial factors and gingival crevicular fluid aspartate aminotransferase levels

Abstract. The aim of this cross-sectional study was to investigate the clinical application of chairside tests for gingival crevicular fluid (GCF) aspartale amino-transferase (AST) levels and plaque BANA hydrolysis activity with the presence of the periodontal pathogens Porphyromonas gingivalis and Actinohacillus action-mycetemcomitans. The study comprised 100 periodontitis sites (pocket depths≥4 mm. GI = 3) from 10 patients with chronic adult periodontitis and 100 control sites (pocket depths <4 mm. GI<3) from 10 periodontally healthy patients comprising 55 healthy sites (pocket depths <4 mm. GI=0) and 45 gingivitis sites (pocket depths <4 mm, GI=1 or 2). The values for both BANA hydrolysis and AST levels were significantly higher in samples from periodontitis compared with gingivitis and healthy sites (p<0.001), A. actinomycetemcomitans was identified in 45% and P. gingivalis in 17% of periodontitis sites but neither pathogen was recovered from control sites and there was no significant correlation with (he clinical parameters measured. There was no significant relationship between the presence of P. gingivalis and/or A. actinmycetemcomitans with BANA hydrolysis or AST levels. A significant correlation (p=0.0017) was observed between BANA hydrolysis and pocket depth and between AST hydrolysis and the GI (p=0.01). This study failed to demonstrate a positive association between chairside analysis of GCF metabolites for AST levels and/or BANA hydrolysis with P. gingivalis and A. actinomycetemcomitans. However, the GCF metabolites had a significant correlation with periodontally diseased sites in patients with chronic adult periodontitis and may help confirm clinical observations.     

Crevicular and serum levels of monocyte chemoattractant protein-4 and high-sensitivity C-reactive protein in periodontal health and disease

ObjectivesChemokines are chemotactic cytokines that are involved in destruction of the periodontal structures. The aim of this study is to determine the presence of MCP-4 and high sensitivity C reactive protein (hsCRP) levels in gingival crevicular fluid (GCF) and serum in periodontal health and disease and to find a correlation between MCP-4 and hsCRP in GCF and serum.Methods40 subjects (20 males and 20 females) were selected and divided into three groups based on clinical parameters and radiologic parameters: Group 1 (10 healthy); Group 2 (15 gingivitis subjects) and Group 3 (15 chronic periodontitis subjects). The levels of serum and GCF MCP-4 were determined by ELISA and hsCRP levels were determined by immunoturbidimetry method.ResultsThe mean GCF and serum concentration of MCP-4 were the highest for group 3 followed by group 2 and least in group 1. Similarly, the mean hsCRP concentrations were highest for group 3 and least in group 1. Moreover, a significant positive correlation was found between serum and GCF MCP-4 and hsCRP levels and periodontal parameters.ConclusionThe levels of MCP-4 and hsCRP increased from healthy to periodontitis. It can be proposed that MCP-4 and hsCRP are the potential biomarkers of inflammation in periodontal health and disease.