Clinical usability of aspartate aminotransferase to evaluate the prognosis of periodontal regeneration therapies: prospective, longitudinal study.

To evaluate the degree of periodontal tissue destruction, aspartate aminotransferase (AST) levels in the gingival crevicular fluid (GCF) are utilized as a predictor of periodontal therapy. We have previously shown that the usefulness of AST activities [periodontal tissue monitor (PTM) values] using a PTM-kit to evaluate the effects of initial periodontal therapy and periodontal regeneration therapy by enamel matrix derivative (EMD). This prospective, longitudinal study was conducted using 38 healthy and 80 periodontitis sites with probing depth (PD) of 5-10 mm for guided tissue regeneration (GTR) and EMD from 36 patients. GCF samples were used to evaluate PTM values at base line (BL) and after 6 months of surgeries (re-evaluation: RE), and periodontal examinations were performed concurrently. PTM values at BL were statistically improved at RE, accompanied by the improvement of periodontal parameters in both groups. PTM values and PD, and the clinical attachment level (CAL) showed high correlations. PD, CAL and bleeding on probing (BOP) were highly correlated with PTM values in both groups, whereas only PD showed a significant correlation with PTM values at RE in the GTR group. Change in the amounts of PD, CAL and BOP between BL and RE in both groups showed no correlation with PTM values. In the negative PTM value sites at BL in EMD group, the mean PD was significantly reduced at RE compared with positive PTM sites at BL. PTM values are able to be utilized as the biochemical predictor of prognosis after periodontal regeneration therapy.

IPLA2 mRNA expression by human neutrophils in type 2 diabetes and chronic periodontitis.

Type 2 diabetes mellitus (T2D) is becoming increasingly prevalent worldwide and complications of T2D cause significant systemic and dental morbidity in the susceptible individual. Although T2D has been linked as a significant risk factor for chronic periodontitis (CP), molecular mechanisms explaining the pathogenesis and inflammatory impact of CP in T2D are lacking. iPLA2 is the calcium-independent form of phospholipase A2. In previous studies, we demonstrated that iPLA2 enzyme activity is altered in T2D. The purpose of this study was to elucidate the level of the iPLA2 abnormality in T2D by measuring messenger RNA levels in T2D-associated CP. A total of 53 healthy and T2D subjects with CP were recruited for this study. The clinical periodontal exam included probing pocket depth, clinical attachment levels and bleeding on probing. Peripheral venous blood was collected and neutrophils were isolated. Real time polymerase chain reaction was used to quantify iPLA2 mRNA in neutrophils from healthy controls and people with diabetes. Results revealed that the prevalence of moderate to severe CP was increased in people with T2D. The iPLA, mRNA levels in diabetics with different severity of CP were not significantly different compared to healthy controls; 1.07 vs 0.97 (mild CP), 1.07 vs 0.85 (moderate CP) and 1.07 vs 1.05 (severe CP). Collectively, the data suggest that levels of iPLA2 mRNA in T2D are not different than in health and are not directly influenced by periodontal disease status. The impact of inflammation on iPLA2 regulation is at the level of activation of the enzyme rather than expression at the mRNA level.

Differential expression of mitogen activating protein kinases in periodontitis.

AIM: Following toll-like receptor (TLR) engagement, lipopolysaccharide (LPS) can stimulate the expression of pro-inflammatory cytokines thus activating the innate immune response. The production of inflammatory cytokines results, in part, from the activation of kinase-induced signalling cascades and transcriptional factors. Of the four distinct classes of mitogen-activated protein kinases (MAPK) described in mammals, p38, c-Jun N-terminal activated kinases (JNK1-3) and extracellular activated kinases (ERK1,2) are the best studied. Previous data have established that p38 MAPK signalling is required for inflammation and bone loss in periodontal disease pre-clinical animal models. MATERIALS & METHODS: In this study, we obtained healthy and diseased periodontal tissues along with clinical parameters and microbiological parameters. Excised fixed tissues were immunostained with total and phospho-specific antibodies against p38, JNK and ERK kinases. RESULTS: Intensity scoring from immunostained tissues was correlated with clinical periodontal parameters. Rank correlations with clinical indices were statistically significantly positive (p-value < 0.05) for total p38 (correlations ranging 0.49-0.68), phospho-p38 (range 0.44-0.56), and total ERK (range 0.52-0.59) levels, and correlations with JNK levels also supported association (range 0.42-0.59). Phospho-JNK and phospho-ERK showed no significant positive correlation with clinical parameters of disease. CONCLUSION: These data strongly implicate p38 MAPK as a major MAPK involved in human periodontal inflammation and severity.

Treponema denticola associates with increased levels of MMP-8 and MMP-9 in gingival crevicular fluid.

OBJECTIVES: The aim was to assess the association between the presence of site-specific subgingival micro-organisms and the levels of matrix metalloproteinases-8 and matrix metalloproteinases-9 (MMP-8 and MMP-9) in gingival crevicular fluid (GCF). MATERIALS AND METHODS: The patient group consisted of 56 subjects with periodontitis and the control group of 43 subjects without periodontitis. GCF samples from four test sites for each subject were collected. Polymerase chain reaction was used to detect the presence of Aggregatibacter actinomycetemcomitans, Porphyromonas gingivalis, Prevotella intermedia, Tannerella forsythia, and Treponema denticola. MMP-8 concentrations were analyzed by a time-resolved immunofluorometric assay, and MMP-9 levels were determined by enzyme-linked immunosorbent assay. Student's unpaired t-test, chi-square test, and Fisher's exact P-value were calculated. RESULTS: The presence of T. denticola in the test sites was significantly higher in the patient group than in the control group. The presence of T. forsythia and T. denticola was associated with increased levels of MMP-8 in the test sites. Respectively, site-specific presence of T. denticola was associated with an increase in MMP-9 levels in three of the four test sites. CONCLUSIONS: The presence of subgingival micro-organisms in GCF, particularly T. denticola, appeared to induce a host response with an increased release of MMP-8 and MMP-9 in the test sites.

Protease inhibitor levels in periodontal health and disease.

BACKGROUND AND OBJECTIVE: Our previous study showed that protease inhibitors were attenuated by the periodontal pathogen Porphyromonas gingivalis in cultured gingival epithelial cells. We hypothesize that fewer protease inhibitors would be present in more advanced periodontal disease sites, where the level of P. gingivalis may be high. The goal of this study was to investigate the relationship between the protease inhibitor [secretory leukocyte protease inhibitor (SLPI), elastase-specific inhibitor (ELAFIN) and squamous cell carcinoma antigen (SCCA)] levels in gingival crevicular fluid and the number of P. gingivalis micro-organisms in subgingival plaque. MATERIAL AND METHODS: Plaque samples from subjects without (n = 18) and with moderate to advanced periodontitis (n = 41) were used to quantify P. gingivalis using real-time PCR. Protease inhibitor levels in the gingival crevicular fluid of all the subjects were determined by ELISA. RESULTS: P. gingivalis was detected in 68.3% of patients with periodontitis, while 16.7% of subjects without periodontitis had a detectable level of P. gingivalis. Patients with periodontitis and P. gingivalis in their plaque exhibited lower SLPI and ELAFIN levels (p < 0.001) compared with control subjects without periodontitis. Secretory leukocyte protease inhibitor was also reduced (p < 0.05) in gingival crevicular fluid of periodontitis patients without a detectable level of P. gingivalis. Periodontitis patients with high vs. low levels of P. gingivalis exhibited reciprocal mean levels of SLPI and ELAFIN concentrations. CONCLUSION: The reduced concentrations of SLPI and ELAFIN may contribute to the loss of host protective capacity and increase susceptibility to breakdown from chronic infection. The work of this investigation may aid in finding diagnostic and prognostic markers in periodontal health and disease and may also help in finding pharmacological targets directed against periodontal inflammation.

MMP3 and TIMP1 variants contribute to chronic periodontitis and may be implicated in disease progression.

AIM: Matrix metalloproteinases (MMPs) play a key role in the tissue destruction characteristic of chronic periodontitis. The purpose of this study was to investigate the association of MMP and TIMP polymorphisms with chronic periodontitis in two populations. MATERIAL AND METHODS: A total of 34 polymorphisms spanning 12 MMP and 2 TIMP genes were genotyped in 401 individuals from Brazil (99 cases with chronic periodontitis and 302 controls), and 274 individuals from the US (70 cases and 204 controls). Individuals were considered cases if presenting at least three teeth exhibiting sites of clinical attachment loss ≥ 5 mm in two different quadrants. Controls were characterized by absence of clinical attachment loss and no sites with probing depth >3 mm. MMP3 and TIMP1 mRNA expression was evaluated in healthy and diseased periodontal tissues. RESULTS: TIMP1 showed association with chronic periodontitis in the Brazilian population (for rs5906435, p = 0.0004), whereas MMP3 showed association in the US population (for rs679620, p = 0.0003; and rs650108, p = 0.002) and in the Brazilian population (for rs639752, p = 0.005). MMP3 and TIMP1 mRNA expression was significantly higher in diseased tissues when compared to control tissues. CONCLUSIONS: Our results further support a role for variations in MMP3 in chronic periodontitis and report a novel association with TIMP1. These genes may be considered additional candidate genes for chronic periodontitis.

Longitudinal study of salivary proteinases during pregnancy and postpartum.

BACKGROUND AND OBJECTIVE: Matrix metalloproteinases (MMPs) and their regulators are connected to periodontal inflammation and destruction. However, the presence and role of the salivary MMPs in pregnancy-related gingivitis are not well known. Our longitudinal study aimed to monitor salivary proteinase levels and possible changes, and relate them to periodontal status during pregnancy and postpartum. MATERIAL AND METHODS: Salivary samples were collected from 30 periodontally healthy pregnant women five times (once during each trimester, 4-6 wk after delivery and after lactation) and, as their controls, from 24 non-pregnant women three times (during successive months). Periodontal examination included visible plaque index, bleeding on probing, probing pocket depth and clinical attachment level measurements. Matrix metalloproteinase-8 levels were measured by immunofluorometric assay, and MMP-2 and MMP-9 levels and molecular forms by gelatin zymography. Salivary elastase, myeloperoxidase and tissue inhibitor of matrix metalloproteinase-1 levels were measured by ELISA. RESULTS: Elastase concentrations maintained stable during the follow-up, while myeloperoxidase concentrations increased significantly after delivery. During pregnancy, MMP-8 concentrations were significantly lower than postpartum concentrations, being lowest during the second trimester and highest after delivery, and varying inversely to pregnancy gingivitis, observed as elevated percentages of bleeding on probing and probing pocket depth during the second and third trimester. In pregnant women, the highest MMP-2 and MMP-9 levels were found in saliva after lactation. In the control group, both clinical and enzymological findings remained stable during the follow-up period. CONCLUSION: Our results suggest that hormonal changes during pregnancy induce or enhance susceptibility to gingivitis, while salivary proteinase and myeloperoxidase levels are reduced.

Crevicular fluid matrix metalloproteinase-8, -13, and TIMP-1 levels in type 2 diabetics.

OBJECTIVES: To evaluate whether type 2 diabetes mellitus (DM) enlarged and if so the quantum of such increase in the gingival crevicular fluid (GCF) levels of matrix metalloproteinase-8 (MMP-8), MMP-13 and tissue inhibitor of metalloproteinases-1 (TIMP-1). METHODS: Subjects (n = 73) were divided into five groups as follows: 12 DM patients with gingivitis (DM-G), 12 DM patients with periodontitis (DM-P), 12 systemically healthy patients with gingivitis (H-G), 13 systemically healthy patients with periodontitis (H-P) and 24 periodontally, systemically healthy volunteer subjects (H-C). Full-mouth clinical periodontal measurements were performed at six sites per tooth. Gingival crevicular fluid samples were obtained from two sites representing the clinical periodontal diagnosis in single-rooted teeth. Gingival crevicular fluid levels of MMP-8, MMP-13 and TIMP-1 were analysed by immunofluorometric MMP assay (IFMA), enzyme-linked immunosorbent assay (ELISA). Data were tested statistically by parametric tests. RESULTS: All clinical periodontal measurements were similar in both diabetic and systemically healthy patients with periodontal disease (all P > 0.05). Total amounts of MMP-8 in GCF samples were significantly lower in H-C group than DM-G, DM-P, H-P groups (all P < 0.05). Matrix metalloproteinase-13, TIMP-1 total amounts were similar in study groups (P > 0.05). Diabetes mellitus patients exhibited similar levels of MMP-8, MMP-13, TIMP-1 with systemically healthy gingivitis/periodontitis patients (P > 0.05). CONCLUSIONS: Within the limits of this study, DM does not seem to significantly affect GCF levels of MMP-8, MMP-13, TIMP-1 or clinical periodontal status.

Role of systemic and local administration of selective inhibitors of cyclo-oxygenase 1 and 2 in an experimental model of periodontal disease in rats.

BACKGROUND AND OBJECTIVE: Periodontal disease is an inflammatory condition of tooth-supporting tissues. Arachidonic acid metabolites have been implicated in development of periodontal disease, especially those derived from the cyclo-oxygenase (COX) pathway. This study investigated the role of inhibitors of cyclo-oxygenases (COX-1 and COX-2) in a model of periodontal disease in rats. MATERIAL AND METHODS: A ligature was placed around the molar of rats. Losses of fiber attachment and of alveolar bone were measured morphometrically in histologically prepared sections. Infiltration of cells into gingival tissue surrounding the ligated tooth was also determined. RESULTS: Systemic and local administration of non-selective and selective COX-2 inhibitors, preventively, resulted in significant reduction of the losses of fiber attachment and alveolar bone, as well as decreased leukocyte numbers in gingival tissue. Preventive selective inhibition of COX-1 was as effective as COX-2 inhibition in reducing local fiber attachment loss and cell migration, but did not prevent alveolar bone loss. CONCLUSION: Our results provide evidence for participation of COX-1 and COX-2 in early stages of periodontal disease in rats. Furthermore, local administration of COX inhibitors reduced the signs of periodontal disease to the same extent as systemic treatment. Therapeutic approaches incorporating locally delivered anti-inflammatory drugs could be of benefit for patients suffering from periodontal disease.

The defensive role of lysozyme in human gingiva in inflammatory periodontal disease.

BACKGROUND AND OBJECTIVE: The presence of lysozyme in human gingiva has not previously been demonstrated. In this study, we looked for evidence for the potential role of lysozyme as a protector of gingival elastic fibres. The objective of this study was also to determine the ex vivo susceptibility to hydrolysis of gingival elastic fibres from patients with or without periodontal disease by human leukocyte elastase and by human cathepsin G. MATERIALS AND METHODS: Using gingival tissue sections from eight control, 10 gingivitis and 10 periodontitis patients, we evaluated the area fraction occupied by gingival elastic fibres (after selective staining) by the use of automated image analysis. In the ex vivo experiments, serial tissue sections from four control, four gingivitis, four young periodontitis and four aged periodontitis patients were submitted to the action of human leukocyte elastase and cathepsin G, after which enzymatic activities were determined by image analysis. Indirect immunodetection of lysozyme was also done on tissue sections for all patients included in this study. RESULTS: Large variations of the area fraction occupied by elastic fibres were observed in human gingiva from young and aged patients with and without periodontal disease. In control and gingivitis patients, leukocyte elastase and cathepsin G had high comparable elastin solubilizing activities. With young and aged periodontitis patients, the two serine proteinases had weak elastin solubilizing activities. Lysozyme appeared to be present at the periphery of gingival elastic fibres in periodontitis patients. CONCLUSION: Lysozyme can be considered an important natural protector of elastic fibres in pathological gingiva.

MMP-13 promoter polymorphisms in patients with chronic periodontitis: effects on GCF MMP-13 levels and outcome of periodontal therapy.

AIM: The aims of this study were to investigate (a) the matrix metalloproteinase-13 (MMP-13) promoter polymorphisms in severe, generalized chronic periodontitis (CP), (b) the relationship of periodontal therapy outcome with these genotypes and (c) gingival crevicular fluid (GCF) MMP-13 level-MMP-13 genotype correlation. MATERIALS AND METHODS: Genomic DNA was obtained from peripheral blood of 102 patients with severe, generalized CP, and 98 periodontally healthy subjects. MMP-13 -77A/G and 11A/12A polymorphisms were determined by the polymerase chain reaction-restriction fragment length polymorphism and DNA sequencing methods, respectively. Fifty-eight CP patients received non-surgical periodontal therapy and were followed for 6 months. Clinical periodontal parameters and GCF samples were collected at baseline and at 6 months. GCF MMP-13 levels were analysed by an enzyme-linked immunosorbent assay. RESULTS: The distribution of MMP-13 -77AG genotypes and allele frequencies did not differ significantly between study groups (p>0.05). Study subjects, except 3, had the 11A/11A genotype. MMP-13 -77G allele carriers had similar GCF MMP-13 levels and clinical periodontal parameters compared with AA genotypes after non-surgical periodontal therapy (p<0.05). CONCLUSIONS: These data suggest that the -77A/G and 11A/12A polymorphisms of MMP-13 gene are not associated with susceptibility to severe, generalized CP in a Turkish population. It seems that -77G allele carriage may not influence the outcome of periodontal therapy.

Total antioxidant capacity and superoxide dismutase activity levels in serum and gingival crevicular fluid in pregnant women with chronic periodontitis.

BACKGROUND: There is evidence of reduced antioxidant (AO) defense in periodontitis and pregnancy and adverse interactions between periodontitis and pregnancy. METHODS: In this study, serum and gingival crevicular fluid (GCF) total AO capacity (TAOC) and superoxide dismutase (SOD) enzyme concentrations in pregnant patients with chronic periodontitis (CP) were compared to those in non-pregnant patients. Periodontal examinations were performed and GCF/serum samples were obtained from 33 pregnant patients with CP (PCP), 18 pregnant patients with gingivitis (PG), and 21 periodontally healthy pregnant controls (P-controls), monitored in the first and third trimesters; 27 non-pregnant women with CP; and 25 non-pregnant control women. The concentrations of TAOC (automated measurement method) and SOD (spectrophotometric method) were determined. RESULTS: Periodontal parameters were higher in pregnant patients versus non-pregnant patients and in the CP group compared to controls, whereas TAOC and SOD concentrations were lower (P <0.05). All parameters, except plaque index, increased in pregnant subjects in the third trimester compared to the first trimester, whereas TAOC and SOD levels decreased (P <0.05). Periodontal parameters were highest and TAOC and SOD levels were lowest in the PCP group in the third trimester (P <0.05). CONCLUSIONS: Systemic and local GCF AO levels decreased in pregnancy and periodontitis, and AO defense reached the lowest levels in the last phase of pregnancy, whereas periodontal status deteriorated. These results suggest that reduced AO capacity may be associated with adverse periodontitis-pregnancy interactions, and each situation can be a provocative risk factor for the other.

Maternal periodontal disease and soluble fms-like tyrosine kinase-1 expression.

BACKGROUND: This study was conducted to examine the relationship between maternal periodontal disease and plasma angiogenic factor expression of soluble fms-like tyrosine kinase (sFlt)-1. METHODS: This was a nested case-control study of 220 women, including 45 healthy women with evidence of active periodontal disease, 98 women without evidence of active periodontal disease, 13 women with fetal exposure to oral pathogens, and 64 women without fetal exposure to oral pathogens. Active periodontal disease was defined as the presence of moderate/severe periodontal disease and evidence of periodontal disease progression. Fetal exposure to oral pathogens was determined by fetal immunoglobulin M (IgM) umbilical cord seropositivity. Maternal plasma was collected at <26 weeks of gestation; umbilical cord blood was collected at delivery. sFlt-1 was measured with an immunoradiometric assay. Demographic and medical data were chart abstracted. Maternal variables and sFlt-1 concentrations were compared between cases and controls using the Student t and chi(2) tests and analysis of variance. RESULTS: The median sFlt-1 concentration at the time of enrollment for all women was 2,374 pg/ml (interquartile range [IQR]: 1,504 to 3,194 pg/ml). Women with evidence of fetal exposure to oral pathogens had significantly higher sFlt-1 concentrations compared to IgM-negative fetuses (3,383 pg/ml [IQR: 2,610 to 4,244 pg/ml] versus 2,123 pg/ml [IQR: 1,456 to 3,011 pg/ml]; P = 0.03). CONCLUSION: Fetal exposure to oral pathogens was associated with increased plasma concentrations of sFlt-1 early in pregnancy.

Associations of chairside salivary aMMP-8 findings with periodontal parameters, potentially periodontal pathogenic bacteria and selected blood parameters in systemically healthy adults.

The aim of this cross-sectional study was to investigate associations between salivary active matrix-metalloproteinase 8 (aMMP-8) and periodontitis severity, potentially periodontal pathogenic bacteria as well as blood parameters in generally healthy participants. Therefore, 188 participants with a mean age of 48.9 ±â€¯8 years were examined. The periodontitis severity was assessed based on periodontal probing depth and clinical attachment loss. Both, aMMP-8 and microbiological analysis were performed using a validated, commercially available test system. Blood values were utilized from regular differential blood count. The aMMP-8 findings were associated with the periodontitis severity (P < 0.01), as well as with the prevalence of Porphyromonas gingivalis, Tannerella forsythia, Prevotella intermedia, Parvimonas micra, Camphylobacter rectus and Eubacterium nodatum (Pi < 0.05). No associations between aMMP-8 and the examined blood parameters were found (Pi > 0.05). In conclusion, salivary aMMP-8 findings seem to reflect periodontal disease severity as a result of an immunoreaction, especially against bacteria with high periodontal pathogenic potential.

Tumor necrosis factor-alpha-converting enzyme (TACE) levels in periodontal diseases.

Tumor necrosis factor-alpha-converting enzyme (TACE) is a metalloprotease which can shed several cytokines from the cell membrane, including receptor activator of NF-kappaB ligand (RANKL). This study aimed to investigate the hypothesis that TACE would be elevated in the gingival crevicular fluid (GCF) of persons with periodontitis. Total TACE amounts in GCF were higher in persons with chronic and aggressive periodontitis than in those with gingivitis or in healthy persons. TACE concentrations in GCF were higher in persons with chronic and aggressive periodontitis than in those with gingivitis, although not significantly higher than in healthy persons. Persons with chronic periodontitis receiving immunosuppressive treatment exhibited over 10-fold lower TACE levels than the other periodontitis groups. TACE was positively correlated with probing pocket depth, clinical attachment levels, and RANKL concentrations in GCF. In conclusion, the increased GCF TACE levels in persons with periodontitis and their positive correlation with RANKL may indicate an association of this enzyme with alveolar bone loss, and may warrant special attention in future therapeutic approaches.

Matrix metalloproteinase-8 concentration in shallow crevices associated with the extent of periodontal disease.

OBJECTIVE: The aim of this study was to analyse the association between matrix metalloproteinase-8 (MMP-8) concentration in shallow, mostly non-bleeding gingival crevices, and the extent of periodontal disease. MATERIAL AND METHODS: Plaque, bleeding on probing (BOP), probing pocket depth (PPD) and attachment level (AL) were assessed clinically in 48 patients with chronic periodontitis. MMP-8 concentrations in gingival crevicular fluid (GCF) from four shallow (PPDor=4 mm (p=0.028) and AL>or=6 mm (p<0.001). CONCLUSION: The above association between MMP-8 concentration in shallow crevices and attachment loss provides a new aspect to future studies of MMP-8 as a prognostic marker for periodontal disease.

The effect of non-surgical periodontal therapy on peroxidase activity in diabetic patients: a case-control pilot study.

OBJECTIVE: To evaluate the effect of periodontal therapy on clinical parameters as well as on total salivary peroxidase (TSP) activity and myeloperoxidase (MPO) activity in the gingival crevicular fluid (GCF) of patients with type 2 diabetes mellitus (DM2) and of systemically healthy individuals. MATERIAL AND METHODS: Twenty DM2 subjects with inadequate metabolic control (test group) and 20 systemically healthy individuals (control group), both groups with chronic periodontitis, were enrolled. Periodontal clinical parameters, namely periodontal probing depth (PD), clinical attachment level (CAL), visible plaque index (VPI), bleeding on probing (BOP), gingival bleeding index (GBI) and presence of suppuration (SUP), as well as TSP activity and GCF MPO activity, were assessed before and 3 months after non-surgical periodontal therapy. RESULTS: At baseline and 3 months post-treatment, the test group presented a higher percentage of sites with VPI and BOP (p<0.01). MPO activity in the GCF presented lower values (p<0.05) for the test group at both baseline and the post-treatment period. The periodontal treatment resulted in a significant improvement of most clinical and enzymatic parameters for both groups (p<0.05). CONCLUSIONS: In both groups, the periodontal therapy was effective in improving most clinical parameters and in reducing salivary and GCF enzymatic activity. The diabetic individuals presented lower MPO activity in the GCF.

Gingival crevicular fluid alkaline phosphatase activity reflects periodontal healing/recurrent inflammation phases in chronic periodontitis patients.

BACKGROUND: Roles for host enzymes as diagnostic indicators of periodontal status in gingival crevicular fluid (GCF) have been proposed. One of these host enzymes is alkaline phosphatase (ALP), the GCF activity of which has been associated with periodontal inflammation. Thus, the present study aimed to improve our understanding of how the healing of chronic periodontitis following scaling and root planing (SRP) affects GCF ALP activity after 15 and 60 days. METHODS: Sixteen systemically healthy subjects (aged 35 to 61 years) with moderate to advanced generalized chronic periodontitis were recruited. In each subject, paired pockets with probing depths (PDs) > or =4 mm that were located in two symmetric quadrants were chosen. These sites were randomized at the split-mouth level, with half receiving SRP treatment and the other half left untreated. Ninety-two pockets were included in the study. Clinical examinations were performed at baseline (prior to SRP) and after 15 and 60 days; information recorded included the presence of plaque, PD, clinical attachment level (CAL), and bleeding on probing. GCF was collected from each pocket included in the study at the three time points. RESULTS: A large and significant decrease in GCF ALP activity was seen 15 days after SRP, concomitant with an improvement in clinical parameters. After 60 days, an increase in GCF ALP activity back to baseline levels was recorded along with further improvements in clinical parameters. Moreover, in the SRP pockets with initial PDs >6 mm, the CAL gains between days 15 and 60 were significantly associated with changes in GCF ALP activity over the same time interval. CONCLUSIONS: The decrease in GCF ALP activity at 15 days corresponded to a decrease in clinical signs of inflammation; in contrast, the increase in GCF ALP activity at 60 days seemed to be related to subclinical recurrent inflammation or further healing/remodeling of the periodontal tissue. Therefore, GCF ALP reflects the short-term periodontal healing/recurrent inflammation phases in chronic periodontitis patients.

Biomarkers of periodontitis in oral fluids.

Matrix metalloproteinases (MMPs), the key enzymes responsible for matrix degradation, are derived from polymorphonuclear leukocytes during the early stages of periodontitis. The present study determined the levels of GCF matrix metalloproteinase-2 (MMP-2) and metalloproteinase-9 (MMP-9) and salivary MMP-8 in patients with gingivitis and periodontitis and in healthy controls. Levels of crevicular MMP-2, MMP-9 and salivary MMP-8 were determined by ELISA in subjects with healthy gingiva (n = 15), gingivitis (n = 18) and periodontitis (n = 20). Significantly higher salivary MMP-8 and crevicular MMP-9 were observed in cases of periodontitis compared to gingivitis and healthy adults. On the other hand, crevicular MMP-2 levels in periodontitis subjects were lower than those in gingivitis and healthy subjects. The three MMP levels were highly correlated to probing depth, and bleeding on probing. Salivary MMP-8, crevicular MMP- 2 and 9 may serve as biomarkers of periodontal disease and aid in early detection of periodontitis or gingivitis.

Characteristics of collagenase-2 from gingival crevicular fluid and peri-implant sulcular fluid in periodontitis and peri-implantitis patients: pilot study.

OBJECTIVE: To compare collagenase activity and collagenolytic matrix metalloproteinase (MMP) levels in gingival crevicular fluid (GCF) and in peri-implant sulcular fluid (PISF) in gingivitis (G), chronic periodontitis (CP), and peri-implantitis (PI) human subjects. MATERIAL AND METHODS: GCF and PISF were collected on filter paper strips, volume was determined, and samples were extracted in buffer containing general proteinase but not MMP inhibitors. Collagenase activity was measured using a DNP-synthetic octapeptide, and molecular and activation forms of collagenase-2 by Western immunoblotting. RESULTS: GCF from CP and G sites exhibited elevated collagenase activity and flow, but collagenase concentrations expressed per microl were not significantly different between the healthy and G sites. Minimal fluid was obtained from healthy PISF, and collagenase concentration was the same or lower than in healthy GCF. Although PISF flow was 34% lower than GCF flow in CP subjects, collagenase concentration in CP and in PI sites was 78% and 971% greater, respectively, than in the appropriate healthy sites. Western immunoblot revealed MMP-8 in both PISF and GCF; fibroblast-type MMP-8 was not detected in healthy GCF and PISF. Immunoreactivity level and inactive and activated forms of PMN-type MMP-8 in GCF and PISF increased with the severity of periodontitis and peri-implantitis. Enhanced levels of fibroblast-type MMP-8 in active form were detected only in severe CP GCF and PI PISF. CONCLUSIONS: Peri-implantitis PISF contained higher collagenase-2 levels and activity than GCF from similar deep CP sites. GCF and PISF from severe CP and PI exhibited the highest activation of MMP-8 isoenzymes species (PMN and fibroblast-type).