Healing of extraction sockets in collagenase-2 (matrix metalloproteinase-8)-deficient mice.

Matrix metalloproteinase-8 (MMP-8) participates in skin wound healing and inflammation. We hypothesized that MMP-8 plays a role in wound healing after tooth extraction and in periapical inflammation. Bone formation, collagen metabolism, and inflammation in tooth extraction socket and in periapical lesions were analyzed in wild-type mice and in MMP-8-deficient (MMP-8(-/-)) mice. New trabecular bone area in the extraction sockets and in periapical lesions were similar in both groups. In extraction sockets significantly more type III procollagen was synthesized, and the neutrophil and MMP-9 levels were lower in MMP-8(-/-) mice. The amount of Fas ligand, identified as a substrate for MMP-8, was lower in alveolar mucosa but higher in alveolar bone of MMP-8(-/-) mice. These results indicate that MMP-8 can modulate inflammation and collagen metabolism of alveolar bone and mucosa.

Expression of hypoxia-induced semaphorin 7A correlates with the severity of inflammation and osteoclastogenesis in experimentally induced periapical lesions.

OBJECTIVE: While hypoxia and inflammation are intimately linked, the effects of inflammatory hypoxia on the pathogenesis of periapical lesions remain largely unknown. The aim of this study was to examine hypoxia during the progression of experimentally induced rat periapical lesions, and to derive correlations between hypoxia-induced Semaphorin 7A (Sema7a) expression, severity of inflammation, and osteoclastogenesis in the lesions. DESIGN: Periapical lesions were developed after mandibular first molar pulp exposure in forty Sprague-Dawley rats. The animals were randomly divided into four groups and sacrificed at 0, 7, 14, and 28days after pulpal exposure. The bilateral mandibles containing the first molar were obtained and routinely prepared for histological, immunohistochemical, enzyme histochemical analyses and quantitative polymerase chain reaction detecting Sema7a mRNA expression. Data were analysed by one-way analysis of variance and the Pearson's correlation and linear tendency test. RESULTS: Periapical tissues become hypoxic during the development of experimentally induced periapical lesions, with steadily increasing numbers of HIF-1α-positive cells that positively correlate with the expression of Sema7a mRNA in the lesions. Furthermore, significant positive correlates were derived for the expression of Sema7a and the degree of inflammatory infiltration and osteoclast number, respectively. CONCLUSIONS: Hypoxia-induced Sema7a participates in the pathogenesis of periapical lesions, providing a novel therapeutic target for the treatment of this inflammatory disease in the future.

Metalloproteinases 2 and 9 Immunoexpression in Periapical Lesions from Primary Endodontic Infection: Possible Relationship with the Histopathological Diagnosis and the Presence of Pain.

INTRODUCTION: The aim of this study was to evaluate the possible associations among the histopathological diagnosis, the inflammatory infiltrate profile, the presence of pain, and the immunoexpression of matrix metalloproteinases MMP-2 and MMP-9 in periapical lesions from primary endodontic infection. METHODS: Fifty-one primary periapical lesions obtained from extracted teeth were selected for this study. Patients were previously evaluated for the presence of pain and sinus tract related to the tooth to be extracted. Tissues were processed for microscopic examination and MMP-2 and MMP-9 immunoexpression. Microscopically, samples were classified as periapical granulomas or periapical cysts and the inflammatory infiltrate as chronic or mixed. The percentage of immunopositive cells for MMP-2 and MMP-9 of each case was performed based on 10 consecutive microscopic fields. The Student t or chi-square tests were used in the statistical analysis. RESULTS: Of the total, 28 cases were classified as periapical granulomas (54.90%) and 23 cases as periapical cysts (45.10%). Seventeen patients (33.33%) reported pain associated with the extracted tooth, with 12 cases of periapical granulomas (70.58%) and 5 cases of periapical cysts (29.42%). All cases showed immunopositivity for MMP-2 and MMP-9 in a high percentage of cells, mainly in the cytoplasm of the leukocytes. MMP-2 was expressed more in periapical granulomas than periapical cysts (P < .05) and in symptomatic cases (P < .05). CONCLUSIONS: According to the results, we may conclude that MMP-2 and MMP-9 are highly expressed in periapical lesions from a primary endodontic infection. Moreover, we may suggest MMP-2 is expressed more in periapical granuloma and in cases associated with pain.

Differential Proteinase Patterns among Candida albicans Strains Isolated from Root Canal and Lingual Dorsum: Possible Roles in Periapical Disease.

INTRODUCTION: Proteinases play pivotal roles in Candida albicans infections. Although the yeast can colonize the pulpal environment, there is no information about the enzymatic profile of this organism. This in vitro study aimed to determine the proteolysis levels and to investigate differences in the expression of aspartyl proteinase genes (Sap 1, Sap 2, and Sap 4) among various root canal strains and clinical isolates from the lingual dorsum. METHODS: The extracellular proteinase activity of 104 C. albicans samples isolated from the lingual dorsum and from necrotic root canals was measured with respect to bovine serum albumin degradation after 5 days of incubation at 37°C. We used reverse-transcription polymerase chain reaction, a highly sensitive method, to detect messenger RNA transcripts of aspartyl proteinase genes (Sap 1, Sap 2, and Sap 4). The C. albicans strain SC 5314 was used as a positive control for both experiments because it is recognized as being highly proteolytic. All tests were performed in triplicate. RESULTS: Regardless of the isolation site, all C. albicans strains produced an opaque precipitation halo around the colonies, indicating some proteinase activity. However, the production of proteinase on the plates was significantly greater (P < .05) by the endodontic samples. Sap 2 was the most commonly expressed gene in all samples. Among the root canal samples, the detection of Sap 1 transcripts was always associated with the expression of Sap 2 and Sap 4. Sap 4 gene expression was detected in all root canal samples. The simultaneous expression of the 3 investigated Sap genes (Sap 1, Sap 2, and Sap 4) was more common in strains isolated from the lingual dorsum (50%) than in those isolated from root canals (29.4%). CONCLUSIONS: The increased proteolytic activity as well as the distinct pattern of Sap expression observed among the root canal samples may suggest a pathogenic role for C. albicans in endodontic infections.

Analysis of arylsulfatases A and B, acid phosphatase, lactate dehydrogenase, and aspartate transaminase in chronic periapical lesions of endodontic origin.

Attempts were made to detect and measure the activities of arylsulfatases. A&B acid phosphatase, lactate dehydrogenase, and glutamate oxaloacetate transaminase (aspartate transaminase) enzymes in human chronic lesions of endodontic origin. Thirteen periapical lesions of endodontic origin and 11 noninflamed control periapical tissues were obtained. The specimens were carried to the laboratory on liquid nitrogen and kept at -70 degrees C. Samples were thawed, homogenized, and then assayed for enzyme activities. The specific activities of arylsulfatase A (nmol/hr/mg protein) were 55.0+/-10.7 (chronic lesions) vs. 3.4+/-2.2 (controls) (p < 0.01). Arylsulfatase B specific activities (nmol/hr/mg protein) were 50.3+/-6.4 (chronic lesions) vs 91.8+/-18.4 (controls). Total acid phosphatase activities (mU/mg protein) were 45.8+/-6.6 (chronic lesions) vs. 26.8+/-3.1 (controls). Lactate dehydrogenase activities (Berger-Broida units/mg protein) of the chronic periapical lesions were significantly higher than the control group (362+/-63.2) vs. (140+/-46.0) (p < 0.05). There was no significant difference between the specific activities of aspartate transaminase in chronic lesions and the control group (68.0+/-14.5) vs. (53.0+/-10.4) mU/mg protein).

Tissue levels of matrix metalloproteinases in pulps and periapical lesions.

The purpose of this study was to evaluate the tissue levels of matrix metalloproteinase (MMP)-1, -2, -3 and their distributions in inflamed human dental pulps and periapical lesions. Samples were subjected to enzyme-linked immunosorbent assay and/or immunohistochemistry by using specific antibodies to MMP-1, -2, and -3. Results from enzyme-linked immunosorbent assay were analyzed by using the Mann-Whitney U test and presented as p values. The concentrations of MMP-1 in all experimental groups were significantly higher than in the control (p < 0.05). The acute pulpitis and control groups were significant different in terms of their MMP-2 levels (p < 0.05). The concentration of MMP-3 in acute pulpitis was significantly higher than the control and chronic pulpitis groups (p < 0.05). Immunohistochemically, MMP-1 and MMP-3 were localized in the infiltrating neutrophils, macrophages, and extracellular matrix of the acute pulpitis group. These results suggest that MMPs play an important role in the pulp tissue destruction of acute, inflamed pulp.

Matrix metalloproteinases (MMP-1, -8, -13) in chronic periapical lesions.

BACKGROUND/AIM: Matrix metalloproteinases (MMPs) are proteolytic enzymes capable of degrading almost all extracellular matrix and basement membrane components in many destructive pathological processes, such as chronic inflammation and bone-destructive lesions. The aim of this study was to determinate the correlation between concentration of collagenases (MMP-1, -8, -13) in chronic periapical lesions and their dimension calculated with software predilection through X-ray. METHODS: Chronic periapical tissues were collected by periapical surgery from 60 teeth with clinically and radiographically verified different chronic periapical lesions (20 granulomas, 20 diffuse periapical lesions, 10 cysts). Ten normal pulps used as controls were obtained by extirpation of the pulp of impacted third molars after their surgery. For rapid analysis of MMP-1, -8, -13 collagenase activities in the examined material Chemicon Collagenase Activity Assay Kit were used. From the X-ray trough software predilection (Image Tool3 Program) of the volume of chronic periapical tissue, correlation between concentration of MMPs in the periapical lesions and their dimension was confirmed. RESULTS: Different concentrations of collagenases (MMP-1, -8 and -13) in chronic periapical process from different inflammation types showed different activity of MMPs. The obtained results showed the highest values of collagenases concentration (MMP-1, -8, -13) in chronic diffuse lesions (5.39 ng/ml). Low values of concentration of MMPs accompanied less serious lesions, whereas chronical periapical lesions of large dimension had high concentration of MMPs, which was proportional to progression of the lesion and destruction of bone tissue. CONCLUSIONS: This study confirmed the destructive role of collagenases (MMP-1, -8 and -13) in inflammation process, which directly depends on the concentration of MMPs in pathologically changed tissue.

Activation of p38 mitogen-activated protein kinase in rat periapical lesions.

The purpose of this study was to investigate the activation of p38 mitogen-activated protein kinase (MAPK) during the development of periapical lesions in rats. Periapical lesions developed within 28 days after pulp exposure of mandibular first molars in Wistar rats. The animals were sacrificed randomly at 0, 7, 14, 21, and 28 days after pulpal exposure. The jaws that contained the first molar were obtained and routinely prepared for immunohistochemistry and enzyme histochemistry. A few phosphorylated p38 MAPK (P-p38)-positive cells and osteoclasts could be observed on day 7; both peaked in number on day 14. In the 21- and 28-day samples, the P-p38 MAPK expression decreased and fewer osteoclasts were observed. From day 7 to day 28, a significant positive correlation was found between P-p38 MAPK expression and osteoclasts. These findings showed that the activation of p38 MAPK might be associated with bone resorption in periapical lesions.

Sequential expressions of MMP-1, TIMP-1, IL-6, and COX-2 genes in induced periapical lesions in rats.

To elucidate the pathogenesis of periapical lesion-associated bone resorption, a disease model of Wistar rat molar was employed. After lesion induction, the mRNAs encoding for matrix metalloproteinase-1 (MMP-1), tissue inhibitor of metalloproteinase-1 (TIMP-1), interleukin-6 (IL-6), and cyclooxygenase-2 (COX-2) in the developing lesions were detected by in situ hybridization at day 5, 10, 15 and 20, respectively. At day 5, MMP-1, IL-6 and COX-2 mRNAs appeared predominantly in macrophages. During day 15 to day 20, increased expressions of these mediators were also found in osteoblasts but to a lesser extent compared with those in macrophages. MMP-1 mRNA was also detected in osteoclasts. In contrast, expression of the TIMP-1 gene was noted primarily in osteoblasts and was less pronounced compared with that of MMP-1. The mediator-expressing cells aggregated in the vicinity of bone resorption areas and their numbers increased with time. These data suggest that macrophages and osteoblasts are involved in the development of periapical lesions, and that they promote bone resorption by producing MMP-1, IL-6 and COX-2. In addition, administration of a specific COX-2 inhibitor, meloxicam, reduced the extent of periapical bone resorption by 43% and simultaneously diminished the numbers of cells synthesizing MMP-1 and IL-6 mRNAs. These results further elucidate the significance of COX-2 in disease progression of periapical lesions as it modulates indirectly the production of MMP-1 and IL-6.