| Objective: Experimental periodontitis model of rats was established to observe the expression of ASC, caspase-1 and IL-1βin rat periodontium, analysis their relevance with the development of periodontitis. Provide experimental evidence for further research about the inflammation mechanism of periodontitis.Method: Forty 6-month-old male Wistar rats were randomly divided into four groups: normal group, ligation at 2 weeks group (group 2W), ligation at 4 weeks group (group 4W) and ligation at 8weeks group (group 8W). Each group had 10 rats. Rats of group 2W, group 4W, group 8W were anesthesiaed, then ligated the bilateral maxillary first molar teeth of cervical using a wire diameter of 0.20mm. High-sugar (glucose 100g/L) soft food (sugar water immersion) was used after the operation. After ligated 2 weeks, 4 weeks, 8 weeks, killed the rats under general anesthesia, taken maxillary, put into 10% formaldehyde buffer solution for fixing 48 hours, then decalcified by 10% EDTA solution for 8 weeks, periodontal tooth continuous slices was made for HE staining and immunohistochemistry staining (Envision method). The morphological changes and the expression and distribution of ASC, caspase-1 and IL-1βin periodontium of each group were observed under light microscope. The date was processed for statistical analysis.Results:1 Estabilishment of the rat periodontitis model:Taken the made maxillary first molar periodontal slices, under optical microscope observed: when ligated in 4 weeks, obvious gingival flammation infiltrated, epithelial nail apparent elongated and increased, vessels of periodontal membeancer expanded, shown inflammatory cells infiltration, junctional epithelium move to the root, periodontal pocket was already to shamed, the absorbed and destructured alveolar bone. All of them have indicated that: periodontitis of experimental rats'model had been successfully established.2 Observation of the rat periodontal tissue morphology by the HE staining:Normal group: gingival epithelium had no significant infiltration of inflammatory cells; junctional epithelium was located in the cemento-enamel junction boundaries. Gingival fibers and periodontal ligament fibers arranged regularly and orderly. The frame of alveolar bone was integrity.2 weeks group: gingival epithelial nail stretched and proliferated to connective tissue, neutrophils, lymphocytes infiltrated. The large number of epithelial neutrophil infiltrated in the connective tissue beneath. The frame of alveolar bone was integrity.4 weeks group: a large number of inflammatory cells infiltrated in the gingival epithelial subcutaneous connective tissue, the majority of which was neutrophils and lymphocyte. The junctional epithelium proliferated to the root side, extended than formed a periodontal pocket, periodontal ligament space widened, the height of the alveolar bone was significantly loss.8 weeks group: the gingival junctional epithelium further to proliferated to the root side, formed a deep periodontal pocket, osteoclasts formed, shown extensive infiltration of inflammatory cells, some were plasma cells and lymphocyte predominance, some were neutral PMN-based, and some even formed abscesses. The alveolar bone was absorpted and can be seen clearly. 3 Immunohistochemical detection of the expression of ASC, caspase-1 and IL-1βin rat periodontal tissue:3. 1 The ASC expressed in rat periodontal tisuue:The positive staining of ASC was yellow or brown granules, and mainly located in the cytoplasm of the epithelial cells, neutrophils, monocytes- macrophages, vascular endothelial cells, fibroblasts and other cells. ASC in the normal group of periodontal tissues showed weak and positive expression; in periodontal tissues of 2 weeks group, 4 weeks group and 8 weeks group showed positive expression; in inflammatory cells infiltration showed strong positive expression of lesion. The OD values of the four groups were: the normal group: 0.0188; the 2 weeks group: 0.0291; the 4 weeks group: 0.0390; the 8 weeks group: 0.0529.3. 2 The caspase-1expressed in rat periodontal tisuue:The positive staining of caspase-1 was yellow or brown granules, and mainly located in the cytoplasm of the epithelial cells, neutrophils, monocytes- macrophages, vascular endothelial cells, fibroblasts and other cells. Caspase-1 in the normal group of periodontal tissues showed weak and positive expression; in periodontal tissues of 2 weeks group, 4 weeks group and 8 weeks group showed positive expression; in inflammatory cells infiltration showed strong positive expression of lesion. The OD values of the four groups were: the normal group: 0.0219; the 2 weeks group: 0.0327; the 4 weeks group: 0.0489; the 8 weeks group: 0.0604.3. 3 The IL-lβexpressed in rat periodontal tisuue:The positive staining of IL-lβwas yellow or brown granules, and mainly located in the epithelial cells, neutrophils, monocytes-macrophages, vascular endothelial cells, fibroblasts and other cells cytoplasm and matrix. IL-lβin the normal group of periodontal tissues showed weak and positive expression; in periodontal tissues of 2 weeks group, 4 weeks group and 8 weeks group showed positive expression; in inflammatory cells infiltration was strongly positive expression of lesion. The OD values of the four groups were: the normal group: 0.0222; the 2 weeks group: 0.0296; the 4 weeks group: 0.0439; the 8 weeks group: 0.0558.4 The result of statistical analysis about intensity of positive signal (average optical density value):Compared the OD value of the ASC expression in the four groups: the 8 weeks group> the 4 weeks group> the 2 weeks group>the normal group (P<0.05).Compared the OD value of the caspase-1 expression in the four groups: the 8 weeks group> the 4 weeks group> the 2 weeks group>the normal group (P<0.05). Compared the OD value of the IL-1βexpression in the four groups: the 8 weeks group> the 4 weeks group> the 2 weeks group>the normal group. (P<0.05).5 Correlation analysis of the three:The correlations of ASC and caspase-1 in the four groups were: 0.772, 0.898, 0.840, 0.848, and all of them have a positive correlation.The correlations of ASC and IL-1βin the four groups were: 0.770, 0.893, 0.795, 0.812, and all of them have a positive correlation.The correlations caspase-1 of and IL-1βin the four groups were: 0.847, 0.863, 0.803, 0.804, and all of them have a positive correlation.Conclusion:1 Periodontal ligation successfully established a model of experimental rat periodontitis. And the inflammation severity increased as the ligation time prolonged.2 In each group, ASC, caspase-1 and IL-1βexpression were detected in rat periodontal tissues.3 With the increasing degree of periodontal disease, the expression of ASC, caspase-1 and IL-1βwere increased accordingly in rat periodontal tissue. The result indicates that: the ASC/caspase-1/IL-1βplayed an important role in the process of rat periodontitis.4 The expression of ASC, caspase-1 and IL-1βhad a highly positive correlation in normal and periodontitis tissue. The result indicates that: the ASC/caspase-1/IL-1βplayed an synergetic role in the process of rat periodontitis. |
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