The Primary Study Of Adiponectin In The Mechanism Of Periodontitis

Objective:By investigating the role of adiponectin in the machanism of periodontitis,we performed the primary culture of human gingival fibroblasts,performed the regulation of adiponectin in P.g LPS induced inflammation in cultured HGFs,so that we can preliminarily reveal the role of Adiponectin and gingival fibroblasts in the development of periodontal inflammation,and to provide a theoretical basis for further discovering the pathogenesis of periodontitis and adjuvant drug therapy.Methods:In this study,we selected patients for the removal of the third molar impacted teeth in Tianjin Medical Oral Hospital(age 20-26 years,the average age of 24 years)from August 2017 to October 2017,excluding systemic diseases such as diabetes,pregnancy,smoking,access to the patient informed consent,In the operation,we collected healthy and no obvious inflammation gingival tissue for our study.(1)By using tissue block primary culture,healthy human gingival fibroblasts were cultured in vitro,the growth status was observed under microscope,well-known cells were obtained after generation,and the 3rd generation cells were used for immunohistochemical staining to identify the source,and 4-6 generations of cells were selected for the experiment.(2)APN,P.g LPS or APN combined with P.g LPS stimulated HGFs for 24 h.Cell stimulation was as follows: 1μg/ml APN,2μg/ml P.g LPS,2μg/ml P.g LPS+1μg/ml APN,blank control(10% FBS medium).(3)After stimulating with above methods,the Interleukin-6(IL-6),Interleukin-10,(IL-10)、 Human recombinant heme oxygenase-1(HMOX-1)、cycloxygenase-2(COX-2)in HGFs at different time points(6,8,24 hours)was detected by real-time PCR.(4)Using SPSS 23.0 statistical software for data analysis :the measurement data was expressed as mean ± standard deviation;One-factor ANOVA analysis and factorial design using factorial analysis,p <0.05 was considered statistically significant.Result:(1)HGFs ware successfully cultured from tissue block by using tissue block method.Under the microscope,the cells were cultured from the tissue block after 6-8 days,and the passage of cell morphology are the consistent long spindle.Immuno-histochemical results showed that the cell anti keratin staining was negative,and the resistance to wave-like silk protein staining was positive,which confirmed that our cultured cells were human gingival fibroblasts from the middle germ layer and did not mix epithelial cells.(2)Compared with the blank control group,P.g LPS promoted secretion of anti-inflammatory media IL-10 and HMOX-1,and there are statistical differences(p<0.05),and P.g LPS significantly inhibited HGFS secretion of inflammatory media IL-6 and COX-2,and there were statistical differences(p<0.05),APN plays an anti-inflammatory role in the periodontal inflammation process.(3)Compared with the blank control group,APN significantly inhibited HGFS secretion of inflammatory media IL-6 and COX-2,and there were statistical differences(p<0.05),but promoted HGFs secretion of anti-inflammatory media IL-10 and HMOX-1,and there are statistical differences(p<0.05),APN plays an anti-inflammatory role in the periodontal inflammation process.(4)In each best stimulation point,P.g LPS stimulates HGFs secrete IL-6 and COX-2,while APN obviously suppresses this role;P.g LPS inhibits HGFs secretion of anti-inflammatory factors IL-10 and HMOX-1,of which APN plays an antagonistic role in IL-10.Conclusion:(1)P.g LPS,as the main pathogenic factor of periodontitis,can stimulate the synthesis and secretion of various cytokines in the gingival fibroblasts,which can be used as an effective inducer in the vitro model of periodontal inflammation.(2)APN can inhibit HGFs secretion of inflammatory media IL-6 and COX-2,promote HGFs secretion of anti-inflammatory media IL-10 and HMOX-1,play an anti-inflammatory role.(3)APN can play an anti-inflammatory role in the development of periodontitis by regulating proinflammatory factors and anti-inflammatory factors.