The Primary Study Of Adiponectin And Its Receptors In The Development Of Periodontitis

Objective:By investigating the role of adiponectin and its receptors in the development of periodontitis,we performed the primary culture of human gingival fibroblasts determined the expression of adiponectin receptors in HGFs under normal conditions and mimic periodontitis in vitro,performed the regulation of adiponectin in P.g LPS induced inflammation in cultured HGFs,so that we can provide basis for further revealing intrinsic mechanism in the relationship of obesity-related diseases and periodontitis and the medicine for patients with periodontitis and obesity.Methods:In this study,after informed consent from every patient?aged 18-24 years,mean age 22 years?who performed the extraction of wisdom teeth at the maxillofacial department of Stomatology Hospital of Tianjin Medical University without systemic diseases such as diabetes and cardiovascular disease,pregnancy,smoking and history of antibiotics within 3 months.?1?The human gingival fibroblsts were obtained from healthy gingival tissue and cultured in vitro.The cells at 4-6^thh passage were used in this study.?2?Total RNA was extracted from HGFs,and cDNA was reverse transcribed.The mRNA expression of AdipoR1 and AdipoR2 in HGFs was detected by reverse transcription polymerase chain reaction?RT-PCR?.?3?HGFs were stimulated with P.g LPS or TNF-?or APN for 24 h.Cell stimulation was as follows:TNF-?50ng/ml,P.g LPS 0.1?g/ml,APN 1?g/ml,blank control?10%FBS medium?.?4?After stimulating with above methods,real-time PCR was used to detect adiponectin receptor expression in HGFs at different time point.?5?P.g LPS or combined with APN stimulated HGFs for 24h.Cell stimulation was as follows:2?g/ml P.g LPS,2?g/ml P.g LPS+1?g/ml APN,blank control?10%FBS medium?.?6?After stimulating with above methods,the tumor necrosis factor-?TNF-??,inteleu-kin-6?IL-6?,prostaglandin-E2?PGE₂?,and matrix metalloproteinase-1?MMP-1?inHGFs at different time points?6,8,24 hours?was detected by real-time PCR.?7?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 were obtained.immunohistochemistry showed that the passaging cells were negative for anti-keratin staining and positive for anti-silk fibroin staining,confirming the mesodermal-derived fibroblasts.?2?AdipoR1 and AdipoR2 were expressed in human gingival fibroblasts by RT-PCR.The brightness of AdipoR2 band was higher than that of AdipoR1,suggesting that the fibroblast receptors in gingival connective tissue may be mainly AdipoR2,further siRNA experiments and protein experiments are needed to confirm the result.?3?TNF-?down-regulated the expression of AdipoR2,up-regulated the expression of AdipoR1,APN and P.g LPS can up-regulated the expression of AdipoR1/R2.?4?P.g LPS can promote the secretion of PGE₂,IL-6,MMP-1 and TNF-?,APN can inhibit significantly the secretion of IL-6 and MMP-1 induced by P.g LPS in HGFs?p<0.05?,can not promote significantly the secretion of TNF-?and PGE₂induced by P.g LPS in HGFs?p>0.05?.Conclusion:?1?AdipoR1 and AdipoR2 were mainly expressed in human gingival tissues,adiponectin may act on periodontal tissues by endocrine mainly via AdipoR2.?2?APN can inhibit the secretion of IL-6 and MMP-1 induced by P.g LPS,therefore inhibiting the role of pathogenic microorganisms in pathogenic periodontitis.?3?In the condition of periodontal inflammation,the efficiency of adiponectin and its receptors declines,causing further existing periodontal inflammation,therefore timely debridement therapy and auxiliary APN local application to prevent disease progression may be of great significance.