The Study Of FimA Genetic Polymorphism Of Porphyromonas Gingivalis And Its Pathogenicity

Porphyromonas gingivalis (P. gingivalis) is considered to be the most important infectious agent causing chronic periodontitis. Several studies have demonstrated heterogenic virulence properties among strains of P. gingivalis; and P. gingivalis strains can be divided into virulent strains and avirulent strain by animal and experimental model systems; the expression of heterogenic virulence properties is dependent on its clonal diversity. Identifying virulent strains of periodontal pathogens is one of the focuses of the etiology study of periodontal disease, therefore, the search for virulent strains of P. gingivalis and its pathogenesis will be helpful to prevent and cure periodontitis. Adhesion to host tissues is the first step for pathogens to cause infection, and fimbriae are the major adhesion structure of P. gingivalis, which possess massive biological activities. The fimA gene encoding fimbriae has wide clonal variations, it has been classified into five groups (I to V) on the basis of nucleotide sequences of opening reading frame. Epidemical studies show that P. gingivalis strains with II fimA and IV fimA were closely associated with the initiation and progression of periodontitis in Japanese,while strains with I fimA were associated with health periodontal status, which indicted that specific fimA genotypes were associated with the initiation and progression of periodontitis, therefore fimA gene may be the key factor which leads to virulence diversities of P. gingivalis. Now virulence diversities among different fimA genotypes have been one of the hot spots of periodontitis pathogenesis. Our study aims to find the distribution of fimA genotypes of P. gingivalis in Chinese chronic periodontitis, and try to investigate the pathogenesis of P. gingivalis with fimA genotypes closely associated with Chinese chronic periodontitis.In the first part, the occurrence of P. gingivalis in 101 chronic periodontitis patients was determined by species-specific 16S rRNAPCR and culture method; distribution of fimA genotypes was assessed in P.gingivalis positive samples by fimA type-specific PCR to investigate the distribution of fimA genotype of P. gingivalis in chronic periodontitis patients and try to find the relationship between fimA genotypes and periodontitis.In the second part, cytokine and adhesion molecular induction of oral epithelial cells by type I fimA P. gingivalis and type IV fimA P. gingivalis were compared on the two levels of mRNA and protein by RT-PCR, ELISA and FCM.And the results are as follows:(1) The detective ratio of P. gingivalis by species-specific 16S rRNA PCR and culture method was 88.1%, 38.6%, respectively, which indicates that species-specific 16S rRNA PCR was more sensitive than culture method.(2) Multiple fimA genotypes were detected in subgingival sample of Chinese periodontitis patients. Type II and type IV P. gingivalis strainswere more prevalent, and the detective ratio of type II and type IV was 43.8% and 40.4%, respectively, which indicates type II and type IV P. gingivalis were closely associated with the initiation and progression of periodontitis.(3) The regulation effects on IL-6 expression by type IV P. gingivalis were stronger than type I, which happened on transcription and post-transcription levels. The IL-6 mRNA up-regulation effects caused by type IV P. gingivalis were higher than type I P. gingivalis. The production of IL-6 protein was not consistent with its mRNA; Under the stimulation of type IV P. gingivalis, IL-6 protein level in epithelial cell culture supernants was lower than that of type I P. gingivalis, which indicates that the protease proteolysis on IL-6 of type IV P. gingivalis was stronger than that of type I P. gingivalis.(4) The regulation effects on IL-8 expression by type IV P. gingivalis were stronger than type I, which happened on post-transcription levels. The IL-8 mRNA up-regulation effects caused by type IV P. gingivalis were similar with type I P. gingivalis. The production of IL-8 protein was not consistent with its mRNA; Under the stimulation of type IV P. gingivalis, IL-8 protein level in epithelial cell culture supernants was lower than that of type I P. gingivalis, which indicates that the protease proteolysis on IL-8 of type IV P. gingivalis was stronger than that of type I P. gingivalis.(5) Under the environments of our experiment, the expression of IL-ip protein and mRNA was low, which indicates the induction of IL-ip of oral epithelial cell was not sensitive to the stimulation of P. gingivalis.(6) The expression of ICAM-1 mRNA was up-regulated by P. gingivalis stimulation, ICAM-1 expression on epithelial cell membrane was intensified;ICAM-1 up-regulation effects caused by type IV P. gingivalis was stronger than type I P. gingivalis.Based on these findings, we concluded that P. gingivalis strains with II fimA and IV fimA were closely associated with the initiation and progression of chronic periodontitis in Chinese. The regulation effects of the cytokines and adhesion molecules production of oral epithelial cells by P. gingivalis strains with IV fimA were stronger than type I P. gingivalis, which indicates the virulence and pathogenicity of P. gingivalis was related with its fimA genotypes.