The Study Of Correlation Between TRPC1 And Gingival Overgrowth Induced By Nifedipine And Cyclosporine A

Objective To investigate the correlation between classical transient receptor potential channel 1(TRPC1)and nifedipine(Nif)and cyclosporine A(CsA)alone or in combination iuduced gingival overgrowth.Methods Human gingival fibroblasts(HGF)were isolated and cultured by tissue block culture,and cells were identified by immunocytochemical staining using vimentin(Vim)and cytokeratin(CK).The certain concentration of Nif and CsA was used alone or in combination to stimulate human gingival fibroblasts(HGF)in vitro.After 1 day,3days,5 days,and 7 days,CCK8 method was used to detect the proliferation and cell viability of HGF.At the same time,the mRNA and total protein of each group were extracted after 1 day,3 days,5 days,and 7 days.The mRNA and protein expression of TRPC1,proliferating cell nuclear antigen(PCNA)and apoptosis marker molecule B-cell lymphoma-2(Bcl-2)were detected by Real-Time qPCR and Western blot.The expression of TRPC1 in each group was detected by ICC method.The expression and difference of TRPC1 in the connective tissue of the rat gingival tissue sections which have been modeled by the same research group were detected by IHC.Statistical analysis was performed using SPSS19.0,and the test data were all expressed sx ±.The difference between the multiple groups was analyzed by the variance analysis of the two-factor repeated measurement data,and the subsequent pairwise comparison was performed by the least significant difference method(LSD).The difference was statistically significant at P < 0.05.Results HGF was isolated and cultured in vitro,and about 6-8 days,cells were observed to climb out from the tissue block.The cells were mostly fusiform and arranged in a spiral shape.The results of immunocytochemical staining showed that the HGF surface antigen Vim antibody was positive expression,CK antibody was negative expression,indicating that the cultured cells are gingival fibroblasts.CCK8 results showed that Nif,CsA and Nif+CsA could increase the vitality of HGF and promote the proliferation of HGF cells(P<0.05);Real-Time qPCR and Western blot results showed that Nif and CsA alone or in combination could up-regulated the expression of Bcl-2and TRPC1,but not PCNA.ICC results suggested that the expression of TRPC1 in HGF stimulated by Nif and CsA alone or in combination was significantly increased.Immunohistochemical results showed that the expression of TRPC1 was significantly increased in Nig and CsA alone and in combined stimulation of gingival connective tissue.Conclusion Under action of Nif and CsA combined and alone,the proliferation of HGF cells can not changed significantly but the expression of Bcl-2 increased significantly;And the expression of TRPC1 in rat gingival connective tissue and HGF increased significantly,suggesting that TRPC1 may play a role in the pathogenesis of drug-induced gingival overgrowth.