A Preliminary Study On The Effect Of IL-17A On Inflammatory Cytokines In Rat Bronchial Fibroblasts By Autophagy

Objective:To explore whether IL-17 A can influence the expression of inflammatory cytokines by regulating autophagy of rat bronchial fibroblasts.Methods: 1.RBFs was cultured by enzyme digestion combined with tissue sticking method,and the morphology of the 3 ～ 4th generation was observed under microscope.The Vimentin of RBFs marker protein was identified by immunofluorescence.2.The early experimenters of our research group used rapamycin to induce the autophagy model of RBFs,and the optimal concentration and time of IL-17 A and rapamycin were detected by CCK8 method.20 ng/ml for 24 hours is the best intervention concentration and intervention time for IL-17 A.2 nmol/ml for 24 hours is the best intervention concentration and intervention time for rapamycin.3.Group: control group,IL-17 A group,rapamycin group and rapamycin +IL-17 A group.4.The expression of LC3II/I protein after autophagy of RBFs was detected by Western Blot.5.ELISA was used to detect the contents of IL-6,IL-8 and CCL20 in supernatant after autophagy of RBFs.Results: 1.RBFs was successfully cultured by enzyme digestion combined with tissue sticking method.The third generation RBFs was identified by immunofluorescence chemistry,and then photographed under inverted fluorescence microscope,and Vimentin antibody was used as identification antibody.RBFs was successfully identified by immunofluorescence.The cytoplasm of vimentin expressed in cells was dyed green and the nucleus was dyed blue.It is proved that the research object of this experiment is bronchial fibroblasts.2.WB results: Compared with the control group,IL-17 A intervention down-regulated the expression of LC3Ⅱ/I protein(P<0.05).After rapamycin pretreatment and IL-17 A intervention,the expression of LC3Ⅱ/I protein decreased compared with the rapamycin group(P<0.05).3.ELISA results: compared with the control group,the levels of IL-6,IL-8 and CCL20 in IL-17 A group,rapamycin group and rapamycin +IL-17 A group increased(P<0.05).Compared with IL-17 A group and rapamycin group,the expression levels of IL-6,IL-8 and CCL20 in rapamycin +IL-17 A group decreased(P<0.05).Conclusion: 1.IL-17 A could inhibit the autophagy activity of RBFs.2.Bronchial fibroblasts could produce IL-6,IL-8,CCL20,and IL-17 A can influence their secretion by inhibiting autophagy.3.Moderate up-regulation of autophagy may improve the imbalance of inflammatory factors in RBFs induced by IL-17 A.