Effect Of SiRNA Targeted Inhibition Of Caveolin-1 On The Ultraviolet-induced Damage And Aging Of Human Skin Fibroblasts And Related Mechanisms

Objective To study whether small interferingRNA(siRNA)down-regulates the ex pression of caveolin-1(Caveolin-1)in human skin fibroblasts(HSF)in hum an skin fibroblasts(HSF),and to detect the proliferation and apoptosis of HSF cells through experiments,and it is clear The relationship between C aveolin-1 protein content and the decline of human skin fibroblasts.In con structing a UVA-induced HSF cell damage model,we further explored the mechanism of siRNA targeting to inhibit Caveolin-1 to protect UVA-induced human skin fibroblast damage and aging.Methods The human skin fibroblasts HSF were resuscitated and cultured in complete medium.The cultured HSF cells were divided into normal group,UVA group,siRNA-Cav-1 group,siRNA-Cav-1+UVA group after different treatments.The expression levels of Caveolin-1 mRNA and protein in each group of cells and the expression of related senescent proteins were detected by PCR and Western blot.The processed cells of each group were tested by MTT method for cell viability,flow cytometry to detect cell apoptosis,DCFH-DA staining to detect ROS level,and SA-?-GAL staining to observe cell senescence.Results Real-time fluorescent quantitative PCR and Western blotting method showed that the relative expression of Caveolin-1 mRNA and protein in the UVA group cells increased significantly,which was significantly different from the control group(P<0.05).The transfected siRNA-Cav-1 group had two The relative expression of these indicators decreased significantly(P<0.05).After UVA irradiation,the expression of p16,p21,p53 protein and the phosphorylation of p38 MAPK increased significantly.After treatment with 10?mol/L p38 MAPK inhibitor SB203580,the expression of p16,p21,p53 protein and the phosphorylation of p38 MAPK were 48 h after treatment.The level was significantly down-regulated(P<0.05),and the cells in the siRNA-Cav-1+UVA group also showed the same results as the SB203580+UVA group.The MTT test showed that the cell survival rate of the UVA group was significantly decreased,which was significantly different from that of the control group(P<0.05).The cell survival rate was significantly improved after transfection.Flow cytometry showed the same test results.The results of DCFH-DA staining showed that the green fluorescence of the UVA group was significantly enhanced,suggesting that the ROS level was significantly different from that of the control group(P<0.05).After transfection,the green fluorescence was significantly weakened,which can significantly reduce the ROS level.The results of SA-?-GAL staining showed that senescent cells increased significantly after UVA irradiation,while senescent cells in the transfected siRNA-Cav-1+UVA group decreased significantly(P<0.05).Conclusions After UVA-induced damage and aging of human skin fibroblasts,the targeted inhibition of Caveolin-1 expression can protect cells.The mechanism is related to reducing intracellular ROS levels,inhibiting p16/p21/p53 pathway activation and p38 MAPK phosphorylation Level related.