gilz对巨噬细胞功能影响及其在小鼠肝脏缺血再灌注中保护作用-effect of gilz on macrophage function and its protective effect on hepatic ischemia-reperfusion in mice.docx

AbstractPartⅠtransfect GILZand siGILZ tomouseperitoneal macrophages in vitroObjective: GILZ have ability of anti-inflammation and regulation of epithelial sodium channels and many other effects.It’s a new important mediator of glucocorticoid action.SiRNA was looked as a revolutionary breakthrough in the history of genetic research, has the specificity and efficiency of the characteristics in inhibition of target mRNA . SiRNA technique is a powerful ways to inhibit gene expression specifically. The present study, GILZ and siGILZ plasmid vector that has been constructed transfect to the mouse peritoneal macrophages with sufficient quantity and purity in vitro , and detect expression of GILZ mRNA and protein levels.The model of high expression and silence of GILZ has been established for further study of the genetic characteristics of GILZ in macrophages. METHODS: Cultured sufficient quantity and purity of mouse peritoneal macrophages, siGILZ and GILZ transfected into mouse peritoneal macrophages by the French company Polyplus-transfections transfection reagents INTERFERinTM and jetPEIT M respectively. The interference test is divided into a blank Control group, siC group and siGILZ group and 1 hour, 3 hours, 6 hours, 12 hours and 24 hours were taken as the observation point. GILZ gene transfer trials were divided into control group, empty vector AAV group and GILZ group, were also taken after 1 hour, 3 hours, 6 hours, 12 hours and 24 hours for observation. Realtime PCR detected the mRNA expression levels of GILZ in each treatment group, Western-blot detected expression of the protein GILZ of cells in each treatment group. Results: Compared with the control group and siC group, siGILZ significantly decreased the mRNA level and protein levels of GILZ after transfected for 6 hours (p 0.05); GILZ gene transfer experiments, compared with the control group and empty plasmid AAV group, GILZ was significantly increased the expression of GILZ in mRNA and protein le