Effect Of Glycyeehizin On Inflammatory Factors And Fibrotic Factors In Glomerular Mesangial Cells Through HIF-1α/VEGF Under High Glucose Condition

Objective To investigate whether Glycyrrhizic acid(GA)can inhibit the expression of inflammatory factors and fibrotic factors in mesangial cells induced by high glucose by affecting the expression of HIF-1α(Hypoxia inducible factor-1 alpha,HIF-1α),thus providing a theoretical basis for the treatment of diabetic nephropathy.Methods The murine mesangial cell line was SV40 MES 13 in vitro.The experiment was divided into two parts.In the first part,SV40 MES 13 cells were divided into three groups: NG group,HG group,and HG+GA group.The growth and morphological changes of the cells in different groups were observed by microscope,and IL-1β,IL-6,IL-8 were detected by Western blot,immunofluorescence and Elisa.Expression of TNF-1α and α-SMA.In the second part,1,SV40 MES13 cells were divided into three groups: NG group,HG group and HG+GA group.The expression levels of HIF-1α and VEGF in different groups were verified by Western blot and immunofluorescence assay.2.SV40 MES 13 cells were divided into NG group,HG group,HG+GA group,HG+ KC7F2 group,HG+GA+ KC7F2 group,HG+DMOG group,and HG+GA+DMOG group.The cell proliferation level was detected by CCK-8 and Western blot.Elisa was used to detect the expression of IL-1β,IL-6,IL-8,TNF-1α and α-SMA in different groups.Results In the first part,1,Western blot and immunofluorescence experiments showed that compared with NG group,the expression levels of inflammatory factorrelated proteins IL-1β,TNF-α,IL-6 and IL-8 in HG group were increased(P<0.05).The intracellular fluorescence intensity of IL-1β and TNF-α was increased in HG group(P<0.05),while the expression level of inflammatory factor protein was decreased in HG+GA group than that of HG group(P<0.01).The fluorescence intensity of IL-1β and TNF-α were decreased in HG+GA group(P<0.05).The supernatant contents of inflammatory factor were shown by Elisa to be consistent with the WB results.2.Western blot and immunofluorescence experiments showed that the expression level of fibrosis factor α-SMA in HG group was significantly higher than that of HG+GA group(P<0.05),and GA significantly inhibited the expression level of fibrosis factor α-SMA compared with HG group.In the second part 1,Western blot and immunofluorescence experiments confirmed that GA could inhibit the expression level of HIF-1α/VEGF factor in signaling pathway(P<0.05).2.Mouse mesangial cells cultured with high glucose in vitro were treated with 2.5,5,7.5 μmol/L HIF-1α activator DMOG and5,15,25 μmol/L HIF-1α inhibitor KC7F2 respectively,and the results showed that DMOG and KC7F2 activated or inhibited HIF-1α expression in a concentrationdependent manner.Mouse mesangial cells were treated with 7.5μmol/L activator DMOG and 25μmol/L inhibitor KC7F2,respectively.CCK8 assay showed that HIF-1αup-regulation promoted the proliferation of mouse mesangial cells.Glycyrrhizic acid could further inhibit cell proliferation induced by the activator DMOG,and glycyrrhizic acid could cooperate with the inhibitor KC7F2 to further inhibit the proliferation of glomerular mesangial cells.Western blot results showed that glycyrrhizic acid could inhibit the HIF-1α/VEGF protein level induced by DMOG,and could also cooperate with the inhibitor KC7F2 to down-regulate the expression of HIF-1α/VEGF.Western blot and Elisa results showed that glycyrrhizic acid inhibited DMOg-induced IL-1β in mouse mesangial cells through the intervention of activator DMOG and inhibitor KC7F2.The expression of TNF-α,IL-6,IL-8 and fibrosis factor α-SMA can also cooperate with KC7F2 to inhibit the expression of IL-1β,TNF-α,IL-6,IL-8 and fibrosis factor α-SMA.Conclusion 1.Glycyrrhizic acid can inhibit high glucose-induced expression of inflammatory and fibrotic factors in mesangial cells.2.The mechanism by which glycyrrhizic acid inhibits the expression of inflammatory and fibrotic factors in mesangial cells under conditions of high glucose may be related to the abnormal signaling pathway of HIF-1α/VEGF.Glycyrrhizic acid may play a protective role in high glucose induced mesangial cells by down-regulating the expression of HIF-1α.