Effects Of Zaohuang Mixture Contained Serum On Proliferation And The Expression Of CTGF、 MCP-1in Human Mesangial Cells Cultured In High Glucose

[Objective] To investigate the inhibition of zaohuang mixture contained serum (ZHM) on the proliferation of human mesangial cells (HMC) induced by high glucose.To investigate the effect of ZHM on the expression of connective tissue growth factor (CTGF) and monocytes chemotactic protein1(MCP-1) expression in HMC.To explore the significance of the prevention and treatment in diabetic kidney disease (DKD)fom cellular and molecular level.[Methods] HMC was cultured in vitro and identified.Serums containing different concentration of ZHM and haikunshenxi were prepared by serum pharmacology methodHMC were incubated in DMEM medium with10%contained serum and high glucose.Low-glucose group is the control group.The proliferation of mesangial cells was detected by cell counting kit (CCK-8) after incubation at different times (24h,48h,72h).Select a new time point48h.The level of CTGF protein and mRNA expression was measured by immunocytochemistry and RT-PCR respectively.The level of MCP-1protein and mRNA expression was measured by ELISA and RT-PCR technology.[Results](1) HMC were successfully cultured.The cellular markers including actin,yosin,vimentin, desmin were expressed,and the expressions of factor Ⅷ and cytokeratin were negative by immunofluorescence method.(2) Compared with the group of low glucose the proliferation of HMC in high glucose group was significantly higherComparing with the group of high glucose the ZHM groups with different concentration and haikunshenxi group could inhibit the HMC proliferation induced by high glucose and showed dose dependent manner.(3) With the stimulation of high glucose,the expression of CTGF protein and mRNA was increased significantly(P<0.01),which then were decreased by the treatrnent of ZHM and showed a dose-dependent manner.The most obvious inhibition of group is the ZHM high-dosage group and haikunshenxi group.Compared with haikunshenxi group CTGF protein and mRNA expression in the ZHM middle-dosage group and high-dosage group was not statistically significant(P>0.05).(4) With the stimulation of high glucose,the expression of MCP-1protein and mRNA was also increased significantly(P<0.01),which then were decreased by the treatment of ZHM and showed a dose-dependent marnnerThe most obvious inhibition of group is the ZHM high-dosage group and haikunshenxi group.Compared with haikunshenxi group MCP-1protein and mRNA expression in the ZHM middle-dosage group and high-dosage group was not statistically significant (P>0.05).[Conclusion](1) Cultivate high purity of HMC successfully.Found out the optimum condition for HMC culture.For further research of ZHM intervention HMC laid experimental basis.(2) The proliferation of HMC may be induced by high glucose and ZHM can inhibit the proliferation.(3) The expression of CTGF and MCP-1in HMC may be induced by high glucose and ZHM can inhibit the expression.(4) ZHM can restrain cell proliferation and CTGF、MCP-1secretion increased,in order to reduce the accumulation of extracellular matrix and prevent glomerular sclerosis and glomerular fibrosis events.This may be the mechanism of ZHM in prevention and treatment of DKD.