The Effect Of Qijishenkang Decoction On The Proliferation Of Human Glomerular Mesangial Cells Induced By Ang? And The Expression Of Col-IV, IL-6 And Smads

Purpose:In this study,on the premise of revealing the mechanism of Qijishenkang Tang(Patent number: ZL 2011 1 0103377.6)in the prevention and treatment of Ig A nephropathy,to explore the inhibitory effect of Qijishenkang Tang on the proliferation of human Mesangial cells induced by angiotensin II and to observe the effect on type IV collagen,IL-6,the effect of Qijishenkang Tang on the proliferation of human Mesangial cells induced by angiotensin II was discussed.Effect of Smads Family proteins.Method:1.To prepare the drug-containing serum:40 male Sprague-Dawley(SD)rats were randomly divided into 5 groups: normal group,western medicine control group,low-dose group of Qijishenkang Tang,high-dose group of Qijishenkang Tang,and high-dose group of Qijishenkang Tang.After one week of continuous gastric administration,the blood was collected from the abdominal aorta.The blood was centrifuged for 20 minutes using a centrifuge 3000 r,the supernatant was pipetted,and treated with water bath at a temperature of 56 & deg;C for 30 min.After filtration and sterilization with a filter,the blood was stored at-20.degree.C.for later use.2.Culture,proliferation and grouping of human mesangial cells: inoculating human mesangial cells in a 75cm2 culture flask,adding RPMI-1640 culture solution containing 10% FBS,standing at 37 & deg;C,and culturing in a 5% CO2 incubator;and using angiotensin II as an induction factor to promote the proliferation of mesangial cells;The group of mesangial cells is as follows:(1)normal group,(2)model group,(3)western medicine control group,(4)low-dose group of traditional Chinese medicine,(5)middle-dose group of traditional Chinese medicine,and(6)high-dose group of traditional Chinese medicine.3.Test method:(1)the proliferation of cells was observed by optical microscope and(MTT)assay,(2)to detect Col-? in the supernatant of each group by enzyme-linked immunosorbent assay(ELISA),and to detect the content of Col-? in the supernatant of each group by enzyme-linked immunosorbent assay(Elisa).(3)Real-time fluorescence quantitative PCR was used to detect the expression of IL-6m RNA and Smad2 m RNA,Smad7 m RNA.Result:1.Microscopic observation showed that the proliferation of HMC was obvious.2.The results of MTT showed that the proliferation of HMC was decreased in each dose group of Qijishenkang Tang but there was no significant difference between the low dose group and the model group.Compared with the western medicine control group there was a significant difference(p < 0 01)between the low dose group and the model group(P < 0 05)and the western medicine control group.3.The results of ELISA showed that the content of Col-? in the supernatant of proliferative HMC in each group was significantly higher than that in the control group,and the difference was significant(p < 0 01).Western medicine control group,Qijishenkang Tang low,middle,high dose group compared with the model group,there was significant difference(p < 0 01);Western medicine control group and Qijishenkang Tang low,middle and high dose groups have difference(p < 0 05).In the middle and high dose groups of traditional Chinese medicine,the content of Col-? is higher than that of traditional Chinese medicine group(p < 0 05).The content of COL-? in the high-dose group was the least than that in the western medicine control group,and there was a significant difference between the groups of Qijishenkang Tang and the high-dose group(p < 0.01).4.The results of real-time fluorescence quantitative PCR showed that:(1)compared with the model group,there was significant difference in the expression of IL-6m RNA,Smad2 m RNA,but the expression of Smad7 m RNA was significantly higher in each dose group than that in the model group(p < 0.01),but there was a significant difference in the expression of Smad7 m RNA in each dose group(p < 0.01).(2)IL-6m RNA expression: compared with the western medicine control group,there was no significant difference in the expression of IL-6m RNA between the low dose group and the high dose group(p > 0.05),but there was a difference in the expression of IL-6m RNA in the middle dose group(p < 0.05);(3),but there was no significant difference in Smad2 expression between the low dose group and the high dose group.MRNA expression: compared with the western medicine control group,there was no significant difference in the expression of Smad2 m RNA between the low-dose group and the high-dose group(p > 0.05),but there was significant difference in the middle-dose group(p < 0.05).(4)expression of Smad7 m RNA:compared with the western medicine control group,there were significant differences in each dose group of Qijishenkang Tang(p < 0.05),and the most significant difference was found in the middle-dose group(p < 0.01).Conclusion:1.Ang II can induce the proliferation of human Mesangial cells.2.It is suggested that the target of Qijishenkang Tang in inhibiting the proliferation of human Mesangial cells may be Col-IV,IL-6 and Smads family proteins.3.To elucidate the mechanism of Qijishenkang Tang in the prevention and treatment of Ig AN: by inhibiting the expression of Smad2 m RNA,promoting the expression of Smad7 m RNA,inhibiting the signal transduction pathway of TGF-?1/Smads,decreasing the content of Col-IV in HMC,inhibiting the expression of IL-6m RNA,inhibiting the proliferation of HMC and changing the pathological manifestation of Ig AN.To prevent the further deterioration of Ig AN.