Tan ⅡA Of TGF-β1-induced HK-2 Cells TGF-β1 / Smads Signaling Pathway In Experimental Research Impacts

Background:Renal interstitial fibrosis(RIF) is one of the important manifestations of the development end-stage chronic kidney disease. Researches showed that in various causes of kidney disease transdifferentiation of renal tubular epithelial cells into Epithelial-mesenchymal transition(EMT) is an important factor.Excessive proliferation of renal interstitial fibroblasts lead to the accumulation of extracellular matrix(ECM).ECM components mainly collagen Ⅰ,collagen Ⅲ and collagen Ⅳ.Transforming growth factor is the recognized cause of fibrosis factor,can start and finish EMT.Smad proteins found in recent years are the transduction molecules of TGF-beta family signals from the receptors to the nuclear in the cell.TGF-beta/Smad signaling pathways play an important role in the development of renal fibrosis.So to cut off the signal transduction of TGF-betal effectively is the key to terminate and reduce the renal fibrosis.We choose tanshinone ⅡA as experimental material.It from huaxuehuayu Chinese herb Salvia.In past surveys,tanshinone IIA has been confirmed with resistance to pulmonary fibrosis,liver fibrosis and renal fibrosis.With regard to renal fibrosis,the Specific work stage is not fully clear.It Needs our further experimental studies.Objective:Cells were cultured under vitro condition.To observe Influence of Tanshione ⅡA on proliferation of human renal epithelial and secretion of ECM and expression of smad2、 smad3protein in Human renal tubular epithelial induced by transforming growth factors1(TGF-β1).To explore the mechanism of preventing and treating renal fibrosis by Tanshione IIA. To provide theoretical basis for its clinical application.Method:Human renal tubular epithelial (HK-2) cells were divided into five groups as follows:blank control group,TGF-β110ng/ml group,low dose group(Tanshione IIA1μg/ml+TGF-β110ng/ml),middle dose group (Tanshione ⅡA10μg/ml+TGF-β110ng/ml),high dose group(Tanshione IIA10μg/ml+TGF-β110ng/ml)1、Using inverted microscope to observe the morphological changes of cells in every group2、 Using MTT assays to observe the influence of Tanshione ⅡA on cell proliferation3、The content of collagen Ⅰ and Ⅲ was measured by ELISA method4、Expression of smad2and smad3and smad7protein was detected after cell lysis using western-blot methodResults:1、 the cells morphology were changed into fibroblast by TGF-β110ng/ml under inverted microscope2、 MTT results suggest that the proliferation of HK-2cells was induced significantly by TGF-β110ng/ml with a significant difference compared with the blank control group 3、(p<0.05)when TGF-β1and Tanshione ⅡA were combined and affected on the cells,the proliferation of HK-2cells were restrained,and it presents dose-effect relationship.4、Elisa showed that The content of collagen Ⅰ and Ⅲ were significantly raised in HK-2cells stimulated by TGF-β110ng/ml compared with the blank control group (p<0.05),but when joined with Tanshione ⅡA (1ug/ml-10μg/ml),the content of collagen Ⅰ and Ⅲ was obvious decreased,The concentration dependent was obvious(p<0.05).4, Western results proved that the expression of smad2、smad3protein in TGF-β110ng/ml group was significantly higher than the blank control group,when joined with Tanshione IIA (1μg/ml-10μg/ml),The expression of smad2、samd3protein was obvious decreased.The expression of smad7in TGF-β110ng/ml group was significantly higher than the blank control group,when joined with Tanshione ⅡA (1μg/ml-10μg/ml),The expression of smad7protein was rise.Between the different dose of Tanshione ⅡA was not obvious dose-effect relationship.Conclusion:Tanshione ⅡA can maintain the morphological stability of HK-2cell, prevent cells from fibrosis,and inhibit cell proliferation and secretion of collagen Ⅰ and Ⅲ induced by TGF-β1.It can also regulate the renal tubular epithelial cell signaling pathway that mediated by the TGF-betal mediated. Tanshione ⅡA can inhibit the renal interstitial fibrosis to a certain extent.