Role of Ubiquitinproteasome Pathway in TGF-beta/Smad Signaling Pathway in Human Proximal Tubular Epithelial Cells

Objective: To identify how Smad7 changing and Smad signaling regulated by ubiquitinproteasome pathway(UPP) in tubulointerstitial on human proximal epithelial cells.Methods: Human kidney proxima tubular cells HK2 were divided into three groups according to the differnet treatment: control group,TGF-β1(5 μg/L) group,TGF-β1(5 μg/L)+MG-132(5 mmol/L) group.Western blot were performed to determine the changes of Smad7 protein and Smad2/3 phosphorylation,and semi-quantitive RT-PCR were performed to test diversify of Smad7 mRNA.Results: Compared with that in TGF-β1 group,the expression of phospho-Smad2/3 decreased by 90% in MG-132+TGF-β1 group(P0.01).The level of Smad7 protein decreased progressively in tubular cells after TGF-β1 inducing,Smad7 protein expression decreased by 20% at 30 min(P0.05) and almost showed no expression at 24 h(P0.01).Compared with that in TGF-β1 group(P0.01),Smad2/3 phosphorylation and significantly decreasing of Smad7 protein was hindered by addition of MG-132 in MG-132+TGF-β group.Semi-quantitative RT-PCR showed in contrast to expression of Smad7 protein,Smad7 mRNA was up-regulated rapidly by TGF-β1 inducion and no difference was found between MG-132+TGF-β1 group and control group.Conclusion: Different expression between Smad7 protein and Smad7 mRNA after TGF-β1 stimulation is discovered in this study.Up-regulation of Smad7 mRNA and UPP degradating Smad7 protein are both involved in this change.