DKK3在慢性肾小球肾炎中表达及诱导肾小球足细胞迁移的机制研究

Expression of DKK3 in chronic glomerulonephritis tissues and its induction of migration of glomerular podocytes

目的研究DKK3在慢性肾小球肾炎中的表达,分析DKK3在慢性肾小球肾炎发生、发展中的意义。方法选取人正常肾组织20例、慢性肾小球肾炎组织80例为研究材料,通过实时定量PCR和免疫蛋白印迹实验验证DKK3在人正常肾组织、慢性肾小球肾炎组织中的相对表达;实时定量PCR分析β-catenin mRNA在人正常肾组织、慢性肾小球肾炎组织中的相对表达,Pearson相关性分析DKK3与β-catenin在慢性肾小球肾炎组织中的表达相关性;选取肾小球足细胞MPC5进行DKK3基因过表达,transwell实验验证实验组和对照组中细胞迁移和侵袭的影响;免疫蛋白印迹实验验证DKK3对细胞迁移和侵袭的影响机制。结果与正常肾组织相比,DKK3在慢性肾小球肾炎组织中表达上调(P<0.01),同时DKK3的表达与β-catenin呈正相关性,并参与慢性肾小球肾炎的进展(P=0.0002,r^2=0.1651);DKK3基因过表达后能明显促进细胞迁移和侵袭(P<0.01);免疫蛋白印迹实验显示DKK3基因过表达后能上调Vimentin、N-cadherin、MMP7、β-catenin蛋白的表达,下调E-cadherin蛋白的表达水平(P<0.01)。结论 DKK3在慢性肾小球肾炎组织中表达上调,可能是重要的调控因子参与慢性肾小球肾炎的进展。

Objective To investigate the expression of DKK3 in chronic glomerulonephritis tissues,and to discuss the roles of DKK3 in development and progression of chronic glomerulonephritis. Methods As the study object,20 cases of normal human nephridial tissues,80 cases of human chronic glomerulonephritis tissues were collected.With real-time quantitative PCR and Western blotting,relative expression of DKK3 in these normal human nephridial tissues and human chronic glomerulonephritis ones were examined..With Pearson correlation test,the correlation of DDK3 with expression of β-catenin in the human chronic glomerulonephritis tissues were investigated.Glomerular podocyte line MPC5 was selected to conduct over-expression of the DDK3 gene,and through Transwell migration assay effects of cell migration and invasion were explored in the test group and the control group.With Western blotting,induction of DDK3 on glomerular podocyte migration and invasion was analyzed. Results Compared to the normal nephridial tissues,expression of DKK3 was upregulated in the human chronic glomerulonephritis tissues ( P <0.01),and meanwhile exhibited positive correlation with expression of β-catenin and involved in progression of chronic glomerulonephritis( P =0.0002,r 2 =0.1651).Transwell migration and invasion assays showed that over-expression of DKK3 in the cell line MPC5 significantly promoted migration and invasion of podocytes ( P <0.001).Furthermore,Western blotting assay indicated that over-expression of DDK3 upregulated expressions of some proteins including Vimentin,N-cadherin,MMP7 and β-catenin,and down-regulated expression of E-cadherin. Conclusions Up -regulation of DKK3 in human chronic glomerulonephritis tissues suggests that DDK3 may be a key regulator to involve in development and progression of chronic glomerulonephritis.