Research On The Expression Of Podocyte-Associated Molecules Of Adriamycin Nephropathy

Objective:The prevention of chronic renal failure requires therapies that specifically interfere with the pathogenesis of the various glomerular diseases. Recent discoveries in podocyte proteins involved in the renal filtration barrier have shed new light on the ultrastructure of the kidney filter and pathogenesis of proteinuria. We design the study to investigate the expression of podocyte-associated molecules in rats with adriamycin nephropathy, and to explore their possible actions in development of proteinuria. Another aim of this study is to explore the effects of mycophenolate mofetil(MMF) and leflunomide(LEF) on proteinuria and expression of podocyte-associated molecules in rats with adriamycin nephropathy.Methods and Results:1. By semiquantitative RT-PCR and western blotting, the mRNA or protein levels of several key proteins were compared between the adriamycin nephropathy groups and the controls. The molecules studied included nephrin, podocin, CD2-associated protein(CD2AP), WT1 and a-actinin-4.In the nephropathy groups, nephrin mRNA level decreased by 50% at the 2ndweek, and the control level was reached at the 4th week, and it showed 2- and 1.6-fold up-regulation at the 6th and 8th week, respectively. The mRNA level of podocin in the 2nd week nephropathy group was 1.4 times of that in normal control group, and it did not show significant changes at other time points. CD2AP mRNA levels in the 4th, 6th and 8th week nephropathy groups were 1.7, 2.5 and 1.9 times of that in normal control group, respectively. The protein levels of CD2AP in the 2nd, 4th and 6th week groups were 3.0, 1.8 and 1.2 times of that in normal control group, respectively. Compared with the control group, the protein levels of WT1 showed 1.5-, 2.7-, 3.1- and 4.3-fold up-regulation in the 2nd, 4th, 6th and 8th week nephropathy groups, respectively. No appreciable alterations were seen in the expression level of a-actinin-4 at any time point.2. Proteinuria of MMF-treated, LEF-treated and non-treated nephropathy rats were measured with BCA protein assay kit. By semiquantitative RT-PCR and western blotting, the mRNA or protein levels of above podocyte proteins were compared between the MMF-treated, LEF-treated and non-treated nephropathy rats.Proteinuria in MMF-treated or LEF-treated nephropathy rats was significantly lower than that in non-treated nephropathy rats. In MMF-treated or LEF-treated nephropathy rats, the protein expression of CD2AP, a-actinin-4 and WT1 did not show remarkable change compared with non-treated nephropathy rats. Neither did the level of CD2AP mRNA. In MMF-treated and LEF-treated nephropathy rats, nephrin mRNA levels in the 8th week group were 1.4 and 1.3...