Effect of Wenyang-Huoxue-Lishuiformula on Cytoskeleton and Relatedproteins of Podocytes

Abstract：Objective: This study aimed to investigate the effect of warming yang serum on correlative proteins of podocytes in podocytes of mouse with puromycin-induced injury,including the CathepsinL,transcription factor Dendrinon,membrane-associated guanylate kinase inverted 2,synaptopodin and RhoA as well as the distribution of cytoskeletal protein actin and intermediate filament in the podocyte. Methods: Mouse podocytes were challenged with puromycin aminonucleoside(PAN)alone in the present of dexamethasone or serum containing traditional Chinese drugs at 10％ or 20％ concentration respectively for 12h. The protein expression of MAGI-2, synaptopodin, CatL and RhoA was quantitatively measured by western blotting, and the mRNA expression of CatL and synaptopodin were detected by fluorescence quantitative PCR. Indirect cell immunofluorescence staining was used to display the location of dendrin in podocytes. FITC-conjugated phalloidinand Cy3-conjugated rabbit-anti-vimentin were used to observe the distribution of F-actin and intermediate filament. Results: Afterp PAN-induced podocyte injury for 12 hours, the percentage of nuclear translocation of dendrin was elevated (compared with that of normal group, P ＜ 0. 01). The protein expression of MAGI-2, RhoA, and synaptopodin were decreased significantly (compared with normal group,all P ＜ 0. 01). However,there was no change in the mR- NA expression of synaptopodin (compared with model group, P ＞ 0. 05). The mRNA and protein expression of CatL were significantly increased (compared with normal control group, all P ＜ 0. 01). Podocyte pseudopodia were retracted and shortened; the structure and arrangement of microfilament and intermediate filament were almost completely disrupted. After treatment with dexamethasone, 10％ and 20％ of Chinese medicine,the mRNA and protein expression of CatL were decreased(compared with model group,all P ＜ 0. 01). The protein expression of MAGI-2, RhoA, and synaptopodinwere increased (compared with model group, all P ＜ 0. 01). The percentage of nuclear translocation of dendrin was declined(compared with model group, P ＜ 0. 01). The cytoskeleton structure was restored in each treatment group. Conclusion: Wenyang-Huoxue-Lishui formula is able to upregulate the expression of MAGI-2, and then to decrease nuclear import of dendrin, in turn todecrease CatL level, consequently to increase synaptopodin and RhoAandstabilize the cytoskeletal structure, ultimately to decrease foot process fusion and proteinuria.