抑制Rho—kinase在柔红霉素诱导的肾小球硬化中的作用及机制

目的：研究抑制Rho—kinase对柔红霉素诱导的肾小球硬化的调控作用，并探讨Rho—kinase抑制剂法舒地尔改善肾小球硬化的作用机制。方法：36只雄性SD大鼠，随机分为三组：假手术（Sham）组，单侧肾脏切除＋柔红霉素（模型）组，单侧肾脏切除＋柔红霉素＋法舒地尔（干预）组，每组各12只。模型组和干预组大鼠在切除左肾后第7、14天，从尾静脉各注射柔红霉素5mg／kg 1次，同时Sham组大鼠以等剂量的生理盐水尾静脉注射。干预组在第2次注射柔红霉素后从腹腔每天注射法舒地尔3mg／kg，完成上述处理后的第2、4周，随机取各组大鼠6只处死留取肾标本，处死前收集24h尿液检测尿蛋白排泄，用HE，PAS染色，免疫组化，透射电镜进行肾组织病理学分析，应用RT-PCR检测Rho—kinase，P27的核酸表达水平。结果：（1）模型组较Sham组24h尿蛋白明显升高（P〈0．01），而法舒地尔干预后，干预组24h尿蛋白较模型组显著降低（P〈0.05）。（2）模型组大鼠足细胞第4周时出现了弥漫性足突融合，系膜基质增殖明显，出现典型的肾小球节段性硬化，免疫组化PCNA表达升高，P27表达下降；而干预组可以改善柔红霉素诱导的肾小球硬化大鼠足细胞的病理改变，系膜细胞增生受抑制，系膜基质蓄积减少，较模型组PCNA表达下降，P27升高。（3）模型组较Sham组Rho—kinase的mRNA表达升高，细胞周期抑制蛋白P27 mRNA的表达下降，而干预组较模型组Rho—kinase的mRNA表达显著降低，P27 mRNA的表达明显升高。结论：（1）抑制Rho—kinase的表达能明显改善柔红霉素诱导的大鼠的肾小球硬化。（2）法舒地尔的作用机制可能是通过改善足细胞的病理改变，减少蛋白尿，升高细胞周期抑制蛋白P27的表达，从而抑制系膜基质增殖，延缓肾小球硬化的进展。

Inhibitition of Rho-kinase Retards the Daunorubicin Induced Progressive Glomerulosclerosis

Objective：Objective：To study the effects of inhibiting the Rho- kinase in Daunorubicin induced progressive glomerulosclerosis and to explore the mechanism of Fasudil on ameliorating glomerulosclerosis. Methods：Thirty six male SD rats were randomly allocated into sham operation group （Sham, n = 12）, unilateral nephrectomy（UNX） ＋ daunorubicin（DRB） group （Model,n = 12）, UNX＋ DRB ＋ Fasudil group （Treat, n = 12）. The rats in Model group and Treat group were injected into rat＇s tail with DRB（5 mg/kg） on day 7 and day 14 after UNX. The rats in Treat group were administrated with Fasudil （3 mg.kg^-1.d^-1） intraperitoneally, At week 2,4 respectively following animal model, 6 rats in each group were taken randomly for determining 24 hour urine protein excretion,sections of kidneys were examined by HE and PAS staining, immunohistochemistry and transmission electric microscope （TEM）. The expression of Rho- kinase mRNA and P27 mRNA in kidney were examinedby RT - PCR. Results： （1）The 24 hour urine excretion in Model group were increasing marldy when compared with Sham group（ P 〈0.01 ）. but this increase was significantly suppressed by fasudil （P 〈 0.05）. （2） At 4 week, the foot process effacement in podocytes , mesangial pro- liferation and ECM accumulation, showing focal segmental glomerulosclerosis in Model group. But in Treat group, the fasudil improved glomerular injury scores with parallel amelioration of proliferating cell nuclear antigen- positive cell infiltration and increased the expression of P27, a cyclin- dependent kinase inhibitor. （3） The expression of Rho- kinase mRNA was rernarkablely enhanced in Model group and was suppressed in Treat group. Finally, fasudil up - regulated the mRNA expression of p27. Conclusion： （ 1 ）The glomerulosclerosis was markly ameliorated by inhibititing the expression of Rho- kinase. （2） Rho- kinase pathway is involved in the pathogenesis of renal injury. Furthermore, the inhibition of Rho- kinase may co