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作 者:张挺[1] 徐东亮[1] 周强平[1] 鲁佩[1] 殷长军[1] 张炜[1] 徐正铨[1] 顾民[1]
机构地区:[1]南京医科大学第一附属医院泌尿外科肾移植中心,江苏南京210029
出 处:《南京医科大学学报(自然科学版)》2011年第6期818-823,共6页Journal of Nanjing Medical University(Natural Sciences)
摘 要:目的:探讨骨桥蛋白(osteopontin,OPN)的shRNA干扰质粒在大鼠慢性移植肾肾病(chronic allograft nephropathy,CAN)模型中对大鼠移植肾上皮细胞向间充质转分化(epithelial to mesenchymal transition,EMT)的作用,初步探讨OPN在慢性移植物肾病中的作用及机制。方法:以F344大鼠为供体,Lewis大鼠为受体,行原位异体肾移植术,建立大鼠CAN模型(Allo组),以Lewis大鼠做供、受体作为同系移植对照(Syn组),以基于流体力学为基础的肾脏基因转染方法转染质粒到移植肾(RNAi组),采用空质粒作为对照(EV组),于术后12周处死各组动物,取移植肾标本进行HE和马松染色,观察组织形态学变化。采用免疫组织化学方法对移植肾组织α-平滑肌肌动蛋白(α-SMA)、E-钙黏蛋白(E-Cadherin)的表达进行观察。同时行Western blot检测移植肾组织OPN的表达,并对其灰度比作统计分析。结果:RNAi组OPN表达显著低于Allo组。Allo组和EV组大鼠移植肾组织形态学符合慢性移植肾肾病改变。RNAi组移植肾形态学变化较Allo组减轻。免疫组化结果显示,与Syn组相比,Allo组主要在肾小管上皮及肾间质强表达α-SMA,而肾小管上皮E-Cadherin表达较Syn组明显下降。与之相比,RNAi组α-SMA表达下调,E-Cadherin表达上调。结论:OPN在大鼠CAN模型移植肾中表达上调,OPN体内RNA干扰能够减少OPN在移植肾的表达,并且OPN的RNA干扰能缓解移植肾的病理学改变,抑制移植肾小管上皮细胞EMT。提示OPN与CAN的发生有潜在相关性,其可能通过促进肾小管上皮发生EMT途径影响CAN发病。Objective:To investigate the effect of in vivo shRNA plasmid vector gene transfer to silence OPN expression upon ep-ithelial to mesenchymal transition of the tubular epithelial cells(TECs)in rat chronic allograft nephropathy(CAN)models,and to explore the possible role of OPN in CAN.Methods:Orthotopic renal-transplantation using F344 rats as donors and Lewis rats as re-cipients was operated to establish Allo group,while transplantation from Lewis to Lewis rats as Syn group.Hydromechanics based gene transfer method was used to establish RNAi and EV groups,for which OPN-shRNA or empty vector was injected into the renal vein.Rats in each group were sacrificed 12 weeks after the surgery.Allograft samples were collected and sectioned for HE staining,masson’s trichrome staining,immunohistochemical study and Western blot assay.Results:The morphological changes of allograft in Allo /EV group consist with CAN.Compared with the Syn group,Western blot showed a significant rise of OPN expression.In the RNAi group,the OPN expression was significantly reduced(P 〈 0.05)compared with Allo /EV group.The RNA interference inhibit-ed EMT of TECs and the pathogenesis of CAN,which were confirmed by immunohistochemistry and HE /Masson’s trichrome lightscopy.Conclusion:The OPN expression in rat CAN model was significantly up-regulated.The in vivo RNA interference inhibited the OPN expression.Inhibition of OPN can protect the allograft from pathogenesis of EMT and IF /TA in CAN.
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