中华病理学杂志
中華病理學雜誌
중화병이학잡지
Chinese Journal of Pathology
2015年
11期
778-782
,共5页
黄平%张月娟%蒋涛%张农
黃平%張月娟%蔣濤%張農
황평%장월연%장도%장농
醛还原酶%转化生长因子β1%纤连蛋白%肾小球系膜细胞
醛還原酶%轉化生長因子β1%纖連蛋白%腎小毬繫膜細胞
철환원매%전화생장인자β1%섬련단백%신소구계막세포
Aldehyde reductase%Transforming growth factor beta1%Fibronectins%Mesangial cells
目的 探讨p38 MAPK信号通路与醛糖还原酶(AR)对转化生长因子β1(TGF-β1)诱导人系膜细胞(HMC)细胞外基质成分纤连蛋白(FN)表达的影响.方法 应用醛糖还原酶抑制剂和p38 MAPK信号通路抑制剂、转染及RNA干扰技术分别作用于HMC后,再用TGF-β1刺激,观察刺激前后HMC表达FN的情况.Western blot和免疫荧光检测HMC的FN、AR、p38的变化及real-time聚合酶链反应(PCR)鉴定转染和RNA干扰效果.结果 HMC在TGF-β1作用后,AR和FN蛋白表达升高;单独使用醛糖还原酶抑制剂Sorbinil、Zopolrestat并不能下调FN蛋白的表达;但预先使用Sorbinil或Zopolrestat孵育后,再用TGF-β1刺激,FN蛋白表达下降(P＜0.05);单独使用p38 MAPK信号通路抑制剂后,FN蛋白表达下降;各组细胞预先使用Sorbinil、Zopolrestat、SB203580孵育后,再用TGF-β1刺激,FN蛋白表达下降(P＜0.05);转染pCDNA3-AR后,HMC中AR mRNA及FN蛋白表达增多,再用TGF-β1刺激,FN蛋白表达明显上调(P＜0.05);用AR-siRNA干扰AR基因后,HMC中AR mRNA及FN蛋白表达减少,即使用TGF-β1刺激,FN蛋白表达仍然下降(P＜0.05).结论 AR基因作为TGF-β1反应性基因,参与了HMC中TGF-β1诱导的FN表达的调控,p38 MAPK信号通路在这一过程中起着重要作用.
目的 探討p38 MAPK信號通路與醛糖還原酶(AR)對轉化生長因子β1(TGF-β1)誘導人繫膜細胞(HMC)細胞外基質成分纖連蛋白(FN)錶達的影響.方法 應用醛糖還原酶抑製劑和p38 MAPK信號通路抑製劑、轉染及RNA榦擾技術分彆作用于HMC後,再用TGF-β1刺激,觀察刺激前後HMC錶達FN的情況.Western blot和免疫熒光檢測HMC的FN、AR、p38的變化及real-time聚閤酶鏈反應(PCR)鑒定轉染和RNA榦擾效果.結果 HMC在TGF-β1作用後,AR和FN蛋白錶達升高;單獨使用醛糖還原酶抑製劑Sorbinil、Zopolrestat併不能下調FN蛋白的錶達;但預先使用Sorbinil或Zopolrestat孵育後,再用TGF-β1刺激,FN蛋白錶達下降(P＜0.05);單獨使用p38 MAPK信號通路抑製劑後,FN蛋白錶達下降;各組細胞預先使用Sorbinil、Zopolrestat、SB203580孵育後,再用TGF-β1刺激,FN蛋白錶達下降(P＜0.05);轉染pCDNA3-AR後,HMC中AR mRNA及FN蛋白錶達增多,再用TGF-β1刺激,FN蛋白錶達明顯上調(P＜0.05);用AR-siRNA榦擾AR基因後,HMC中AR mRNA及FN蛋白錶達減少,即使用TGF-β1刺激,FN蛋白錶達仍然下降(P＜0.05).結論 AR基因作為TGF-β1反應性基因,參與瞭HMC中TGF-β1誘導的FN錶達的調控,p38 MAPK信號通路在這一過程中起著重要作用.
목적 탐토p38 MAPK신호통로여철당환원매(AR)대전화생장인자β1(TGF-β1)유도인계막세포(HMC)세포외기질성분섬련단백(FN)표체적영향.방법 응용철당환원매억제제화p38 MAPK신호통로억제제、전염급RNA간우기술분별작용우HMC후,재용TGF-β1자격,관찰자격전후HMC표체FN적정황.Western blot화면역형광검측HMC적FN、AR、p38적변화급real-time취합매련반응(PCR)감정전염화RNA간우효과.결과 HMC재TGF-β1작용후,AR화FN단백표체승고;단독사용철당환원매억제제Sorbinil、Zopolrestat병불능하조FN단백적표체;단예선사용Sorbinil혹Zopolrestat부육후,재용TGF-β1자격,FN단백표체하강(P＜0.05);단독사용p38 MAPK신호통로억제제후,FN단백표체하강;각조세포예선사용Sorbinil、Zopolrestat、SB203580부육후,재용TGF-β1자격,FN단백표체하강(P＜0.05);전염pCDNA3-AR후,HMC중AR mRNA급FN단백표체증다,재용TGF-β1자격,FN단백표체명현상조(P＜0.05);용AR-siRNA간우AR기인후,HMC중AR mRNA급FN단백표체감소,즉사용TGF-β1자격,FN단백표체잉연하강(P＜0.05).결론 AR기인작위TGF-β1반응성기인,삼여료HMC중TGF-β1유도적FN표체적조공,p38 MAPK신호통로재저일과정중기착중요작용.
Objective To study the effects of p38 MAPK signaling pathway and aldose reductase (AR) on the transforming growth factor (TGF)-31-induced expression of fibronectin (FN).Methods Human mesangial cells (HMCs) were cultured, and transfected with pCDNA3-AR.AR gene silencing was induced by small interfering RNA (siRNA).AR expression in HMCs was examined by immunofluorescence analysis.RT-PCR and real-time PCR were performed to detect the mRNA expression of AR in the HMCs and Western blotting was used to detect the protein expression of AR, FN and p38.AR inhibitors (ARIs),Sorbinil and Zopolrestat were added and co-incubated, followed by addition of TGFq1.Western blotting was used to document protein expression of FN and p38 mitogen-activated protein kinases (p38 MAPKs) in the HMCs.Results Immunofluorescence analysis showed a stronger expression of AR in HMCs transfected with AR than that of normal HMCs and HMCs transfected with blank vector.In comparison with normal HMCs and those transfected with blank vector, HMCs transfected with AR showed stronger protein expression of FN (P ＜ 0.05).After incubation of ARIs, protein expression of FN decreased in HMCs transfected with AR (P ＜ 0.05).Mter stimulation of TGF-β1, FN protein expression increased in both normal HMCs and those transfected with AR (P ＜ 0.05).Mter preincubation with ARI, FN protein expression in HMCs transfected decreased significantly (P ＜0.05).Mter stimulation of TGF-β1, nave HMCs showed increased expression of phosphor-p38.In contrast, HMCs preincubated with ARIs showed reduced expression of phosphor-p38, and HMCs transfected with AR showed increased expression of phosphor-p38 (P ＜ 0.05).Conclusions AR regulates the expression of FN through the stimulation of TGF-β1, which may involve the activation of p38-MAPK signaling pathway.AR may play a role in the pathogenesis of glomerulosclerosis.
