中华肾脏病杂志
中華腎髒病雜誌
중화신장병잡지
2012年
6期
484-488
,共5页
曹陶%曹师荣%李辉雁%熊丽萍%范瑾瑾%余学清%毛海萍
曹陶%曹師榮%李輝雁%熊麗萍%範瑾瑾%餘學清%毛海萍
조도%조사영%리휘안%웅려평%범근근%여학청%모해평
热休克蛋白72%转化生长因子β1%肾小管上皮细胞%肾小管上皮细胞向间质细胞转分化
熱休剋蛋白72%轉化生長因子β1%腎小管上皮細胞%腎小管上皮細胞嚮間質細胞轉分化
열휴극단백72%전화생장인자β1%신소관상피세포%신소관상피세포향간질세포전분화
Heat shock protein 72%Transforming growth factor betal%Renal tubular epithelial cells%Epithelial to mesenchymal transition
目的 探讨热休克蛋白72肽结合区在肾小管上皮间质转分化(EMT)过程中的作用和可能机制.方法 应用质粒转染方法分别诱导热休克蛋白72(HSP72)野生型、肽结合区缺失型(HSP72-△PBD)和肽结合区(PBD)的表达.用转化生长因子β1(TGF-β1)刺激大鼠肾小管上皮细胞(NRK-52E)48 h,Western印迹和免疫荧光染色检测细胞E-钙黏蛋白(cadherin),α-平滑肌肌动蛋白(SMA),HSP72和Smad3/磷酸化(p)-Smad3蛋白表达.结果 TGF-β1(10 μg/L)刺激NRK-52E细胞48 h后上调α-SMA和下调E-cadherin蛋白表达水平.Western印迹及细胞免疫荧光显示,过表达HSP72和PBD能明显减轻TGF-β1诱导的NRK-52E细胞E-cadherin蛋白表达下调和α-SMA蛋白表达上调,而过表达HSP72-△PBD不能改变上述蛋白的表达.此外,过表达HSP72和PBD显著抑制Smad3的磷酸化.结论 HSP72抑制Smad3活化和EMT的发生可能与PBD的功能有关.
目的 探討熱休剋蛋白72肽結閤區在腎小管上皮間質轉分化(EMT)過程中的作用和可能機製.方法 應用質粒轉染方法分彆誘導熱休剋蛋白72(HSP72)野生型、肽結閤區缺失型(HSP72-△PBD)和肽結閤區(PBD)的錶達.用轉化生長因子β1(TGF-β1)刺激大鼠腎小管上皮細胞(NRK-52E)48 h,Western印跡和免疫熒光染色檢測細胞E-鈣黏蛋白(cadherin),α-平滑肌肌動蛋白(SMA),HSP72和Smad3/燐痠化(p)-Smad3蛋白錶達.結果 TGF-β1(10 μg/L)刺激NRK-52E細胞48 h後上調α-SMA和下調E-cadherin蛋白錶達水平.Western印跡及細胞免疫熒光顯示,過錶達HSP72和PBD能明顯減輕TGF-β1誘導的NRK-52E細胞E-cadherin蛋白錶達下調和α-SMA蛋白錶達上調,而過錶達HSP72-△PBD不能改變上述蛋白的錶達.此外,過錶達HSP72和PBD顯著抑製Smad3的燐痠化.結論 HSP72抑製Smad3活化和EMT的髮生可能與PBD的功能有關.
목적 탐토열휴극단백72태결합구재신소관상피간질전분화(EMT)과정중적작용화가능궤제.방법 응용질립전염방법분별유도열휴극단백72(HSP72)야생형、태결합구결실형(HSP72-△PBD)화태결합구(PBD)적표체.용전화생장인자β1(TGF-β1)자격대서신소관상피세포(NRK-52E)48 h,Western인적화면역형광염색검측세포E-개점단백(cadherin),α-평활기기동단백(SMA),HSP72화Smad3/린산화(p)-Smad3단백표체.결과 TGF-β1(10 μg/L)자격NRK-52E세포48 h후상조α-SMA화하조E-cadherin단백표체수평.Western인적급세포면역형광현시,과표체HSP72화PBD능명현감경TGF-β1유도적NRK-52E세포E-cadherin단백표체하조화α-SMA단백표체상조,이과표체HSP72-△PBD불능개변상술단백적표체.차외,과표체HSP72화PBD현저억제Smad3적린산화.결론 HSP72억제Smad3활화화EMT적발생가능여PBD적공능유관.
Objective To investigate the effects of peptide-binding domain (PBD) of heat shock protein (HSP) 72 on epithelial to mesenchymal transition (EMT) in rat renal tubular epithelial cells.Methods The expressions of wild-type HSP72,mutant of HSP72 lacking peptide binding domain (HSP72-△PBD) and HSP72-PBD were induced by plasmid transfection.NRK-52E ceils were stimulated by TGF-β1 for 48 h.The expressions of α-smooth muscle actin (α-SMA),E-cadherin,HSP72 and Smad3/p-Smad3 were detected by Western blot and immunofluorescence.Results After NRK-52E cells were stimulated by TGF-β 1 (10 μg/L) for 48 h,the expression of α-SMA was increased and the protein level of E-cadherin was decreased.Western blotting and immunofluorescence showed that over-expression of both HSP72 and PBD inhibited TGF-β1-induced up-regulation of protein α-SMA expression,down-regulation of protein E-cadherin.However,overexpression of HSP72-△PBD did not change the protein level of E-cadherin and α-SMA.In addition,over-expression of HSP72 and PBD significantly inhibited the phosphorylation of Smad3.Conclusion Inhibition of Smad3 activation and EMT by HSP72 is associated with the function of PBD.
