EGF和AngⅡ激活人肾上腺皮质癌H295R细胞ERK通路

Activation of ERK pathway by EGF and angiotensin Ⅱ in adrenocortical carcinoma H295R cells

目的研究表皮生长因子(EGF)和血管紧张素Ⅱ(AngⅡ)联合作用对肾上腺皮质癌H295R细胞的细胞外信号调节蛋白激酶(ERK)信号通路的影响。方法用不同剂量的EGF和AngⅡ刺激H295R细胞,或用特异性EGF受体酪氨酸激酶抑制剂AG1478预处理后再加入AngⅡ刺激细胞,用Western blot法检测磷酸化ERK1/2;EGF及AngⅡ单独或联合刺激细胞,检测ERK1/2、P90RSK和Bad的磷酸化水平。结果 EGF和AngⅡ均刺激细胞ERK1/2和P90RSK磷酸化,在刺激ERK1/2磷酸化上呈剂量依赖性,两者联合处理其激活ERK1/2和P90RSK的作用产生叠加,AngⅡ和EGF单独及联合处理细胞,ERK1/2磷酸化水平分别是对照组的6.3±1.4、2.2±0.6及10.1±1.1倍。AngⅡ刺激细胞Bad磷酸化,而EGF无此作用。AG1478不能阻断AngⅡ诱导的Erk1/2激活。结论EGF和AngⅡ明显激活H295R细胞的ERK信号通路,两者作用有叠加效应。

Objective To find potential effects of EGF and angiotensinⅡ on ERK pathways in adrenocortical carcinoma H295R Cells.Methods Treated with EGF and angiotensin Ⅱ with different concentrations,phosphorylation of ERK1/2 was examined by Western blot.The effect of Angiotensin Ⅱ on phospho-ERK1/2 was also observed when pretreated with the specific inhibitor of the EGF receptor tyrosine kinase,AG1478.Treated with EGF and angiotensin Ⅱ alone or both,phosphorylation of ERK1/2,P90RSK and Bad was detected.Results Both EGF and angiotensin Ⅱ increased phospho-ERK1/2 in a dose-dependent manner,and they also promoted phospho-P90RSK.EGF and angiotensin Ⅱ showed an additive effect on phosphorylation of ERK1/2 and P90RSK.Treated by Angiotensin Ⅱ and EGF separately or both,phospho-ERK1/2 were 6.3±1.4、2.2±0.6 and 10.1±1.1,respectively,relatived to the control.Angiotensin Ⅱ,but not EGF,stimulated phospho-Bad.The effects of angiotensin Ⅱ on phospho-ERK1/2 was not blocked by AG1478.Conclusions Both EGF and Angiotensin Ⅱ promotes ERK pathway with an additive effect in H295R cells.