抑癌蛋白PTEN在雌激素和胰岛素样生长因子1诱导子宫内膜癌细胞ERK活化中的作用

背景与目的:雌激素和胰岛素样生长因子1(insulinlikegrowthfactor1,IGF-1)在子宫内膜癌细胞中的信号转导机制尚不清楚。本文拟研究雌激素和IGF-1在子宫内膜癌细胞信号转导中激活细胞外信号调节激酶(extracellularsignal-regulatedkinase,ERK)的情况,探讨与细胞骨架蛋白同源的磷酸酶(phosphataseandtensinhomologue,PTEN)在雌激素和IGF-1诱导的ERKs活化中的作用。方法:构建野生型PTEN和突变型PTEN(G129E)的逆转录病毒载体,体外转染Ishikawa细胞,用G418筛选稳定表达的细胞系。Westernblot检测PTEN基因在Ishikawa细胞中的表达,并检测和观察17-β-雌二醇和IGF-1对细胞系Ishikawa-EGFP(对照)、Ishikawa-PTEN和Ishikawa-PTEN(G129E)ERKs活化的浓度效应和时相特点,以及17-β-雌二醇在瞬时转染pCXN2hERα和pCXN2hERβ的NIH3T3细胞中激活ERK的情况。结果:IGF-1可激活ERK,以ERK2为主。不同浓度IGF-1作用于Ishikawa-EGFP和Ishikawa-PTEN时,对ERK2活化无明显区别。40μg/L的IGF-1作用显示5min时ERK2活化最明显。PTEN在IGF-1作用Ishikawa30min、2h和24h时均能抑制ERKs活化,而在5min时没有明显作用。40μg/LIGF-1刺激Ishikawa-PTEN(G129E)5min,和Ishikawa-EGFP相比,无明显改变。17-β-雌二醇可激活ERK,以ERK2为主。不

Role of suppressor encoprotein PTEN in IGF-1 induced activation of ERK in endometrial carcinoma cells

BACKGROUND & OBJECTIVES: The machanism of signal transduction of insulin-like growth factor (IGF-1) in endometrial carcinoma is still unknow. The objective of this paper was to study extracellular signal-regulated kinase(ERK) activation in endometrial carcinoma cell line Ishikawa under the stimulation of IGF-1, and to elucidate the role of suppressor encoprotein phosphatase and tensin homologue(PTEN) in activation of ERK. METHODS: Retrovirus vector of PTEN and PTEN (G129E) was constructed. Ishikawa was transfected in vitro. Expressionstable cell line was screened by G418. Western blot was applied to examine PTEN expression in Ishikawa cells after transfection. Optimal concentration and time of IGF-1 and 17-beta-estrodial which activated ERK in Ishikawa-PTEN and Ishikawa-PTEN(G129E) cells were detected. Western blot was applied to examine ERK activation under the stimulation of 17-beta-estrodial in NIH3T3 fibroblasts after transient transfection of pCXN2hER alpha and pCXN2hER beta. RESULTS: IGF-1 activated ERK cascades(mainly ERK2) in Ishikawa cells. There was no obvious difference in ERK activation among different doses of IGF-1 (20, 40, and 80 micrograms/L), But the maximal activations of ERK2 took place at 5 min after stimulation with IGF-1. The activation of ERK2 was inhibited obviously by PTEN at 30 min, 2 h, and 24 h. There was no obvious difference in ERK activation between Ishikawa-PTEN(G129E) and Ishikawa-EGFP. 17-beta-estrodial activated ERK cascades (mainly ERK2) in Ishikawa cells. The activation of ERK was increased with increasing of concentration. The maximal activations of ERK2 took place at 5 min after stimulation with 17-beta-estrodial. The activation of ERK2 was inhibited obviously by PTEN, not by PTEN(G129E). 17-beta-estrodial activated ERKs cascades in NIH3T3 fibroblasts after transient transduction of pCXN2h-ER alpha. CONCLUSIONS: 17-beta-estrodial and IGF-1 activated ERK cascades in Ishikawa cells. Lipid phosphatase of PTEN had an inhibitory role in the activation of ERK under the stimulation of 17-beta-estrodial and IGF-1.