Jin Yi, Hu Jian-Li, Xiao Lan, Cui Wen
Center of Oncology, Union Hospital, Huazhong University of Science and Technology, Wuhan, Hubei, 430022, P. R. China.
Ai Zheng. 2008 Dec;27(12):1239-43.
BACKGROUND & OBJECTIVE: PTEN plays a critical role in the development and progression of many cancers. PTEN may act directly or indirectly to integrate complex signal network system, to influence the downstream signal pathway and target molecules. This study was to investigate the impact of PTEN gene knockout in human breast carcinoma MCF-7 cells on the activity of JNK pathway.
MCF-7 cells were transfected with PTEN antisense oligonucleotide, or treated with 10 micromol/L SP600125, or received both transfection and SP600125 treatment. The expression of PTEN protein was detected by confocal spectral microscopy. The early stage cell apoptosis and cell cycle were detected by flow cytometry (FCM). The proliferation of MCF-7 cells was determined by MTT assay. The phosphorylated JNK and downstream substrate ATF-2, C-Jun protein were assayed by Western blot.
The expression of PTEN protein in MCF-7 cells was effectively blocked by PTEN antisense oligonucleotide. Early apoptosis of MCF-7 cells was induced by SP600125 after knocking down PTEN, with an early apoptosis rate of (32.4+/-2.4)%; MCF-7 cells were arrested at G1 phase. The proliferation rate was significantly lower in combination group than in SP600125 group and antisense oligonucleotide group (P < 0.05). The phosphorylation levels of JNK, ATF-2 and C-Jun was down-regulated in combination group.
PTEN plays an important role in the activation of JNK pathway in MCF-7 cells. Loss of PTEN results in the activation of JNK pathway in MCF-7 cells, and improves sensitivity of MCF-7 cells to JNK inhibitor.
PTEN在多种癌症的发生发展过程中发挥关键作用。PTEN可能直接或间接作用于整合复杂的信号网络系统，影响下游信号通路及靶分子。本研究旨在探讨人乳腺癌MCF-7细胞中PTEN基因敲除对JNK通路活性的影响。
用PTEN反义寡核苷酸转染MCF-7细胞，或用10 μmol/L SP600125处理，或同时进行转染和SP600125处理。采用共聚焦光谱显微镜检测PTEN蛋白表达。用流式细胞术（FCM）检测早期细胞凋亡及细胞周期。采用MTT法测定MCF-7细胞的增殖情况。用蛋白质免疫印迹法检测磷酸化JNK及下游底物ATF-2、C-Jun蛋白。
PTEN反义寡核苷酸有效阻断了MCF-7细胞中PTEN蛋白的表达。敲低PTEN后，SP600125诱导MCF-7细胞早期凋亡，早期凋亡率为（32.4±2.4）%；MCF-7细胞停滞于G1期。联合组的增殖率显著低于SP600125组和反义寡核苷酸组（P<0.05）。联合组中JNK、ATF-2和C-Jun的磷酸化水平下调。
PTEN在MCF-7细胞JNK通路激活中起重要作用。PTEN缺失导致MCF-7细胞JNK通路激活，并提高MCF-7细胞对JNK抑制剂的敏感性。
