TF-1细胞增殖特性与信号蛋白JAK2/STAT3表达研究

目的 探索JAK／STAT途径活化与细胞增殖和凋亡之间的关系。方法 通过表达内源性GM-CSF、受体的TF-1细胞，观察细胞增殖与凋亡；建立免疫沉淀和Western blot印迹方法，检测经GM-CSF100 ng／ml瞬时诱导的TF-1细胞信号蛋白JAK2和STAT3酪氨酸磷酸化情况。结果 经过GM-CSF刺激，细胞不但免于凋亡，TF-1细胞还呈现剂量与时间信赖性的增殖反应。当耗竭细胞因子6～12h后，倒置显微镜下TF-1细胞呈现折光性改变，细胞开始凋亡，呈现凋亡特征性形态改变和出现DNA梯形条带，而加入0．1ng／ml的GM-CSF，TF-1细胞即开始进入增殖状态，提示GM-CSF对细胞具有凋亡保护作用。经过100ng／ml GM-CSF的瞬时诱导，在分子量130kd区域见到JAK2蛋白条带，显示GM-CSF可诱导TF-1细胞磷酸化JAK2表达；而在分子量90kd区域未见到STAT3蛋白条带，显示无磷酸化STAT3表达；而未经GM-CSF诱导的TF-1细胞，既无磷酸化JAK2也无磷酸化STAT3表达。结论 GM-CSF、为TF-1细胞增殖诱导和凋亡保护所必需；GM-CSF诱导的TF-1细胞生物学效应涉及到胞浆信号分子JAK2磷酸化而非STAT3磷酸化。

The Proliferation and Expression of Signal Molecules JAK2/STAT3 in TF-1 Cell Line

Objective To investigate the relationship between the activated JAK/STAT proteins and the proliferation of myeloid leukemic cells.Methods In human myeloid TF 1 cell line expressing the endogenous GM CSF receptor,immunoprecipitation and Western blot analysis were used to determine if there ware tyrosine phosphorylated JAK2 and STAT3 stimulated by GM CSF;Simultaneously the apoptotic and proliferative status was observed on the due time. Results In TF 1 cell line,the activation of JAK2 but not STAT3 was detected induced by GM CSF;Moreover this cell line was observed to proliferate in a dose and time dependent manner induced by GM CSF;In addition,GM CSF could protect TF 1 cells from apoptosis demonstrated by morphological alteration and DNA fragmentization. Conclusion GM CSF was essential for the protection of cells from apoptosis and the promotion of proliferation of TF 1 cells; Moreover,GM CSF induced biological effect was associated with the activation of JAK2; In addition,activated JAK/STAT signal pathway may be involved in the apoptosis and proliferation,which further confirmed the autocrine or paracrine mechanism of leukemogenesis.