摘要
目的探讨白血病抑制因子(LIF)受体α亚基胞内区在细胞内游离存在时,对HL-60细胞增殖分化的调节及相关信号转导机制。方法鉴定已构建的重组质粒pCDNA3.0-190CT2+3和PCDNA3.0-190CT3;用脂质体将质粒转染HL-60细胞,细胞培养基中加入G-418来筛选阳性克隆。用免疫细胞化学和免疫印迹法检测转染组细胞中目的蛋白的表达。绘制细胞生长曲线。免疫印迹法检测各组细胞增殖细胞核抗原(PCNA)的表达和信号分子STAT3的磷酸化水平。结果免疫印迹法检测及目的蛋白条带,获得稳定表达目的蛋白的细胞株。与野生组细胞相比,190CT2+3和190CT3组的细胞体积增大,分叶核细胞占总细胞数的比例增加,细胞增殖减慢,PCNA的表达减少,STAT3的磷酸化水平升高。结论LIF受体α亚基胞内区游离片段促进了HL-60细胞的分化,抑制了增殖,激活了信号分子STAT3。
Objective ： To investigate the regulatory effect of dissociative intracytoplasmic domain of leukemia inhibitory factor receptor α （LIFRα） subunit on the growth of HL-60 cells and the related signal transduction mechanism. Methods： The constructed recombinant plasmids pcDNA3.0-190CT2 ＋ 3 and cDNA3.0-190CT3 were identified and transfected into HL-60 cells with liposome FuGene-6. Then the cells were cultured in a medium containing G-418 and the positive clones were selected. Immunohistochemical method and Western blotting were used to detect the expression of the target protein in HL-60 cells and the growth curve of the cells was plotted. The expression of proliferating cell nuclear antigen （PCNA） and the phosphorylation levels of signal transducer and activator of transcription 3 （Stat3） were assayed by Western blotting. Results： Western blotting detected the band of target protein and the cell line stably expressing the target protein was obtained. Compared with that of the wild type group, the cell sizes in 190CT2＋3 and 190CT3 group were enlarged. The ratio of leaf cell number to the total cell number increased and the cell proliferation was slowed down. We also found that the expression of PCNA was decreased and the phosphorylation level of STAT3 was increased. Conclusion： The dissociative intracytoplasmic domain of LIFRa subunit can accelerate the differentiation of HL-60 cells, inhibit the proliferation of HL-60 cells, and activate signal molecule STAT3.
出处
《第二军医大学学报》
CAS
CSCD
北大核心
2006年第12期1277-1280,共4页
Academic Journal of Second Military Medical University
基金
国家自然科学基金(30170468)~~
关键词
白血病抑制因子
受体
增殖细胞核抗原
信号转导
STAT3
leukemia inhibitory factor
receptor
proliferating cell nuclear antigen
signal transduction
STAT3
