摘要
背景与目的:蛋白激酶FAK部分介导肿瘤细胞在悬浮培养下细胞积聚体的形成,从而阻止细胞发生凋亡和促进细胞增殖,但是参与细胞积聚体形成的FAK下游通路尚不清楚。本研究旨在研究蛋白激酶FAK及其可能的下游分子ERK、AKT和SRC活化在悬浮状态肺癌GLC-82细胞积聚中的作用。方法:用polyHEMA悬浮培养观察肺正常细胞HBE和肿瘤细胞GLC-82积聚体形态;细胞凋亡用DNA琼脂糖凝胶电泳检测梯状DNA分析;细胞转化能力用软琼脂糖集落形成实验分析;磷酸化蛋白激酶水平用免疫印迹实验检测;RNA干扰实验降低FAK蛋白水平的表达。结果:GLC-82肺腺癌细胞在polyHEMA悬浮状态下能够存活和形成积聚体,肺正常细胞HBE在polyHEMA悬浮状态下发生凋亡和不形成积聚体。磷酸化FAK、ERK、AKT和SRC的水平和GLC-82积聚体有关。沉默FAK蛋白的表达,能够部分地阻断肿瘤细胞积聚体的形成,降低磷酸化ERK和AKT水平,以及降低软琼脂集落形成能力。FAKRNA干扰前后的GLC-82细胞软琼脂集落形成率分别为(12.2±1.1)%和(3.6±0.7)%(P=0.001)。结论:GLC-82肺癌细胞积聚体的形成部分由FAK信号通路介导,ERK和AKT是FAK的下游通路蛋白。
BACKGROUND ＆ OBJECTIVE： Our previous study revealed that tyrosine kinase FAK can partly mediate the aggregation of tumor cells in suspension culture, therefore, suppress cell apoptosis and promote cell proliferation. However, the downstream pathway of FAK in the formation of cell aggregation is unclear. This study was to investigate the roles of FAK and its potential downstream molecules ERK, AKT, and SRC in mediating aggregation of lung adenocarcinoma GLC-82 cells in suspension culture. METHODS： Morphology of the aggregations of GLC-82 cells and normal lung HBE cells in polyHEMA suspension culture was observed. Cell apoptosis was studied by DNA electrophoresis on agarose gel. The ability of cell transformation was studied by colony formation assay. The phosphorylation of the protein kinases was investigated by Western blot. FAK was silenced by RNA interference. RESULTS： GLC-82 cells survived and aggregated in suspension culture; while apoptosis occurred in HBE cells which formed no aggregation in suspension culture. Phosphorylation levels of FAK, ERK, AKT, and SRC were related to GLC-82 cell aggregation. FAK silencing partly blocked the formation of GLC-82 cell aggregation, and decreased the phosphorylation levels of ERK and AKT; the colony formation rate in soft agar was significantly lower in GLC-82 cells with FAK silencing than in GLC-82 cells without FAK silencing E（12.2±1.1 ）% vs. （3.6±0.7）%, P=0.001 ]. CONCLUSION： The formation of GLC-82 cell aggregation is partly mediated by FAK signal pathway, and ERK and AKT are the downstream molecules of FAK.
出处
《癌症》
SCIE
CAS
CSCD
北大核心
2005年第10期1206-1212,共7页
Chinese Journal of Cancer
基金
国家自然科学基金(No.30371621)
广东省科技计划项目(No.2004-139)~~
