PTEN对SMMC—7721人肝癌细胞迁移增殖及粘着斑激酶磷酸化水平的影响

抑癌基因PTEN编码产物具有双专一性磷酸酶活性 ,并与细胞骨架张力蛋白同源。PTEN可参与粘着斑的形成和解聚而影响细胞迁移。现用PTEN表达质粒转染SMMC 772 1肝癌细胞 ,研究SMMC 772 1细胞运动能力的变化及PTEN与粘着斑激酶 (FAK)酪氨酸磷酸化水平之间的关系。PTEN过表达能够显著抑制细胞在Fn基质上的活动 :细胞在Fn基质上的迁移下降了 35 % ;在 30min和 6 0min两个时间点 ,Fn基质上细胞铺展分别降低了 2 9%和 2 6 % ;而在多聚赖氨酸基质上细胞铺展并没有变化。运用免疫沉淀和Western印迹方法 ,分析FAK及其酪氨酸磷酸化水平 ,发现PTEN过表达不影响FAK表达 ,但显著降低Fn诱导的FAK酪氨酸磷酸化水平 ,两者水平呈负相关。流式细胞仪分析细胞周期结果表明 ,PTEN抑制细胞 ,S期细胞下降了 16 %。上述结果提示 ,PTEN抑制肝癌细胞迁移铺展和增殖 ;PTEN对细胞运动的影响可能通过调节FAK酪氨酸磷酸化水平而实现。

The Effects of PTEN Gene on Migration and FAK Phosphorylation of SMMC-7721 Human Hepatocarcinoma Cell Line

PTEN is a major tumor suppressor gene that encodes a dual-specificity phosphatase with high sequence similarity to the cytoskeletal protein tensin. PTEN may be involved in the formation and disassembly of focal adhesion and affect cell migration. In the present study, PTEN expression plasmid was constructed and transfected into the hepatoma cell line SMMC-7721 to analyze the alterations of cell motility and FAK tyrosine phosphorylation. It was observed that the overexpression of PTEN gene significantly inhibited cell motility on extracellular matrix (Fn), and the cell migration on fibronectin was reduced by 35%. Similarly, at 30-min and 60-min, the cell spreading on Fn but not on polylysine was inhibited by 29% and 26% respectively. The data obtained from immunoprecipitation and immunoblotting analyses showed that the overexpression of PTEN did not affect FAK expression but resulted in a decrease in FAK tyrosine phosphorylation. The level of FAK phosphorylation was inversely correlated with the level of PTEN protein in three cell lines. It was also found that the overexpression of PTEN led to growth inhibition, with the number of cells in S phase reduced by 16%. These results indicate that PTEN exerts its tumor-suppressive effects on hepatocellular carcinoma cells through the inhibition of cell motility and cell cycle progression.