PTEN基因对胃癌细胞株SGC7901生物学行为及血管内皮生长因子和基质金属蛋白酶2及9表达的影响

目的探讨PTEN基因对胃癌细胞生物学行为的影响及其机制。方法脂质体转染并经免疫细胞化学、Western blot鉴定得到稳定高表达PTEN蛋白的SGC7901细胞克隆;细胞倍增时间测定、平板克隆形成试验及流式细胞术分析转染前后细胞增殖能力及细胞周期与凋亡率的变化;应用ELISA法、明胶酶谱分析、免疫细胞化学及Western blot等技术分别检测转染前后细胞中血管内皮生长因子（VEGF）、基质金属蛋白酶（MMP）-9与MMP-2蛋白的分泌与表达。结果成功地建立了稳定高表达PTEN蛋白的SGC7901细胞模型。PTEN—SGC7901细胞（PTEN基因转染细胞组）的倍增时间较SGC7901（未转染细胞组）、PBP—SGC7901细胞（空白质粒转染细胞组）显著延长（P〈0．05）。PTEN-SGC7901细胞的克隆形成率明显低于对照组,PTEN—SGC7901细胞对SGC7901、PBP-SGC7901细胞的集落抑制率分别为69．8％、64．8％。PTEN—SGC7901组细胞的G1期细胞含量明显较对照组增多（P〈0．05）,但三组细胞之间的凋亡率无明显差别（P〉0．05）。PTEN—SGC7901细胞VEGF与MMP-9蛋白的表达和分泌明显低于对照组（P〈0．05）,但MMP-2蛋白在三组间的表达无差异（P〉0．05）。结论PTEN可抑制胃癌细胞株SGC7901细胞的生长与增殖;PTEN可能通过下调SGC7901细胞株VEGF及MMP-9的表达与分泌来抑制肿瘤浸润与转移的发生。

Biological implication of PTEN gene expression in human gastric cancer and related molecular mechanisms

OBJECTIVE: To study the role of PTEN gene involved in the biological behavior of human gastric carcinoma cells and underlying molecular mechanisms. METHODS: Gastric carcinoma cell line, SGC7901, was transfected with plasmid PBP-PTEN and stable high PTEN expression clones were selected by Western blot and cell immunohistochemistry screening. Cell proliferation rate and apoptosis index of transfected cells were investigated by growth curve analysis, colony-formation assay and flow cytometry (FCM). Expressions of vascular endothelial growth factor (VEGF), matrix metalloprotease-2 (MMP-2) and MMP-9 proteins in cell culture supernatant and cytoplasm were determined by ELISA, gelatin zymogram, Western blot and cell immunohistochemistry. RESULTS: Stable clone with high level expression of PTEN was successfully established (PTEN-SGC7901). Cell doubling time of PTEN-SGC7901 was longer than that of the control cells (P < 0.05). The size and colony-forming efficiency of PTEN-SGC7901 cells deceased compared with those of the control. The relative colony-inhibition efficiency of PTEN-SGC7901 to SGC7901 (na?ve untransfected) and PBP-SGC7901 (control vector transfected) cells were 69.8% and 64.8%, respectively. PTEN-SGC7901 clone had more cells at G1 phase (P < 0.05) compared with that of the control. However, the apoptosis index did not show significant differences among the three groups (P > 0.05). There were significantly less VEGF and MMP-9 protein expressions in the PTEN-SGC7901 culture supernatant and cytoplasm (P < 0.05). In contrast, the MMP-2 expression among three cell groups had no significant difference (P > 0.05). CONCLUSIONS: PTEN expression suppresses the growth and proliferation of gastric carcinoma cell SGC7901, possibly through an inhibition of the expressions of VEGF and MMP-9.