体外血管内皮生长因子单克隆抗体对C6胶质瘤细胞增殖、侵袭和迁移力的影响

目的:在体外观察血管内皮生长因子(VEGF)单克隆抗体对C6胶质瘤细胞增殖、侵袭和迁移力的影响。方法:采用氯化钴(Co Cl2)模拟缺氧环境,予以0、0.1、1、10μg/m L的VEGF抗体,对常氧或缺氧条件下对C6胶质瘤细胞进行处理。通过MTT实验检测细胞增殖,Transwell实验检测细胞的侵袭和迁移能力,Western blotting检测HIF-1α蛋白、FAK/Pyk2总蛋白及磷酸化FAK-Tyr397/Pyk2-Tyr402表达水平变化。结果:200μmol/L Co Cl2明显抑制C6细胞增殖(P<0.05),10μg/m L VEGF抗体可明显抑制C6细胞增殖(P<0.05)。与常氧相比,缺氧可使C6细胞的迁移、侵袭能力增强(P<0.05)。常氧和缺氧下,VEGF抗体的干预均增强C6细胞的迁移、侵袭能力(P<0.05),且相对于常氧,缺氧可进一步增强VEGF抗体促C6细胞迁移、侵袭的作用(P<0.05)。Co Cl2干预可增加C6细胞中HIF-1α的含量;在常氧和缺氧情况下,只有1μg/m L VEGF抗体可降低FAK的总蛋白量(P<0.05);在缺氧情况下,VEGF抗体可明显增高Pyk2-Tyr402位点的磷酸化水平。结论:在缺氧情况下,VEGF抗体的干预可使Pyk2-Tyr402位点的磷酸化水平增高,引起C6细胞的侵袭迁移能力进一步增强。

Effect of VEGF Monoclonal Antibody on Proliferation,Invasion and Migration of C6 Glioma Cells in vitro

Objective:To observe the effect of vascular endothelial growth factor(VEGF) monoclonal antibody on C6 glioma cells' proliferation,migration and invasion in vitro.Methods:CoCl2 was added in culture medium to imitate hypoxic culture condition.C6 glioma cells were treated by VEGF antibody with different concentrations (0.1 μg/mL and 1 μg/mL) under normoxic or hypoxic conditions.Cell proliferation was detected by MTT assay.The ability of cell migration and invasion were evaluated by Transwell testing assay.The expression of HIF-1 α protein,FAK/Pyk2 total protein and level of FAK-Tyr397/Pyk2-Tyr402 phosphorylation were detected by Western blot.Results:Treatment with 200 μmol/L CoCl2 significantly inhibited the proliferation of C6 glioma cells (P＜0.05).10 μg/mL VEGF antibody significantly inhibited the proliferation of C6 glioma cells (P＜ 0.05).Compared to the normoxic condition,hypoxiia increased the migration and invasion of C6 cells (P＜0.05).Under the normoxic and hypoxic condition,the VEGF antibody can increase the migration and invasion of C6 (P＜0.05).Compared to the normoxic condition,hypoxia can enhance the migration and invasion by VEGF antibody to C6 cell (P＜0.05).The expression of total FAK protein was decreased in C6 cells by VEGF antibody only in the 1 μg/ml concentration (P＜0.05).Under hypoxic condition,the phosphorylation levels of Pyk2-Tyr402 was increased in C6 cells after VEGF antibody treatment (P＜0.05).Conclusion:Under hypoxic condition,VEGF antibody can increase the phosphorylation levels of Pyk2-Tyr402,which contribute to the invasion and migration of C6 cells.