肝细胞黏附分子调节蛋白激酶C ε对肾癌786-O细胞增殖和迁移的影响

Effect of hepatocyte cell adhesion molecule regulating protein kinase C ε on proliferation and migration of renal carcinoma 786-O cells

摘要:

目的：研究肝细胞黏附分子（hepatocyte cell adhesion molecule，HepaCAM）对肾癌786-O细胞增殖及迁移的影响。方法：以786-O为目的细胞，通过感染HepaCAM腺病毒（Ad-GFP-HepaCAM）和空载病毒（Ad-GFP），Western blot检测各处理组蛋白激酶C ε（protein kinase C ε，PKCε）膜蛋白、浆蛋白和总蛋白的变化，Ad-GFP-HepaCAM和10 μmol/L PKCε特异性转位抑制剂（εV1-2）处理24 h后基质金属蛋白酶（matrix metalloproteinase，MMP）-9和细胞周期蛋白cyclinD1的变化；平板克隆和划痕实验检测Ad-GFP-HepaCAM及εV1-2对786-O增殖和迁移的影响。结果：感染Ad-GFP-HepaCAM后与对照组比较，PKCε膜蛋白降低（F=82.724，P=0.000；P=0.000）而浆蛋白升高（F=537.006，P=0.000；P=0.000），总蛋白无统计学差异（F=0.548，P=0.605；P=0.805）。Ad-GFP-HepaCAM和εV1-2处理后，MMP-9降低（F=33.109，P=0.001，P=0.000；F=96.868，P=0.000，P=0.000），cyclinD1降低（F=562.626，P=0.000，P=0.000；F=159.433，P=0.000，P=0.000）。Ad-GFP-HepaCAM组比εV1-2组克隆形成率降低（F=259.918，P=0.000；P=0.000），迁移率降低（F=550.169，P=0.000；P=0.000）。结论：HepaCAM可能通过阻止PKCε从胞浆转位到胞膜而部分失活来抑制786-O细胞增殖及迁移。

Abstract:

Objective：To investigate the effect of re-expression of hepatocyte cell adhesion molecule（HepaCAM） on the proliferation and migration ability of renal carcinoma 786-O cells. Methods：Protein kinase C ε（PKCε） membrane protein level，PKCε cytosolic protein level and total protein level were determined by Western blot after 786-O cell being infected with Ad-GFP-HepaCAM and Ad-GFP. Matrix metalloproteinase（MMP）-9 and cyclinD1 protein levels were also detected by Western blot after being treated with Ad-GFP-HepaCAM and εV1-2（10 μmol/L，24 h）. The proliferation ability and migration ability of 786-O cells were analyzed by colony formation assay and wound healing assay，respectively. Results：When compared with those of control group，PKCε membrane protein level was decreased（F=82.724，P=0.000；P=0.000） and cytosolic protein level was increased（F=537.006，P=0.000；P=0.000），but PKCε total protein level was not changed（F=0.548，P=0.605；P=0.805） after being infected with Ad-GFP-HepaCAM. After expo－sure to Ad-GFP-HepaCAM and εV1-2，MMP-9 protein level was down-regulated（F=33.109，P=0.001，P=0.000；F=96.868，P=0.000，P=0.000） and cyclinD1 protein level was also down-regulated（F=562.626，P=0.000，P=0.000；F=159.433，P=0.000，P=0.000） when compared with those of control group. The colony formation rate was decreased（F=259.918，P=0.000；P=0.000） and migration ratio was also reduced（F=550.169，P=0.000；P=0.000） in Ad-GFP-HepaCAM group compared with those of εV1-2 group. Conclu－sion：HepaCAM could inhibit 786-O cell proliferation and mi－gration via blocking PKCε translocating from cytoplasm to membrane and inactivating PKCε partially.