沉默血管内皮钙黏蛋白基因对人高转移肝癌HCCLM3细胞增殖及迁移的影响

Effect of silencing vascular endothelial cadherin gene on proliferation and migration of human high metastatic hepatocellular carcinoma cells

目的探讨沉默血管内皮钙黏蛋白(vascular endothelial cadherin,VE-cadherin)基因对人高转移肝癌HCCLM3细胞增殖及迁移的影响。方法将携带3种不同抑制靶点的VE-cadherin基因沉默重组质粒GV248-shRNA-VEcadherin(1)、(2)、(3)分别转染至人高转移肝癌HCCLM3细胞,同时设阴性对照组(转染空载体质粒GV248-shRNANC)和空白对照组(未转染)。采用qRT-PCR及Western blot法分别检测各组细胞中VE-cadherin基因mRNA转录及蛋白的表达水平,同时筛选出最有效的靶点。MTT法测定各组细胞的体外增殖能力;Transwell小室试验检测各组细胞的迁移能力。结果与空白对照组及阴性对照组比较,3组转染VE-cadherin-shRNA干扰质粒的HCCLM3细胞的VE-cadherin基因mRNA转录及蛋白表达水平显著降低(P<0.05),其中GV248-shRNA-VE-cadherin(1)的VEcadherin基因抑制率最高(P<0.05),筛选为最佳靶点;最佳靶点干扰组HCCLM3细胞的增殖及迁移能力均明显降低(P<0.05)。结论沉默VE-cadherin基因可显著抑制人高转移肝癌HCCLM3细胞的增殖及迁移。

Objective To investigate the effect of silencing vascular endothelial cadherin（VE-cadherin） gene on the proliferation and migration of highly metastatic human hepatocellular carcinoma cell line HCCLM3. Methods Three sh RNA plasmids targeting VE-cadherin gene, GV248-shRNA-VE-cadherin（1）,（2）and（3）, were constructed, with which HCCLM3 cells were transfected, using those transfected with empty vector GV248-shRNA-NC as negative control and those untransfected as blank control. The mRNA transcription and protein expression levels of VE-cadherin gene were determined by qRT-PCR and Western blot respectively, while the most effective targets were screened. The cell proliferation in vitro was determined by MTT assay, while the migration ability by Transwell assay. Results The m RNA transcription and protein expression levels of VE-cadherin gene in HCCLM3 cells transfected with GV248-shRNA-VEcadherin（1）,（2） and（3） decreased significantly as compared with those in blank and negative control groups（P〈0. 05）. GV248-shRNA-VE-cadherin（1） showed the highest inhibitory rate（P〈0. 05） to VE-cadherin gene, which contained the most effective target. Both the proliferation and migration abilities of HCCLM3 tranfected with the plasmid decreased significantly（P〈0. 05）. Conclusion The silencing of VE-cadherin gene inhibited the proliferation and migration of HCCLM3 cells significantly.