整合素α5的表达下调抑制肝癌Bel-7404细胞的增殖、侵袭及转移

Integrin α5 silencing inhibits proliferation,invasion and metastasis of human liver cancer Bel-7404 cells in vitro

目的分析整合素α5(ITGA5)与肝癌分级及肝癌患者总体生存率之间的关系,进一步研究整合素ITGA5基因沉默后对人肝癌Bel-7404细胞增殖、侵袭迁移能力的影响及其作用机制探讨。方法(1)利用UALCAN分析ITGA5在肝癌组织及正常组织中的表达,以及在不同分级肝癌组织中的表达及其差异统计;通过GEPIA在线分析ITGA5表达量与肝癌患者生存情况的关系。(2)培养前期已经构建的ITGA5沉默细胞系,采用Western blot方法检测ITGA5蛋白水平的表达,以鉴定ITGA5基因沉默的效果。(3)采用平板克隆形成实验、Transwell等实验方法检测ITGA5基因沉默后Bel-7404细胞的增殖、侵袭迁移能力的变化。(4)通过Oncomine癌标本数据库分析肝癌组织及对照组织中ITGA5与PI3K表达及分析其相关性。结果(1)ITGA5在肝癌中的表达显著高于正常组,差异有统计学意义(P<0.05);在不同肝癌分级中,Grade 1的ITGA5表达量显著低于Grade 2,Grade 2的ITGA5表达量显著低于Grade 3,差异均有统计学意义(P<0.05)。ITGA5高表达组的总体生存率(OS)显著低于低表达组,差异有统计学意义(P<0.05)。(2)Western blot鉴定ITGA5基因沉默组的蛋白质表达水平比空白、阴性对照组明显降低,差异具有统计学意义(P<0.05)。(3)平板克隆形成实验结果显示,基因沉默组比空白、阴性对照组的克隆形成率降低,差异具有统计学意义(P<0.05);Transwell结果显示,基因沉默组细胞穿膜数均比空白及阴性对照组显著降低,差异具有统计学意义(P<0.05)。(4)ITGA5和PI3K在肝癌组织中高表达且呈正相关,差异具有统计学意义(P<0.05)。结论ITGA5与肝癌进展及肝癌患者预后密切相关,其表达下调抑制肝癌细胞的增殖、侵袭迁移;ITGA5可能通过调控PI3K信号通路促进促进肝癌增殖及转移。

Objective To analyze the association of integrinα5(ITGA5)with grading of liver cancer and the overall patient survival and investigate the effects of integrinα5(ITGA5)silencing on the proliferation,invasion and migration abilities of human liver cancer Bel-7404 cells.Methods UALCAN was used to analyze the expression of ITGA5 in liver cancer tissues and normal tissues,and expression in different grades of liver cancer tissues.GEPIA was used to analyze the relationship between ITGA5 expression and the survival of liver cancer patients through.The ITGA5 shRNA lentiviral vector was used to infect Bel-7404 cells to establish a cell line with stable ITGA5 silencing verified by Western blotting.Plate clone formation assay and Transwell assay were used to detect the proliferation,invasion and migration of Bel-7404 cells.The correlation between ITGA5 and PI3K in liver cancer tissues and control tissues was analyzed using Oncomine cancer specimen database.Results The expression of ITGA5 was significantly higher in liver cancer than in normal tissues(P<0.05).The expression of ITGA5 was significantly lower in grade 1 than in grade 2 liver cancer,and also lower in grade 2 than in grade 3 liver cancer(P<0.05).The patients with high ITGA5 expression had a significantly lower overall survival rate than those with low ITGA5 expression(P<0.05).Plate clone formation assay showed that the clone formation rate was significantly lowered in Bel-7404 cells with ITGA5 silencing compared with the blank and negative control cells(P<0.05).ITGA5 silencing significantly attenuated the migration of Bel-7404 cells as shown by Transwell assay(P<0.05).ITGA5 and PI3K were both highly expressed and showed a positive correlation in liver cancer tissues(P<0.05).Conclusion ITGA5 is closely related to the progression of liver cancer and the patients'prognosis.ITGA5 silencing inhibits the proliferation,invasion and migration of liver cancer cells.ITGA5 promotes the liver cancer growth and metastasis possibly by regulating the PI3K signaling pathway.