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目的:探讨ANGPTL4基因及其缺失突变体对肝癌细胞SMMC-7721生长的影响。方法:构建ANGPTL4基因全长及其功能结构域缺失突变体的融合表达载体,即ANGPTL4-GFP-N1、ANGPTL4-del1-GFP-N1、ANGPTL4-del2-GFP-N1、ANGPTL4-del3-GFP-N1,脂质体法将EGFP-N1空载体、ANGPTL4基因及其缺失突变体分别转染肝癌细胞SMMC-7721,G418筛选,建立稳定细胞系,用MTT试验法、细胞集落形成试验检测细胞体外生长活性和增殖能力;流式细胞仪检测ANGPTL4基因对细胞凋亡的影响;裸鼠移植瘤试验检测转染细胞的体内成瘤特性。结果:ANGPTL4基因及其缺失突变体体外对肝癌细胞SMMC-7721的生长、集落形成具有明显的抑制作用(均为P<0.01);ANGPTL4全长基因及其缺失突变体ANGPTL4-del1-GFP-N1和ANGPTL4-del2-GFP-N1均对SMMC-7721细胞体内成瘤有明显抑制作用(均为P<0.01)。但转染ANGPTL4全长基因及其缺失突变体的SMMC-7721细胞的细胞凋亡发生率与转染空载体组相比没有明显差别(均为P>0.05)。结论:ANGPTL4基因及其缺失突变体对肝癌SMMC-7721细胞体内、外生长均具有明显抑制作用,但该抑制作用不是通过促进细胞凋亡实现的。
Objective: To investigate the effect of ANGPTL4 gene and its deletion mutant on the growth of hepatocellular carcinoma cell line SMMC-7721. METHODS: ANGPTL4-GFP-N1, ANGPTL4-del1-GFP-N1, ANGPTL4-del2-GFP-N1 and ANGPTL4-del3-GFP-N1 were constructed by constructing the full-length ANGPTL4 gene and its functional domain deletion mutants The EGFP-N1 empty vector, ANGPTL4 gene and their deletion mutants were transfected into SMMC-7721 and G418 cells by lipofectamine respectively to establish stable cell lines. MTT assay and colony formation assay were used to detect the in vitro growth activity And proliferation ability. The effect of ANGPTL4 gene on apoptosis was detected by flow cytometry. The in vivo tumorigenicity of transfected cells was detected by transplanted tumor in nude mice. Results: The ANGPTL4 gene and its deletion mutant significantly inhibited the growth and colony formation of hepatocellular carcinoma cell line SMMC-7721 in vitro (all P <0.01). ANGPTL4 full-length gene and its deletion mutant ANGPTL4-del1-GFP-N1 And ANGPTL4-del2-GFP-N1 both inhibited tumorigenesis of SMMC-7721 cells in vivo (all P <0.01). However, the rate of apoptosis of SMMC-7721 cells transfected with the full-length ANGPTL4 gene and its deletion mutant was not significantly different from that of the blank vector transfected group (all P> 0.05). CONCLUSION: ANGPTL4 gene and its deletion mutant have significant inhibitory effects on hepatocellular carcinoma SMMC-7721 cells both in vitro and in vivo, but this inhibition is not achieved by promoting apoptosis.