KAI1基因对胃癌细胞体外生长及迁移的影响

Effect of KAI1 gene on growth and migration of gastric cancer cells in vitro

目的通过基因转染改变胃癌细胞中KAI1基因表达量,进一步观察KAI1基因对胃癌细胞体外生长及迁移能力的影响。方法将人胃癌细胞BGC823分为过表达KAI1的实验组(BGC823-OE组)、空载体的阴性对照组(BGC823-NC组)及空白组(BGC823-Ctrl组);同时将人胃癌细胞MKN28分为沉默KAI1的实验组(MKN28-OE组)、空载体的阴性对照组(MKN28-NC组)及空白对照组(MKN28-Ctrl组),并建立稳定转染细胞株;采用Western blot和RT-PCR技术检测两种稳转细胞株中KAI1蛋白和mRNA相对表达量;四唑盐比色法(MTT)检测两种稳转细胞株的体外增殖情况;Transwell实验检测两种稳转细胞株体外迁移情况;流式细胞术检测两种稳转细胞株细胞周期变化。结果慢病毒感染72 h后两种胃癌细胞OE组和NC组观察到大量荧光表达,加药筛选扩增后成功构建稳转细胞株。BGC823-OE组的KAI1 mRNA和蛋白相对表达量高于BGC823-NC组及BGC823-Ctrl组(P<0.05),而MKN28-OE组的KAI1 mRNA和蛋白相对表达量低于MKN28-NC组及MKN28-Ctrl组(P<0.05)。BGC823-OE组24、48、72h的相对增殖率低于BGC823-NC组及BGC823-Ctrl组(P<0.05),而MKN28-OE组24、48、72h的相对增殖率高于MKN28-NC组及MKN28-Ctrl组(P<0.05)。BGC823-OE组细胞迁移数目、S期占比均少于BGC823-NC组及BGC823-Ctrl组(P<0.05),而MKN28-OE组迁移数目、S期占比均多于MKN28-NC组及MKN28-Ctrl组(P<0.05)。结论成功构建过表达KAI1基因的人胃癌细胞BGC823细胞株和沉默KAI1基因的人胃癌细胞MKN28细胞株;KAI1基因对不同胃癌细胞体外生长、迁移能力有负向调控作用。

Objective To change the expression of KAI1 gene in gastric cancer cells by gene transfection,and further observe the effects of KAI1 gene on the growth and migration of gastric cancer cells in vitro.Methods Human gastric cancer cell line BGC823 was divided into experimental group(BGC823-OE group)infected with KAI1 lentivirus overexpression vector,negative control group(BGC823-NC group)infected with empty vector and blank group(BGC823-Ctrl group).Human gastric cancer cell line MKN28 was also divided into experimental group(MKN28-OE group)infected with KAI1 RNAi lentiviral vector,negative control group(MKN28-NC group)infected with empty vector and blank control group(MKN28-Ctrl group).Western blot and RT-PCR were used to detect the relative expression of KAI1 protein and mRNA in the cells after transfection.The proliferation of the two cells was detected by tetrazolium salt colorimetry(MTT).The migration of cells in vitro was detected by Transwell assay.Flow cytometry was used to detect changes in the cell cycle.Results After 72 hours of lentivirus infection,a large amount of fluorescent expression was observed in the OE group and the NC group.After the drug screening and amplification,the stable cell line was successfully constructed.The relative expression of KAI1 protein and mRNA in BGC823-OE group was higher than that in BGC823-NC group and BGC823-Ctrl group(P<0.05),while the relative expression of KAI1 protein and mRNA in MKN28-OE group was lower than MKN28-NC group and MKN28-Ctrl group(P<0.05).The relative proliferation rate of BGC823-OE group at 24,48 and 72 h was lower than that of BGC823-NC group and BGC823-Ctrl group(P<0.05),while the relative proliferation rate of MKN28-OE group at 24,48 and 72 h was higher than that of MKN28-NC group and MKN28-Ctrl group(P<0.05).The number of cell migration and S phase ratio in BGC823-OE group were lower than those in BGC823-NC group and BGC823-Ctrl group(P<0.05),while the migration number and S phase ratio in MKN28-OE group were higher than those in MKN28-NC group and MKN28-Ctrl group(P<0.05).Conclusion The human gastric cancer cell line BGC823 over expressing KAI1 gene and the human gastric cancer cell line MKN28 silencing KAI1 gene were successfully constructed.KAI1 gene has a negative regulation effect on the growth and migration ability of different gastric cancer cells in vitro.