垂体肿瘤转化基因调控结直肠癌细胞凋亡及其与PI3K/AKT信号通路的关系

Tumor transforming gene regulates colorectal cancer cell apoptosis and its relationship with PI3K/AKT signaling pathway

目的:探讨垂体肿瘤转化基因(PTTG1)调控结直肠癌细胞凋亡及其与磷脂酰肌醇-3-激酶(PI3K)/丝氨酸-苏氨酸蛋白激酶(AKT)信号通路的关系。方法:使用慢病毒感染系统构建PTTG1基因过表达和干扰表达的HCT116人结直肠癌细胞稳定细胞株,随机分为空白对照组(A组),阴性对照组(B组),PTTG1敲除组(C组)和PTTG1过表达组(D组)。采用免疫印迹实验(Western blotting法)及实时RT-PCR检测不同稳定细胞株中PTTG1及PTTG1 mRNA的表达,分别采用CCK-8、流式细胞术、Transwells小室对细胞的增殖、凋亡、迁移和侵袭情况进行检测。比较各组PTTG1及PTTG1 mRNA水平的表达、细胞的增殖、凋亡、迁移和侵袭情况,并分析PTTG1对PI3K/AKT信号通路的影响。结果:与A组比较,C组的PTTG1 mRNA表达明显降低,D组的明显升高(P<0.05)。C组PTTG1蛋白表达低于B组,D组高于B组(P<0.05)。A组、B组、C组的细胞增殖能力比较,差异无统计学意义(P>0.05);D组的细胞增殖能力明显低于其他3组(P<0.05)。D组凋亡的细胞数均明显高于其他3组(P<0.05)。D组的细胞迁移个数和侵袭个数均明显低于其他3组(P<0.05)。PTTG1上调可抑制PI3K表达,造成PI3K/AKT通路失活。进一步使用PI3K激动剂(740Y-P)和AKT激动剂(PDGF)反向逆转,发现PTTG1对PI3K/AKT信号通路同样存在抑制作用。结论:PTTG1基因过表达能够抑制结直肠癌细胞增殖,促进凋亡,并抑制肿瘤细胞的浸润和转移,且这一过程与PTTG1基因对PI3K/AKT信号通路的抑制相关。

Objective:To investigate the regulation of pituitary tumor transforming gene(PTTG1)on colorectal cancer cell apoptosis and its relationship with phosphatidylinositol-3-kinase(PI3K)/serine-threonine protein kinase(AKT)signaling pathway.Methods:The lentiviral infection system was used to construct a stable HCT116 human colorectal cancer cell line with overexpression and interference expression of PTTG1 gene,and randomly divided into group A(blank control group),group B(negative control group),group C(PTTG1 knockout group),and group D(PTTG1 overexpression group).Western blotting was used to detect the expression of PTTG1 and PTTG1 mRNA in different stable cell lines.CCK-8,flow cytometry,and Transwells chamber were used to detect cell proliferation,apoptosis,migration and invasion,respectively.The expression of PTTG1 and PTTG1 mRNA levels,cell proliferation,apoptosis,migration,and invasion were compared,and the influence of PTTG1 on PI3K/AKT signaling pathway was analyzed.Results:Compared with group A,the expression of PTTG1 mRNA in group C was significantly decreased,and the mRNA expression of PTTG1 in group D was significantly increased(P<0.05).The protein expression of PTTG1 in group C was lower than that in group B,and group D was higher than group B(P<0.05).The cell proliferation ability of group A,group B,and group C was not statistically different(P>0.05).The cell proliferation ability of group D was significantly lower than other three groups(P<0.05).The apoptotic cells of group D were more than other three groups,while the migration and invasion cells were significantly less(P<0.05).Up-regulation of PTTG1 could inhibit the expression of PI3K and cause the inactivation of the PI3K/AKT pathway.Use of PI3K agonist(740Y-P)and AKT agonist(PDGF)to reverse the reversal and found that PTTG1 also had an inhibitory effect on the PI3K/AKT signaling pathway.Conclusion:Overexpression of PTTG1 gene can inhibit tumor cell proliferation in colorectal cancer cells,promote apoptosis,and inhibit tumor cell infiltration and metastasis,and this process is related to the inhibition of PI3K/AKT signaling pathway by PTTG1 gene.