摘要
目的为探索胃癌细胞增殖的分子机制,研究多聚嘧啶区结合蛋白1(PTBP1)和细胞分裂周期5样(裂殖酵母)(CDC5L)基因结合促进胃癌细胞增殖凋亡的影响。方法GEPIA2数据库预测PTBP1和CDC5L在胃癌细胞中的表达;RT⁃qPCR和Western blot检测PTBP1和CDC5L的表达水平;GEPIA2数据库预测CDC5L和PTBP1之间的相关性,并通过免疫蛋白沉淀实验检测验证。通过转染,干扰PTBP1和CDC5L的表达。随后,克隆实验检测细胞增殖;流式细胞术检测细胞周期;TUNEL荧光染色检测细胞凋亡;Western blot检测凋亡蛋白Bax、Bcl⁃2和caspase3的表达。结果GEPIA2数据库预测所示PTBP1和CDC5L在胃癌中表达差异有统计学意义(P<0.05),且高表达。与癌旁组织中蛋白表达相比,PTBP1和CDC5L在胃癌组织中表达提高(P<0.01);比正常细胞中蛋白表达相比,PTBP1和CDC5L在胃癌细胞中表达均上调(P<0.01);GEPIA2数据库显示PTBP1与CDC5L具有正向线性相关性;免疫沉淀实验结果表明PTBP1和CDC5L可相互结合。细胞克隆实验表明PTBP1和(或)CDC5L被抑制后,胃癌细胞增殖水平下降(P<0.05);且敲低PTBP1和/或CDC5L后胃癌细胞周期G0/G1期受阻滞(P<0.001)。细胞凋亡水平检测显示,与对照组相比,干扰PTBP1和/或CDC5L后,胃癌细胞凋亡增多(P<0.01);相关凋亡蛋白差异具有统计学变化(Bcl⁃2,P<0.05;Bac,P<0.01;cleaved⁃caspase3/caspase3,P<0.05)。结论PTBP1和CDC5L具有结合关系,敲低PTBP1和CDC5L的表达具有抑制胃癌细胞增殖,促进细胞凋亡作用。
Objective To explore the molecular mechanism of gastric cancer cell proliferation,we investigated the effect of PTBP1 and CDC5L protein binding on the proliferation and apoptosis of gastric cancer cells. Methods The expression of PTBP1 and CDC5L in gastric cancer cells was predicted by the GEPIA2 database;the expression levels of PTBP1and CDC5L were detected by RT-qPCR and Western blot;the correlation between CDC5L and PTBP1 was predicted by the GEPIA2 database and verified by immunoprecipitation assay. The expression of PTBP1 and CDC5L was disturbed by transfection. Subsequently,cell proliferation was detected by cloning assay;cell cycle was detected by flow cytometry;apoptosis was detected by TUNEL fluorescence staining;and expression of apoptotic proteins Bax,Bcl-2,and caspase3 was detected by Western blot. Results The GEPIA2 database predicted that PTBP1 and CDC5L were statistically different(P<0.05)and highly expressed in gastric cancer. PTBP1 and CDC5L expression was elevated in gastric cancer tissues compared with protein expression in paraneoplastic tissues(P<0.01);both PTBP1 and CDC5L expression was upregulated in gastric cancer cells compared with protein expression in normal cells(P<0.01);the GEPIA2 database showed a positive linear correlation between PTBP1 and CDC5L;the results of immunoprecipitation assay showed that PTBP1 and CDC5L could bind to each other. Cell cloning assay showed that the proliferation level of gastric cancer cells decreased after PTBP1 and(or)CDC5L were inhibited(P<0.05);and the G0/G1phase of gastric cancer cell cycle was blocked after knocking down PTBP1and/or CDC5L(P<0.01). Apoptosis assay showed increased apoptosis in gastric cancer cells after disturbance of PTBP1 and/or CDC5L compared with the control group(P<0.01);there were statistically different changes in related apoptotic proteins(Bcl-2,P<0.05;Bac,P<0.001;cleaved-caspase3/caspase3,P<0.05). Conclusion PTBP1 and CDC5L had a binding relationship,and knockdown of PTBP1 and CDC5L expression had the effect of inhibiting the proliferation of gastric cancer cells and promoting apoptosis.
作者
郭应伟
刘通
GUO Ying-wei;LIU Tong(De-partment of Gastrointestinal Surgery,Baise People's Hospital,Baise,Guangxi 533000,China)
出处
《解剖学研究》
CAS
2022年第5期429-436,共8页
Anatomy Research
基金
广西壮族自治区卫计委项目(Z2016713)。
关键词
胃癌细胞
多聚嘧啶区结合蛋白1
细胞分裂周期5样(裂殖酵母)基因
增殖
凋亡
Gastric cancer cells
Recombinant polypyrimidine tract binding protein 1(PTBP1)
Cell division cycle 5-like(s.pombe)(CDC5L)gene
Proliferation
Apoptosis
