同型异型盒C11基因在结肠癌细胞增殖中的作用及其机制

Role and mechanism of homebox C11 in proliferation of colon cancer

目的探讨同型异型盒C11(HOXC11)基因在结肠癌增殖中的作用及其机制。方法在肿瘤基因组图谱(TCGA)数据库检索结肠癌相关基因进行分析。定量聚合酶链反应(qPCR)法检测河北医科大学第四医院胃肠外科2019年1月至2019年12月行结肠癌根治术的70例临床结肠癌组织、癌旁正常黏膜组织中HOXC11表达;检测结肠癌细胞株人类肿瘤细胞系116(HCT116)、人类大肠癌H-29细细胞(HT-29)、人结肠癌SW480细胞(SW480)及肠上皮细胞株新型人肠胚胎细胞模型(HIEC)(均购自中国科学院上海细胞资源中心)中HOXC11表达,对数据库结果进行验证。用HOXC11-小干扰RNA(siRNA)转染SW480细胞,检测转染对SW480细胞活性及增殖的影响;同时检测抑制HOXC11后SW480细胞增殖相关基因表达。使用STRING数据库对结肠癌中HOXC11的功能进行蛋白相互作用(PPI)网络分析及富集分析。两组间比较采用独立样本t检验,多组间比较采用单因素方差分析。结果TCGA数据库结果显示HOXC11在结肠癌高表达(|logFC|>2,P<0.05),高表达HOXC11是患者预后差的标志(χ^(2)=3.92,P<0.05),验证结果与数据库结果一致。结肠癌细胞株HOXC11水平在SW480为1.073±0.190、HT-29为0.743±0.029,明显高于HIEC(0.247±0.042),SW480细胞中HOXC11水平最高(F=40.221,P<0.05),差异均有统计学意义。PPI网络分析显示HOXC11与其相邻节点呈现出共表达趋势,功能富集分析显示HOXC11的靶基因影响了生长、发育、增殖等很多生物学过程。HOXC11-siRNA转染后SW480细胞活性低于对照物及无关序列组,细胞活性在对照组、无关序列组、HOXC11-siRNA组分别为0.993±0.114、0.980±0.077、0.468±0.077(F=65.011,P<0.05)、细胞周期处于G0/G1期的细胞比例高于对照物及无关序列组,处于S期的比例明显低于对照物及无关序列组,处于G0/G1期3组数值分别为53.767±9.150、53.433±6.240、79.300±6.252(F=12.248,P<0.05);处于S期3组数值分别为33.200±7.529、36.167±7.731、13.367±4.565(F=10.071,P<0.05);G2/M期3组数值分别为12.967±3.584、10.367±1.620、7.300±2.022(F=3.700,P<0.05)。HOXC11-siRNA转染后SW480细胞Cyclin D1、CDK4表达分别为1.032±0.142、0.967±0.058、0.439±0.101;1.028±0.139、0.964±0.091、0.467±0.075明显低于对照组及无关序列组(F=18.798,P<0.05;F=17.011,P<0.05),而p21的表达分别为0.457±0.105、0.486±0.132、0.847±0.060(F=8.932,P<0.05),明显高于对照组及无关序列组,差异均有统计学意义。结论HOXC11在结肠癌中表达增强与预后有关,并可能通过促进细胞增殖导致肿瘤进展。

Objective To explore the role and molecular mechanism of homebox C11(HOXC11)in proliferation of colon cancer cells.Methods The cancer genome atlas(TCGA)database was used to search colon cancer-related genes.Quantitative polymerase chain reaction(qPCR)was applied to detect the expression of HOXC11 in 70 cases of colon cancer and normal mucosal tissues,and HOXC11 in colon cancer cell lines human colon cancer cell line(HCT116),Human colon cancer cells(HT-29),colorectal carcinoma cell lines(SW480)and intestina epithelial cell line new human intestinal embryonic cell model(HIEC)was also detected.SW480 cells were transfected with HOXC11-small interfering RNA(siRNA),and viability and proliferation were tested.Proliferation-related genes were detected in SW480 cells after transfection.STRING database and protein-protein interaction(PPI)enrichment analysis were used to investigate function of HOXC11 in colon cancer.Results Results of TCGA database showed that HOXC11 was over-expressed in colon cancer(|logFC|>2,P<0.05),and over-expression of HOXC11 predicted poor prognosis(χ^(2)=3.92,P<0.05).The verification results were consistent with the database results.The level of HOXC11 in SW480(1.073±0.190)and HT-29(0.743±0.029)was higher in colon cancer cells than in HIEC(0.247±0.042),and the highest level of HOXC11 was found in SW480 cells(F=40.221,P<0.05).PPI network analysis showed a trend of co-expression between HOXC11 and its neighboring nodes.Functional enrichment analysis showed that the target genes of HOXC11 might affect many biological processes such as growth,development,and proliferation,etc.After HOXC11-siRNA transfection,the activity of SW480 cells decreased,the proportion of cells in the G0/G1 phase increased significantly(F=65.011,P<0.05),and the proportion of cells in the G2/M phase decreased significantly(G0/G1 phase,F=12.248,P<0.05;S phase,F=10.071,P<0.05;G2/M phase,F=3.700,P>0.05).Cyclin D1 and cyclin dependent kinase 4(CDK4)expression in SW480 cells decreased significantly after transfection(F=18.798,P<0.05;F=17.011,P<0.05),while the expression of p21 increased significantly(F=8.932,P<0.05).Conclusion HOXC11 expression was up-regulated in colon cancer,which was correlated to the prognosis;HOXC11 might promote cancer progression by regulating cell proliferation.