TRPV1对肝细胞性肝癌生物学行为的影响及可能的机制预测

Effects of TRPV1 on biological behavior of hepatocellular carcinoma and prediction of possible mechanisms

目的明确瞬时受体电位香草醛亚家族1(transient receptor potential vanilloid 1,TRPV1)在肝细胞性肝癌(hepatocellular carcinoma,HCC)中的表达及与预后的关系,探讨其对HCC细胞生物学行为的影响。方法利用癌症基因组图谱(the cancer genome atlas,TCGA)、基因型组织表达(genotype-tissue expression,GTEx)及基因表达综合数据库(gene expression omnibus,GEO)验证TRPV1在HCC中的表达及与预后的关系。通过CCK-8实验、划痕实验、Transwell实验、流式细胞术检测TRPV1对HCC细胞增殖、迁移、侵袭和细胞周期的影响,蛋白质印迹法(western blot,WB)检测细胞周期调节蛋白激酶4(cyclin-dependent kinase 4,CDK4)、细胞周期素依赖激酶6(cyclin-dependent kinase 6,CDK6)的表达,裸鼠皮下成瘤实验检测肿瘤增殖能力。最后,构建TRPV1共表达网络并进行富集分析。结果TCGA、GTEx和GEO数据库中,TRPV1在HCC组织中表达水平均下调(均P<0.001),且与患者总生存时间更短有关(P=0.032),受试者工作特征(receiver operating characteristic,ROC)曲线下面积为0.91。过表达TRPV1可抑制HCC细胞增殖、迁移和侵袭能力,阻滞细胞周期进展,CDK4和CDK6蛋白表达水平下降(均P<0.05),敲低TRPV1则相反(均P<0.05)。稳定过表达TRPV1后裸鼠皮下瘤体体积减小(P=0.015),质量减轻(P=0.033),Ki-67蛋白阳性表达水平下调(P=0.010)。TRPV1共表达基因主要参与核糖核酸(ribonucleic acid,RNA)剪接、细胞周期蛋白结合、细胞增殖等过程。结论TRPV1抑制了HCC细胞的增殖、迁移和侵袭能力,并阻滞细胞周期的进程,在HCC中发挥抑癌作用,为HCC预后分析、治疗提供了新方向。

Objective To clarify the expression of transient receptor potential vanilloid subfamily 1(TRPV1)in hepatocellular carcinoma(HCC)and its relationship with prognosis,and to explore its effect on the biological behavior of HCC cells.Methods The cancer genome atlas(TCGA),genotype-tissue expression(GTEx)and gene expression ominbus(GEO)databases were utilized to verify the expression of TRPV1 in HCC and its relationship with prognosis.The effects of TRPV1 on the proliferation,migration,invasion,and cell cycle of HCC cells were detected by CCK-8 assay,scratch assay,Transwell assay,and flow cytometry.The expression of cyclin-dependent kinase 4(CDK4)and cyclin-dependent kinase 6(CDK6)was detected by Western blot,and the tumor proliferation ability was detected by subcutaneous tumor formation assay in nude mice.Finally,the TRPV1 co-expression network was constructed and analyzed for functional enrichment.Results The expression level of TRPV1 in HCC tissues was down-regulated in TCGA,GTEx,and GEO databases(P<0.001)and was associated with a shorter overall survival time of patients(P=0.032),with an area under the ROC curve of 0.91.Overexpression of TRPV1 suppressed the proliferation,migration,and invasion ability of HCC cells,blocked cell cycle progression,and CDK4 and CDK6 protein expression levels decreased(both P<0.05),and the opposite was true for knockdown of TRPV1(both P<0.05).After stable overexpression of TRPV1,the subcutaneous tumor volume of nude mice was reduced(P=0.015),the mass of nude mice was reduced(P=0.033),and the positive expression level of Ki-67 protein was down-regulated(P=0.010).TRPV1 co-expressed genes were mainly involved in the processes of RNA splicing,cell cycle protein binding,and cell proliferation.Conclusions TRPV1 inhibited the proliferation,migration,and invasion of HCC cells and blocked cell cycle progression,which exerted an oncogenic role in HCC and provided a new direction for the prognostic analysis and treatment of HCC.