多巴胺受体4对肝癌细胞增殖转移的影响及预后

Effect of Dopamine Receptor 4 on Proliferation and Metastasis of Hepatocellular Carcinoma Cells and Its Prognostic Significance

【目的】探讨多巴胺受体4(DRD4)在肝细胞癌组织以及癌旁组织的表达,分析其表达情况与肝癌术后患者预后相关临床病理因素的关系和预后意义;探讨DRD4激动剂PD-168077以及阻滞剂L-745870对肝癌细胞增殖、迁移及侵袭的影响,并分析其可能的机制。【方法】通过免疫组化以及实时定量PCR(qPCR)技术检测肝癌以及癌旁组织DRD4蛋白和mRNA表达情况,并分析DRD4表达的预后意义;采用Cell Counting Kit 8(CCK8)和Transwell实验检测PD-168077以及L-745870对肝癌细胞增殖、迁移及侵袭的作用,蛋白免疫印迹(Western blot)技术检测信号通路蛋白,并使用通路抑制剂验证。【结果】在196例肝癌组织中有85例(43.4%)有DRD4蛋白高表达,在74对肝癌以及非癌组织中,非癌组织的DRD4 mRNA高表达,非肝癌细胞株Miha表达较肝癌细胞系高;DRD4高表达患者包括无复发生存期(RFS)和总生存期(OS)较长,并且DRD4是肝癌切除术后患者RFS和OS的独立预后因素(P<0.05);PD-168077显著抑制肝癌细胞增殖、迁移和侵袭,并呈剂量依赖性(P<0.05),而1 nmol/L浓度的L-745870能显著增强肝癌细胞增殖、迁移和侵袭(P<0.05);PD-168077抑制ERK蛋白的磷酸化,而L-745870则可促进ERK蛋白的磷酸化,ERK抑制剂PD98059可以逆转L-745870的作用。【结论】DRD4高表达的患者预后较好,有可能成为预测肝癌术后患者预后的分子标志物;DRD4可能成为肝癌治疗的潜在靶点。

【Objective】To explore the correlation of dopamine receptor 4(DRD4)expression with clinicopathological characteristics and the prognostic significance after examining the expression of DRD4 in hepatocellular carcinoma(HCC)and non-tumor tissues;to show the effect of DRD4 agonist PD-168077 and inhibitor L-745870 on proliferation,migration and invasion of HCC cells,and to reveal its possible mechanism.【Methods】Immunohistochemistry and real time qPCR were implemented to detect the expression of DRD4 protein and mRNA in tumor and non-tumor tissues and prognostic significance of DRD4 expression was analyzed.To reveal the impact of PD-168077 and L-745870 on HCC cells,CCK8 and Transwell were performed;the signaling pathway was monitored by Western blot,and was verified with a specific inhibitor.【Results】Eighty-five(43.4%)of 196 HCC tissues had high expression of DRD4 protein;non-tumor tissues had higher expression of DRD4 mRNA in 74 pairs tissues,and DRD4 mRNA was highly expressed in non-tumor cell line Miha compared with HCC cell lines.Patients with high DRD4 expression significantly had longer recurrence free survival(RFS)and overall survival(OS),and DRD4 was an independent prognostic factor for RFS and OS in patients after HCC resection(P<0.05).PD-168077 showed a dose-dependent inhibition on proliferation,migration and invasion of HCC cells(P<0.05),while 1nmol/L of L-745870 significantly enhanced the proliferation,migration and invasion of HCC cells(P<0.05).PD-168077 inhibited the phosphorylation of ERK,while L-745870 promoted the phosphorylation of ERK.The ERK inhibitor PD98059 could reverse the effect of L-745870.【Conclusions】HCC patients with high DRD4 expression had better prognosis and DRD4 might be a biomarker for prognosis of HCC patients.DRD4 might become a target of targeted therapy for HCC.