DUSP1在药物诱导的肝癌细胞衰老中的表达变化及其作用研究

目的:在肝癌的治疗中,化学疗法是重要的治疗手段,尤其在结合外科手术切除的联合治疗中,化疗被广泛的应用于临床。而体内试验证实,化疗药物诱导的细胞衰老在治疗肿瘤的过程中,普遍存在并发挥重要作用,我们着重探讨在药物诱导的人肝癌细胞衰老中,细胞内重要信号途径的变化,发现并利用对细胞衰老有重要意义的蛋白,增强化学治疗肿瘤的效果。在本课题中,我们利用化疗药物阿霉素(doxorubicin)和蛋白酶体抑制剂MG132诱导的人衰老肝癌细胞模型,对MAPK信号途径的负调控因子DUSP家族的表达变化进行检测,筛选表达水平发生显著变化的双特异性磷酸酶(DUSP)家族成员,并初步探讨其在细胞衰老中的作用。方法:利用低剂量的阿霉素和蛋白酶体抑制剂MG132诱导人肝癌细胞衰老;通过Real time RT-PCR和Western blotting检测DUSP家族基因mRNA和蛋白质水平的变化;利用siRNA干扰技术敲降DUSP1,检测药物诱导肝癌细胞衰老水平的变化。结果:经低剂量的阿霉素和蛋白酶体抑制剂短时间处理后,人肝癌细胞系Huh7和SMMC-7721细胞发生明显的细胞衰老;在诱导的衰老细胞中,DUSP1mRNA和蛋白质水平呈显著升高。在敲降DUSP1后,MG132诱导的细胞衰老得到显著的缓解。结论:DUSP1在药物诱导的人肝癌细胞衰老中具有重要的作用,为研究临床治疗中化疗药物引起肿瘤细胞衰老的具体机制带来启发。

Expression Patterns and Functional Analysis of DUSP1 in Drug-induced Cellular Senescence in Human Liver Cancer Cells *

Objective： Chemotherapy plays an important role and is widely used in clinical by combining with surgical operation excision in the treatment of HCC. Current research suggests that tumor cellular senescence that induced by Chemotherapy contributes to treatment outcome in vivo. Here, we focused on screening and detecting the changed signaling pathways in human senescent liver cancer cell induced by chemotherapy drug. In this study, we screened of the DUSP family members which are the cardinal negative regulators of MAPK with altered expression in doxorubicin-and MG132-induced senescence of human liver cancer cells. Methods： The chemotherape- utic drug doxorubicin and proteasome inhibitor MG132 were used for inducing senescence in human liver cancer cells. The expression levels of DUSP members in the senescent cells were measured by RT-PCR and Western blotting. The proportion of senescent cells was detected by SA-13-gal staining in the cells treated with or without the drug when DUSP1 was knocked down by siRNA. Results： Doxorubicin and MG132 robustly elicited senescence in human liver cancer cells. DUSP1 was significantly up-regulated in the senescent Huh7 and SMMC-7721 cells induced by doxorubiein and MG132. The inhibition of DUSPI by siRNA was able to suppress MG132- but not doxorubicin-induced cellular senescence. Conclusions： DUSP1 was up-regulated mostly in 10 DUSP members in drug-induced cellular senescence. The DUSP1 expression has an important function in drug-induced senescence.