Liu Jian, Liu Jing, Li Sheng-Ze, Zheng Ying-Ao, Guo Su-Yang, Wang Xiu
Bengbu Medical College, Bengbu 233030, China. E-mail:
Nan Fang Yi Ke Da Xue Xue Bao. 2016 Apr 20;37(4):475-481. doi: 10.3969/j.issn.1673-4254.2017.04.09.
To investigate the relationship between sensitivity to cisplatin (DDP) and the expression of HSP70 in cervical cancer cells in vitro.
Cervical cancer Hela229 cells treated with different concentrations of DDP and the HSP70 inhibitor (PFT-µ) were examined for cell viability using MTT assay and colony forming ability. The cell apoptosis was analyzed by flow cytometry with propidium iodide staining and DAPI staining, and JC-1 staining was used to determine mitochondrial membrane potential. The expressions of HSP70, Bcl-2, Bax and caspase-3 were measured with Western blotting. A nude mouse model bearing Hela229 cell xenograft was used to evaluate the effect of DDP and PFT-µ on tumor growth.
Hela229 cells expressed a higher level of HSP70 than normal cervical cells. The combined use of PFT-µ significantly enhanced the inhibitory effect of DDP (P<0.01) and increased the cell apoptosis in Hela229 cells. JC-1 staining demonstrated that DDP combined with PFT-µ more obviously reduced mitochondrial membrane potential. DDP combined with PFT-µ more strongly lowered Bcl-2 expression and increased the expressions of casepase-3 and Bax than DDP alone. In the nude mouse model, PFT-µ significantly enhanced DDP sensitivity of Hela229 cell xenografts (P<0.01).
Inhibition of HSP70 expression can enhance the sensitivity of cervical cancer cell to DDP both in vivo and in vitro possibly by promoting cell apoptosis, suggesting the potential of HSP70 as a new target for gene therapy of cervical cancer.
探讨体外培养的宫颈癌细胞对顺铂(DDP)的敏感性与热休克蛋白70(HSP70)表达之间的关系。
采用MTT法和集落形成能力检测不同浓度DDP及HSP70抑制剂(PFT-μ)处理后的宫颈癌Hela229细胞的活力。通过碘化丙啶染色和DAPI染色的流式细胞术分析细胞凋亡情况,并用JC-1染色法检测线粒体膜电位。采用蛋白质免疫印迹法检测HSP70、Bcl-2、Bax和半胱天冬酶-3的表达。利用荷Hela229细胞异种移植瘤的裸鼠模型评估DDP和PFT-μ对肿瘤生长的影响。
Hela229细胞中HSP70的表达水平高于正常宫颈细胞。联合使用PFT-μ可显著增强DDP的抑制作用(P<0.01),并增加Hela229细胞的凋亡。JC-1染色显示,DDP与PFT-μ联合使用更明显地降低了线粒体膜电位。与单独使用DDP相比,DDP与PFT-μ联合使用更强烈地降低了Bcl-2的表达,并增加了半胱天冬酶-3和Bax的表达。在裸鼠模型中,PFT-μ显著增强了Hela229细胞异种移植瘤对DDP的敏感性(P<0.01)。
抑制HSP70表达可能通过促进细胞凋亡,在体内和体外均可增强宫颈癌细胞对DDP的敏感性,提示HSP70有望成为宫颈癌基因治疗的新靶点。