调控线粒体通透转运孔道对人肝癌耐药细胞SK-Hep1/CDDP多药耐药的影响

Multidrug Resistance in Hepatoma Cell Line SK-Hep1/CDDP by Modulation of the Mitochondrial Permeability Transition Pore

采用大剂量冲击、间歇诱导法获得人肝癌顺铂(cisplatin,CDDP)多药耐药细胞系SK-Hep1/CDDP。利用线粒体通透转运孔道(mitochondrial permeability transition pore,mPTP)开放剂苍术苷(atractyloside,ATR)和抑制剂环孢素A(cyclosporin,CsA)分别干预SK-Hep1和SK-Hep1/CDDP细胞;免疫印迹法检测多药耐药基因Mdr-1和Bax表达水平;采用Annexin V/PI双标记法检测细胞凋亡率;采用荧光探针JC-1检测线粒体膜电位△Ψm的变化。探讨调控线粒体mPTP对人肝癌耐药细胞SK-Hep1/CDDP多药耐药的影响。结果提示,ATR可促进mPTP开放,加速△Ψm下降,同时降低Bax活性,增加SK-Hep1/CDDP细胞凋亡;CsA抑制mPTP开放,可减轻并延迟线粒体膜电位下降,使多药耐药细胞SK-Hep1/CDDP对CDDP诱导凋亡的耐受能力提高,同时增加Bax活性。但mPTP活性的变化对细胞Mdr-1蛋白表达水平无影响。mPTP的激活可能成为增加肿瘤细胞对化疗药物敏感性及逆转肿瘤细胞多药耐药的新方法。

SK-Hepl/CDDP cell line was induced by pulse treament with a high concentration of cisplatin （CDDP） in vitro. SK-Hepl and SK-Hepl/CDDP were treated by mitochondrial permeability transition pore （mPTP） reactivator atractyloside （ATR） and mPTP inhibitor cyclosporin （CsA） respectively. Expressions of Mdr-1 and Bax were detected by Western blotting. Apoptosis of the cells was assessed with FITC-Annexin V. Mitochondrial inner membrane potential （△ψm） were monitored with fluorescence dry Jc-1. The results showed that mPTP agonist ATR promoted the opening of the mPTP, accelerated the loss of △ψm, enhanced apoptosis induced by CDDP, and also increased Bax activity, Whereas mPTP blocker CsA blocked the mPTP, delayed the loss of △ψm, inhibited apoptosis induced by CDDP, and also inhibited Bax activity. Meanwhile there was no influence on mdr-1 expression. The results suggested that activation of mPTP might be efficient in clinic trials for cancer treatment and reversal of multidrug resistance.