Augmentation of Transport for Cisplatin-glutathione Adduct in Cisplatinresistant Cancer Cells

我们研究了顺铂(CDDP) -谷胱甘肽（GSH）加合物（DDP-GSH）在抗CDDP癌细胞中的向外转运。在CDDP存在下培养细胞导致GSH形成加合物并随后运输到细胞外。我们以对CDDP敏感（HCT8）和耐药（HCT8DDP）的人结肠癌细胞和对CDDP敏感（A2780）和耐药（A2780DDP）的人卵巢癌细胞为材料。耐药细胞内GSH浓度较高（分别为118.7<正负>5.9nmol/10^6 HCT8DDP vs 19.0<正负>1.0nmol/10^6 HCT8和24.1<正负>1.2nmol/10^6 A2780DDP vs 9.4<正负>0.5nmol/10^6 A2780）。GSH合成酶γ -谷氨酰半胱氨酸合成酶（γ - gcs）活性在抗cddp细胞中较高（分别为7.1<±>0.2 mmol /10^6 HCT8DDP vs 2.2<±>0.1 mmol /10^6 HCT8和2.9<±>0.1 mmol /10^6 A2780DDP vs 1.4<±>0.1 mmol /10^6 A2780）。此外，与GSH浓度的变化一致，抗cddp细胞中γ - gcs的免疫水平和γ - gcs mRNA的表达均高于对照组。DDP-GSH转运在cdp耐药结肠癌细胞中增加219%(324<正负>12fmol/10^6 HCT8细胞/min vs 148<正负>11fmol/10^6 HCT8细胞/min)，在cdp耐药卵巢癌细胞中增加126%（127<正负>7fmol/10^6 A2780DDP细胞/min vs 101<正负>8fmol/10^6 A2780细胞/min）。DDP-GSH的运输也通过这些细胞的内-外囊泡来估计。HCT8DDP中DDP-GSH的主动转运量为HCT8的243%，A2780DDP中A2780的121%。这些数据表明，在癌细胞中获得抗cddp的部分原因是由于DDP-GSH在细胞外的转运增加以及GSH浓度的增加。对膜蛋白对人红细胞谷胱甘肽偶联物刺激的Mg^<2+>- atp酶血清的免疫评价没有发现明显的交叉反应性，这表明DDP-GSH存在多种转运系统。少

We studied the outward transport of cisplatin (CDDP) -glutathione (GSH) adduct (DDP-GSH) in CDDP-resistant cancer cells. Incubating the cells in the presence of CDDP resulted in the formation of an adduct with GSH and subsequent transport outside the cells. We used human colonic cancer cells sensitive (HCT8) and resistant (HCT8DDP) to CDDP,and human ovarian cancer cells sensitive (A2780) and resistant (A2780DDP) to CDDP as materials.The concentration of intracellular GSH was higher in the resistant cells (118.7<plus-minus>5.9nmol/10^6 HCT8DDP vs 19.0<plus-minus>1.0nmol/10^6 HCT8 and 24.1<plus-minus>1.2nmol/10^6 A2780DDP vs 9.4<plus-minus>0.5nmol/10^6 A2780, respectively). The activity of the GSH synthesizing enzyme, gamma-glutamylcysteine synthetase (gamma-GCS) was higher in the CDDP-resistant cells (7.1<plus-minus>0.2milliunits/10^6 HCT8DDP vs 2.2<plus-minus>0.1milliunits/10^6 HCT8, and 2.9<plus-minus>0.1milliunits/10^6 A2780DDP vs 1.4<plus-minus>0.1milliunits/10^6 A2780 respectively) … More . Furthermore, immunological levels of gamma-GCS and the expression of gamma-GCS mRNA were higher in these CDDP-resistant cells than those in the control cells in accordance with the change in the concentration of GSH.DDP-GSH transport increased in the CDDP-resistant colonic cancer cells by 219% (324<plus-minus>12fmol/10^6 HCT8DDP cells/min vs 148<plus-minus>11fmol/10^6 HCT8 cells/min) and the CDDP-resistant ovarian cancer cells by 126% (127<plus-minus>7fmol/10^6 A2780DDP cells/min vs 101<plus-minus>8fmol/10^6 A2780 cells/min).DDP-GSH transport was also estimated using inside-out vesicles from these cells. The active transport of DDP-GSH was 243% of HCT8 in HCT8DDP and 121% of A2780 in A2780DDP.These data suggest that the acquisition of CDDP-resistance in cancer cells is due partly to the increase in the transport of DDP-GSH outside the cells as well as the increase in the concentration of GSH.Immunological estimation of the membrane proteins against human erythrocyte glutathione Sconjugate-stimulated Mg^<2+>-ATPase sera resulted in no apparent cross reactivity, suggesting that there are several transport systems for DDP-GSH. Less