pH and drug resistance.: I.: Functional expression of plasmalemmal V-type H+-ATPase in drug-resistant human breast carcinoma cell lines
pH and drug resistance.: I.: Functional expression of plasmalemmal V-type H+-ATPase in drug-resistant human breast carcinoma cell lines
复制标题
DOI：
10.1016/s0006-2952(99)00022-2
复制
发表时间：
1999-05-01
影响因子：
5.8
通讯作者：
Gillies, RJ
中科院分区：
文献类型：
--
作者：
Martínez-Zaguilán, R;Raghunand, N;Gillies, RJ
有效治疗癌症的主要障碍是许多肿瘤细胞表现出的多药耐药（MDR）现象。许多，但不是所有的MDR细胞表现出膜相关的P-糖蛋白（P-gp），药物外排泵。然而，大多数MDR的机制是复杂的，采用P-gp与其他不明确的活动相结合。改变的细胞溶质pH（pH（i））已被认为在耐药性中起作用。在目前的研究中，我们研究了药物敏感（MCF-7/S）和耐药人乳腺癌细胞中pH（i）调节的机制。在耐药细胞系中，一个含有P-gp（MCF-7/DOX;也称为MCF-7/D40），一个不含P-gp（MCF-7/MITOX）。三种细胞系的静息稳态pH（i）相似。此外，在所有细胞系中，HCO3-轻微酸化pH（i），并增加酸负荷后pH（i）恢复的速率，表明存在阴离子交换剂（PIE）活性。这些数据表明，在这些细胞系中，Na +/H+交换和AE都没有差异表达。然后研究了这些细胞系中质膜液泡型H +-ATP酶（pmV-ATP酶）活性的存在。在缺乏Na+和HCO3-的情况下，MCF-7/S细胞不能从酸负荷中恢复，而MCF-7/MITOX和MCF-7/DOX细胞可以。此外，pH（i）的恢复被巴夫洛霉素A（1）和NBD-Cl（有效的V-ATP酶抑制剂）抑制。试图本地化V-ATP酶免疫细胞化学在这些细胞的质膜是不成功的，表明V-ATP酶是不是静态驻留在质膜。与此相一致的是，与药物敏感细胞相比，耐药细胞中内体捕获的葡聚糖的释放更快。此外，耐药细胞将阿霉素截留到胞内囊泡中，而药物敏感细胞则没有。因此，假设耐药细胞中测得的pmV-ATP酶活性是内膜快速更新的结果。这种行为对耐药性的潜在影响在配套手稿中进行了研究。(C)1999 Elsevier Science Inc.
A major obstacle for the effective treatment of cancer is the phenomenon of multidrug resistance (MDR) exhibited by many tumor cells. Many, but not all, MDR cells exhibit membrane-associated P-glycoprotein (P-gp), a drug efflux pump. However, most mechanisms of MDR are complex, employing P-gp in combination with other, ill-defined activities. Altered cytosolic pH (pH(i)) has been implicated to play a role in drug resistance. In the current study, we investigated mechanisms of pH(i) regulation in drug-sensitive (MCF-7/S) and drug-resistant human breast cancer cells. Of the drug-resistant lines, one contained P-gp (MCF-7/DOX; also referred to as MCF-7/D40) and one did not (MCF-7/MITOX). The resting steady-state pH(i) was similar in the three cell lines. In addition, in all the cell lines, HCO3- slightly acidified pH(i) and increased the rates of pH(i) recovery after an acid load, indicating the presence of anion exchanger (PIE) activity. These data indicate that neither Na+/H+ exchange nor AE is differentially expressed in these cell lines, The presence of plasma membrane vacuolar-type H+-ATPase (pmV-ATPase) activity in these cell lines was then investigated. In the absence of Na+ and HCO3-, MCF-7/S cells did not recover from acid loads, whereas MCF-7/MITOX and MCF-7/DOX cells did. Furthermore, recovery of pH(i) was inhibited by bafilomycin A(1) and NBD-Cl, potent V-ATPase inhibitors. Attempts to localize V-ATPase immunocytochemically at the plasma membranes of these cells were unsuccessful, indicating that V-ATPase is not statically resident at the plasma membrane. Consistent with this was the observation that release of endosomally trapped dextran was more rapid in the drug-resistant, compared with the drug-sensitive cells. Furthermore, the drug-resistant cells entrapped doxorubicin into intracellular vesicles whereas the drug-sensitive cells did not. Hence, it is hypothesized that the measured pmV-ATPase activity in the drug-resistant cells is a consequence of rapid endomembrane turnover. The potential impact of this behavior on drug resistance is examined in a companion manuscript. (C) 1999 Elsevier Science Inc.