Hamilton G, Cosentini E P, Teleky B, Koperna T, Zacheri J, Riegler M, Feil W, Schiessel R, Wenzi E
Department of Surgery, AKH, University School of Medicine, Vienna, Austria.
Anticancer Res. 1993 Nov-Dec;13(6A):2059-63.
In this study we have investigated the effects of the multidrug-resistance (MDR) modifiers verapamil (VPM), cyclosporin A (CsA) and tamoxifen (TMX) on the intracellular pH(pHi) of four colon carcinoma-derived cell lines with low P-glycoprotein expression (CaCo-2, HT-29, SW 620 and SW 480). Addition of VPM (1 mu M), CsA (1 microgram/ml) or TMX (2 microM) in HEPES- or bicarbonate/CO2-buffered Ringer's solution was followed by dose-dependent and reversible decreases of the pHi (0.1-0.3 units) of all cell lines, as measured ratiometrically by the changes in the pH-dependent fluorescence of bis(carboxyethyl)carboxyfluorescein (BCECF). Testing the effects of the resistance modifiers on the Na+/H+ antiporter and bicarbonate trans-porters under appropriate buffer conditions and addition of inhibitors (amiloride, DIDS) revealed that the chemomodulator-induced acidification does not interfere with the function of these major pHi-regulating acid-base transporters. The induction of changes in pHi shows no correlation with MDR-reversing activity of the drugs and our data do not support the P-gp-inhibition-mediated accumulation of acidic substrates as underlying mechanism. In addition to the P-gp-directed MDR-reversal, chemomodulator-induced intracellular acidification may enhance the chemosensitivity of the cells especially under alkaline extracellular conditions, and contribute to the decreased efficacy of MDR-modifiers in acidic extracellular environments and to the chemosensitising effect of VPM in P-gp-negative cell lines.
在本研究中，我们研究了多药耐药（MDR）调节剂维拉帕米（VPM）、环孢素A（CsA）和他莫昔芬（TMX）对四种低P-糖蛋白表达的结肠癌衍生细胞系（CaCo-2、HT-29、SW 620和SW 480）细胞内pH（pHi）的影响。在HEPES或碳酸氢盐/CO2缓冲的林格氏溶液中加入VPM（1μM）、CsA（1μg/ml）或TMX（2μM）后，所有细胞系的pHi均出现剂量依赖性和可逆性降低（0.1 - 0.3个单位），通过双（羧乙基）羧基荧光素（BCECF）的pH依赖性荧光变化进行比率测定。在适当的缓冲条件下测试耐药调节剂对Na+/H+反向转运体和碳酸氢盐转运体的影响，并加入抑制剂（氨氯吡咪、二异丙基氨基磺酸钠），结果显示化学调节剂诱导的酸化并不干扰这些主要的pHi调节酸碱转运体的功能。pHi变化的诱导与药物的MDR逆转活性无关，我们的数据不支持将P-糖蛋白抑制介导的酸性底物积累作为潜在机制。除了针对P-糖蛋白的MDR逆转外，化学调节剂诱导的细胞内酸化可能会增强细胞的化学敏感性，特别是在碱性细胞外条件下，并导致MDR调节剂在酸性细胞外环境中的疗效降低以及VPM在P-糖蛋白阴性细胞系中的化学增敏作用。
