Saengkhae Chantarawan, Loetchutinat Chatchanok, Garnier-Suillerot Arlette
Laboratoire de Physicochimie Biomoléculaire et Cellulaire, Université Paris Nord, Bobigny, France.
Biophys J. 2003 Sep;85(3):2006-14. doi: 10.1016/S0006-3495(03)74628-1.
Characterization of rhodamine 123 as functional assay for MDR has been primarily focused on P-glycoprotein-mediated MDR. Several studies have suggested that Rh123 is also a substrate for MRP1. However, no quantitative studies of the MRP1-mediated efflux of rhodamines have, up to now, been performed. Measurement of the kinetic characteristics of substrate transport is a powerful approach to enhancing our understanding of their function and mechanism. In the present study, we have used a continuous fluorescence assay with four rhodamine dyes (rhodamine 6G, tetramethylrosamine, tetramethylrhodamine ethyl ester, and tetramethylrhodamine methyl ester) to quantify drug transport by MRP1 in living GLC4/ADR cells. The formation of a substrate concentration gradient was observed. MRP1-mediated transport of rhodamine was glutathione-dependent. The kinetics parameter, k(a) = V(M)/k(m), was very similar for the four rhodamine analogs but approximately 10-fold less than the values of the same parameter determined previously for the MRP1-mediated efflux of anthracycline. The findings presented here are the first to show quantitative information about the kinetics parameters for MRP1-mediated efflux of rhodamine dyes.
若丹明123作为多药耐药（MDR）功能检测方法的特性研究主要集中在P-糖蛋白介导的MDR上。多项研究表明，若丹明123也是多药耐药相关蛋白1（MRP1）的底物。然而，截至目前，尚未对MRP1介导的若丹明流出进行定量研究。测量底物转运的动力学特性是增强我们对其功能和机制理解的有力方法。在本研究中，我们使用了一种连续荧光检测法，用四种若丹明染料（若丹明6G、四甲基若丹明、四甲基若丹明乙酯和四甲基若丹明甲酯）来定量MRP1在活的GLC4/ADR细胞中的药物转运。观察到底物浓度梯度的形成。MRP1介导的若丹明转运依赖于谷胱甘肽。四种若丹明类似物的动力学参数k(a) = V(M)/k(m)非常相似，但比先前确定的MRP1介导的蒽环类药物流出的相同参数值小约10倍。此处呈现的研究结果首次展示了关于MRP1介导的若丹明染料流出动力学参数的定量信息。
