Role of P-glycoprotein in cyclosporine cytotoxicity in the cyclosporine-sirolimus interaction
Role of P-glycoprotein in cyclosporine cytotoxicity in the cyclosporine-sirolimus interaction
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DOI：
10.1038/sj.ki.5001649
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发表时间：
2006-09-01
影响因子：
19.6
通讯作者：
Thervet, E.
中科院分区：
文献类型：
--
作者：
Anglicheau, D.;Pallet, N.;Thervet, E.
环孢素肾毒性仍然是实体器官移植中的主要副作用，并且可通过西罗莫司伴随给药而加重。环孢素和西罗莫司是P-糖蛋白（Pgp）底物。我们假设Pgp活性水平可能通过干扰Pgp从肾小管细胞内清除环孢素的能力而影响环孢素的细胞毒性，环孢素和西罗莫司对Pgp功能的相互作用可能解释西罗莫司增强环孢素肾毒性。通过细胞活力和细胞毒性测定，在正常人肾上皮细胞（HREC）的原代培养物中评价环孢素的细胞毒性。维拉帕米、奎宁、PSC 833和PGP-4008被用作Pgp抑制剂。罗丹明-123（R-123），Pgp的荧光底物，用于评估Pgp介导的转运。通过高效液相色谱-串联质谱法测定细胞内环孢素浓度。Pgp表达和功能在HREC中得到证实，并且在该模型中显示环孢霉素和西罗莫司是Pgp抑制剂。维拉帕米对Pgp表达的抑制可使细胞内环孢素浓度显著升高（P < 0.05）。环孢素对HREC产生浓度依赖性细胞毒作用，抑制Pgp活性可显著增加细胞毒作用。西罗莫司对HREC中的R-123外排产生抑制作用，并以剂量依赖性方式增加细胞内环孢素浓度。这些数据表明，Pgp在保护肾上皮细胞免受环孢素毒性中起着关键作用。西罗莫司对Pgp介导的外排和环孢素细胞浓度的抑制作用可以解释临床观察到的环孢素肾毒性加重。
Cyclosporine nephrotoxicity remains a major side effect in solid organ transplantation, and can be exacerbated by concomitant administration of sirolimus. Cyclosporine and sirolimus are P-glycoprotein (Pgp) substrates. We hypothesized that the Pgp activity level may affect cyclosporine cytotoxicity by interfering with the ability of Pgp to remove cyclosporine from within tubular cells, and that an interaction between cyclosporine and sirolimus on Pgp function may explain the enhancement of cyclosporine nephrotoxicity by sirolimus. Cyclosporine cytotoxicity was evaluated in primary cultures of normal human renal epithelial cells (HRECs) by cell viability and cytotoxicity assays. Verapamil, quinine, PSC833, and PGP-4008 were used as Pgp inhibitors. Rhodamine-123 (R-123), a fluorescent substrate of Pgp, was used to assess Pgp-mediated transport. Cellular cyclosporine concentration was measured by high-performance liquid chromatography coupled to tandem mass spectrometry. Pgp expression and function were confirmed in HRECs and cyclosporine and sirolimus were shown to be Pgp inhibitors in this model. Verapamil-induced inhibition of Pgp led to a significant increase in cellular concentration of cyclosporine (P < 0.05). Cyclosporine exerted a concentration-dependent cytotoxic effect on HRECs that was significantly increased by inhibition of Pgp activity. Sirolimus exerted an inhibitory effect on R-123 efflux in HRECs and increased cellular cyclosporine concentrations in a dose-dependent manner. These data demonstrate that Pgp plays a critical role in protecting renal epithelial cells from cyclosporine toxicity. The inhibitory effect of sirolimus on Pgp-mediated efflux and the cellular concentration of cyclosporine could explain the exacerbation of cyclosporine nephrotoxicity observed clinically.