9-硝基喜树碱在Caco-2细胞模型中的体外摄取、转运及外排动力学

The in vitro kinetics of uptake, transport and efflux of 9-nitrocamptothecin in Caco-2 cell model

目的 研究9-硝基喜树碱(9-NC)的细胞摄取、转运及外排特性。方法 一种体外培养的人小肠上皮细胞模型Caco-2应用于9-NC的小肠七皮细胞的摄取、跨膜转运及外排动力学研究。评价了时间、温度、pH,P-糖蛋白(P-glycoprotein,P-gp)抑制剂对细胞摄取的影响。采用HPLC测定药物含量。结果 9-硝基喜树碱以被动扩散为主要方式被细胞摄取和转运。药物的摄取与时间呈正相关,与温度、pH呈负相关。P-gp抑制剂环孢菌素和维拉帕米增加9-NC细胞摄取(P<0.05)。药物从Basolateral(B,基底面)到Apical(A,肠腔面)的渗透系数Papp大于A到B(2.6—6.9倍)。9-NC外排符合二级外排动力学过程,A侧m0(148.0±2.2)pmol·cm-2和外排速率(41.1 pmol·cm2·min-1)高于B侧的m0[(121±7)pmol·cm-2(P<0.05)和外排速率(29.2 pmol·cm2·min-1)(P<0.01)。结论9-NC是以被动扩散方式为主要方式被小肠上皮细胞摄取和转运,并受到P-糖蛋白强烈的外排作用。

Aim To study the kinetics of uptake, transepithelial transport and efflux of 9-nitrocamptothecin (9-NC). Methods A human intestinal epithelial cell model Caco-2 cell in vitro cultured had been applied to study the kinetics of uptake, transport and efflux kinetics of 9-NC at small intestine. The effects of time, pH, temperature and P-glycoprotein inhibitors on the uptake of 9-NC were investigated. The determination of 9-NC was performed by HPLC. Results The uptake and absorption of 9-NC were passive diffusion as the dominating process. The uptake of 9-NC is positively correlated to uptake time, and negatively correlated to pH and temperature. The inhibitors, cyclosporine A and verapamil, significantly enhanced the uptake amount of 9-NC ( P < 0. 05 ) . Papp of Basolateral to Apical was much more than that of Apical to Basolateral (2.6 -6.9 fold). The efflux of 9-NC was fitted to apparent two-order process. The m0 [ ( 148. 0 ±2. 2) pmol · cm-2 ] and the efflux rate (41. 1 pmol · cm2 · min-1 ) on Apical side were higher than the m0 [ (121 ±7) pmol · cm-2 ] ( P < 0. 05 ) and the efflux rate (29.2 pmol ·cm2 · min-1) on Basolateral side (P < 0. 01). Conclusion The uptake and absorption of 9-NC were passive diffusion as the dominating process. P-glycoprotein had strong efflux effects on the uptake and transepithelial transport of 9-NC.