白坚木碱类二聚体在Caco-2细胞模型中的吸收机制研究

Absorption Mechanism of 10-Dehydroxyl-12-Demethoxy-Conophylline in a Caco-2 Monolayer Cell Model

目的考察白坚木碱二聚体(10-dehydroxyl-12-demethoxy-conophylline,wtdr-11)在人类结肠癌细胞系Caco-2细胞模型中吸收和转运机制。方法建立Caco-2单层细胞完整模型,采用超高效液相色谱-荧光法(ultra-high performance liquid chromatography-fluorescence,UPLC-FLR)测定细胞模型中wtdr-11的浓度并计算表观渗透系数(apparent permeability coefficient,P_(app));分别考察不同浓度、时间、温度、pH值、P-糖蛋白(P-glycoprotein,P-gp)抑制剂(维拉帕米和环孢素A)对wtdr-11在Caco-2细胞内摄取的影响;考察时间变化对wtdr-11在Caco-2细胞中双向转运分别从细胞刷状缘侧(apical,AP)→细胞基底侧(basolateral,BL)和BL→AP方向的影响。结果wtdr-11的吸收随着时间和药物浓度的增加而增加,但温度、pH值和P-gp抑制剂等因素对Caco-2细胞吸收该药物的影响无统计学意义(P>0.05)。跨膜转运实验中,P_(app)>10^(-6) cm/s,外排率<1.5。结论wtdr-11的主要吸收机制可能是被动扩散,wtdr-11可能不是P-gp转运蛋白的底物。

Objective To investigate the absorption and transport mechanisms of 10-dehydroxyl-12-demethoxy-conophylline(wtdr-11)in human colon cancer Caco-2 cells.Methods A Caco-2 monolayer cell model was established.Ultra-high performance liquid chromatography-fluorescence was used to measure the concentration of wtdr-11 in the cell model and calculate their apparent permeability coefficient(P_(app)).The influence of concentration,time,temperature,pH,and P-glycoprotein(P-gp)inhibitors(verapamil and cyclosporin A)on wtdr-11 absorption in Caco-2 cells were analyzed,as well as the influence of time change on the bidirectional transport of wtdr-11 in Caco-2 cells[from apical(AP)side to basolateral(BL)side and from BL side to AP side].Results The absorption of wtdr-11 increased with the increase in time and concentration,while the factors such as temperature,pH,and P-gp inhibitors had no significant influence on the absorption of the drug by Caco-2 cells(P>0.05).The transmembrane transport experiment showed a P_(app) of>10^(-6) cm/s and an efflux rate of<1.5.Conclusion Passive diffusion might be the main absorption mechanism of wtdr-11,and wtdr-11 may not be the substrate of P-gp transporter protein.