毛蕊异黄酮体外转运研究及对多药耐药基因MDR1表达的影响

Transport Characteristics of Calycosin and Its Effect on Multidrug Resistance Gene Expression in Vitro

目的研究毛蕊异黄酮在体外模型Caco-2模型的转运特征,及对多药耐药基因MDR1表达的影响。方法采用MTT法确定毛蕊异黄酮对Caco-2模型作用的适用浓度范围;考察浓度、p H、温度及P-糖蛋白(P-gp)抑制剂(维拉帕米)和能量代谢抑制剂(叠氮化钠)对毛蕊异黄酮转运的影响;抑制多药耐药基因MDR1对毛蕊异黄酮转运的影响。结果从上层(AP)到下层(BL),毛蕊异黄酮的Papp>10-6cm/s,表明吸收性良好;毛蕊异黄酮的转运与其浓度和时间呈正相关,4℃条件下毛蕊异黄酮的Papp与25、37℃下的Papp差异有统计学意义[AP→BL:(1.85±0.16)×10-6cm/s比(3.05±0.13)×10-6cm/s、(3.89±0.31)×10-6cm/s,P<0.01;BL→AP:(1.94±0.29)×10-6cm/s比(3.49±0.18)×10-6cm/s、(4.06±0.26)×10-6cm/s,P<0.01];p H=9时的Papp与p H=5、7.4时的Papp值差异有统计学意义[(1.13±0.16)×10-6cm/s比(3.46±0.32)×10-6cm/s、(2.72±0.26)×10-6cm/s,P<0.01]。毛蕊异黄酮的转运不受MDR1基因的影响;毛蕊异黄酮的BL侧到AP侧和AP侧到BL侧的Papp比值<1.5,不受能量代谢影响。结论毛蕊异黄酮在Caco-2模型的转运方式以被动扩散为主,不受P-gp和能量代谢的影响,且毛蕊异黄酮在低温和碱性环境下不利于其吸收。

Objective To investigate the transport characteristics of calycosin across the Caco-2 cell monolayer and its effect on multidrug resistance （MDR1） gene expression. Methods Therapeutic concentration range of caly-cosin against Caco-2 cell monolayer model was determined by MTT method. The effects of concentration, pH, tem-perature, P-gp inhibitor （verapamil） and energy metabolism inhibitor （sodium azide） on the absorption of calycosin were assessed. The effect of the suppression of MDR1 on calycosin transport was evaluated. Results The Papp of calycosin transport from apical （AP） side to basolateral （BL） side was over 10-6 cm/s, which showed a good ab-sorption. The transport of calycosin was positively correlated with time and concentration. The transport of calycosin was influenced by the change of temperature [AP to BL：（1.85±0.16）í10-6cm/s at 4℃ vs （3.05±0.13）í10-6cm/s at 25℃ and （3.89±0.31）í10-6cm/s at 37℃, P〈0.05; BL to AP：（1.94±0.29）í10-6cm/s at 4℃ vs （3.49±0.18）í10-6cm/s at 25℃ and （4.06±0.26）í10-6cm/s at 37℃, P〈0.05] and pH[（1.13±0.16）í10-6cm/s at pH=9 vs （3.46±0.32）í10-6cm/s at pH=5 and （2.72±0.26）í10-6cm/s at pH=7.4, P〈0.05]. The transport of calycosin was not affected by MDR1 gene expression. The number of PappBL→AP/PappAP→BL was less than 1.5, indicating that energy metabolism did not play a role in calycosin transport. Conclusion The transport of calycosin across Caco-2 cell monolayer model is passive diffusion, and it is not influenced by the change of P-gp and energy metabolism. Low temperature and alkaline environment is not conducive to the absorption of calycosin.