甘草与甘遂的配伍对大鼠肠黏膜P-gp的影响

Modulation on the P-glycoprotein in the jejunum by combined use of Glycyrrhiza inflata and Kansui

评价甘草与甘遂配伍对大鼠肠黏膜P糖蛋白(P-glycoprotein)的影响。通过对大鼠口服甘草煎液、甘遂煎液、甘草甘遂合煎液及其合并液1周后,使用垂直型扩散池(ussing chamber)技术,体外评价罗丹明123(rhodamine123,R123)和荧光素钠(fluorescein sodium,CF)经大鼠空肠黏膜的经时吸收方向和分泌方向的透过量和表观渗透系数。R123和CF在接受室的浓度用荧光分光光度计检测。应用实时荧光定量聚合酶链式反应技术(real-time fluorescent quantitative polymerase chain reaction)检测mdr1a基因在肠黏膜中的表达。在ussing chamber实验中,4种药液除甘草外均具有增加R123经吸收方向(mucosa to serosa,M-S)透过性和减少分泌方向(serosa to mucosa,S-M)透过性的趋势。甘草只增加了R123吸收方向的透过,但对分泌方向影响不大,且均无统计学意义;甘遂组与合煎组均使R123S-M透过明显减少(P<0.01);合并组明显增加了R123M-S的透过(P<0.05)。各组的泵出比均明显降低(P<0.05)。而rt-pcr实验中,评价甘遂对肠黏膜的P-gp活性的影响时,发现大鼠灌胃7d和14d甘遂后,与对照组相比都有下调mdr1a表达的趋势,但是没有统计学意义。另外,在评价甘草与甘遂对旁细胞转运CF影响的实验中,甘遂组、合煎组和合并组均明显减少了CF S-M的透过(P<0.01);甘遂组、合煎组和合并组则均明显减少了CF M-S的透过(P<0.05),但甘草对CF转运的影响没有统计学意义。甘遂可能是一种P-gp抑制剂,而甘草与甘遂合用后对R123透过的影响与单用甘遂相似,可能是甘草与甘遂有协同作用,使其毒性成分吸收增加,这可能是两者配伍产生毒性的机制之一。

To investigate the modulation on the P-glycoprotein in the jejunum by combined use of Glycyrrhiza inflata and Kansui with ussing chamber and rt-pcr,Rhodamine 123（R123）,a P-gp substrate and fluorescein sodium （CF）,a model drug of non-P-gp substrate transported by a passive diffusion were taken as investigational drugs. Because these two drugs can be easily assayed and widely used in various research fields. The permeability of R123 or CF via Wistar rat jejunum membranes was evaluated by in vitro ussing chamber after oral administration of four different decoctions of Glycyrrhiza inflata and Kansui for 1 week. And the concentration of R123 or CF was determined by the fluorospectrophotometry in the receiving solution. Meanwhile the expression of mdr1a in P-glycoprotein was detected by real-time fluorescent quantitative PCR. After oral administration of combine decoction of the single drug,the absorptive directed permeability of R123 increased significantly （P 〈0.01）. On the other hand,Kansui and combine decoction of the two drugs also decrease the permeability of secretory directed transport （P 〈0.05）. No action of Glycyrrhiza inflata was found on the secretory transport of R123 [Papp = （2.56 ± 0.38） ×10^-5,cm.s^-1] across the jejunum tissues,while Papp of control group was found [Papp = （2.35 ± 0.27） ×10^-5,cm.s^-1]. After oral administration of Kansui decoction for 1 week and 2 weeks,the levels of mdr1a expression in Wistar rats were lower than that of the control group,but there were no significant difference in the results. Meanwhile,Glycyrrhiza inflata had no effect on transport of CF across the jejunum tissues,though the other three groups could decrease the permeability of CF,as compared with control group. Kansui may slightly inhibit P-glycoprotein function in the intestinal membrane. For another,some compositions in Kansui inhibit P-glycoprotein function,and some others strengthen the tight junction between cells in the intestinal membrane to decrease permeability of CF. As the inhibitory action to P-glycoprotein was enhanced by combination of Glycyrrhiza inflata and Kansui,based on the results,it may be one of the mechanisms of creating toxicity once co-administration of Glycyrrhiza inflata and Kansui.