芍药苷-6-O'-苯磺酸酯在Caco-2细胞模型中吸收机制研究
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摘要
目的考察芍药苷-6-O'-苯磺酸酯(代号CP-25)在Caco-2细胞模型中吸收和转运机制。方法体外培养人结肠癌细胞Caco-2,建立高效液相色谱方法(HPLC)测定细胞模型中顶端侧(AP侧)和基底侧(BL侧)中CP-25的浓度并计算表观渗透系数(Papp)和表观渗透比(PDR);考察不同浓度和温度对CP-25转运的影响;分别考察P-糖蛋白(P-gp)抑制剂GF120918、多药耐药蛋白2(MRP2)抑制剂MK571和乳腺癌耐药蛋白(BCRP)抑制剂KO143对CP-25在Caco-2细胞吸收的影响。结果 CP-25(5、10、20μg/ml)从AP侧到BL侧的Papp值随着浓度的增加而增加;在2、5、10、20μg/ml浓度范围内,CP-25的吸收随着浓度的增加而增加,各浓度CP-25 PDR值均小于1. 5;在4℃条件下,CP-25 Papp值与37℃条件下的Papp值相比显著降低; GF-120918和MK571可显著降低CP-25从BL侧到AP侧的Papp值,显著提高从AP侧到BL侧的Papp值; KO143对CP-25双向转运Papp值无影响。结论 CP-25主要吸收机制是被动转运,可能存在主动转运的参与; CP-25是P-gp、MRP2的底物,但并不是BCRP转运蛋白的底物。
Objective To investigate the mechanism of CP-25 absorption and transport in the Caco-2 cell model.Methods Human colon cancer Caco-2 cell line was cultured in vitro. The effect of concentration and temperature on the transport of CP-25 was investigated. A high-performance liquid chromatography(HPLC) methodology was established to detect the concentration of CP-25 in the apical side(AP side) and basolateral side(BL side) and calculate their apparent permeability coefficient(Papp) and apparent permeability ratio(PDR). The effects of different concentration,temperature on CP-25 transport in Caco-2 cells were investigated. The effects of P-glycoprotein(P-gp) inhibitor GF120918,multidrug resistance-associate protein 2(MRP2) inhibitor MK571 and breast cancer resistance protein(BCRP) inhibitor KO143 on CP-25 transport in Caco-2 cells were investigated. Results The Pappvalue of CP-25(5,10,20 μg/ml) from AP surface to BL surface increased with the increasing of concentration; Absorption of CP-25 increased with the concentration increasing(2,5,10,20 μg/ml) and all groups of PDR were less than 1. 5. The Pappvalue of CP-25 significantly decreased at 4 ℃ when compared with the Pappvalue of CP-25 at 37 ℃. GF120918 and MK571 could significantly decrease the Pappvalue of CP-25 transport from the BL side to the AP side and significantly increase the Pappvalue of CP-25 transport from the AP side to the BL side,KO143 has no obvious effect on the bidirectional transport of CP-25. Conclusion Passive transport is the main mechanism of CP-25 absorption,active transport may also be involved. CP-25 is P-gp and MRP2 substrates,but not BCRP substrates.
Objective To investigate the mechanism of CP-25 absorption and transport in the Caco-2 cell model.Methods Human colon cancer Caco-2 cell line was cultured in vitro. The effect of concentration and temperature on the transport of CP-25 was investigated. A high-performance liquid chromatography(HPLC) methodology was established to detect the concentration of CP-25 in the apical side(AP side) and basolateral side(BL side) and calculate their apparent permeability coefficient(Papp) and apparent permeability ratio(PDR). The effects of different concentration,temperature on CP-25 transport in Caco-2 cells were investigated. The effects of P-glycoprotein(P-gp) inhibitor GF120918,multidrug resistance-associate protein 2(MRP2) inhibitor MK571 and breast cancer resistance protein(BCRP) inhibitor KO143 on CP-25 transport in Caco-2 cells were investigated. Results The Pappvalue of CP-25(5,10,20 μg/ml) from AP surface to BL surface increased with the increasing of concentration; Absorption of CP-25 increased with the concentration increasing(2,5,10,20 μg/ml) and all groups of PDR were less than 1. 5. The Pappvalue of CP-25 significantly decreased at 4 ℃ when compared with the Pappvalue of CP-25 at 37 ℃. GF120918 and MK571 could significantly decrease the Pappvalue of CP-25 transport from the BL side to the AP side and significantly increase the Pappvalue of CP-25 transport from the AP side to the BL side,KO143 has no obvious effect on the bidirectional transport of CP-25. Conclusion Passive transport is the main mechanism of CP-25 absorption,active transport may also be involved. CP-25 is P-gp and MRP2 substrates,but not BCRP substrates.
引文
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[3] Chang Y,Jia X Y,Wei F,et al. CP-25,a novel compound,protects against autoimmune arthritis by modulating immune mediators of inflammation and bone damage[J]. Sci Rep,2016,6(10):26239.
[4] Zhang F,Shu J L,Li Y,et al. CP-25,a novel anti-inflammatory and immunomodulatory drug,inhibits the functions of activated human B cells through regulating BAFF and TNF-alpha signaling and comparative efficacy with biological agents[J]. Front Pharmacol,2017,8(22):933-41.
[5] Wang C,Yuan J,Zhang L L,et al. Pharmacokinetic comparisons of Paeoniflorin and Paeoniflorin-6'O-benzene sulfonate in rats via different routes of administration[J]. Xenobiotica,2016,46(12):1142-50.
[6] Yang X D,Wang V,Zhou P,et al. Absorption characteristic of paeoniflorin-6'-O-benzene sulfonate(CP-25)in in situ single-pass intestinal perfusion in rats[J]. Xenobiotica,2016,46(9):775-83.
[7] Yu J,Xiao F,Asenso J,et al. Simultaneous determination of paeoniflorin-6'-O-benzene sulfonate(CP-25)and its active paeoniflorin(Pae)metabolite in rat plasma using UPLC-MS/MS:an application for pharmacokinetic studies[J]. RSC Adv,2016,6(114):113209-18.
[8] Zhu M L,Liang X L,Zhao L J,et al. Elucidation of the transport mechanism of baicalin and the influence of a Radix Angelicae Dahuricae extract on the absorption of baicalin in a Caco-2 cell monolayer model[J]. J Ethnopharmacol,2013,150(2):553-9.
[9] Sha X,Fang X. Transport characteristics of 9-nitrocamptothecin in the human intestinal cell line Caco-2 and everted gut sacs[J]. Int J Pharm,2004,272(12):161-71.
[10] Vaidyanathan J B,Walle T. Transport and metabolism of the tea flavonoid(-)-epicatechin by the human intestinal cell line Caco-2[J]. Pharm Res,2001,18(10):1420-5.
[11] Song L,Xu D. Absorption mechanism of paeoniflorin across Caco-2 monolayer model[J]. Chin Tradit Herbal Drugs,2008,6(39):41-4.
[12] Hidalgo I J,Raub T J,Rorchardt R T. Characterization of the human colon carcinoma cell line(Caco-2)as a model system for intestinal epithelial permeability[J]. Gastroenterology,1989,13(3):736-49.
[13] Gutmann H,Fricker G,Torok M,et al. Evidence for different ABC-transporters in Caco-2 cells modulating drug uptake[J].Pharm Res,1999,16(3):402-7.
[14] Taipalensuu J,Tornblom H,Lindberg G,et al. Correlation of gene expression of ten drug efflux proteins of the ATP-binding cassette transporter family in normal human jejunum and in human intestinal epithelial Caco-2 cell monolayers[J]. J Pharmacol Exp Ther,2001,299(1):164-70.
[15] Tsume Y,Hilfinger J M,Amidon G L. Enhanced cancer cell growth inhibition by dipeptide prodrugs of floxuridine:increased transporter affinity and metabolic stability[J]. Mol Pharm,2008,5(5):717-27.