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Clarification of transcellular transport mechanism of drugs utilizing tissue cultured cell systems

利用组织培养细胞系统阐明药物的跨细胞转运机制

基本信息

批准号：
04452305
负责人：
TSUJI Akira
金额：
$ 4.29万
依托单位：
Kanazawa University
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (B)
财政年份：
1992
资助国家：
日本
起止时间：
1992 至 1993
项目状态：
已结题

项目摘要

Tissue cultured cells, which retain energy-dependent transport and metabolic functions in vivo, are useful as in vitro model to investigate drug transport at the cellular level. The purpose of the present research is to prove, by utilizing proper cultured cell systems, our new hypotheses, which are different from the passive diffusion mechanism for drug transports through the intestinal epithelial cells and brain capillary endothelial cells. The following results including several new findings were obtained :1)Utilizing cultured human intestinal cell line Caco-2, we have successfully proved that characteristics of the apical to basolateral transepithelial transport of [^<14>C]benzoic acid and[^<14>] salicylic acid in Caco-2 are very similar to those obtained for intestinal brush-border membrane vesicle and that the pH dependent intestinal-absorption of weak-acidic drugs is attributed not to the previously believed "pH-partition theory" but to "protoncoupled and carrier-mediated transpo … More rt" for monocarboxylic acids.2)A heterologous gene expression system, Xenopus laevis oocytes injected with mRNA from the intestinal mucosa, were used to prove the carrier-mediated transport of beta-lactam amtibiotics in small intestine. As the results, the intestinal transport of zwitterionic and dicarboxylic-acid beta-lactam antibiotics was confirmed to be mediated by a specialized transport system which is common to dipeptides. beta-lactam antibiotics were also to be transported by carrier-mediated mechanisms in rabbits and humans as well as rats.3)The uptake of [^3H]vincristine (VCR) and [^3H]cyclosporin A(CsA) by cultured monolayrs of bovine brain capillary endothelial cells (BCECs), which express P-glycoprotein (P-gp) at the luminal membrane sruface, was significantly enhanced by treatment of MDR reversing agents, an anti-P-gp monoclonal antibody and metabolic inhibitors. These results indicate that low permeability of VCR and CsA into the brain is caused by the active efflux from BCEC by P-gp and suggest that P-gp functions as blood-brain barrier for lipophilic and cytotoxic comounds. Less

组织培养的细胞在体内保持能量依赖性转运和代谢功能，可用作研究细胞水平药物转运的体外模型。本研究的目的是利用适当的培养细胞系统来证明我们的新假说，即药物通过肠上皮细胞和脑毛细血管内皮细胞转运的被动扩散机制不同。结果表明：1）利用培养的人小肠细胞系Caco-2，我们成功地证明了[^<14>C]苯甲酸和[^<14>]水杨酸在Caco-2中的跨上皮转运特性与在小肠刷状缘膜囊泡中获得的转运特性非常相似，并且弱酸性药物的pH依赖性药物吸收不是以前认为的“pH分配理论”，而是“质子耦合和载体-中介转运 ...更多信息（2）利用外源基因表达系统--非洲爪蟾卵母细胞注射肠粘膜mRNA，证明β-内酰胺类抗生素在小肠中的载体介导转运。结果证实，两性离子和二羧酸β-内酰胺抗生素的肠道转运是由二肽共有的专门转运系统介导的。β-内酰胺类抗生素在兔、人和大鼠中也通过载体介导的机制转运。3）培养的单层牛脑毛细血管内皮细胞（BCEC）对[^3H]长春新碱（VCR）和[^3H]环孢菌素A（CsA）的摄取通过MDR逆转剂处理显著增强，BCEC在管腔膜表面表达P-糖蛋白（P-gp），抗P-gp单克隆抗体和代谢抑制剂。这些结果表明VCR和CsA进入脑的低渗透性是由P-gp从BCEC主动外排引起的，并表明P-gp作为亲脂性和细胞毒性化合物的血脑屏障发挥作用。少