Cocaine permeability and metabolism in colonic T-84 epithelial cell line.
Cocaine permeability and metabolism in colonic T-84 epithelial cell line.
复制标题
结肠 T-84 上皮细胞系中的可卡因渗透性和代谢。
DOI：
10.1016/s0024-3205(01)01430-8
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发表时间：
2001
期刊：
影响因子：
6.1
通讯作者：
Vidyasagar,D
中科院分区：
文献类型：
--
作者：
Bhat,R;Chari,G;Rao,M;Negrusz,A;Vidyasagar,D
我们以结肠T-84单层为模型，研究了可卡因在人体肠道的吸收、转运和代谢。T-84细胞在含6%新生小牛血清的DMEM/HAM‘s F-12培养液中培养，温度为37℃，接种于1.0μm胶原片上。细胞发育成一个偏振的单层，顶面朝向上室，基侧面朝向下室。单层在7天内产生的跨上皮阻力为≥600Ωcm2。将不同浓度的盐酸可卡因加入无血清培养液中，仅对管腔侧进行分析，并在30分钟和60分钟后从管腔和浆膜侧取出样品进行分析。用气相色谱-质谱法测定可卡因及其代谢物，可卡因通过T-84单层膜的转运随可卡因浓度的增加而线性增加，暴露30min和60min之间无显著差异。即使在800毫克可卡因的作用下，跨上皮细胞的阻力也没有改变，这表明这对单层细胞的存活率没有影响。管腔和浆膜侧均可检测到苯甲酰ecGonine(BE)和ecGonine甲酯(EME)的代谢产物，但未检测到去甲可卡因。腔侧BE和EME浓度显著高于浆膜侧。腔内和浆膜侧的可卡因、BE和EME的合计回收率为可卡因总量的52-80%。虽然新鲜的培养基不代谢可卡因，但先前暴露在单层(无细胞培养基)中的培养基会显著分解为BE和EME，这表明酯酶可能被释放到培养基中。这些结果表明，可卡因在该模型肠上皮细胞系中的转运是简单扩散的，并且是浓度依赖的。这些研究表明，羊水中的可卡因可被胎儿胃肠道吸收。
We studied the uptake, transport and metabolism of cocaine in human intestine using the colonic T-84 monolayers as a model. T-84 cells were grown in DMEM/Ham's F-12 medium containing 6 % newborn calf serum at 37 ° C on 1.0 μm collagen inserts. The cells develop into a polarized monolayer with the apical surface facing the upper chamber and the basolateral surface facing the lower. The monolayers develop a transepithelial resistance of ≥ 600 Ω cm2in 7 days. Varying concentrations of cocaine HCl was added in a serum free medium to the luminal side only, and after 30 min and 60 min samples from the luminal and serosal aspect were removed for analysis. Cocaine and its metabolites were measured by G C/MS. Cocaine transport across T-84 monolayers increased linearly with increasing concentration of cocaine, with no significant difference between 30 min and 60 min of exposure. Transepithelial resistance did not change even at 800 ng of cocaine suggesting no effect on monolayer viability. The metabolites, benzoylecgonine (BE) and ecgonine methyl ester (EME) but not norcocaine were detected in both luminal and serosal side. The concentrations of BE and EME in the luminal side were significantly higher than in the serosal. Combined recovery of cocaine, BE and EME from luminal and serosal sides were 52 – 80 % of total added cocaine. While fresh medium did not metabolize cocaine, media previously exposed to the monolayer (cell-free medium) caused a significant breakdown into BE and EME, suggesting that esterases may be released into the medium. These results indicate transfer of cocaine across this model of intestinal epithelial cell line is by simple diffusion and is concentration dependent. These studies imply that cocaine in swallowed amniotic fluid can be absorbed by the fetal gastrointestinal tract.