MDR1 IN HUMAN INTESTINE

MDR1(多药耐药)是一种多功能的ATP依赖泵，它 从细胞质中运输各种无关的物质来减少 它们在细胞内的堆积。多药耐药基因在许多正常组织中的表达 组织，包括肠上皮的管腔表面，在那里我的 数据表明，mdr1是外源物质屏障的一部分。 我发现在个体之间存在着显著的差异 多药耐药基因在小肠组织中的表达。此外，我 已经表明这种可变性在很大程度上解释了 MDR1底物口服药代动力学的患者间差异， 环孢素A。我认为mdr1表达的这种变异很可能是 影响多药耐药底物的口腔清除，并可能代表 以前未被认识到的环境疾病风险因素。这个 我要检验的第一个假设是肠道mdr1的变异 表达在很大程度上是由于非遗传因素。这一假设是 根据我的初步数据显示MDR1在体外和在 利福平在人体肠道内的活体作用。在具体目标1中，我将确定 多药耐药相关基因在小肠和结肠组织中的表达 含有可疑诱导剂的培养的人类肠道外植体。具体而言 目标2，在目标1中确定的诱导剂将通过体内测试 检测小肠和大肠活检组织中多药耐药基因的表达 健康受试者在4天无诱导剂饮食后进行内窥镜检查 在使用诱导剂进行短期治疗后再次出现。肠道外植体 从每个受试者身上提取的蛋白质将与诱导剂同时培养 体内和体外结果的相关性。我还会测试一下 饮食对肠道多药耐药基因表达的影响 志愿者的饮食中含有木炭烤牛肉或与之合作 还有保罗·沃特金斯，西柚汁(类黄酮类)。多药耐药基因mdr1水平 和蛋白质将在获得的小肠和大肠活检中被检测 在研究开始时，在4天无诱导剂饮食后，以及在7天后 测试饮食。具体目标3将直接测试假设 肠道多药耐药水平影响药物在体内的吸收 肠子。我将首先检查mdr1的表达水平是否影响 多药耐药底物的“吸收”阈值和速率， 环孢素A和长春花碱，跨培养的Caco-2细胞单层。 第二，我将确定肠道中的个体间变异 多药耐药基因mdr1的表达与阿司匹林口服药代动力学的相关性 他克莫司、咪达唑仑和乙炔雌二醇(与Paul合作 沃特金斯和艾伦·莱希特曼)。我相信这些数据 研究将引导我提出关于mdr1效应的新假说。 外源生物吸收的表达与环境风险 疾病。

MDR1 (multiple drug resistance) is a versatile ATP-dependent pump which transports a variety of unrelated substances from the cytoplasm to reduce their intracellular accumulation. MDR1 is expressed in many normal tissues including the luminal surface of intestinal epithelia where my data suggest that mdr1 functions as part of a barrier to xenobiotics. I have found there is significant interindividual variation in the expression of mdr1 mRNA and protein in the small bowel. Furthermore, I have shown this variability accounts for a significant portion of the interpatient differences in oral pharmacokinetics of the mdr1 substrate, cyclosporin A. I believe this variation in mdr1 expression is likely to affect the oral clearance of mdr1 substrates and may represent a previously unrecognized risk factor for environmental diseases. The first hypothesis I will test is that variation in intestinal mdr1 expression is largely due to non-genetic factors. This hypothesis is based on my preliminary data showing mdr1 is inducible in vitro and in vivo in human intestine by rifampin. In Specific Aim 1, I will identify inducers of mdr1 mRNA and protein in small bowel and colon by treating cultured human intestinal explants with suspected inducers. In Specific Aim 2, the inducers identified in Aim 1 will be tested in vivo by measuring mdr1 expression in small and large bowel biopsies obtained endoscopically from healthy subjects after a 4 day inducer free diet and again after short term treatment with an inducer. Intestinal explants from each subject will be simultaneously cultured with the inducer for correlation of in vivo and in vitro results. I will also test the effects of diet on intestinal mdr1 expression by placing healthy volunteers on diets containing charcoal broiled beef or, in collaboration with Paul Watkins, grapefruit juice (flavonoids). The level of mdr1 mRNA and protein will be determined in small and large bowel biopsies obtained at study entry, after a 4 day inducer free diet, and after 7 days on the test diet. Specific Aim 3 will directly test the hypothesis that the level of intestinal mdr1 influences the absorption of drugs in the intestine. I will first examine if the level of mdr1 expression affects the threshold and rate of "absorption" of the mdr1 substrates, cyclosporin A and vinblastine, across cultured Caco-2 cell monolayers. Second, I will determine whether interindividual variation in intestinal mdr1 expression correlates with the variable oral pharmacokinetics of tacrolimus, midazolam, and ethinyl estradiol (collaboration with Paul Watkins and Alan Leichtman). I am confident that the data from these studies will lead me to novel hypotheses regarding the effects of mdr1 expression on xenobiotic absorption and the risk of environmental diseases.