REGULATION OF MICROSOMAL HEMOPROTEINS

描述（来自申请人的摘要）： 具有广泛的医学、经济、法律的和政治后果的毒理学 是外推到人类的测试结果的不确定性， 给实验室动物注射高剂量的外源性物质。 我们的拨款申请 将利用分子和细胞生物学的进展来帮助定义 遗传和环境因素，可能有助于宿主的 抵抗对特定个体的不利健康影响的风险。 我们 目前的研究重点是CYP3A基因亚家族的细胞色素P450， 在人肝脏中显著存在的微粒体血红素蛋白， 糖皮质激素以及诸如异生物质的“应激”条件， 苯巴比妥和多卤代芳香族环境化学品。 CYP3A 参与临床重要药物的代谢， 环孢菌素和硝苯地平以及致癌物如 黄曲霉毒素和苯并芘。 利用新定义的系统 大鼠和人肝细胞在Matrigel上的原代培养物， 基底膜，并允许保留分化状态， 在体外，我们将进行近似比较的调节， CYP3A结构基因及其产物的控制DNA片段 动物和人类细胞之间的相互作用 条件 分子分离和表征技术将是 用于指定调控CYP3A基因表达的核心DNA元件， 肝脏和配体依赖性调节转录蛋白因子 与这些DNA反应元件相互作用。 我们还将使用反向 基因分析，以调查以前未被识别的分类， 在相同的“压力”控制下调控的基因， 肝脏协调一致地适应环境压力。 促进 这项工作是一个细胞培养和组织库，提供动物和 这些调查所需的人类材料 经过深思熟虑 结合这些方法，我们希望能够定量地 描述细胞表达变化的分子事件 由于药物、环境化学品、内分泌控制， 等因素 我们将研究这些影响是如何 可能因人而异。 我们完全期望， 了解基因结构、基因表达和疾病结果， 提出的研究方案。

DESCRIPTION (from Applicant's Abstract): A major unsolved problem in toxicology having broad medical, economic, legal, and political consequences is the uncertainty of extrapolation to humans of the results of tests of high doses of xenobiotics given to laboratory animals. Our grant proposal will utilize advances in molecular and cellular biology to help define the genetic and environmental factors that may contribute to the host's resistance to risks for an adverse health effect to a given individual. We are focusing on the cytochromes P450 of the CYP3A gene subfamily, a group of microsomal hemoproteins prominent in human liver which are induced under conditions of "stress" by glucocorticoids and also by such xenobiotics as phenobarbital, and polyhalogenated aromatic environmental chemicals. CYP3A are involved in the metabolism of clinically important drugs such as Cyclosporin and nifedipine and the bioactivation of such carcinogens as aflatoxins and benzo[a]pyrene. Taking advantage of a newly defined system primary culture of rat and human hepatocytes on Matrigel, a reconstituted basement membrane and permits retention of the differentiated state in vitro, we will make proximate comparisons of the regulation of the controlling DNA segments of the CYP3A structural genes and their products between animal and human cells under defined, rigorously controlled conditions. Molecular isolation and characterization techniques will be used to specify the core DNA elements that regulate CYP3A gene expression in the liver and the igand dependent regulatory transcription protein factors that interact with these DNA response elements. We will also use reverse genetic analysis to investigate a previously unrecognized assortment of genes regulated under the same "stress" controls that extends beyond the liver acting in concert to adapt to environmental stressors. Facilitating this work is a Cell Culture and Tissue Bank which provides the animal and human material needed for these investigations. Through a thoughtful combination of these approaches, we expect to be able to quantitatively describe the molecular events that underlie changes in cellular expression of the CYP3A genes due to drugs, environmental chemicals, endocrine controls and other factors. We will investigate the question of how these effects may differ among individuals. We fully expect that refinements in understanding gene structure, gene expression, and disease outcome can be achieved by the proposed research program.