PAHs: Balance of Detoxication vs Metabolic Activation

描述（由申请人提供）：燃烧产物中存在多环芳烃（PAHs），亚硝胺和n -杂环，例如烧烤食品和香烟烟雾。细胞色素P450 1A1和1B1 （CYP1A1, CYP1B1）负责多种多环芳烃的代谢，其原型是苯并[a]芘（BaP）。CYP1A2负责亚硝胺和n -杂环的代谢，但在较小程度上也负责多环芳烃（特别是BaP）的代谢。使用Cyp1a1（-/-）和Cyp1b1（-/-）敲除小鼠，我们已经证明Cyp1a1在解毒中比代谢激活更重要，而Cyp1b1导致BaP代谢激活到不需要的反应中间体。换句话说，在摄入BaP的完整小鼠中，CYP1A1的益处大于坏处，而在通过各种途径摄入PAHs的完整小鼠中，CYP1B1的益处大于坏处。对于口服BaP而言，肠系膜淋巴系统与门静脉系统（肠系膜血管、肝脏、胆汁）的重要性尚不清楚。这个实验室现在有7个基因敲除cyp1的小鼠系——三个是单基因敲除，三个是双基因敲除，一个是三基因敲除cyp1的小鼠系。我们的假设是：淋巴BaP摄取和远端组织（如免疫细胞、脾脏和骨髓）的CYP1B1是口服BaP毒性的主要原因，而肝脏和肠道中诱导的CYP1A1是BaP解毒的主要原因。因此，在这个拟议的项目中，我们将：[a]鉴定并确定野生型和所有7种Cyp1敲除小鼠系的肠系膜淋巴、门静脉血液、肝脏和胆汁中代谢物与未改变亲本BaP的数量，以及乳糜微粒在将BaP输送到靶器官中的作用和机制；[b]产生肝脏和肠道上皮特异性Cyp1a1条件敲除细胞系；[c]将基因组中的Cyp1b1基因替换为Cyp1a1基因，反之亦然；[d]在这四种新生成的小鼠品系中重复我们的BaP药代动力学研究（见[a]）。了解三种CYP1酶在接受口服BaP的完整小鼠中的组织特异性作用，将使我们更好地了解BaP解毒与代谢激活。我们期望这些知识将为理解摄入BaP的消除和传播机制提供蓝图，并将在临床研究中提供信息，我们将确定这三种人类基因的哪一种单倍型可能与多环芳烃诱导的毒性和癌症的抗性和敏感性相关。

DESCRIPTION (provided by applicant): Polycyclic aromatic hydrocarbons (PAHs), nitrosamines, and N-heterocyclics are present in combustion products, e.g. grilled foods & cigarette smoke. Cytochromes P450 1A1 & 1B1 (CYP1A1, CYP1B1) are responsible for the metabolism of numerous PAHs, the prototype of which is benzo[a]pyrene (BaP). CYP1A2 is responsible for metabolizing nitrosamines and N-heterocyclics, but also PAHs (in particular, BaP) to a lesser extent. Using Cyp1a1(-/-) and Cyp1b1(-/-) knockout mice, we have shown that CYP1A1 is more important in detoxication than metabolic activation, whereas CYP1B1 causes metabolic activation of BaP to unwanted reactive intermediates. In other words, CYP1A1 is more good than bad in the intact mouse ingesting BaP, and CYP1B1 is more bad than good in the intact mouse administered PAHs by various routes. The importance of mesenteric lymphatics vs. the portal system (mesenteric blood vessels, liver, bile) is not known for oral BaP. This lab now has seven-all three single, all three double, and the one triple-Cyp1 knockout mouse lines. Our hypothesis is: lymph BaP uptake and CYP1B1 in distal tissues (e.g. immune cells, spleen, and bone marrow) are the principal causes of oral BaP toxicity, whereas inducible CYP1A1 in liver and intestine is the principal cause of BaP detoxication. In this proposed project, we therefore will: [a] identify and determine the amounts of metabolites vs. unchanged parent BaP in mesenteric lymph, portal vein blood, liver, and bile in wild-type and all seven Cyp1 knockout mouse lines, and the role and mechanism of chylomicrons in delivering BaP to target organs; [b] generate liver- and intestinal epithelium-specific Cyp1a1 conditional knockout lines; [c] replace the Cyp1b1 gene (in the genome) with the Cyp1a1 gene, and vice versa; and [d] repeat our BaP pharmacokinetics studies (see [a]) in these four newly generated mouse lines. Understanding the tissue- specific roles for each of the three CYP1 enzymes in the intact mouse receiving oral BaP will provide us with a greater understanding of BaP detoxification vs. metabolic activation. We expect this knowledge will provide a blueprint for understanding the mechanisms of elimination vs. dissemination of ingested BaP and will be informative in clinical studies in which we would determine which haplotypes of these three human genes might be associated with resistance vs. sensitivity to PAH-induced toxicity and cancer.