Function and Regulation of Human Cytochrome P4502S1

描述(由申请人提供)：本申请代表授予R01ES015384的竞争性修订，以回应通知编号(NOT-OD-09-058)和通知标题：NIH宣布可为竞争性修订申请提供恢复法案资金。在家长赠款中，我们正在分析细胞色素P450 2S1(CYP2S1)的功能和酶活性。在这篇名为“CYP2S1与哮喘”的修订中，我们现在将扩展这些研究，以探讨CYP2S1在哮喘的发生和发展中的作用。我们的一些(未发表的)观察表明，这种酶在这种疾病中发挥了作用，特别是可能作为一种反调节机制来减轻个别哮喘发作的严重性。这些观察结果包括以下几点。(A)Cyp2s1在小鼠(和人)肺中高表达。(B)低氧通过低氧诱导因子(HIF)介导的过程上调Cyp2s1在小鼠和人细胞系以及小鼠肺中的表达。(C)在哮喘小鼠模型中，低氧诱导因子是哮喘样反应所必需的。(D)在哮喘样反应过程中，Cyp2s1在小鼠肺中表达上调。(E)Cyp2s1在二十烷类化合物的降解过程中非常活跃，其中许多已知与哮喘的发生和发展密切相关。具体目标1将检验我们的假设，即Cyp2s1活性降低哮喘反应。为此，将利用一种新开发的Cyp2s1基因敲除小鼠来确定Cyp2s1活性的丧失是否会增强急性期和慢性期哮喘样症状的表现。特殊目的2将检验我们的假设，即Cyp2s1基因敲除小鼠和野生型小鼠在哮喘样反应期间二十烷类化合物的分布不同，从而为前者增强的哮喘样反应提供一个潜在的解释。为此，(A)将检查纯化的小鼠Cyp2s1和上述两种类型小鼠的微粒体对相关二十烷类化合物的代谢，以及(B)将确定这些小鼠的肺、细支气管灌洗液(BAL)和血清中二十烷类化合物的分布。因此，特定的目标2将确定其在小鼠组织中的浓度与哮喘样反应的严重程度相关的二十烷类化合物，并确定这些二十烷类化合物是否是Cyp2s1的底物，从而识别可能负责介导Cyp2s1对哮喘样反应的保护作用的特定二十烷类化合物(S)。我们的研究可能会指向哮喘治疗的新靶点，即CYP2S1。我们之前的研究表明，蔬菜中发现的某些天然化合物可以上调Cyp2s1。因此，这些化合物可能是治疗哮喘的潜在新疗法。 公共卫生相关性：这些研究将提供对一种重要的环境疾病--哮喘--的机械性洞察。此外，他们还承诺将CYP2S1酶确定为治疗、改善或预防哮喘的药物的新分子靶点。这类制剂可能包括某些天然化学物质，这些化学物质会影响蔬菜中发现的CYP2S1的活性。

DESCRIPTION (provided by applicant): This application represents a Competitive Revision to grant R01ES015384, in response to the Notice Number (NOT-OD-09-058) and Notice Title: NIH Announces the Availability of Recovery Act funds for Competitive Revision Applications. In the parent grant, we are analyzing the function and enzymatic activity of the enzyme, cytochrome P450 2S1 (CYP2S1). In this revision, "CYP2S1 and Asthma", we will now extend these studies to investigate the role of CYP2S1 in the initiation and progression of asthma. Several of our (unpublished) observations suggest that this enzyme plays a role in this disease, and in particular, may function as a counter- regulatory mechanism to diminish the severity of individual asthmatic episodes. These observations include the following. (a) Cyp2s1 is highly expressed in mouse (and human) lung. (b) Hypoxia upregulates Cyp2s1 expression in mouse and human cell lines and in mouse lung in a process mediated by Hypoxia Inducible Factor (HIF). (c) HIF is required for the asthma-like response in a mouse model of asthma. (d) Cyp2s1 is upregulated in mouse lung during the asthma-like response. (e) Cyp2s1 is very active in the degradation of eicosanoids, many of which are known to be intimately involved in the initiation and progression of asthma. Specific Aim 1 will test our hypothesis that Cyp2s1 activity diminishes the asthmatic response. To this end, a newly developed knockout mouse for Cyp2s1 will be utilized to ascertain whether loss of Cyp2s1 activity enhances the manifestations of asthma-like symptoms in both the acute and chronic phases. Specific Aim 2 will test our hypothesis that the profile of eicosanoids differs between Cyp2s1 knockout and wild-type mice during the asthma-like response, thus providing a potential explanation for the enhanced asthma-like response in the former. To this end, (a) the metabolism of relevant eicosanoids by purified mouse Cyp2s1 and by microsomes from both types of the above mice treated or untreated with the asthma-inducing protocol, will be examined, and (b) the profiles of eicosanoids in lungs, bronchiolar lavage fluid (BAL) and sera from these mice will be determined. Specific Aim 2 will thereby identify eicosanoids whose concentrations in mouse tissues correlate with the severity of the asthma-like response, and also determine whether these eicosanoids are substrates of Cyp2s1, thus identifying the particular eicosanoid(s) that are likely responsible for mediating the protective effect of Cyp2s1 towards the asthma-like response. Our studies will potentially point to a novel target for asthma therapy, namely CYP2S1. Our previous studies suggest that certain natural compounds found in vegetables can upregulate Cyp2s1. These compounds therefore may represent potential novel therapeutics for asthma. PUBLIC HEALTH RELEVANCE: These studies will provide mechanistic insight into an important environmental disease - asthma. Furthermore, they promise to identify the enzyme CYP2S1 as a novel molecular target for agents for the treatment, amelioration or prevention of asthma. Such agents could include certain natural chemicals that impact the activity of CYP2S1, which are found in vegetables.