In Vivo Role of NO in Mucosal Inflammation and Cancer

该项目将描述一氧化氮（NO）和NO衍生物质在炎症性肠病（IBD）和癌症小鼠模型中的作用。这些研究将使用129/SvEv激酶激活基因2（Rag-2）敲除（KO）小鼠和C57 BL/6 T细胞受体（TCR）α KO小鼠。129/SvEv背景的Rag-2 KO小鼠在感染H.肝组织学病变和巨噬细胞和中性粒细胞浸润的模式将与Rag-2 KO小鼠中DNA和蛋白质的硝化、氧化和卤化的生物标志物相关。这些生物标志物将通过比较携带初始和效应CD45RB高T细胞的Rag-2 KO小鼠中的疾病以及通过用N-甲基精氨酸（NMA）调节NO产生来进一步验证。当感染H.肝与Rag-2 KO小鼠一样，TCR β KO小鼠中的IBD是克罗恩病（CD）的模型，其特征在于活化的巨噬细胞和丰富的干扰素-γ（IFN）和肿瘤坏死因子-α（TNF）。TCR α KO小鼠是溃疡性结肠炎（UC）的模型，并且疾病的发展依赖于白介素-4（Ki-4）而不是IFN或TNF。组织学病变和细胞因子水平将与TCR α和TCR β KO小鼠中的生物标志物相关。为了进一步表征NO和NO衍生物质在IBD中的作用，我们将比较诱导型一氧化氮合酶（iNOS）的药理学抑制与通过产生iNOS缺陷型TCR α KO小鼠的酶的遗传失活。还将进行在存在和不存在NO和NO衍生物质的情况下体内产生的体细胞突变的体外分析。通过使用新型IBD相关癌症模型以及UC和CD模型，我们将能够验证DNA和蛋白质硝化、氧化和卤化的新生物标志物。我们还将更好地了解NO和NO衍生物质在广泛的致癌事件中的作用，包括遗传毒性，细胞增殖，细胞毒性和血管生成。具体目标是： 具体目标#1。在Rag-2 KO小鼠中表征NO和氧化应激在IBD和癌症中的作用 具体目标#2表征NO和氧化应激在TCR α基因敲除小鼠IBD中的作用

This Project will characterize the role of nitric oxide (NO) and NO-derived species in mouse models of inflammatory bowel disease (IBD) and cancer. These studies will use 129/SvEv Recombinase-activating gene 2 (Rag-2) knockout (KO) mice and C57BL/6 T cell receptor (TCR) alphabeta KO mice. Rag-2 KO mice on a 129/SvEv background develop severe IBD and cancer when infected with H. hepaticus. Histopathologic lesions and the pattern of macrophage and neutrophil infiltration will be correlated with biomarkers for nitration, oxidation, and halogenation of DNA and proteins in Rag-2 KO mice. These biomarkers will be further validated by comparing disease in naive and effector CD45RB high T cell-bearing Rag-2 KO mice, and by modulating NO production with N-methylarginine (NMA). TCR alphabeta KO mice develop severe IBD when infected with H. hepaticus. IBD in TCRbeta KO mice, like in Rag-2 KO mice, is a model for Crohn's disease (CD) and features activated macrophages and abundant interferon-gamma (IFN) and tumor necrosis factor-alpha (TNF). TCR alpha KO mice are a model for ulcerative colitis (UC), and are dependent on interleukin-4 (K,-4) but not IFN or TNF for the development of disease. Histopathologic lesions and cytokine levels will be correlated with biomarkers in TCR alpha and TCR beta KO mice. To further characterize the role of NO and NO-derived species in IBD, we will compare pharmacologic inhibition of inducible nitric oxide synthase (iNOS) with genetic inactivation of the enzyme by generating iNOS-deficient TCR alphabeta KO mice. In vitro analysis of somatic mutations arising in vivo in the presence and in the absence of NO and NO-derived species will also be performed. By using a novel IBD associated cancer model and models for UC and CD, we will be able to validate new biomarkers for nitration, oxidation, and halogenation of DNA and proteins. We will also gain a better understanding of the role of NO and NO-derived species across a broad range of carcinogenic events, including genotoxicity, cellular proliferation, cytotoxicity, and angiogenesis. The Specific Aims are: Specific Aim #1. Characterize the role of NO an oxidative stress in IBD and cancer in Rag-2 KO mice Specific Aim #2. Characterize the role of NO and oxidative stress in IBD in TCR alphabeta KO mice