Immunomodulatory effects of arginine supplementation in colitis and colon cancer

描述(申请人提供)：炎症性肠病(IBD)，由溃疡性结肠炎(UC)和克罗恩病组成，是美国140万受影响的人的主要发病率来源，并可发展为结肠癌。它很难治疗，舒适的免疫疗法只在不到一半的病例中诱导缓解。我们一直关注半必需氨基酸L-精氨酸(L-精氨酸)作为补充和替代药物的作用。我们已经证明了L精氨酸在体外和在结肠炎中的有益作用机制。L-精氨酸的诱导性转运体--阳离子氨基酸转运体2和L-精氨酸的摄取在小鼠结肠炎模型中表达上调，口服L-精氨酸对葡聚糖硫酸钠诱导的结肠炎上皮损伤和炎症有有效的治疗作用。UC组织中CAT2表达、L-精氨酸摄取和L-精氨酸水平均降低。在偶氮甲烷(AOM)-DSS结肠炎相关癌(CAC)模型中，CAT2-/-小鼠表现出明显的DSS结肠炎恶化和结肠肿瘤形成。在DSS结肠炎模型中，CAT2-/-小鼠对补充L-精氨酸没有反应，这表明L-精氨酸单独作为人类UC的治疗方法可能不足以克服其受损的组织转运。在创伤修复模型中，结肠上皮的重建依赖于CAT2，这是由于精氨酸酶利用L-精氨酸产生L-鸟氨酸(L-On)，该酶可被鸟氨酸脱羧酶(ODC)代谢生成多胺，或被鸟氨酸转氨酶(OAT)代谢生成L-Pro(L-Pro)。当抑制精氨酸酶或敲除精氨酸酶1(Arg1)可阻止这种恢复，但在L-Arg存在下，加入L-Orn或L-Pro可完全恢复这种恢复。敲除OAT，而不是ODC，阻止了L-Arg对修复的有利影响，暗示OAT参与维持上皮功能。此外，ODC+/-小鼠在DSS结肠炎中表现出改善，组织巨噬细胞NO产生增加，调节性T细胞和巨噬细胞反应增强，表明多胺在该模型中起有害作用。我们推测，L精氨酸在结肠炎和结肠炎相关肿瘤形成中的益处依赖于CAT2及其下游效应分子改善上皮修复、天然免疫功能和获得性免疫。在我们的特定目标中，我们将确定：1)由于CAT2缺失和L-Arg可用性的丧失而导致的DSS结肠炎的恶化 模拟人类UC是由于上皮细胞或巨噬细胞缺陷，如果通过补充L-奥恩或L-Pro与L-精氨酸联合改善这一点；ODC+/-小鼠DSS结肠炎的改善是由于上皮细胞或巨噬细胞效应，如果这是由于提高了L-Arg对诱导型一氧化氮合酶和/或燕麦的利用率；CAT2缺失的加速肿瘤发生是由于上皮细胞或巨噬细胞的缺陷，如果这一过程可以通过下调ODC值或补充L-奥恩或L-Pro联合L-精氨酸进行有益的调节。通过对上皮和免疫功能的研究，这些研究试图为IBD的治疗和结肠炎相关性异型增生和癌症的预防提供新的基于常量营养素的策略。

DESCRIPTION (provided by applicant): Inflammatory bowel disease (IBD), consisting of ulcerative colitis (UC) and Crohn's disease, is a source of substantial morbidity for 1.4 million people affected in the USA, and it can progress to colon cancer. It is difficult to treat, with cosly immunotherapies only inducing remission in less than half of cases. We have been focusing on the role of the semi-essential amino acid, L-arginine (L-Arg) as a complementary and alternative medicine. We have demonstrated mechanisms for beneficial effects of L-Arg in vitro and in colitis. Cationic amino acid transporter 2 (CAT2), the inducible transporter of L-Arg, and uptake of L-Arg are upregulated in murine colitis, and oral L-Arg supplementation is effective as a treatment for epithelial injury and inflammation induced by dextran sulfate sodium (DSS), a model that mimics UC. CAT2 expression, L-Arg uptake, and L-Arg levels are all decreased in UC tissues. CAT2-/- mice exhibit marked exacerbation of DSS colitis, and colon tumorigenesis in the azoxymethane (AOM)-DSS model of colitis-associated carcinoma (CAC). CAT2-/- mice do not respond to L-Arg supplementation in the DSS colitis model, suggesting that L-Arg alone as a therapy for human UC may not be sufficient to overcome its impaired transport into tissues. Colonic epithelial restitution in a wound repair model is dependent on CAT2 and this is due to utilization of L-Arg by arginase that generates L-ornithine (L-Orn), which can be metabolized by either ornithine decarboxylase (ODC) to generate polyamines, or ornithine aminotransferase (OAT) to generate L-proline (L-Pro). While inhibition of arginase or knockdown of arginase 1 (Arg1) prevented restitution, this could be completely restored, in the presence of L- Arg, when either L-Orn or L-Pro was added. Knockdown of OAT, but not ODC, prevented beneficial effects of L-Arg on restitution, implicating OAT in the maintenance of epithelial function. Additionally, ODC+/- mice exhibit improvement in DSS colitis, with increased tissue macrophage NO production, and enhanced regulatory T cell and macrophage responses, indicating a deleterious role for polyamines in this model. We hypothesize that benefits of L-Arg in colitis and colitis-associated tumorigenesis depend on CAT2 and downstream effectors to improve epithelial restitution, innate immune function, and adaptive immunity. In our Specific Aims we will determine if: 1) exacerbation of DSS colitis due to deletion of CAT2 and loss of L-Arg availability that mimics human UC is due to an epithelial or macrophage defect and if this is ameliorated by supplementation of L-Orn or L-Pro in combination with L-Arg; 2.) improvement in DSS colitis in ODC+/- mice is due to an epithelial or macrophage effect and if it results from enhanced L-Arg availability for iNOS and/or OAT; 3.) accelerated tumorigenesis with CAT2 deletion is due to an epithelial or macrophage defect and if this process can be beneficially modulated by downregulation of ODC or supplementation of L-Orn or L-Pro in combination with L- Arg. Through investigation of epithelial and immune function, these studies seek to provide new macronutrient- based strategies for treatment of IBD and prevention of colitis-associated dysplasia and carcinoma.