Immunomodulatory effects of arginine supplementation in colitis and colon cancer

描述（由申请方提供）：炎症性肠病（IBD）由溃疡性结肠炎（UC）和克罗恩病组成，是美国140万受影响患者的主要发病原因，并且可进展为结肠癌。它很难治疗，cosly免疫疗法只能在不到一半的病例中诱导缓解。半必需氨基酸L-精氨酸（L-Arg）作为补充和替代药物的作用一直是我们关注的焦点。我们已经证明了L-Arg在体外和结肠炎中的有益作用的机制。阳离子氨基酸转运蛋白2（CAT 2），L-Arg的诱导型转运蛋白和L-Arg的摄取在小鼠结肠炎中上调，口服L-Arg补充剂可有效治疗葡聚糖硫酸钠（DSS）诱导的上皮损伤和炎症，DSS是一种模拟UC的模型。UC组织中CAT 2表达、L-Arg摄取和L-Arg水平均降低。CAT 2-/-小鼠在结肠炎相关癌（CAC）的氧化偶氮甲烷（AOM）-DSS模型中表现出DSS结肠炎和结肠肿瘤发生的显著恶化。CAT 2-/-小鼠在DSS结肠炎模型中对L-Arg补充没有反应，表明单独L-Arg作为人UC的治疗可能不足以克服其受损的转运到组织中。伤口修复模型中的结肠上皮恢复依赖于CAT 2，这是由于产生L-鸟氨酸（L-Orn）的精氨酸酶利用L-Arg，L-鸟氨酸可以通过鸟氨酸脱羧酶（ODC）代谢以产生多胺，或通过鸟氨酸氨基转移酶（OAT）代谢以产生L-脯氨酸（L-Pro）。虽然抑制腺苷酸酶或敲低腺苷酸酶1（Arg 1）阻止了恢复，但在L-Arg存在下，当加入L-Orn或L-Pro时，这可以完全恢复。敲低OAT，而不是ODC，阻止了L-Arg对恢复的有益作用，暗示OAT在上皮功能的维持中。此外，ODC+/-小鼠表现出DSS结肠炎的改善，组织巨噬细胞NO产生增加，调节性T细胞和巨噬细胞应答增强，表明多胺在该模型中的有害作用。我们假设L-Arg在结肠炎和结肠炎相关肿瘤发生中的益处取决于CAT 2和下游效应子，以改善上皮恢复、先天免疫功能和适应性免疫。在我们的具体目标中，我们将确定是否：1）由于CAT 2缺失和L-Arg可用性丧失导致DSS结肠炎恶化 模拟人UC的疾病是由于上皮或巨噬细胞缺陷，并且如果这通过补充L-Orn或L-Pro与L-Arg的组合来改善; 2.）ODC+/-小鼠中DSS结肠炎的改善是由于上皮或巨噬细胞作用，并且如果其是由iNOS和/或OAT的增强的L-Arg可用性引起的; 3.）CAT 2缺失的加速肿瘤发生是由于上皮或巨噬细胞缺陷，并且如果该过程可以通过下调ODC或补充L-Orn或L-Pro与L-Arg的组合来有利地调节。通过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的异型增生和癌症的预防提供新的基于大量营养素的策略。

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.