T cell activation and crypt cell apoptosis

描述（由申请人提供）：IBD患者的慢性肠道组织炎症水平与隐窝细胞凋亡增加和结直肠癌发展风险增加相关。我们的研究结果表明p53激活是介导肠道炎症状态下隐窝细胞凋亡的关键步骤。p53的主要作用是通过修复DNA损伤和/或诱导细胞凋亡来保护发育中的细胞。观察到p53突变在慢性UC患者早期（甚至在发育不良之前）被检测到，而在散发性结肠癌中，p53突变发生相对较晚，这突出了p53在肠道中的重要性。具体来说，70%的UC相关癌症和20%的UC发育不良病变包含p53突变。这些临床观察增加了在组织炎症过程中发现p53诱导和激活机制的重要性。本研究将利用抗cd3单克隆抗体处理的T细胞诱导隐窝细胞凋亡小鼠模型。基于初步结果，我们假设上皮细胞中的上皮TNF受体1和2信号诱导上皮p53表达，而bm来源细胞中TNF诱导的iNOS表达释放NO，激活p53蛋白并诱导下游p53靶基因表达参与隐窝细胞凋亡。本提案将探讨这一途径的要素。首先，我们将研究在tell诱导的p53激活过程中涉及TNF受体信号传导和iNOS诱导的细胞和分子途径。这些研究将限制TNFR-1、TNFR-2和iNOS在上皮细胞和bm来源细胞中的表达，并研究p53的激活、p53靶点的表达和隐窝细胞凋亡的诱导。接下来，我们计划研究p53的下游效应物。我们将使用特异性基因敲除小鼠（bax/bak和bid-/-）来解决基于野生型和p53无基因小鼠p53靶基因分析的假设。我们之前的研究显著推进了我们对T细胞诱导的隐窝细胞凋亡的关键步骤的理解，但仍有一些关键问题尚未得到解答。目前的建议将使我们更接近于理解肠道炎症中诱导隐窝细胞凋亡所需的细胞和分子事件。这些研究将增强我们对正常隐窝细胞死亡相关机制的理解，并深入了解IBD诱导肠道癌变的途径。

DESCRIPTION (provided by applicant): Chronic levels of intestinal tissue inflammation in IBD are associated with increased crypt cell apoptosis and an increased risk for development of colorectal cancer. Our results implicate p53 activation as a key step in mediating crypt cell apoptosis in states of intestinal inflammation. The primary role of p53 is to protect developing cells by repairing DNA damage and/or inducing apoptosis. The importance of p53 in the intestine is highlighted by observations that p53 mutations are detected early in patients with chronic UC (even before dysplasia), whereas in sporadic forms of colon cancer, p53 mutations occur relatively late. Specifically, 70% of UC-associated cancers and 20% of dysplastic lesions analyzed in UC contain p53 mutations. These clinical observations increase the importance of discovering mechanisms for p53 induction and activation during tissue inflammation. Studies in the present aim will utilize the anti-CD3 mAb-treated mouse model of T cell-induced crypt cell apoptosis. Based on preliminary results, we hypothesize that epithelial TNF receptor 1 and 2 signaling in epithelial cells induce epithelial p53 expression whereas TNF-induced iNOS expression in BM-derived cells releases NO that activates p53 protein and induces expression of downstream p53 target genes involved in crypt cell apoptosis. The elements of this pathway will be explored in the current proposal. First, we will examine the cellular and molecular pathways involved in TNF receptor signaling and iNOS induction during Tell-induced activation of p53. These studies will restrict expression of TNFR-1, TNFR-2, and iNOS to epithelial Vs BM-derived cells and examine p53 activation, expression of p53 targets and induction of crypt cell apoptosis. Next, we plan to examine the downstream effectors of p53. We will use specific gene knockout mice (bax/bak and bid-/-) to address hypotheses based on analysis of p53 target genes in wild type and p53 null mice. Our previous studies significantly advanced out understanding of the critical steps in T cell-induced crypt cell apoptosis, yet there are key questions that remain unanswered. The current proposal will move us closer to understanding the cellular and molecular events required for the induction of crypt cell apoptosis in intestinal inflammation. These studies will enhance our understanding of the mechanisms relevant to normal crypt cell death and add insight into the pathways involved in the induction of intestinal carcinogenesis in IBD.