Tissue-specific contribution of Selenoprotein P in colitis and oxidative damage

 说明(申请人提供)：硒是一种必需的微量元素，以硒半胱氨酸的形式被结合到硒蛋白中，如硒蛋白P(SEPP1)。一些流行病学研究表明，营养硒状况与癌症风险呈负相关，尤其是在结肠癌中。由于SEPP1是唯一一种含有一种以上硒半胱氨酸的硒蛋白，SEPP1被假设为向各种组织供应硒，以允许原位生成含硒蛋白。然而，SEPP1也含有一个氧化还原结构域，并具有广泛的抗氧化功能。这些活动表明SEPP1可能在癌症预防中发挥重要作用，确实SEPP1信息在结直肠癌中下调。此外，我们还发现，SEPP1的全局减少增加了服用结肠炎相关癌症方案的小鼠的肿瘤发生。但是SEPP1是如何在肿瘤形成中介导这些变化的呢？由于SEPP1的大部分被认为是在肝脏中合成的，我们之前培育了一个肝脏特异性的Sepp1基因敲除小鼠。令人惊讶的是，肝脏中SEPP1的缺失对结肠炎评分或肿瘤形成没有影响，这表明SEPP1的局部来源可能改变或影响肿瘤的发生。这种局部来源的SEPP1的两个可能来源包括1)肠上皮细胞和2)渗入的免疫细胞。这两种类型的细胞都能产生SEPP1，但它们在炎症损伤和癌症中的单独作用尚不清楚。因此，我们将分别删除两个细胞群中的SEPP1，以确定SEPP1在炎症性癌变中的组织特异性贡献。为了确定SEPP1如何影响肿瘤的形成，我们将对上皮细胞和髓系细胞群体采用体内和体外相结合的方法。利用结肠炎和异型增生的小鼠模型，肠样和巨噬细胞/肠样共培养，以及骨髓巨噬细胞培养，我们将确定组织特异性SEPP1如何影响肠道稳态、肠道损伤、氧化应激和巨噬细胞功能，以及每种功能如何在结肠炎和相关的异型增生中起作用。有趣的是，我们发现，在体外肠道培养模型中，Sepp1缺失会增加氧化损伤，而结肠炎相关癌的特征是氧化应激增加。为了具体研究氧化损伤的作用及其对SEPP1表型的机制贡献，我们将利用氧化加合物的抑制剂水杨胺以及氧化还原缺陷的SEPP1变体。总之，这些实验将阐明硒和SEPP1在结肠疾病和疾病进展中的作用。

 DESCRIPTION (provided by applicant): Selenium is a necessary trace element that is incorporated as selenocysteine into selenoproteins, such Selenoprotein P (SEPP1). Several epidemiological studies have inversely correlated nutritional selenium status and cancer risk, particularly in colon cancer. Because SEPP1 is the only selenoprotein to contain more than one selenocysteine, SEPP1 is hypothesized to supply various tissues with selenium to allow in situ generation of selenium containing proteins. However, SEPP1 also contains a redox domain and fulfills a generalized antioxidant function. Such activities suggest that SEPP1 could play a significant role in cancer prevention, and indeed SEPP1 message is downregulated in colorectal cancers. Furthermore, we have found that global SEPP1 reduction increases tumorigenesis in mice placed on a colitis associated carcinoma protocol. But how is SEPP1 mediating these changes in tumor formation? As the majority of SEPP1 is thought to be synthesized in the liver, we previously generated a liver- specific Sepp1 knock-out mouse. Surprisingly, loss of SEPP1 in the liver had no effect on colitis score or tumor formation, suggesting a local source of SEPP1 might modify or influence tumorigenesis. Two possible sources for this locally-derived SEPP1 include 1) intestinal epithelial cells and 2) infiltrating immune cells. Both cell types are known o produce SEPP1, but their separate contributions to inflammatory injury and cancer are yet unknown. Therefore, we will delete SEPP1 in both cell populations individually to define tissue-specific contributions of SEPP1 to inflammatory carcinogenesis. To determine how SEPP1 influences tumor formation, we will utilize a combination of in vivo and ex vivo approaches for both epithelial and myeloid cell populations. Using mouse models of colitis and dysplasia, intestinal enteroid and macrophage/enteroid co-culture, and bone marrow macrophage culture we will determine how tissue-specific SEPP1 affects intestinal homeostasis, intestinal injury, oxidative stress, and macrophage function, and how each function contributes to colitis and associated dysplasia. Interestingly, we have found that Sepp1 loss increases oxidative damage in ex vivo intestinal culture models, and colitis- associated carcinoma is characterized by increased oxidant stress. To specifically investigate the role of oxidative damage and its mechanistic contribution to the SEPP1 phenotype, we will utilize the compound salicylamine, an inhibitor of oxidative adducts, as well as redox-deficient SEPP1 variants. Together, these experiments will elucidate the roles of selenium and SEPP1 in colon disease and disease progression.