Role of Selenium in Development and Health

硒是人类和其他哺乳动物饮食中必不可少的微量营养素。许多健康益处都要归功于硒，它们包括预防各种形式的癌症(如结肠癌、前列腺癌、肺癌和肝癌)、心脏病和其他心血管和肌肉疾病、抑制病毒表达、延缓人类免疫缺陷病毒(HIV)阳性患者的获得性免疫缺陷综合征(AIDS)的进展、减缓衰老过程以及在哺乳动物发育，包括男性生殖和免疫功能中发挥作用。近年来，人们进行了大量的人类临床试验，以评估这种元素在癌症预防、延缓艾滋病进展等方面的作用，耗资数十亿美元，但人们对硒如何在哺乳动物的代谢水平上发挥这些健康益处的机制知之甚少。我们提出，硒的健康益处在很大程度上是由于它以含硒氨基酸--硒半胱氨酸(SEC)的形式存在于硒蛋白中。因此，我们的计划主要集中在：1)建立小鼠模型，以评估硒和硒蛋白在癌症预防和发展中的作用；2)表征和阐明各种硒蛋白的功能及其在癌症预防和发展中的作用；3)确定SEC的生物合成和整合到蛋白质中的方法。这里讨论的项目检查了我们关于开发各种小鼠模型的研究，以确定硒在癌症、癌症预防和发展中的作用。在过去的几年里，我们专注于完成对小鼠模型的研究：1)敲除SEC tRNA基因(指定为TrSP)，从而导致(A)T细胞、(B)巨噬细胞、(C)皮肤组织和(D)肝脏中硒蛋白表达的丧失，以及随后野生型或突变型TrSP转基因对硒蛋白表达的挽救或部分挽救。在过去的一年里，我们在这个项目中的重点几乎完全转移到了建立小鼠模型上，这些模型可以处理硒蛋白在癌症进展中的作用，并有针对性地移除单个硒蛋白，而不是通过移除TrSP来敲除所有的硒蛋白。在上一财年，我们完成了对硒蛋白在细胞免疫中的分子机制的研究，并在过去的一年里，我们强调提供足够的小鼠来完成一项与Melinda Beck博士的合作研究，以确定缺乏、足够和补充水平的硒摄入对T细胞介导的免疫反应的影响，这些小鼠不在T细胞中表达硒蛋白，或者只表达关键的管家硒蛋白(但不表达非必需的应激相关硒蛋白)。我们继续了一项针对不同组织中仅清除硫氧还蛋白还原酶1(TR1)和/或谷胱甘肽过氧化物酶4(Gpx4)的研究，以考察这两种必需的硒蛋白在几个器官和组织中的作用。在表皮组织(这里指皮肤)中Gpx4的丢失几乎模拟了所有硒蛋白的丢失，只是小鼠在第10天后恢复，当时它们通常在Trsp基因敲除小鼠中死亡，并似乎过着几乎正常的生活。Gpx4基因敲除的小鼠有脱发，皮肤剥落和脆弱，组织学研究显示表皮增生，毛囊外观发生变化。这些观察证明了Gpx4在皮肤发育中先前未知的作用，提供了进一步的证据，证明这种硒蛋白在细胞发育中具有许多基本功能。靶向去除皮肤中的TR1似乎没有任何表型变化。在我们的体内结肠癌发生研究中，我们能够证明，与野生型或杂合子对照相比，Sep15基因敲除小鼠似乎能够防止由偶氮甲烷诱导的肿瘤前病变(异常隐窝病灶)的形成。此外，使用微阵列以及实时RT-PCR对结肠粘膜的分析表明，鸟氨酸结合蛋白1在9月15基因敲除小鼠中高度上调。这种干扰素-伽马调节蛋白和结肠癌中15 kDa硒蛋白之间的可能联系将在未来的实验中进行研究。此外，我们还将进一步阐明膳食硒对9月15基因敲除小鼠结肠癌模型的影响。此外，我们的另一个小鼠研究模型，包括使用肝脏TR1缺陷小鼠暴露于肝脏致癌物质二乙基亚硝胺(DEN)，在过去一年取得了成果。我们发现，肝脏特异的TR1基因敲除小鼠比对照小鼠更容易发生肿瘤(肝腺瘤、肝癌和肝胆管细胞腺瘤)。18只基因敲除小鼠中有16只发生了肿瘤，而19只对照小鼠中有2只发生了肿瘤。此外，我们发现在TR1缺陷的肝脏中NRF2调节的基因上调，包括几个谷胱甘肽转移酶和硒蛋白谷胱甘肽过氧化物酶2，它在肿瘤中进一步上调。

Selenium is an essential micronutrient in the diet of humans and other mammals. Many health benefits have been attributed to selenium that include preventing various forms of cancer (e.g., colon cancer, prostate cancer, lung cancer and liver cancer), heart disease and other cardiovascular and muscle disorders, inhibiting viral expression, delaying the progression of acquired immunodeficiency syndrome (AIDS) in human immunodeficiency virus (HIV)-positive patients, slowing the aging process, and having roles in mammalian development, including male reproduction and immune function. Numerous human clinical trails have been undertaken in recent years to assess the role of this element in cancer prevention, delaying the progression of AIDS, etc., at a cost of billions of dollars, but little is known about the mechanism of how selenium acts at the metabolic level in mammals to exert these many health benefits. We have proposed that the health benefits of selenium are due largely to its presence in selenoproteins as the selenium-containing amino acid, selenocysteine (Sec). Our program therefore focuses on: 1) developing mouse models to assess the role of selenium and selenoproteins in cancer prevention and development, 2) characterizing and elucidating the function of various selenoproteins and their roles in cancer prevention and development, and 3) identifying the means by which Sec is biosynthesized and incorporated into protein. The project discussed herein examines our research on the development of various mouse models for determining the role of selenium in cancer, cancer prevention and development. In the past several years, we have focused on completing our studies on characterizing mouse models: 1) that knockout the Sec tRNA gene (designated Trsp) and consequently result in the loss of selenoprotein expression in (a) T cells, (b) macrophage, (c) epidermal skin tissue, and (d) liver, and subsequent rescue or partial rescue of selenoprotein expression with a wild-type or mutant Trsp transgene. Our direction of focus in this project has shifted in the past year almost totally towards generating mouse models that deal with the role of selenoproteins in the progression of cancer and the targeted removal of individual selenoproteins instead of knocking out all selenoproteins through the removal of Trsp. In an earlier fiscal year, we completed our study on examining the molecular mechanism of selenoproteins in cell immunity and in the past year, we emphasized providing sufficient mice to complete a collaborative study with Dr. Melinda Beck in determining the effects of deficient, adequate, and supplemental levels of selenium intake on the immune response mediated by T cells in mice that do not express selenoproteins in T cells or express only the critical, housekeeping selenoproteins (but not the non-essential, stress-related selenoproteins). We have continued a study on targeting the removal of only thioredoxin reductase 1 (TR1) and/or glutathione peroxidase 4 (GPx4) in various tissues to examine the role of these two essential selenoproteins in several organs and tissues. Loss of GPx4 in epidermal tissue (designated skin herein) virtually mimicked the loss of all selenoproteins with the exception that the mice recover after day 10, when they routinely died in Trsp knockout mice, and appear to live a virtually normal life. The GPx4 knockout mice had alopecia along with a flaky and fragile skin and histological studies revealed epidermal hyperplasia along with changes in hair follicle appearance. These observations demonstrate a prior unknown role of GPx4 in cutaneous development providing further evidence that this selenoprotein has many essential functions in cell development. The targeted removal of TR1 in skin did not appear to have any alteration in phenotype. In our in vivo colon carcinogenesis study, we were able to demonstrate that Sep15 knockout mice, compared to wild type or heterozygote littermate controls, appeared to be protected against azoxymethane-induced formation of pre-neoplastic lesions (aberrant crypt foci). Furthermore, using microarrays as well as real-time rtPCR analysis of the colonic mucosa showed that guanylate-binding protein 1 was highly up-regulated in Sep15 knockout mice. The possible connection between this interferon-gamma-regulated protein and the 15kDa selenoprotein in colon cancer will be investigated in future experiments. In addition, the possible influence of dietary selenium in our colon cancer model using Sep15 knockout mice will be further elucidated. Furthermore, our other mouse study model, which involved using liver TR1 deficient mice exposed to the liver carcinogen, diethylnitrosamine, (DEN), came to fruition in the past year. We found that liver-specific TR1 knockout mice developed tumors (hepatic adenomas, hepatic carcinomas and hepatocholangiocellular adenomas) much more readily than control mice. 16 of 18 knockout mice developed tumors compared to 2 of 19 control mice. In addition, we found an upregulation of Nrf2-modulated genes in the TR1 deficient livers, including several of the glutathione transferases and the selenoprotein, glutathione peroxidase 2, which was even further upregulated in tumors.