mt-Nd2 and Resistance to Autoimmune Diabetes

描述（由申请人提供）：线粒体（mt）在细胞能量产生和细胞死亡中起关键作用。β细胞功能与线粒体紧密相关;因为胰岛素合成和葡萄糖刺激的胰岛素分泌都需要线粒体ATP产生。在这种情况下，与人类糖尿病（T2 D）家系相关的线粒体DNA（mtDNA）突变报告占人类T2 D的1%。然而，线粒体DNA突变通常与自身免疫性1型糖尿病（T1 D）无关，尽管导致mt-ND 2基因中亮氨酸取代为蛋氨酸的C至A颠换与高危人群以及T1 D易感NOD与T1 D耐药ALR小鼠的交叉中的T1 D保护有关。本申请的目的是了解这种单一氨基酸变化如何改变对T1 D的抗性。 T1 D易感性的遗传分析将注意力集中在控制异常免疫细胞功能的候选基因上，而很少关注可能在细胞水平上导致易感性或抗性的基因。来自ALR小鼠品系的胰岛对自身免疫效应物的功能损伤和杀伤保持不寻常的遗传抗性。我们的结果将一些ALR衍生的T1 D抗性与mt-Nd 2a联系起来。虽然该等位基因在渗入NOD遗传背景中时不会阻止自发性T1 D的发展，但ALR衍生的遗传变体实际上确实保护β细胞免受细胞毒性T淋巴细胞以及组合细胞因子的破坏。我们已经确定这种抗性源于促凋亡应激不能诱导mt活性氧。我们的目标是了解这个基因在细胞抵抗自身免疫性杀伤中的作用。在具体目标1中，我们将确定 mt-Nd 2a编码细胞和原代胰岛抵抗肿瘤坏死因子受体家族成员TNFR 1和Fas介导的破坏的机制。目的2将进一步研究mt-Nd 2a编码的胰岛细胞避免CTL裂解的机制。具体目标3将使用人类胰岛来确认对细胞破坏的抵抗机制，以及cybrid人类细胞来测试mt-ND 2的人类等位基因是否改变人类细胞死亡的特征。这些研究将提供有关导致人类β细胞死亡的早期凋亡信号以及ND 2a如何保护人类β细胞的有意义的数据。 公共卫生相关性：对自身免疫性T1 D的易感性与抗性的遗传学主要关注控制免疫系统功能的候选基因，而不是靶β细胞。假设产生胰岛素的β细胞表达靶抗原的共同库，但在T1 D发病机制中除尸体外不起其他作用。与这种普遍持有的观点相反，我们的工作清楚地表明，遗传学在靶组织抵抗导致1型糖尿病的自身免疫性破坏方面发挥着重要作用。目前的提议利用了一种遗传差异，这种差异为人类和小鼠提供了对自身免疫介导的破坏的抵抗力。该提案的目的是确定该基因变体用于防止β细胞死亡的机制，并确定暴露于自身免疫效应物的β细胞中发生的早期促死亡信号。这项工作的长期目标是确定mt-ND 2a等位基因如何在人群中提供针对T1 D的保护。人类胰岛可以用来抵御自身免疫介导的破坏机制的保护因子对于移植、干细胞工程和未来的遗传以及药物糖尿病预防疗法具有重要的影响。

DESCRIPTION (provided by applicant): Mitochondria (mt) play key roles in cellular energy production and cell death. Beta cell function is tightly linked to mitochondria; as both insulin synthesis and glucose stimulated insulin secretion require mitochondrial ATP production. In this context, reports of mitochondrial DNA (mtDNA) mutations associated with diabetes (T2D) pedigrees in humans account for up to 1% of human T2D. However, mutations in mtDNA are not commonly associated with autoimmune Type 1 diabetes (T1D), although a C to A transversion resulting in a leucine to methonine substitution in the mt-ND2 gene has been associated with protection from T1D in both an at risk human population and in crosses of the T1D-prone NOD with T1D-resistant ALR mice. The goal of this application is to understand how this single amino acid change modifies resistance to T1D. Genetic analysis of T1D susceptibility has focused attention on candidate genes controlling aberrant immune cell function with little focus on genes that may contribute susceptibility or resistance at the ¿ cell level. Pancreatic islets from the ALR mouse strain maintain an unusual genetic resistance to functional impairment and killing by autoimmune effectors. Our results have linked some of the ALR-derived T1D resistance to mt- Nd2a. While this allele does not prevent the development of spontaneous T1D when introgressed into the NOD genetic background, the ALR-derived genetic variant does in fact protect beta cells from destruction by cytotoxic T lymphocytes as well as combined cytokines. We have determined that this resistance stems from the inability of pro-apoptotic stress to induce mt reactive oxygen species. Our goal is to understand the role this gene plays in ¿ cell resistance to autoimmune killing. In specific aim 1 we will determine the mechanism by which mt-Nd2a encoding ¿ cells and primary islets resist destruction mediated by members of the Tumor Necrosis Factor Receptor family, TNFR1 and Fas. Aim 2 will extend the studies to determine the mechanisms utilized by mt-Nd2a encoding islet cells to avert lysis by CTL. Specific aim 3 will use human islets to confirm mechanisms of resistance to ¿ cell destruction as well as cybrid human ¿ cells to test if the human alleles of mt-ND2 alter the characteristics of human ¿ cell death. The studies, as proposed, will provide meaningful data on both the early apoptotic signals that result in human beta cell death as well as insights into how ND2a protects human beta cells. PUBLIC HEALTH RELEVANCE: The genetics of susceptibility versus resistance to autoimmune T1D has primarily focused attention on candidate genes controlling function of the immune system rather than on the target beta cell. The assumption has been that insulin producing beta cells express a common repertoire of target antigens, yet play no other role in T1D pathogenesis than the corpse. In contrast to this commonly held view, our work has clearly shown that genetics play an important role in resistance of the target tissue to the autoimmune destruction leading to Type 1 Diabetes. The current proposal exploits a genetic difference that provides resistance to autoimmune- mediated destruction in both human and mouse. The objectives of this proposal are to determine the mechanism this gene variant employs to prevent beta cell death and also to identify the early pro-death signals that occur in beta cells exposed to autoimmune effectors. The long-term goal of this work is to identify how the mt-ND2a allele provides protection against T1D in human populations. Protective factors human pancreatic islets can employ to ward off autoimmune-mediated destruction mechanisms have important ramifications for transplantation, stem cell engineering, and future genetic as well as pharmacological diabetes preventative therapies.