GAD RELEASE FROM BETA CELLS IN TYPE 1 DIABETES

GAD65的循环自身抗体是导致I型糖尿病的自身免疫过程中最重要的标志之一。GAD65是谷氨酸脱羧酶的MR 65000亚型。越来越多的证据表明，GAD65和小鼠的GAD67在自身免疫过程中是启动自身抗原的关键。以前，人们认为GAD65是人类胰岛中唯一表达的GAD亚型。然而，我们已经证明，作为另一种剪接事件的结果，人类胰岛也产生GAD67变体GAD25。初步数据显示，某些新诊断为糖尿病的幼儿会产生GAD25自身抗体。其他数据表明，β细胞释放GAD65，可能与GAD25一起，可能有助于触发胰岛自身免疫。我们的中心假设是，GAD65在1型糖尿病中作为主要的自身抗原发挥作用，因为它的囊泡关联使其容易受到应激诱导的释放，而且GAD25可能作为自身抗原发挥作用，因为它也可以被释放。这一建议的具体目的是：1)检验GAD65和GAD25是由生理应激源唯一地从人胰岛β细胞释放出来的假说；2)检验膜相关GAD是释放GAD分子的细胞内来源的假说，并开始表征释放机制；3)检验GAD25通过其定位和调节是1型糖尿病潜在自身抗原的假说。这项研究的长期目标是更好地了解GAD65早期免疫反应的机制以及GAD25所起的作用。了解促进胰岛自身免疫的因素对于开发预防或抑制β细胞破坏的治疗方法至关重要，并可能导致新的治疗方法，以帮助保护移植的胰岛免受免疫介导的破坏。这项研究将为申请者作为I型糖尿病领域的独立研究员的学术生涯做好准备。他将获得分子生物学方面的进一步培训，学习进行基于群体的研究，并熟悉用于研究胰岛细胞生物学和生理学以及非生理性自身反应的关键方法和途径。向独立性的过渡将得到特别丰富的培训环境的推动，首先是由Ake Lernmark领导的R.H.威廉姆斯实验室，研究将在那里进行，其次是周围大量高效的糖尿病研究人员，所有这些都在附近，威廉姆斯实验室是其中的一员。

Circulating autoantibodies to GAD65, the Mr 65,000 isoform of glutamic acid decarboxylase, are one of the most important markers of the autoimmune process that results in type I diabetes mellitus. A growing body of evidence implicates GAD65, and, in mice, GAD67, as key initiating autoantigens in the autoimmune process. Previously it was thought that GAD65 was the only GAD isoform expressed in human islets. We have shown, however, that human islets also produce a GAD67 variant, GAD25, as the result of an alternative splicing event. Preliminary data suggest that certain young children with newly diagnosed diabetes produce autoantibodies to GAD25. Other data suggest that release of GAD65 from beta cells, perhaps along with GAD25, may help trigger islet autoimmunity. Our central hypothesis is that GAD65 functions as a major autoantigen in type 1 diabetes because its vesicular association makes it susceptible to stress-induced release and also that GAD25 may function as an autoantigen because it, too, can be released. The specific aims of this proposal are 1) to test the hypothesis that GAD65 and GAD25 are uniquely released from human islet beta cells by physiologic stressors; 2) to test the hypothesis that membrane-associated GAD is the intracellular source of released GAD molecules and begin to characterize the mechanism of release and 3) to test the hypothesis that GAD25, by virtue of its localization and regulation, is a potential autoantigen in type 1 diabetes. The long-term objective of this research is to better understand the mechanisms underlying the early immune reaction to GAD65 and the role played by GAD25. Knowledge of the factors that precipitate islet autoimmunity will be vital for the development of therapies to prevent or inhibit beta cell destruction and could result in new treatments to help protect transplanted islets from immune-mediated destruction. This research will prepare the applicant for an academic career as an independent investigator in the field of type I diabetes. He will obtain further training in molecular biology, learn to conduct population-based studies and become acquainted with the key methodologies and approaches used to study islet cell biology and physiology and diabetogenic autoreactivity. The transition to independence will be facilitated by the especially rich training environment afforded, first, by the R.H. Williams Laboratory, headed by Ake Lernmark, where the research will be carried out and, second, by the large surrounding community of highly productive diabetes researchers, all in close proximity, of which the Williams Laboratory is a member.