Visualzing Lymphocyte Attack on Pancreas B Cells

1型糖尿病（T1 D）是由自身反应性糖尿病（auto-reactive diabetes，AD）驱动的产生胰岛素的β细胞的破坏引起的。 T淋巴细胞。在小鼠和人类中，T1 D是一种受管制的疾病，具有重要的检查点： 当炎性细胞进入病变时引发自身免疫，导致胰岛炎;后来，在 这种胰岛炎的后果，它可以持续很长一段时间，或相反地导致p细胞破坏， 很短的订单。胰岛炎的细胞动力学和调控机制还不清楚 防止自身反应细胞造成晚期损伤我们建议分析均衡 自身免疫效应细胞（Teff）和调节细胞（Treg）之间的关系， 由我们的MGH同事（Projectl，CoreA）实施。这些研究将使用T1 D的NOD模型， 其转基因衍生物与其中细胞通过转基因表达荧光标记的小鼠进行比较， 蛋白质，在胰腺β细胞或T细胞亚群中。它们围绕着三个问题： 1.有组织的胰岛炎是自我维持的吗？何时以及通过何种途径是新的效应器， 募集调节性T细胞？胰岛炎组分的群体动力学的基本方面将首先是 通过流式细胞术和常规显微镜分析。然后我们将使用活体显微镜来追踪 自身反应性Teff或Treg细胞的胰岛途径。介观荧光断层扫描（MFT）将在 用于对胰腺中T细胞的全球分布进行成像，与Project 2合作进行组合 炎症和P细胞的分析。我们还将尝试动态定量炎症细胞的流入， 通过体内非侵入性荧光蛋白断层扫描（FPT）在疾病过程中检测细胞。 2.效应细胞和调节细胞之间的相互作用如何影响种群动态？ Treg细胞对自身免疫性糖尿病的影响已经得到了很好的证实，但关于Treg细胞对自身免疫性糖尿病的影响存在重大争议。 在何处以及如何干扰自身免疫发病机制。使用相同的成像工具，我们将 分析胰岛病变的群体动力学，其中TdR变化的条件下（特别是随着时间的推移， 控制的Treg细胞消融），并使用双光子活体显微镜观察 Teff、Teff和胰岛DC或β细胞。 3.什么时候获得促炎或调节转录细胞因子程序？我们将 在IFN γ、IL 17和IL 10的控制下编码荧光蛋白的回交报告转基因 基因座，并使用MFT来可视化和定量这些关键细胞因子基因座的活性的演变，在 在NOD小鼠中展开糖尿病发生的过程。 这些研究将使用最先进的成像工具来可视化胰岛素的自身免疫攻击- 产生自身免疫性糖尿病的细胞。它们将加深我们对糖尿病病因的理解， 为高危个体的诊断或预后成像以及治疗靶点开辟了几条途径。

Type-1 diabetes (T1D) results from the destruction of insulin-producing p-cells driven by autoreactive T lymphocytes. In both mice and humans, T1D is a regulated disease, with important checkpoints: at the initiation of autoimmunity when inflammatory cells enter the lesion, resulting in insulitis; later, in the consequences of this insulitis, which can persist for long periods, or conversely result in p-cell destruction in short order. The cellular dynamics of the insulitis are poorly understood, as are the regulatory mechanisms that prevent autoreactive cells from wreaking terminal damage. We propose to analyze the equilibrium between autoimmune effector cells (Teff) and regulatory cells (Treg) using the powerful imaging technologies implemented by our MGH colleagues (Projectl, CoreA). The studies will use the NOD model of T1D and transgenic derivatives thereof, with mice in which cells are labeled by transgenic expression of fluorescent proteins, in pancreatic p-cells or in T cell subpopulations. They revolve around three questions: 1. Is the organized insulitis self-sustaining, when and by which route are new effector and regulatory T cells recruited? Basic aspects of the population kinetics of insulitis components will first be analyzed by flow cytometry and conventional microscopy. We will then use intravital microscopy to track the path to the islets of autoreactive Teff or Treg cells. Mesoscopic Fluorescence Tomography (MFT) will be used to image the global distribution of T cells in the pancreas, collaborating with Project2 for the combined analysis of inflammatory and p-cells. We will also attempt to quantitate dynamically the influx of inflammatory cells over the course of the disease by non-invasive Fluorescent Protein Tomography (FPT) in vivo. 2. How are population dynamics affected by the interplay between effector and regulatory cells? The impact of Treg cells on autoimmune diabetes is well established, but there is significant debate as to where and how Tregs interfere with autoimmune pathogenesis. Using the same imaging tools, we will analyze the population dynamics of insulitic lesions in conditions where Tregs vary (in particular with time- controlled Treg cell ablation), and use 2-photon intravital microscopy to visualize the interactions between Teff, Tregs, and islets DCs or p-cells. 3. When are pro-inflammatory or regulatory transcriptional cytokine programs acquired? We will backcross reporter transgenes encoding fluorescent proteins under the control of the IFNy, IL17 and IL10 loci, and use MFT to visualize and quantitate the evolution of the activity of these key cytokine loci, over the course of unfolding diabetogenesis in NOD mice. These investigations will use cutting edge imaging tools to visualize the autoimmune attack of insulin- producing cells in autoimmune diabetes. They will deepen our understanding of the causes of diabetes and open several avenues for diagnostic or prognostic imaging in at-risk individuals, and for treatment targets.