Survival and potential of insulin-deficient beta cells in type 1 diabetes

1 型糖尿病中胰岛素缺乏的 β 细胞的存活率和潜力

• 批准号: 10705841
• 负责人: DESMOND Arthur SCHATZ
• 金额: $ 70万
• 依托单位: UNIVERSITY OF FLORIDA
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-09-20 至 2026-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10705841
• 关键词: Acceleration; Agonist; Autoantibodies; Autoimmunity; Automobile Driving; Beta Cell; Biological Assay; Biological Models; Biological Products; Blood Vessels; Cell physiology; Cells; Cellular Metabolic Process; Characteristics; Chromogranin A; Cytometry; DNA; DNA Methylation; Data; Detection; Diabetes Mellitus; Disease; Down-Regulation; Endocrine; Endoplasmic Reticulum; Extravasation; Functional disorder; GCG gene; Gene Expression; Genetic Transcription; Genetically Engineered Mouse; Hormones; Human; Hyperglycemia; Image; Immune; Immune system; Immunofluorescence Immunologic; Immunologics; In Situ; In Vitro; Inflammation; Inflammatory; Insulin; Insulin deficiency; Insulin-Dependent Diabetes Mellitus; Interferons; Islet Cell; Islets of Langerhans; Lesion; Lobular; Longevity; Lymphatic; Maps; Membrane Glycoproteins; Metabolic; Metabolic Pathway; Methods; Methylation; Modality; Modeling; Molecular; Mus; Natural History; Non-Insulin-Dependent Diabetes Mellitus; Organ Donor; Pancreas; Pathway interactions; Patients; Phenotype; Pilot Projects; Play; Process; Production; Proinsulin; Proteins; Quality of life; Residencies; Role; STAT5B gene; Sampling; Signal Pathway; Signal Transduction; Slice; Stains; System; Tamoxifen; Testing; Therapeutic; Tissues; WFS1 gene; autoimmune pathogenesis; cytokine; drug candidate; effective intervention; endoplasmic reticulum stress; experimental study; immunoreactivity; improved; in vivo; inhibitor; insight; insulin dependent diabetes mellitus onset; insulin secretion; islet; laser capture microdissection; methylation pattern; mimetics; mouse model; mutant; mutant mouse model; new technology; non-diabetic; novel; repository; response; senescence; targeted treatment; therapeutic candidate; therapeutic development; therapeutic evaluation; therapeutic target; trafficking; transcriptome sequencing; type I and type II diabetes

SUMMARY / ABSTRACT Recent evidence put forth by our group and others suggests that the pancreas of patients with recent-onset type 1 diabetes (T1D) contains a significant number of “empty” beta cells that have lost the ability to secrete mature insulin but retained other hallmark features such as proinsulin expression. The mechanisms that underlie formation of such cells and their fate remain poorly understood. We hypothesize that empty beta cells, which are invisible to standard insulin immunostaining and potentially also to the immune system, play important roles in the pathophysiology of T1D while holding therapeutic potential for reversion of diabetes. We propose in depth experiments to characterize empty beta cells, using MACSima ultra high-content immunofluorescence imaging to assess in situ expression of 78 immune and pancreas cell markers, including vascular and lymphatic annotation with signatures of inflammation, extravasation/trafficking, and immune cell residency in autoantibody positive (AAb+) and T1D donors as compared to donors with type 2 diabetes (T2D) and non-diabetic controls available through the Network for Pancreatic Organ donors with Diabetes (nPOD) repository. We will compare insulin containing versus insulin negative islets, within and across donors, to identify empty beta cells and determine how they correlate with islet, acinar, and immune cell phenotypes. These data will serve as a template for a serial section to undergo laser capture microdissection (LCM) of insulin containing islets, insulin negative islets, and acinar tissue regions, which will be subjected to bulk RNAseq as well as our novel method for quantifying beta cells based on DNA methylation patterns. In addition, we will use a novel mouse model (beta cell-specific, tamoxifen-inducible Adar1-mutant) and cultured human pancreas slices to functionally interrogate molecular pathways underlying the formation of empty beta cells. Specifically, we propose to leverage these two model systems to test therapeutic candidates— including an incretin mimetic (GLP1), an endoplasmic reticulum (ER) stress inhibitor (ISRIB), and multiple biologics targeting specific immune subsets— for their ability to modulate empty beta cells, their insulin content and insulin secretion. We will then correlate these functional data with molecular and cellular features of hormone negative islet cells via MACSima, RNAseq, and DNA methylation. These studies are expected to yield insights into a fundamental yet little understood process taking place in human T1D. We anticipate that empty beta cells can be re-functionalized, paving the way for therapeutic development to restore endogenous beta cell function in T1D. Ultimately, when combined with effective interventions to constrain autoimmunity, it is our hope that the metabolic modalities explored here could provide a means to reduce insulin requirements or even achieve insulin independence after T1D onset, dramatically improving longevity and quality of life for these patients.

摘要/摘要 我们小组和其他人提出的最新证据表明，新发型患者的胰腺 1 型糖尿病 (T1D) 含有大量“空”β 细胞，这些细胞已丧失分泌成熟细胞的能力 胰岛素，但保留了其他标志性特征，例如胰岛素原表达。其背后的机制 这些细胞的形成及其命运仍然知之甚少。我们假设空的β细胞， 标准胰岛素免疫染色不可见，免疫系统也可能不可见，发挥重要作用 参与 T1D 的病理生理学研究，同时具有逆转糖尿病的治疗潜力。我们深入建议 使用 MACSima 超高内涵免疫荧光成像表征空 β 细胞的实验 评估 78 种免疫和胰腺细胞标记物的原位表达，包括血管和淋巴细胞标记物 自身抗体中炎症、外渗/运输和免疫细胞驻留特征的注释 与 2 型糖尿病 (T2D) 捐献者和非糖尿病对照捐献者相比，阳性 (AAb+) 和 T1D 捐献者 可通过糖尿病胰腺器官捐献者网络 (nPOD) 存储库获取。我们会比较 供体内部和之间的含胰岛素胰岛与胰岛素阴性胰岛的比较，以识别空的β细胞和 确定它们如何与胰岛、腺泡和免疫细胞表型相关。这些数据将作为模板 对于含有胰岛素的胰岛进行激光捕获显微切割 (LCM) 的连续切片，胰岛素阴性 胰岛和腺泡组织区域，将进行批量 RNAseq 以及我们的新方法 根据 DNA 甲基化模式量化 β 细胞。此外，我们将使用一种新颖的小鼠模型（beta 细胞特异性、他莫昔芬诱导型 Adar1 突变体）和培养的人胰腺切片以进行功能性询问 空β细胞形成的分子途径。具体来说，我们建议利用这两个 用于测试候选治疗药物的模型系统——包括肠促胰岛素模拟物 (GLP1)、内质网 (ER) 应激抑制剂 (ISRIB) 和多种针对特定免疫子集的生物制剂——因为它们能够 调节空β细胞、其胰岛素含量和胰岛素分泌。然后我们将关联这些功能数据 通过 MACSima、RNAseq 和 DNA 具有激素阴性胰岛细胞的分子和细胞特征 甲基化。这些研究预计将深入了解一个基本但鲜为人知的过程 在人类 T1D 中的位置。我们预计空的β细胞可以重新功能化，为治疗铺平道路 恢复 T1D 内源性 β 细胞功能的开发。最终，当与有效的 限制自身免疫的干预措施，我们希望这里探索的代谢方式可以提供 一种在 T1D 发病后显着减少胰岛素需求甚至实现胰岛素独立的方法 提高这些患者的寿命和生活质量。