β cell miRNAs Function as Molecular Hubs of Type 1 Diabetes Pathogenesis

β 细胞 miRNA 作为 1 型糖尿病发病机制的分子中心

• 批准号: 10321295
• 负责人: Carmella Evans-Molina
• 金额: $ 42.69万
• 依托单位: INDIANA UNIV-PURDUE UNIV AT INDIANAPOLIS
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-05-01 至 2025-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10321295
• 关键词: 3' Untranslated Regions; Address; Autoantibodies; Autoimmunity; Beta Cell; Binding; Biological Markers; Cell Death; Cell Survival; Cell physiology; Cells; Cellular Stress; Cessation of life; Childhood; Chronic; Clinical; Complex; Cues; Data; Data Set; Detection; Development; Diabetes Mellitus; Diabetes autoantibodies; Disease; Evolution; Exhibits; Exposure to; Female; Fluorescent in Situ Hybridization; Functional disorder; Gene Expression; Genetic Transcription; Genetic Translation; Goals; High Fat Diet; Homicide; Human; Immune; Immune system; Inflammatory; Insulin; Insulin-Dependent Diabetes Mellitus; Interferon Type II; Interleukin-1 beta; Invaded; Islets of Langerhans; Knowledge; Measures; Mediating; Mediator of activation protein; Messenger RNA; Metabolic Diseases; MicroRNAs; Modeling; Molecular; Mus; Non-Insulin-Dependent Diabetes Mellitus; Organ Donor; Pancreas; Pancreatic Injury; Pathogenesis; Pathway interactions; Pattern; Phenotype; Plasma; Play; Porifera; Process; Quantitative Reverse Transcriptase PCR; Research; Role; Signal Pathway; Small RNA; Streptozocin; Stress; Testing; Therapeutic Intervention; Tissues; Tumor-infiltrating immune cells; Untranslated RNA; Up-Regulation; Validation; Work; base; chemokine; clinical biomarkers; cytokine; diabetes pathogenesis; diabetes risk; experimental study; extracellular vesicles; imaging approach; injured; insulin dependent diabetes mellitus onset; insulin secretion; intercellular communication; islet; mRNA Transcript Degradation; male; mouse model; non-diabetic; novel; overexpression; programs; response; sexual dimorphism; single molecule; transcriptome sequencing

Type 1 diabetes (T1D) is a chronic metabolic disorder that is characterized by immune-mediated β cell destruction, resulting in the lifelong need for exogenous insulin therapy. Historically, T1D has been considered a disease of β cell homicide. However, recent evidence suggests that the β cell actively contributes to its own demise in T1D through engagement of cell intrinsic stress pathways that both hasten cell death and exacerbate autoimmunity. microRNAs (miRNAs, 18-25 nt) are a class of small non-coding RNAs that post transcriptionally modulate gene expression by binding the 3’untranslated region of a target mRNA to either inhibit mRNA translation or cause mRNA degradation. In addition to these regulatory roles within their cells of origin, miRNAs can be packaged and released within extracellular vesicles (EVs), which can be transferred to recipient cells to both facilitate intercellular communication and promote disease pathogenesis. While miRNAs have been shown to regulate several key processes within the β cell and have been implicated as potential mediators of a dialogue between the immune system and the β cell in diabetes, a full understanding of the role of miRNAs in T1D pathophysiology remains elusive. To this end, we profiled changes in miRNA expression patterns in human islets and islet-derived extracellular vesicles (EV’s) in response to IL-1β and IFN-γ, two cytokines selected to model the inflammatory intra-islet milieu observed in T1D. Our initial small RNA sequencing and additional preliminary data has shown that islet miRNA expression patterns are responsive to inflammatory extrinsic cues and that miRNAs appear to be selectively packaged into islet-derived EV’s in response to cytokine treatment. Moreover, islet and islet-derived EV miRNAs exhibited striking sexually dimorphic expression patterns under basal conditions and following cytokine treatment. Interestingly, miR-155-5p and miR-146-5p were the only two miRNAs that were coordinately upregulated in cytokine-treated islets and islet-derived EVs from both male and female donors, and upregulation of these miRNAs was associated with detrimental changes in β cell function and survival. Based on these findings, we hypothesize that β cell miR-155-5p and miR-146-5p function as key molecular hubs during the evolution of T1D, playing a role in disease pathogenesis, while also having potential utility as diabetes biomarkers. To test this hypothesis, we aim to: 1) define how miRNA signatures change in human β cells during T1D progression using state-of-the art smFISH imaging approaches, focused on miR-155 and 146 and a panel of additional miRNAs predicted to have either common or distinct expression patterns in males and females; 2) elucidate the mechanisms underlying miR-155-5p and miR-146-5p upregulation under inflammatory conditions; 3) define how miR-146 and miR-155 regulate β cell function and survival, leveraging both ex vivo human islet models and novel mouse models of β cell specific deletion of miR-155; and 4) test the utility of EV miRNA signatures informed from islet RNA sequencing as clinical biomarkers using plasma EVs and β cell enriched plasma EVs collected from pediatric subjects with recent onset T1D or autoantibody positivity.

1型糖尿病（T1 D）是一种以免疫介导的β细胞凋亡为特征的慢性代谢紊乱性疾病。 破坏，导致终身需要外源性胰岛素治疗。历史上，T1 D被认为是β细胞杀人疾病。然而，最近的证据表明，β细胞通过参与细胞内在应激途径（既加速细胞死亡又加剧自身免疫），积极促进T1 D中自身的死亡。微小RNA（miRNA，18-25 nt）是一类小型非编码RNA，通过结合靶mRNA的3 '非翻译区来抑制mRNA翻译或引起mRNA降解，从而转录后调节基因表达。除了在其起源细胞内的这些调节作用外，miRNA还可以在细胞外囊泡（EV）内包装和释放，其可以转移到受体细胞以促进细胞间通讯并促进疾病发病机制。虽然miRNAs已被证明可以调节β细胞内的几个关键过程，并被认为是对话的潜在介质， 尽管miRNAs在糖尿病中与免疫系统和β细胞之间的作用密切相关，但对miRNAs在T1 D病理生理学中的作用的充分理解仍然是难以捉摸的。为此，我们分析了人类胰岛和胰岛衍生的细胞外囊泡（EV）中响应IL-1β和IFN-γ的miRNA表达模式的变化，IL-1β和IFN-γ是选择用于模拟T1 D中观察到的炎症性胰岛内环境的两种细胞因子。我们最初的小RNA测序和额外的初步数据表明，胰岛miRNA表达模式对炎症外部线索有反应，并且miRNA似乎选择性地包装到胰岛来源的EV中以响应细胞因子治疗。此外，胰岛和胰岛衍生的EV miRNA在基础条件下和细胞因子处理后表现出显著的性二态表达模式。有趣的是，miR-155- 5 p和miR-146- 5 p是仅有的两种miRNA，它们在经精氨酸处理的胰岛和来自男性和女性的胰岛衍生EV中协同上调。 女性供体，这些miRNA的上调与β细胞功能和存活的有害变化相关。基于这些发现，我们假设β细胞miR-155- 5 p和miR-146- 5 p在T1 D的演变过程中作为关键的分子枢纽发挥作用，在疾病发病机制中发挥作用，同时也具有作为糖尿病生物标志物的潜在效用。为了验证这一假设，我们的目标是：1）使用最先进的smFISH成像方法来确定T1 D进展期间人类β细胞中miRNA标签如何变化，重点是miR-155和146以及一组预测在男性和女性中具有共同或不同表达模式的额外miRNA; 2）阐明炎症条件下miR-155- 5 p和miR-146- 5 p上调的潜在机制; 3）确定miR-146和miR-155如何调节β细胞功能和存活，利用离体人胰岛模型和miR-155的β细胞特异性缺失的新型小鼠模型;以及4）使用血浆EV和 从近期发作T1 D或自身抗体阳性的儿科受试者中采集的β细胞富集血浆EV。