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: 10615586
• 负责人: Carmella Evans-Molina
• 金额: $ 42.82万
• 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/10615586
• 关键词: 3' Untranslated Regions; Address; Autoantibodies; Autoimmunity; Beta Cell; Binding; Biological Markers; Cell Death; Cell Survival; Cell physiology; Cells; Cellular Stress; Childhood; Chronic; Clinical; Complex; Cues; Data; Data Set; Detection; Development; Diabetes Mellitus; Disease; Evolution; Exhibits; Exposure to; Female; Fluorescent in Situ Hybridization; Functional disorder; Gene Expression; Genetic Transcription; 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; 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; Untranslated RNA; Up-Regulation; Validation; Work; chemokine; clinical biomarkers; cytokine; diabetes pathogenesis; diabetes risk; experimental study; extracellular vesicles; imaging approach; immune cell infiltrate; injured; insulin dependent diabetes mellitus onset; insulin secretion; intercellular communication; islet; mRNA Transcript Degradation; mRNA Translation; male; mouse model; non-diabetic; novel; overexpression; posttranscriptional; 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(T1D)是一种以免疫介导的β细胞为特征的慢性代谢紊乱 破坏，导致终生需要外源性胰岛素治疗。从历史上看，T1D一直被认为是一种β细胞杀伤性疾病。然而，最近的证据表明，β细胞通过参与加速细胞死亡和加剧自身免疫的细胞内在应激途径，积极地促进了自身在T1D的死亡。MicroRNAs(miRNAs，18-25nt)是一类小的非编码RNA，通过与靶mRNA的3‘非翻译区结合，在转录后调节基因表达，从而抑制mRNA翻译或导致mRNA降解。除了在来源细胞内的这些调节作用外，miRNAs还可以在细胞外小泡(EV)内包装和释放，这些EV可以转移到受体细胞，以促进细胞间的沟通和促进疾病的发病。虽然miRNAs已被证明调节β细胞内的几个关键过程，并被认为是潜在的对话调解人 在糖尿病的免疫系统和β细胞之间，对miRNAs在T1D病理生理学中的作用仍然难以完全理解。为此，我们分析了人胰岛和胰岛来源的细胞外小泡(EV‘s)对IL-1β和干扰素-γ的反应中miRNA表达模式的变化，这两种细胞因子被选作T1D中观察到的胰岛内炎症环境的模型。我们最初的小RNA测序和额外的初步数据表明，胰岛miRNA的表达模式对炎症外部线索有反应，并且miRNAs似乎选择性地包装到胰岛来源的EV中，以响应细胞因子的治疗。此外，胰岛和胰岛来源的EV miRNAs在基础条件下和细胞因子处理后表现出显著的性别二型性表达模式。有趣的是，miR-155-5p和miR-146-5p是仅有的两个在细胞因子处理的胰岛和胰岛来源的EV中协同上调的miRNA。 这些miRNA的上调与β细胞功能和存活率的有害变化有关。基于这些发现，我们推测β细胞miR-155-5p和miR-146-5p在T1D的进化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，同时也可能作为糖尿病的生物标志物。为了验证这一假设，我们的目标是：1)使用最先进的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，利用最先进的smFISH成像方法，确定miR-155和miR-146-5p在人β细胞T1D进展过程中的变化；2)阐明炎症条件下miR-155-5p和miR-146-5p上调的机制；3)确定miR-146和miR-155如何调节β细胞的功能和存活，利用体外人类胰岛模型和新颖的β细胞特异性缺失miR-155的小鼠模型；和4)使用血浆EVS和EVS来测试从胰岛RNA测序获得的EV miRNA信号作为临床生物标记物的有效性 从新近发病的T1D或自身抗体阳性的儿科受试者中收集的β细胞浓缩血浆EVS。