Role of dysregulated miRNA in Tregs in Multiple Sclerosis

Tregs 中失调的 miRNA 在多发性硬化症中的作用

• 批准号: 8283087
• 负责人: Amy E Lovett-Racke
• 金额: $ 22.88万
• 依托单位: OHIO STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-05-01 至 2014-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8283087
• 关键词: 3' Untranslated Regions; Affect; Algorithms; Autoimmune Diseases; Biological Assay; CD4 Positive T Lymphocytes; Cell Differentiation process; Cells; Characteristics; Defect; Development; Disease; Disease Progression; Disease susceptibility; Environment; Equilibrium; Figs - dietary; Functional disorder; Gene Expression; Gene Targeting; Genes; Health; Human; Immune; Immune system; Inflammatory; Laboratories; Luciferases; Maintenance; Mediating; MicroRNAs; Multiple Sclerosis; Nerve Degeneration; Neurologic; Pathology; Pathway interactions; Patients; Phenotype; Play; Predisposition; Production; Proteins; RNA Interference Pathway; Regulatory T-Lymphocyte; Relative (related person); Role; Signal Pathway; Signal Transduction; Sorting - Cell Movement; T-Lymphocyte; Th1 Cells; Th2 Cells; Work; central nervous system demyelinating disorder; cytokine; disability; in vivo; inhibitor/antagonist; innovation; memory CD4 T lymphocyte; novel; peripheral blood; receptor; young adult

DESCRIPTION (provided by applicant): Multiple sclerosis (MS) is an immune-mediated demyelinating disease of the central nervous system (CNS) which can result in severe neurological deficits. The cause of the disease is unknown, the immune and neurodegenerative mechanisms underlying the pathophysiology of the disease are poorly understood, and current therapies are only partially effective at slowing disease progression. My laboratory has performed a large miRNA profiling study on naive and effector/memory CD4 T cell in untreated MS patients to determine if miRNA may contribute to disease susceptibility or progression. The TGF¿ signaling pathway was predicted to be suppressed in MS patients by 12 miRNAs identified as being dysregulated in CD4 T cells. TGF¿ has shown to play a vital role in CD4+CD25hiFoxp3+ regulatory T cells (Tregs), and Tregs have been shown to be defective in MS patients. TGF¿ signaling has been shown to be critical for production of iTregs, and the maintenance of suppressor functions and Foxp3 expression in Tregs. We hypothesize that dysregulated miRNA expression in CD4 T cells in MS patients' results in impaired TGF¿ signaling and defective CD4 regulatory T cells, promoting the onset and progression of MS. Aim 1: To determine if miRNAs identified as dysregulated in CD4 T cells in MS patients directly target specific molecules in the TGF ¿ signaling pathway. First, we will determine if there is a correlation in gene expression levels between specific miRNAs and predicted TGF¿ targets in human CD4 T cells from MS patients and healthy controls. Second, we will use luciferase assays to determine if specific miRNAs suppress the 3' UTR sequence of the predicted target genes in the TGF¿ pathway. Aim 2: To determine if the miRNAs that target the TGF¿ pathway alter iTreg development and/or suppressive functions of Tregs. TGF¿ targeting miRNAs will be inhibited or over-expressed in naive CD4 T cells and then cultured under iTreg conditions. The resulting T cells will be phenotyped for iTreg characteristics. In addition, Tregs will be transfected with specific miRNA inhibitors, evaluated for sensitivity to TGF¿, and ability to suppress effector T cells. Aim 3: To determine if there is an association between the level of TGF¿-associated miRNA expression and the number of Tregs in vivo, or an association between TGF¿-associated miRNA expression and suppressive capacity of Tregs in MS patients. Flow cytometric analysis of peripheral blood CD4 T cells from MS patients will evaluate the number of Tregs in MS patients relative to the expression of TGF¿-associated miRNAs. In addition, Tregs will be sorted from peripheral blood of MS patients, evaluated for suppressive capacity, and determine if there is a correlation between TGF¿-associated miRNAs and Treg number and/or function. Given the well-documented role of Tregs in maintaining tolerance to self and the observation that CD4 Tregs are defective in MS patients, this novel study will determine if dysregulated miRNA expression in CD4 T cells underlies this Treg defect in MS patients. Although several studies have identified miRNAs that play a role in Tregs17-22, no studies have identified differentially expressed miRNAs in MS patients that are specifically associated with genes in the TG¿ pathway and their influence on Tregs in an autoimmune disease. Treg cells play an important role in the delicate balance of protection from our environment and protecting ourselves from our own immune system, and TGF¿ has pleiotrophic effects that extend beyond Tregs that have vast importance in health and disease, making this an invaluable study. PUBLIC HEALTH RELEVANCE: Multiple sclerosis (MS) is the leading cause of non-traumatic disability of young adults, affecting over 400,000 people in the USA. The cause of the disease is unknown, the mechanisms that underlie the pathology are poorly understood and treatments are only partially effective, necessitating innovative strategies to understand the disease and susceptibility factors. In the proposed study we will study the role of miRNAs in the development and function of regulatory T cells which have been shown to be defective in MS patients, which may identify a fundamental mechanism underlying disease susceptibility.

描述（由申请人提供）：多发性硬化症（MS）是一种免疫介导的中枢神经系统（CNS）脱髓鞘疾病，可导致严重的神经功能缺损。该疾病的病因尚不清楚，其背后的免疫和神经退行性机制。人们对这种疾病的病理生理学知之甚少，目前的治疗方法只能部分有效地减缓疾病进展。未经治疗的多发性硬化症患者，以确定 miRNA 是否可能导致疾病易感性或进展。据预测，多发性硬化症患者的信号通路会被 CD4 T 细胞中被鉴定为失调的 12 个 miRNA 所抑制。已显示在 CD4+CD25hiFoxp3+ 调节性 T 细胞 (Treg) 中发挥重要作用，而 Treg 已被证明在多发性硬化症患者中存在缺陷。信号转导已被证明对于 iTreg 的产生以及 Tregs 中抑制功能和 Foxp3 表达的维持至关重要，我们发现 MS 患者的 CD4 T 细胞中 miRNA 表达失调会导致 TGF 受损。目标 1：确定 MS 患者 CD4 T 细胞中被鉴定为失调的 miRNA 是否直接靶向 TGF 中的特定分子。首先，我们将确定特定 miRNA 与预测的 TGF 之间的基因表达水平是否存在相关性。其次，我们将使用荧光素酶测定来确定特定 miRNA 是否抑制 TGF¿ 中预测靶基因的 3' UTR 序列。目标 2：确定 miRNA 是否靶向 TGF¿途径改变 iTreg 的发育和/或 Treg 的抑制功能。靶向 miRNA 将在初始 CD4 T 细胞中被抑制或过度表达，然后在 iTreg 条件下培养，并对所得 T 细胞进行 iTreg 特征表型分析。和抑制效应 T 细胞的能力 目标 3：确定 TGF¿ 水平之间是否存在关联。 -相关的miRNA表达和体内Treg的数量，或TGF之间的关联¿ MS 患者中相关的 miRNA 表达和 Tregs 的抑制能力对 MS 患者外周血 CD4 T 细胞进行流式细胞术分析，将评估 MS 患者中 Tregs 的数量相对于 TGF 的表达。此外，将从 MS 患者的外周血中分选 Tregs，评估其抑制能力，并确定 TGF¿相关 miRNA 和 Treg 数量和/或功能 鉴于 Tregs 在维持自身耐受性方面的作用以及对 MS 患者中 CD4 Tregs 缺陷的观察，这项新研究将确定 CD4 T 细胞中 miRNA 表达失调是否是其背后的原因。尽管一些研究已经鉴定出在 Tregs17-22 中发挥作用的 miRNA，但没有研究鉴定出 MS 患者中与 TG 中的基因特异性相关的差异表达 miRNA。在自身免疫性疾病中，Treg 通路及其对 Tregs 的影响在保护我们免受环境影响和保护我们自身免受自身免疫系统和 TGF 侵害的微妙平衡中发挥着重要作用。其多效作用超出了对健康和疾病具有巨大重要性的 Tregs 范围，这使得这项研究变得非常有价值。 公共卫生相关性：多发性硬化症 (MS) 是导致年轻人非创伤性残疾的主要原因，在美国影响着超过 400,000 人。该疾病的病因尚不清楚，其病理机制知之甚少，治疗方法也不清楚。仅部分有效，需要创新策略来了解疾病和易感因素。在拟议的研究中，我们将研究 miRNA 在调节性 T 细胞的发育和功能中的作用，这些作用已被证明是有效的。 MS 患者存在缺陷，这可能确定了疾病易感性的基本机制。