Identical Twins Discordant for Juvenile Dermatomyositis: iPSC-Myogenic Cells

青少年皮肌炎的同卵双胞胎不一致：iPSC-肌源性细胞

• 批准号: 8770388
• 负责人: LAUREN M. PACHMAN
• 金额: $ 20.8万
• 依托单位: LURIE CHILDREN'S HOSPITAL OF CHICAGO
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-09-17 至 2016-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8770388
• 关键词: Adult; Age; Applications Grants; Autoimmune Diseases; Basic Science; Bioinformatics; Biological Assay; Biopsy; Blood; Cardiovascular system; Cells; Characteristics; Child; Childhood; Chronic; Chronic Disease; Data; Dermatomyositis; Development; Diagnosis; Diagnostic; Disease; Exanthema; Experimental Models; Fibroblasts; Functional disorder; Gender; Gene Expression Profile; Genes; Genetic; Germ Lines; Health; Idiopathic Inflammatory Myopathies; Immune; Immune response; In Vitro; Infection; Inflammation; Intervention; Investigation; Laboratories; Lead; Messenger RNA; Methods; MicroRNAs; Mitochondria; Molecular; Mononuclear; Monozygotic Twinning; Monozygotic twins; Mus; Muscle; Muscle Cells; Muscle Fibers; Muscle Weakness; Myositis; Pathway interactions; Patients; Pattern; Peripheral Blood Mononuclear Cell; Phenotype; Plasma; Population; Process; Production; Proteomics; RNA; Race; Rheumatism; Sendai virus; Skeletal Muscle; Somatic Cell; Source; Stem cells; Teratoma; Testing; Transplantation; Tumor Necrosis Factor-alpha; Twin Multiple Birth; Ultraviolet B Radiation; blastocyst; cell type; cytokine; embryonic stem cell; induced pluripotent stem cell; mRNA Expression; neuronal cell body; new technology; novel; peripheral blood; pluripotency; pre-clinical research; precursor cell; premature; research study; season of birth; success; systemic autoimmune disease; trait

DESCRIPTION (provided by applicant): Juvenile Dermatomyositis, the most common of the pediatric inflammatory myopathies, is a systemic pediatric autoimmune disease. There is no experimental model for JDM, greatly limiting our understanding of the pathophysiology of the disease process, which, like other autoimmune diseases, is associated with premature cardiovascular compromise and chronic inflammation. Heritable genetic factors, either unique or shared with other autoimmune diseases, predispose the child to develop JDM in concert with environmental influences, such as UVB exposure, season of birth and antecedent infection. Characterization of muscle-specific molecular pathways is severely constrained by the limited amount of involved muscle from the diagnostic biopsy available for research studies. We have documented dysregulation at the mRNA level and aberrant miRNA control in the muscle of untreated children with JDM, as well as active production of cytokines by the muscle fibers themselves, suggesting that they participate in the immune response, and are not only targets of it. A recent discovery showed that mononuclear cells, present in a small amount (5ml) of peripheral blood, can be "reprogrammed" by infecting them with a non-integrating Sendai virus to create " induced pluripotent stem cells" (iPSC). IPSCs share many traits with embryonic stem cells; they can induce the formation of teratomas when transplanted into mice, and give rise to a wide spectrum of differentiated cell types-among them myogenic progenitor cells, which then develop into muscle. In this study we will derive a single cell population of myogenic precursor cells from iPSCs from 3 sets of monozygotic twins discordant for JDM and their matched healthy controls in order to characterize the genetic contribution of muscle to disease pathophysiology. We hypothesize that the differentiated muscle cells derived from iPSCs will recapitulate molecular differences, associated with phenotype that we have previously identified in the transcriptome (miRNAs and mRNAs) of muscle biopsies from untreated children with JDM vs healthy controls. In Specific Aim #1, we will derive and compare iPSCs from three sets of monozygotic twins: one diagnosed with JDM, one unaffected-- and an age, gender, race matched control in order to: a) Verify their pluripotency by teratoma formation in a murine host and, b) Induce the embryoid body cells to form skeletal muscle in vitro. The unaffected twin will serve as a specific genetic and environmental control. Specific Aim # 2 will characterize the transcriptome differences, using RNAseq and miRNA assays, between: 1) the iPSCs from JDM, unaffected twin and healthy matched control as well as 2) their iPSC-derived myogenic cells to determine differences in IPSc and muscle from each donor as well as between donors. The Bioinformatics analysis will normalize the data and identify differentially expressed genes and their pathways in both the iPSCs and their derived muscle cells for each donor. The success of this project will lead to greater understanding of inflammatory myopathy and will open up a new avenue for pathophysiological investigation of JDM and other autoimmune diseases.

描述（由申请人提供）：青少年皮肌炎是最常见的小儿炎症性肌病，是一种全身性小儿自身免疫性疾病。 JDM 尚无实验模型，极大地限制了我们对该疾病过程病理生理学的理解，与其他自身免疫性疾病一样，JDM 与过早心血管损害和慢性炎症有关。遗传性遗传因素，无论是独特的还是与其他自身免疫性疾病共有的，与环境影响（例如 UVB 暴露、出生季节和既往感染）一起，使儿童容易患上 JDM。肌肉特异性分子途径的表征受到可用于研究的诊断活检中涉及的肌肉数量有限的严重限制。我们已经记录了未经治疗的 JDM 儿童肌肉中 mRNA 水平的失调和异常的 miRNA 控制，以及肌纤维本身积极产生细胞因子，表明它们参与免疫反应，而不仅仅是免疫反应的目标。最近的一项发现表明，存在于少量（5ml）外周血中的单核细胞可以通过用非整合型仙台病毒感染来“重新编程”，以产生“诱导多能干细胞”（iPSC）。 IPSC 与胚胎干细胞有许多共同特征；当它们移植到小鼠体内时，它们可以诱导畸胎瘤的形成，并产生多种分化细胞类型，其中包括肌源祖细胞，然后发育成肌肉。在这项研究中，我们将从 3 组与 JDM 不一致的同卵双胞胎及其匹配的健康对照中的 iPSC 中衍生出肌源性前体细胞的单细胞群，以表征肌肉对疾病病理生理学的遗传贡献。我们假设，源自 iPSC 的分化肌肉细胞将重现分子差异，这些差异与我们之前在未经治疗的 JDM 儿童与健康对照组的肌肉活检的转录组（miRNA 和 mRNA）中鉴定的表型相关。在具体目标 #1 中，我们将从三组同卵双胞胎中获得并比较 iPSC：一组被诊断患有 JDM，一组未受影响，以及年龄、性别、种族匹配的对照，以便：a) 通过畸胎瘤形成验证其多能性小鼠宿主，b) 诱导胚状体细胞在体外形成骨骼肌。未受影响的双胞胎将作为特定的遗传和环境控制。具体目标 # 2 将使用 RNAseq 和 miRNA 测定来表征以下之间的转录组差异：1) 来自 JDM、未受影响的双胞胎和健康匹配对照的 iPSC，以及 2) 其 iPSC 衍生的肌原细胞，以确定来自 JDM 的 iPSC 和肌肉的差异每个捐赠者以及捐赠者之间。生物信息学分析将对数据进行标准化，并识别每个供体的 iPSC 及其衍生肌肉细胞中的差异表达基因及其途径。该项目的成功将加深对炎症性肌病的了解，并为 JDM 和其他自身免疫性疾病的病理生理学研究开辟新途径。