MANIPULATION OF PTEN/AKT SIGNALING IN DUCHENNE MUSCULAR DYSTROPHY AS A MEANS OF T

杜氏肌营养不良症中 PTEN/AKT 信号传导的调控作为 T 的一种手段

• 批准号: 8828567
• 负责人: LOUIS M KUNKEL
• 金额: $ 38.83万
• 依托单位: BOSTON CHILDREN'S HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-04-01 至 2019-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8828567
• 关键词: AKT Signaling Pathway; Affect; Animal Model; Biopsy; Canis familiaris; Cell Culture Techniques; Cell Differentiation process; Cell Line; Cell Survival; Cells; Clinical; Coupled; Data; Disease; Disease Progression; Doxycycline; Duchenne muscular dystrophy; Dystrophin; Future; Gene Targeting; Generations; Genes; Goals; Growth; Health; Human; In Vitro; Inhibition of Apoptosis; Injection of therapeutic agent; Knockout Mice; Laboratories; Lead; Link; MicroRNAs; Microarray Analysis; Modeling; Mus; Muscle; Muscle Cells; Muscular Dystrophies; Mutation; Myopathy; Neuromuscular Junction; Outcome; PTEN gene; Pathogenesis; Pathology; Pathway interactions; Patients; Phenotype; Physiology; Production; Proteins; Proto-Oncogene Proteins c-akt; Publishing; Regulation; Role; Route; Severity of illness; Signal Pathway; Signal Transduction; Skeletal Muscle; Structural Genes; Subfamily lentivirinae; Symptoms; Tamoxifen; Testing; Therapeutic; Therapeutic Uses; Transgenic Mice; Utrophin; Vascularization; Wallerian Degeneration; Wasting Syndrome; Zebrafish; adeno-associated viral vector; boys; cell growth; combinatorial; comparative; disease-causing mutation; dosage; functional improvement; improved; in vivo; mRNA Expression; mouse model; muscle degeneration; muscle hypertrophy; novel; novel therapeutic intervention; overexpression; research study; skeletal; transcriptome sequencing; wasting

DESCRIPTION (provided by applicant): Duchenne Muscular Dystrophy (DMD) is a degenerative muscle wasting disease caused by mutations in the dystrophin gene. The absence of dystrophin protein in muscle results in dysregulated secondary signaling pathways, many of which remain poorly understood. The laboratory of Dr. Mayana Zatz has identified two dystrophin-deficient dogs from a litter of Golden Retriever Muscular Dystrophy (GRMD) dogs that are mildly affected and escape the early lethality and dystrophic phenotype (muscle wasting) that normally occur in severely affected GRMD dogs. Our microarray analysis of these dogs and other GRMD and control dogs, revealed a strong reduction of the Pitpna gene, which is a known regulator of the PTEN/AKT signaling pathway. In parallel studies using microRNA microarrays performed on human biopsies from patients with DMD, we identified a microRNA, miR-486, that is exclusively reduced in DMD muscle and has been shown by us and others to alter PTEN/AKT signaling. We hypothesize that either overexpression of miR-486 and/or inhibition of Pitpna results in decreased PTEN levels while subsequently increasing phosphorylated (activated) AKT, which might lead to reduced severity of disease course. The induction of phosphorylated-AKT (resulting from miR-486 and Pitpna modulation), are predicted to affect several downstream target genes of this pathway which will result in an increase in muscle hypertrophy, muscle structural genes vascularization genes, the increase in the neuromuscular junction number of branch points, inhibition of apoptosis genes, and validated PTEN/AKT downstream target muscle genes (such as Utrophin). We plan to approach our long-term goal of understanding how these two alter signaling pathways in DMD by modulating expression of miR-486 and/or Pitpna in the mouse (mdx5cv) model according to the following three specific aims: 1) Characterization of miR- 486 and Pitpna regulation of PTEN/AKT signaling pathway in vitro. 2) Manipulation of miR-486 expression to modulate PTEN/AKT signaling in dystrophin-deficient muscle for modeling of future therapeutics. 3) Generation of mice with reduced Pitpna levels in muscle and characterization of its effects on PTEN/AKT signaling.

描述（申请人提供）：杜氏肌营养不良症（DMD）是一种由肌营养不良蛋白基因突变引起的退行性肌肉萎缩疾病。肌肉中抗肌营养不良蛋白的缺失会导致次级信号传导途径失调，其中许多信号传导途径仍知之甚少。 Mayana Zatz 博士的实验室从一窝患有轻度肌营养不良症 (GRMD) 的金毛猎犬中鉴定出了两只缺乏抗肌营养不良蛋白的狗，它们避免了通常发生在严重受影响的 GRMD 狗中的早期致死和营养不良表型（肌肉萎缩）。我们对这些狗以及其他 GRMD 和对照狗的微阵列分析显示，Pitpna 基因显着减少，该基因是 PTEN/AKT 信号通路的已知调节因子。在使用 microRNA 微阵列对 DMD 患者的人体活检进行的平行研究中，我们发现了一种 microRNA，即 miR-486，它专门在 DMD 肌肉中减少，并且我们和其他人已证明它可以改变 PTEN/AKT 信号传导。我们假设 miR-486 的过度表达和/或 Pitpna 的抑制会导致 PTEN 水平降低，同时随后磷酸化（激活）AKT 增加，这可能会导致病程严重程度减轻。磷酸化 AKT 的诱导（由 miR-486 和 Pitpna 调节产生）预计会影响该途径的几个下游靶基因，从而导致肌肉肥大、肌肉结构基因血管化基因增加、神经肌肉接头分支点数量增加、凋亡基因抑制以及经过验证的 PTEN/AKT 下游靶肌肉基因（例如 如 Utropin）。我们计划根据以下三个具体目标，通过调节小鼠 (mdx5cv) 模型中 miR-486 和/或 Pitpna 的表达来了解这两种信号通路如何改变 DMD 中的信号通路：1) 体外表征 miR-486 和 Pitpna 对 PTEN/AKT 信号通路的调节。 2) 操纵 miR-486 表达来调节肌营养不良蛋白缺陷肌肉中的 PTEN/AKT 信号传导，用于未来治疗的建模。 3) 肌肉中 Pitpna 水平降低的小鼠的产生及其对 PTEN/AKT 信号传导的影响的表征。