A Novel Xenograft Model of FSHD

一种新型的 FSHD 异种移植模型

基本信息

批准号：
8824131
负责人：
ROBERT J BLOCH
金额：
$ 38.38万
依托单位：
UNIVERSITY OF MARYLAND BALTIMORE
依托单位国家：
美国
项目类别：
财政年份：
2014
资助国家：
美国
起止时间：
2014-09-30 至 2016-09-29
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8824131
关键词：
Affect Biological Markers Biopsy Boston Cell Line Cells Characteristics Chromosomes Communities Contracts D4Z4 Data Development Direct Lytic Factors Disease DsRed Electric Stimulation Engraftment Exhibits Facioscapulohumeral Muscular Dystrophy Family Family member Fiber Fluorescence Genes Genetic Genetic screening method Genomics Goals Growth Hindlimb Human Immunocompromised Host In Vitro Individual Injection of therapeutic agent Label Laboratories Lead Left Link Luciferases Measures Methods Modeling Molecular Monitor Mus Muscle Muscle Fibers Muscular Dystrophies Natural regeneration Neuromuscular Junction PAX7 gene Pathogenesis Patients Pediatric Hospitals Pharmacy (field)Physiological Procedures Production Property Proteins Rag1 Mouse Reagent Relative (related person)Research Schools Scientist Source Staining method Stains Testing Therapeutic Tissues Torque Xenograft Model Xenograft procedure cohort drug testing improved irradiation man mouse model muscle form muscle regeneration neuromuscular novel peroneal nerve polyadenylated messenger RNA precursor cell public health relevance tibialis anterior muscle tool

项目摘要

DESCRIPTION (provided by applicant): Facioscapulohumeral muscular dystrophy (FSHD) is the third most common form of muscular dystrophy and affects 1 in 15,000 to 20,000 people worldwide. Developing and testing therapeutics for FSHD would be significantly advanced if a valid mouse model of the disease were available. Ideally, a murine version of FSHD would reproduce all the features of FSHD muscle, retaining the morphological, physiological and genomic differences found in fresh biopsies. As the pathophysiological mechanisms that lead to FSHD are still unclear, it would be best if such a model were constructed from FSHD tissue itself, rather than by manipulating particular genes or gene products that, although involved, may not be sufficient for pathogenesis. Finally, the mice should be accessible to many laboratories and so should be available in significant numbers. My colleagues and I have been developing such a model. In particular, we have been investigating the use of xenografting to create mice with humanized Tibialis anterior muscles developed from FSHD and control myogenic cell lines. In our preliminary studies, we have used methods that are routine in our laboratory, including X-irradiation of the lower hindlimbs of immunocompromised (NRG) mice and injection of cardiotoxin to eliminate the Tibialis anterior muscles of the mice and their abiliy to regenerate. We have also incorporated new methods, including periodic electrical stimulation of the graft through the peroneal nerve, and unique reagents and methods provided by our collaborators, Drs. W.E. Wright (UT Southwestern), P. Jones (UMass Med School) and L.M. Kunkel (Boston Children's Hospital). Our preliminary results, obtained with a human myogenic cell line that expresses luciferase, to facilitate monitoring by luminometry, show that these cells engraft into the otherwise empty TA compartment of mice, where they survive to form mature muscle fibers that are human in origin and that are essentially free of murine myonuclei. The human fibers are innervated and they contract upon stimulation of the peroneal nerve, to generate a specific force comparable to that of human muscle. As the grafts contain PAX7-positive human cells located adjacent to human myofibers, they are also likely to have the capacity for further growth. Recent data with a cell line derived from an FSHD patient indicate that these cells engraft efficiently as well, forming muscles that specifically express Dux4 and two of its downstream targets. This suggests that our methods preserve the genetic characteristics of myogenic cells as they engraft, grow, and differentiate. We have two specific aims: (i) to optimize the formation of mature human muscle tissue in mice, as a first step in developing a model of FSHD for therapeutic testing; and (ii) to apply our optimized methods to cells from FSHD patients and their unaffected relatives, to develop, characterize, and assess the variability of a novel model of FSHD in mice, for subsequent therapeutic testing. Our studies should therefore generate a model of FSHD muscle in mice that will be an invaluable reagent in identifying and testing drugs to treat FSHD.

描述（由申请人提供）：面肩肱型肌营养不良症 (FSHD) 是第三种最常见的肌营养不良症，全世界每 15,000 至 20,000 人中就有 1 人受到影响，如果该疾病的有效小鼠模型，那么 FSHD 治疗方法的开发和测试将会取得显着进展。理想情况下，FSHD 的小鼠版本将重现 FSHD 肌肉的所有特征，保留形态、由于导致 FSHD 的病理生理学机制仍不清楚，因此最好从 FSHD 组织本身构建这样的模型，而不是通过操纵特定的基因或基因产物，尽管这些基因或基因产物涉及最后，许多实验室都可以使用这些小鼠，因此我和我的同事一直在开发这样的模型，特别是我们一直在研究使用异种移植来创建。在我们的初步研究中，我们使用了实验室常规方法，包括对免疫受损 (NRG) 小鼠的下后肢进行 X 射线照射和注射心脏毒素。我们还采用了新方法，包括通过腓神经定期电刺激移植物，以及提供独特的试剂和方法。由我们的合作者 W.E. Wright 博士（德克萨斯大学西南分校）、P. Jones（麻省大学医学院）和 L.M. Kunkel（波士顿儿童医院）得出，我们的初步结果是通过表达荧光素酶的人类肌原细胞系获得的，以便于通过发光测定法进行监测。，表明这些细胞移植到小鼠原本空的 TA 隔室中，它们在那里存活并形成成熟的肌肉纤维，这些纤维源自人类并且基本上不含小鼠肌核。人类纤维受到神经支配，在腓神经受到刺激时收缩，产生肌肉纤维。由于移植物含有位于人类肌纤维附近的 PAX7 阳性人类细胞，因此它们也可能具有进一步生长的能力。 FSHD 患者表明这些细胞也能有效移植，形成特异性表达 Dux4 及其两个下游靶标的肌肉，这表明我们的方法在移植、生长和分化时保留了肌源性细胞的遗传特征。：（i）优化小鼠成熟人类肌肉组织的形成，作为开发用于治疗测试的 FSHD 模型的第一步；（ii）将我们的优化方法应用于来自 FSHD 患者及其未受影响亲属的细胞，以发展，因此，我们的研究应该建立小鼠 FSHD 肌肉模型，该模型将成为识别和测试治疗 FSHD 药物的宝贵试剂。