Analysis and Therapy of Muscular Dystrophy in Zebrafish

在过去的5年里，我们一直致力于将斑马鱼建立为肌肉发达的模型 营养不良。为此，我们发表了缺乏抗肌营养不良蛋白的斑马鱼的表型和8- 完成了一项大的遗传筛选，以分离更多的营养不良突变体，并鉴定了 Runzel突变体中的突变基因。利用我们在肌肉研究和建立 斑马鱼作为一种疾病模型，我们现在建议用斑马鱼来研究肌肉的发病机制 并评估细胞疗法作为一种潜在的治疗选择。 这个项目的第一个目标是全面描述我们现有的两种营养不良的斑马鱼。 模型(EMZ和SOF)，目的是更好地理解肌营养不良的发病机制。这些 突变体的肌肉退化表型与dystrophin突变体(Saje)非常相似，这表明 这种表型是肌肉营养不良的症状。我们建议确定人类基因组的基因突变 这些突变体使用传统的作图方法，然后对候选基因进行测序来鉴定 特定的突变。如果人类的同源基因目前与肌营养不良症没有关联， 这些基因将被认为是疾病候选基因，并在人类患者中进行测序 肌肉营养不良的原因尚不清楚。因为看似不相关的蛋白质的突变可以表现为 对于肌营养不良，识别更多的基因将有助于建立疾病途径。 其次，我们已经建立了将细胞种群移植到斑马鱼身上的方法 现在建议使用这个系统来识别最有能力的细胞群体 植入并矫正病变肌肉。肌肉移植细胞的基因表达谱 将把群体与非移植细胞进行比较，以确定主要在 移植细胞。差异表达的基因将被认为是潜在的标记，并用于纯化 哺乳动物中类似的细胞群体，用于未来的实验和治疗。最后，我们计划剖析 从斑马鱼的发育潜能分析不同干细胞种群的谱系关系 肌肉祖细胞。这将通过早期移植有限数量的标记细胞来实现。 在发育中，然后随着鱼的成熟而跟随它们的命运。

Over the last 5 years, we have been working to establish the zebrafish as a model for muscular dystrophy. In this capacity, we have published the phenotype of zebrafish lacking dystrophin and 8- sarcoglycan, completed a large genetic screen to isolate additional dystrophic mutants, and identified the mutant gene in the runzelmutant. Using our experiences in muscle research and in establishing the zebrafish as a disease model, we now propose to use the fish to investigate the pathogenesis of muscular dystrophy and evaluate cell therapy as a potential treatment option. The first aim in this project proposes to fully characterize two of our available dystrophic zebrafish models (emz and sof) with the goal of better understanding the pathogenesis of muscular dystrophy. These mutants show a muscle degeneration phenotype very similar to the dystrophin mutant (sapje) suggesting that this phenotype is symptomatic of muscular dystrophy. We propose to identify the genetic mutations in these mutants using a traditional mapping approach and then sequencing candidate genes to identify the specific mutation. If the orthologous human genes are not currently associated with muscular dystrophy, these genes will be considered disease candidate genes and sequenced in human patients for which the cause of muscular dystrophy is unknown. Since mutations in seemingly unrelated proteins can manifest as muscular dystrophy, the identification of additional genes would be helpful for establishing disease pathways. Secondly, we have established methods to transplant cell populations in zebrafish at all developmental stages and now propose using this system to identify the cell population most capable of engrafting into and correcting the diseased muscle. Gene expression profiles of muscle engrafting cell populations will be compared with non-engrafting cells to identify genes expressed predominantly in the engrafting cells. Differentially expressed genes will be considered potential markers and used to purify analogous cell populations in mammals for future experimentation and therapy. Finally, we plan to dissect the lineage relationship of various stem cell populations by assaying the developmental potential of zebrafish muscle progenitor cells. This will be accomplished by transplanting limited populations of labeled cells early in development and then following their fate as the fish matures.