Calcium and oxidative stress in muscular dystrophy

Duchenne肌营养不良症(DMD)是一种致命的遗传病，每3500名男孩中就有一人受到影响。来自多个实验室的越来越多的证据证实，钙调节失调和氧化应激是导致DMD的关键因素，提示钙隔离(CaSeq)或抗氧化(Antiox)途径的上调可能是治疗DMD的靶点。本应用旨在确定对营养不良表型最重要的CaSeq/antiox通路，并评估针对这些通路设计的基因疗法可以实现的治疗潜力。该项目由三个具体目标构成，并将利用两种DMD小鼠模型：MDX和MDX：UTUN-/-菌株。在目标1中，将使用肌肉细胞和分离的肌肉标本来表征单独的CaSeq/antiox通路对营养不良表型的影响。具有最实质性影响的CaSeq/antiox通路将被用作病毒介导的基因治疗的靶点。为了评估这些基因治疗的效果，营养不良小鼠将被静脉注射含有CaSeq/antiox转基因的重组腺相关病毒。我们将确定这些转基因是否可以延长营养不良小鼠的寿命，并纠正与营养不良蛋白缺乏相关的病理生理学。尽管应用范围仍然集中在DMD上，我们预计我们发现的治疗方面将与钙调节失调或氧化应激发挥关键作用的其他疾病的治疗直接相关，如阿尔茨海默氏症、衰老、糖尿病和心血管疾病。在应用的最终目的中，我们通过将MDX小鼠与现有拥有修改的CaSeq/antiox途径的小鼠品系杂交，获得了具有修改的CaSeq/antiox途径的突变的mdx小鼠。这些突变的mdx小鼠将是目前营养不良小鼠模型的有价值的补充，因为它们允许研究人员分离特定的CaSeq/antiox通路对营养不良表型的贡献。

Duchenne muscular dystrophy (DMD) is a lethal genetic disease that affects 1 in 3,500 boys. Accumulating evidence from multiple laboratories corroborate on the involvement of calcium misregulation and oxidative stress as key contributors to the disease, suggesting that upregulation of calcium-sequestering (CaSeq) or anti-oxidant (Antiox) pathways may serve as targets in the treament of DMD. The present application aims to identify the CaSeq/Antiox pathways most significant to the dystrophic phenotype, and assess the therapeutic potential that can be realized by a gene therapy designed to target these pathways. The project is framed by three specific aims and will utilize two murine models of DMD: the mdx and mdx:utrn-/- strains. In Aim 1, muscle cells and isolated muscle preparations will be used characterize the impact of individual CaSeq/Antiox pathways on the dystrophic phenotype. CaSeq/Antiox pathways with the most substantial impact will then be used as targets for viral-mediated gene therapies. To evaluate the efficacy of these gene therapies, dystrophic mice will be injected intravenously with recombinant adeno-associated viruses that contain CaSeq/Antiox transgenes. We will determine whether these transgenes can extend the lifespan of dystrophic mice and correct the pathophyosiology associated with dystrophin-deficiency. Although the scope of the application remains focused on DMD, we expect the therapeutic aspect of our findings to have a direct relevance in the treatment of other diseases where calcium misregulation or oxidative stress play a key role, such as Alzheimer's Disease, aging, diabetes and cardiovascular disease. In the final Aim of the application, we generate mutant mdx mice with modified CaSeq/Antiox pathways by crossing mdx mice with existing strains of mice that possess modified CaSeq/Antiox pathways. These mutant mdx mice will be valuable additions to current dystrophic mouse models, as they allow investigators to isolate the contribution of specific CaSeq/Antiox pathways to the dystrophic phenotype.