Abstract：
Duchenne muscular dystrophy (DMD) is a severe X-linked disorder characterized by dystrophin gene mutations and mitochondrial dysfunction, leading to progressive muscle weakness and premature death of DMD patients. We developed human Dystrophin Expressing Chimeric (DEC) cells, created by the fusion of myoblasts from normal donors and DMD patients, as a foundation for DT-DEC01 therapy for DMD. Our preclinical studies on mdx mouse models of DMD revealed enhanced dystrophin expression and functional improvements in cardiac, respiratory, and skeletal muscles after systemic intraosseous DEC administration. The current study explored the feasibility of mitochondrial transfer and fusion within the created DEC cells, which is crucial for developing new therapeutic strategies for DMD. Following mitochondrial staining with MitoTracker Deep Red and MitoTracker Green dyes, mitochondrial fusion and transfer was assessed by Flow cytometry (FACS) and confocal microscopy. The PEG-mediated fusion of myoblasts from normal healthy donors (MBN/MBN) and normal and DMD-affected donors (MBN/MBDMD), confirmed the feasibility of myoblast and mitochondrial fusion and transfer. The colocalization of the mitochondrial dyes MitoTracker Deep Red and MitoTracker Green confirmed the mitochondrial chimeric state and the creation of chimeric mitochondria, as well as the transfer of healthy donor mitochondria within the created DEC cells. These findings are unique and significant, introducing the potential of DT-DEC01 therapy to restore mitochondrial function in DMD patients and in other diseases where mitochondrial dysfunction plays a critical role.
摘要：
Duchenne型肌营养不良症（DMD）是一种严重的X连锁疾病，其特征是肌营养不良蛋白基因突变和线粒体功能障碍。导致DMD患者进行性肌无力和过早死亡。我们开发了表达人肌营养不良蛋白的嵌合（DEC）细胞，由正常供体和DMD患者的成肌细胞融合产生，作为DT-DEC01治疗DMD的基础。我们对DMD的mdx小鼠模型的临床前研究显示，心肌肌养蛋白表达增强和功能改善，呼吸,全身骨内DEC给药后的骨骼肌。本研究探讨了在所创建的DEC细胞内进行线粒体转移和融合的可行性，这对于开发新的DMD治疗策略至关重要。用MitoTracker深红色和MitoTracker绿色染料进行线粒体染色后，通过流式细胞术（FACS）和共聚焦显微镜评估线粒体融合和转移。PEG介导的来自正常健康供体（MBN/MBN）和正常和受DMD影响的供体（MBN/MBDMD）的成肌细胞的融合，证实了成肌细胞与线粒体融合转移的可行性。线粒体染料MitoTracker深红和MitoTrackerGreen的共定位证实了线粒体嵌合状态和嵌合线粒体的产生，以及健康供体线粒体在创建的DEC细胞内的转移。这些发现是独特而重要的，介绍DT-DEC01治疗在DMD患者和其他线粒体功能障碍中起关键作用的疾病中恢复线粒体功能的潜力。
