St. Boniface Hospital Albrechtsen Research Centre, Regenerative Medicine Program, Department of Physiology and Pathophysiology, Rady Faculty of Health Sciences, Institute of Cardiovascular Sciences, Max Rady College of Medicine, University of Manitoba, Winnipeg, MB R2H 2A6, Canada.
Department of Pediatrics and Child Health, Rady Faculty of Health Sciences, Max Rady College of Medicine, University of Manitoba, Winnipeg, MB R3E 3P4, Canada.
Cells. 2021 Mar 5;10(3):568. doi: 10.3390/cells10030568.
Kearns Sayre syndrome (KSS) is mitochondrial multisystem disorder with no proven effective treatment. The underlying cause for multisystem involvement is the energy deficit resulting from the load of mutant mitochondrial DNA (mtDNA), which manifests as loss of cells and tissue dysfunction. Therefore, functional organ or cellular replacement provides a promising avenue as a therapeutic option. Patient-specific induced pluripotent stem cells (iPSC) have become a handy tool to create personalized cell -based therapies. iPSC are capable of self-renewal, differentiation into all types of body cells including cardiomyocytes (CM) and neural progenitor cells (NPC). In KSS patients, mutations in mtDNA are largely found in the muscle tissue and are predominantly absent in the blood cells. Therefore, we conceptualized that peripheral blood mononuclear cells (PBMNC) from KSS patients can be reprogrammed to generate mutation free, patient specific iPSC lines that can be used as isogenic source of cell replacement therapies to treat affected organs. In the current study we generated iPSC lines from two female patients with clinical diagnosis of classic KSS. Our data demonstrate that iPSC from these KSS patients showed normal differentiation potential toward CM, NPC and fibroblasts without any mtDNA deletions over passages. Next, we also found that functional studies including ATP production, reactive oxygen species generation, lactate accumulation and mitochondrial membrane potential in iPSC, CM, NPC and fibroblasts of these KSS patients were not different from respective cells from healthy controls. PBMNCs from these KSS patients in the current study did not reproduce mtDNA mutations which were present in muscle biopsies. Furthermore, we demonstrate for the first time that this phenomenon provides opportunities to create isogenic mutation free iPSC with absent or very low level of expression of mtDNA deletion which can be banked for future cell replacement therapies in these patients as the disease progresses.
Kearns-Sayre 综合征（KSS）是一种线粒体多系统疾病，目前尚无有效的治疗方法。多系统受累的根本原因是突变线粒体 DNA（mtDNA）负荷导致的能量不足，表现为细胞丢失和组织功能障碍。因此，功能性器官或细胞替代提供了一种有前途的治疗选择。患者特异性诱导多能干细胞（iPSC）已成为创建个性化细胞治疗的有用工具。iPSC 具有自我更新的能力，能够分化为包括心肌细胞（CM）和神经祖细胞（NPC）在内的所有类型的体细胞。在 KSS 患者中，mtDNA 突变主要存在于肌肉组织中，而在血细胞中则主要不存在。因此，我们设想可以将 KSS 患者的外周血单个核细胞（PBMNC）重编程为生成无突变的、患者特异性的 iPSC 系，可作为同源细胞替代疗法的来源，用于治疗受影响的器官。在本研究中，我们从两名具有经典 KSS 临床诊断的女性患者中生成了 iPSC 系。我们的数据表明，这些 KSS 患者的 iPSC 显示出向 CM、NPC 和成纤维细胞正常分化的潜力，且在传代过程中没有任何 mtDNA 缺失。接下来，我们还发现，功能研究包括这些 KSS 患者的 iPSC、CM、NPC 和成纤维细胞中的 ATP 产生、活性氧生成、乳酸积累和线粒体膜电位，与来自健康对照的相应细胞没有差异。来自这些 KSS 患者的 PBMNC 在本研究中并未复制肌肉活检中存在的 mtDNA 突变。此外，我们首次证明，这种现象为创建无 mtDNA 缺失或缺失极低水平表达的同基因无突变 iPSC 提供了机会，可在这些患者的疾病进展过程中为未来的细胞替代疗法储存这些 iPSC。
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