Natural underlying mtDNA heteroplasmy as a potential source of intra‐person hiPSC variability

Abstract

Functional variability among human clones of induced pluripotent stem cells (hiPSCs) remains a limitation in assembling high-quality biorepositories. Beyond inter-person variability, the root cause of intra-person variability remains unknown. Mitochondria guide the required transition from oxidative to glycolytic metabolism in nuclear reprogramming. Moreover, mitochondria have their own genome (mitochondrial DNA [mtDNA]). Herein, we performed mtDNA next-generation sequencing (NGS) on 84 hiPSC clones derived from a cohort of 19 individuals, including mitochondrial and non-mitochondrial patients. The analysis of mtDNA variants showed that low levels of potentially pathogenic mutations in the original fibroblasts are revealed through nuclear reprogramming, generating mutant hiPSCs with a detrimental effect in their differentiated progeny. Specifically, hiPSC-derived cardiomyocytes with expanded mtDNA mutations non-related with any described human disease, showed impaired mitochondrial respiration, being a potential cause of intra-person hiPSC variability. We propose mtDNA NGS as a new selection criterion to ensure hiPSC quality for drug discovery and regenerative medicine.

Random Mutagenesis, Clonal Events, and Embryonic or Somatic Origin Determine the mtDNA Variant Type and Load in Human Pluripotent Stem Cells

“…This diversity in mtDNA variants among lines derived from the same source was also reported in the works of Prigione et al., 2011, Kang et al., 2016b, and Perales-Clemente et al. (2016), all of them suggesting that the variants were already present in the fibroblast population.…”

Human iPSC-Derived Neural Progenitors Are an Effective Drug Discovery Model for Neurological mtDNA Disorders

“…In fact, iPSC-derived NPCs retained the entire mtDNA profile of the initial fibroblasts, including even the hypervariable D-loop region. Studies have indicated that the induction to iPSCs may be associated with heteroplasmic mtDNA alterations (Folmes et al, 2013;H€ am€ al€ ainen et al, 2013;Perales-Clemente et al, 2016;Prigione et al, 2011b). These variations appear to derive from the clonal origin of the iPSCs, because they also occur in clonally expanded fibroblasts (Kang et al, 2016;Ma et al, 2015).…”

In Vitro Tissue‐Engineered Skeletal Muscle Models for Studying Muscle Physiology and Disease

“…Recent studies have demonstrated that hiPSCs accumulate large and varied numbers of mitochondrial mutations within clones of the same isogenic lineage that can significantly impact mitochondrial function. [274,275] Resulting changes in cellular respiration would adversely affect engineered muscle function and outcomes of drug studies, and thus require frequent screening of mitochondrial DNA and metabolic function to prevent such issues. Given the heterogeneous nature of mutations, multiple clones from each hiPSC line should be pretested for resultant muscle function and robust guidelines established to ensure reproducibility and efficacy of the screening procedures.…”