Induced Pluripotent Stem Cells with a Mitochondrial DNA Deletion

Abstract

In congenital mitochondrial DNA (mtDNA) disorders, a mixture of normal and mutated mtDNA (termed heteroplasmy) exists at varying levels in different tissues, which determines the severity and phenotypic expression of disease. Pearson marrow pancreas syndrome (PS) is a congenital bone marrow failure disorder caused by heteroplasmic deletions in mtDNA. The cause of the hematopoietic failure in PS is unknown, and adequate cellular and animal models are lacking. Induced pluripotent stem (iPS) cells are particularly amenable for studying mtDNA disorders, as cytoplasmic genetic material is retained during direct reprogramming. Here we derive and characterize iPS cells from a patient with PS. Taking advantage of the tendency for heteroplasmy to change with cell passage, we isolated isogenic PS-iPS cells without detectable levels of deleted mtDNA. We found that PS-iPS cells carrying a high burden of deleted mtDNA displayed differences in growth, mitochondrial function, and hematopoietic phenotype when differentiated in vitro, compared to isogenic iPS cells without deleted mtDNA. Our results demonstrate that reprogramming somatic cells from patients with mtDNA disorders can yield pluripotent stem cells with varying burdens of heteroplasmy that might be useful in the study and treatment of mitochondrial diseases.

De novo generation of HSCs from somatic and pluripotent stem cell sources

“…V617F myelo-proliferative disorder, 75 dyskeratosis congenita, 76 Pearson syndrome, 77 and others. Hematopoietic differentiation of these iPSCs have recapitulated some aspects of these disorders, 72,77 and uncovered mechanistic links between multiple tissue deficits.…”

“…Hematopoietic differentiation of these iPSCs have recapitulated some aspects of these disorders, 72,77 and uncovered mechanistic links between multiple tissue deficits. 73 However, hematopoietic differentiation of hPSCs yield short-lived progenitors and mature cells 16,78 and without the possibility of generating large numbers of hematopoietic stem and progenitor cells, these protocols limit the scope of experiments that can be performed in vitro and preclude disease modeling in vivo.…”

Mitochondrial Respiratory Defect Causes Dysfunctional Lactate Turnover via AMP-activated Protein Kinase Activation in Human-induced Pluripotent Stem Cell-derived Hepatocytes

“…Decreases in mitochondrial heteroplasmy in some clones of iPSCs have been observed during reprogramming and long term passaging in recent studies (25,26). This phenomenon causes difficulties in the modeling of mitochondrial diseases with iPSCs.…”

“…This reprogramming of mitochondria enables modeling of mitochondrial diseases in specialized cell types because iPSCs are able to differentiate into diverse cell types. Until now, only few mtDNA mutations have been investigated in iPSCs, and any mutation on MTND1 has not been studied in iPSCs (25)(26)(27)(28). Here, we generated disease-specific iPSCs with defective mitochondrial respiratory complex (DMRC-iPSCs) by m.3398T3 C mutation on the MTND1 gene and examined metabolic phenomena in hepatocytes derived from those DMRC-iPSCs.…”

A step forward in disease modelling for mitochondrial diseases

“…This last point is of particular importance, given the significant mtDNA genetic diversity seen in different iPSC generated from the same cellular source. The authors acknowledge that a thorough analysis of the genotype of the cells is imperative, especially since it is known from other studies that iPSC carrying certain mutations show an impaired differentiation and dysfunctional mitochondria, which could severely hamper the reliability of the study (12,13). Also in this line, the authors suggest that, although they only worked on homoplasmic mutations, it should be possible to model in the same way the effect of heteroplasmic variants.…”