Mitochondrial Spare Respiratory Capacity Is Negatively Correlated with Nuclear Reprogramming Efficiency

Abstract

Nuclear reprogramming efficiency has been shown to be highly variable among different types of somatic cells and different individuals, yet the underlying mechanism remains largely unknown. Several studies have shown that reprogramming of fibroblasts into induced pluripotent stem cells (iPSCs) requires remodeling of mitochondria and a metabolic shift from an oxidative state to a more glycolytic state. In this study, we evaluated the nuclear reprogramming efficiency in relation to mitochondrial bioenergetic parameters of fibroblasts from seven different human individuals. Using the Seahorse extracellular energy flux analyzer, we measured oxygen consumption rate (OCR) profiles of the cells, along with their nuclear reprogramming efficiency into iPSCs. Our results showed that fibroblasts with the lowest mitochondrial spare respiratory capacity (SRC) had the highest nuclear reprogramming efficiency, opposed to fibroblasts with the highest mitochondrial SRC, which showed lowest reprogramming efficiency. Furthermore, we found that targeted fluorescent tagging of endogenous genes (MYH6 and COL2A1) by CRISPR/Cas9-mediated homologous recombination was accompanied by an increase in the SRC level of the modified fibroblasts and impaired reprogramming efficiency. Our findings indicate a negative correlation between high mitochondrial SRC in somatic cells and low reprogramming efficiencies. This type of analysis potentially allows screening and predicting reprogramming efficiency before reprogramming, and further suggests that nuclear reprogramming might be improved by approaches that modulate the SRC.

Integrative analysis of Dupuytren's disease identifies novel risk locus and reveals a shared genetic etiology with BMI

“…The upregulated DEG sets for tibial artery and aorta tissues both demonstrated significant ( P DEG = 5.5 x 10 ‐5 and 7.8 x 10 ‐5 , respectively) overlap with the GWAS gene set (Table S8). We also performed a gene property analysis using cell type specific expression data for 5115 study‐defined cell types from 28 scRNA‐seq studies (Alles et al, ; Breton et al, ; Campbell et al, ; Chen et al, ; Darmanis et al, ; Enge et al, ; Furlan et al, ; Gokce et al, ; Haber et al, ; Habib et al, ; Han et al, ; Häring et al, ; Hochgerner et al, , ; Hu et al, ; Joost et al, ; La Manno et al, ; Mohammed et al, ; Romanov et al, ; Saunders et al, ; Tasic et al, ; Usoskin et al, ; Vanlandewijck et al, ; Zeisel et al, , ; Zhong et al, ; Zhou et al, ). While none of the single cell types were significant ( P GP,CT ≤ 0.05/5115), stromal cells and muscle cells were among the top five results (Table S9).…”

“…For the gene property analysis, FUMA tests if the expression of the GWAS gene set in a single tissue or cell type is statistically different than the average expression of the GWAS gene set across all tissues or cell types. We perform this gene property analysis in 53 GTEx (GTEx Consortium, ) tissues () as well as in 5115 study‐defined cell types () using single cell RNA‐seq data from 28 studies (Alles et al, ; Breton et al, ; Campbell et al, ; Chen, Wu, Jiang, & Zhang, ; Darmanis et al, ; Enge et al, ; Furlan et al, ; Gokce et al, ; Haber et al, ; Habib et al, ; Han et al, ; Häring et al, ; Hochgerner et al, ; Hochgerner, Zeisel, Lönnerberg, & Linnarsson, ; Hu et al, ; Joost et al, ; La Manno et al, ; Mohammed et al, ; Romanov et al, ; Saunders et al, ; Tasic et al, ; Usoskin et al, ; Vanlandewijck et al, ; Zeisel et al, , ; Zhong et al, ; Zhou et al, ) as described on the FUMA website (see Web Resources). For the DEG analysis, FUMA defines differentially expressed genes in each tissue by performing a two‐sided t test for that one tissue against all other tissues.…”

mRNA Reprogramming of T8993G Leigh's Syndrome Fibroblast Cells to Create Induced Pluripotent Stem Cell Models for Mitochondrial Disorders

“…We, therefore, expect that LS1-hiPSCs and their differentiated derivatives will exhibit bioenergetics defects due to increased mutation burden. It is widely acknowledged that determining oxygen consumption rate and lactic acid production rate is accurate measures of mitochondrial function and has been extensively used in reprogrammed stem cells, progenitors, and differentiated cells [61][62][63][64]. Our future studies will focus on quantitatively determining correlations between level of mutation burden and alteration in mitochondrial function in the context of LS.…”

Metformin use in pregnancy: promises and uncertainties

“…It should be noted that the ESC line that was used to study potential teratogenic effects of metformin [ 10 ] does not exhibit the responsiveness to high glucose exposure and the high rates of glycolysis relative to oxidative phosphorylation of normal embryos or of ESC derived in physiological glucose media [ 31 ]. While conventional stem cell (embryonic and induced pluripotent) lines exhibit fewer, and less mature, mitochondria compared with more differentiated cells, and high rates of glycolysis relative to oxidative phosphorylation are essential for induction of pluripotency from differentiated cells [ 32 – 34 ], conventional ESCs may be more dependent on mitochondrial metabolism (and less dependent on glycolysis) than the normal mouse embryo in vivo. One can speculate that the endogenous solutes taken up by metformin transporters (OCTs) have a function in mitochondrial metabolism, such that metformin transporters would not be expressed by cells that are highly dependent on glycolysis, but would occur upon maturation of mitochondrial respiration (Fig.…”