Pluripotent stem cell energy metabolism: an update

Abstract

Recent studies link changes in energy metabolism with the fate of pluripotent stem cells (PSCs). Safe use of PSC derivatives in regenerative medicine requires an enhanced understanding and control of factors that optimize in vitro reprogramming and differentiation protocols. Relative shifts in metabolism from naïve through "primed" pluripotent states to lineage-directed differentiation place variable demands on mitochondrial biogenesis and function for cell types with distinct energetic and biosynthetic requirements. In this context, mitochondrial respiration, network dynamics, TCA cycle function, and turnover all have the potential to influence reprogramming and differentiation outcomes. Shifts in cellular metabolism affect enzymes that control epigenetic configuration, which impacts chromatin reorganization and gene expression changes during reprogramming and differentiation. Induced PSCs (iPSCs) may have utility for modeling metabolic diseases caused by mutations in mitochondrial DNA, for which few disease models exist. Here, we explore key features of PSC energy metabolism research in mice and man and the impact this work is starting to have on our understanding of early development, disease modeling, and potential therapeutic applications.

Keywords: differentiation; epigenetics; metabolism; mitochondria; pluripotency.

Figure 1. Influence of energy metabolism on pluripotent status

Naïve human pluripotent stem cells (hPSCs) show an increase in ATP production through oxidative phosphorylation (OXPHOS) compared to more mature, “primed” hPSCs. Primed hPSCs can be converted to the naïve state through ectopic expression of NANOG and KLF4, inhibition of the ERK pathway by two inhibitors (2i), and stimulation with human leukemia inhibitory factor (L) (Takashima et al, 2014). Alternatively, the naïve state can be induced with a cocktail of five inhibitors and growth factors Activin and hLIF (5i/L/A) (Theunissen Thorold et al, 2014). Somatic cells can be reprogrammed with OCT4, SOX2, KLF4, and c-MYC (OSKM). Fibroblasts are more oxidative than primed hPSCs. Factors that activate glycolysis and inhibit OXPHOS promote induced PSC (iPSC) reprogramming. Vitamin C enhances iPSC reprogramming as an antioxidant and as a cofactor for epigenetic enzymes. Rapamycin, an inhibitor of the mTOR pathway, also increases the efficiency of iPSC reprogramming. Withdrawal of methionine from hPSC culture, which is required to maintain DNA and histone methylation, promotes differentiation.

Figure 2. Influence of metabolites on pluripotent stem cell epigenetics

Intermediate metabolism sets and maintains levels of metabolites that serve as substrates or cofactors for epigenetic modifying enzymes. Uptake of threonine and methionine from the culture media is required to maintain S-adenosylmethionine (SAM) levels in mPSCs and hPSCs, respectively. SAM is a methyl donor for histone methyltransferases (HMT) and DNA methyltransferases (DNMTs). Vitamin C is a cofactor for the JMJC family of demethylases and TET methylcytosine dioxygenases (TET). Acetyl-CoA, a TCA cycle intermediate, is an acetyl group donor for histone acetyltransferases (HAT). NAD⁺, generated through glycolysis or by the electron transport chain (ETC), is a cofactor for the sirtuin (SIRT) family of deacetylases.

Figure 3. Somatic cell reprogramming to pluripotency causes a mtDNA “bottleneck”

mtDNA undergoes a genetic bottleneck, or reduction in copy number, during de-differentiation, similar to the mtDNA bottleneck that occurs during normal female germ line oogenesis. This reduction has an unresolved mechanism that results in a shift from a heteroplasmic toward a homoplasmic state with no clear preference for wild-type or mutant mtDNA. This shift does not occur during the continuous culturing of fibroblasts in which characteristic levels of heteroplasmy are maintained over time.