Bioenergetic shifts during transitions between stem cell states (2013 Grover Conference series)

Abstract

Two defining characteristics of stem cells are their multilineage differentiation potential (multipotency or pluripotency) and their capacity for self-renewal. Growth factors are well-established regulators of stem cell differentiation and self renewal, but less is known about the influence of the metabolic state on stem cell function. Recent studies investigating cellular metabolism during the differentiation of adult stem cells, human embryonic stem cells (ESCs), and induced pluripotent stem cells have demonstrated that activation of specific metabolic pathways depends on the type of stem cells as well as the lineage cells are differentiating into and that these metabolic pathways can influence the differentiation process. However, some common patterns have emerged, suggesting that undifferentiated stem cells primarily rely on glycolysis to meet energy demands. Our own data indicate that undifferentiated ESCs not only exhibit a low mitochondrial membrane potential but also express high levels of the mitochondrial uncoupling protein 2 and of glutamine metabolism regulators when compared with differentiated cells. More importantly, interventions that target stem cell metabolism are able to either prevent or enhance differentiation. These findings suggest that the metabolic state of stem cells is not just a marker of their differentiation status but also plays an active role in regulating stem cell function. Regulatory metabolic pathways in stem cells may thus serve as important checkpoints that can be modulated to direct the regenerative capacity of stem cells.

Keywords: UCP2; Warburg; differentiation; embryonic stem cells; glutamine; induced pluripotent stem cells; mesenchymal stem cells; metabolism; mitochondria; stem cells; uncoupling protein 2.

Figure 1

Uncoupled state in undifferentiated human embryonic stem cells (hESCs). Staining of an undifferentiated hESC (H1 cell line) colony (A, left) and adult human aortic smooth muscle cells (SMCs; A, right) with the potentiometric mitochondrial dye JC-1. Scale bar: 25 μm. Green indicates low mitochondrial membrane potential, and red indicates higher mitochondrial membrane potential. Oxygen consumption rate (OCR) is measured by the Seahorse XF-24 analyzer in adherent hESCs and aortic SMCs over time, when exposed to the adenosine triphosphate (ATP) synthase inhibitor oligomycin, the uncoupler carbonyl cyanide 4-(trifluoromethoxy)phenylhydrazone (FCCP), and the respiration inhibitor antimycin (B). Relative expression of the uncoupling protein 2 (UCP-2) in undifferentiated hESCs and SMCs as assessed by quantitative polymerase chain reaction (C). mRNA: messenger RNA.

Figure 2

Glutamine metabolism in human embryonic stem cells (hESCs). hESC (H1 cell line) growth rate decreases rapidly upon removal of glutamine from the culture medium (A). Upon differentiation of hESCs into embryoid bodies, the expression levels of mitochondrial glutaminase (GLS2) and the glutamine transporters SLC1A5 and SLC7A5 as assessed by quantitative polymerase chain reaction are markedly reduced (B). EB: embryoid body; GLS1: glutaminase 1.

Figure 3

Hypoxia induces embryonic transcription factors in adult mesenchymal stem cells (MSCs). Exposure of adult human MSCs to hypoxia (1% for 7 days) increases the expression of the embryonic pluripotency genes Oct-4 and Nanog as assessed by quantitative polymerase chain reaction. Y-axis indicates relative gene expression normalized to normoxic control gene expression levels (A). Immunofluorescence staining for Oct-4 confirms increased nuclear levels of Oct-4 in hypoxia (1% for 7 days) as well as when exposed to the prolyl hydroxylase inhibitor dimethyloxallyl glycine (DMOG; B). ESC: embryonic stem cell. Scale bar: 20 μm.

Figure 4

Metabolic continuum in stem cells. The pluripotency continuum correlates with a metabolic continuum in stem cells. ATP: adenosine triphosphate; ESC: embryonic stem cell; MSC: mesenchymal stem cell; ROS: reactive oxygen species; TCA: tricarboxylic acid.