Abstract 21380: Reduced Mitochondrial Membrane Potential Leads to a Reduced Cytosolic Redox State in Human Embryonic Stem Cells
Introduction: Embryonic stem cells (ESCs) are characterized by pluripotency and unlimited self-renewal, but little is known about the role of metabolism as a regulator of these ESC characteristics. Cancer cells also have high self-renewal rates and accomplish this in part by favoring glycolysis. This lowers the production of mitochondrial reactive oxygen species (ROS) released during mitochondrial oxidation. We therefore investigated whether undifferentiated ESCs also exhibit reduced mitochondrial oxidation.
Methods: Mitochondrial activity and gene expression was compared between the WA01 human ESC line (WiCell) and primary human aortic smooth muscle cells (AoSMCs). Flow cytometry and immunofluorescence were used to evaluate mitochondrial mass (MitoTracker Green and Tomm20) and membrane potential (JC1 staining). Oxygen consumption was measured with an oxygen electrode. Cytosolic and mitochondrial redox status was determined after transfection with redox-sensitive GFP (roGFP) constructs.
Results: Expression of the mitochondrial biogenesis transcription factor PGC1α was 16.29±3.94 fold higher in ESCs when compared to mature vascular cells (P<0.05 vs AoSMCs). In line with this, mitochondria were abundant in ESCs (as shown with Mitotracker Green and Tomm20 stainings) and showed a dynamic mitochondrial network in live cells. Oxygen consumption in ESCs increased only minimally with addition of the mitochondrial uncoupler FCCP, suggesting that ESC mitochondria have a limited reserve capacity. Undifferentiated ESCs showed a reduced cytosolic redox status (0.34±0.02 versus 0.45±0.03 in differentiating cells, P<0.05), while the mitochondrial redox status was similar in both groups.
Conclusions: ESC show high levels of mitochondrial biogenesis with a low mitochondrial membrane potential, possibly due to high levels of the mitochondrial uncoupling protein UCP-2, which is known to lower ROS. In support of this finding, ESCs have a reduced cytosolic redox state compared to differentiating cells. Expression of UCP-2 therefore could be an essential prerequisite to protect ES cells against oxidative damage. Expression of UCP-2 could therefore be considered as a therapy to increase the regenerative potential of adult stem/progenitor cells.
- © 2010 by American Heart Association, Inc.