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INTRODUCTION: Residual pluripotent stem cells (PSC) within differentiated populations are problematic because of their potential to form tumors. Simple methods to reduce their occurrence are needed. METHODS: Here, we demonstrate that control of the oxygen partial pressure (pO2) to physiological levels typical of the developing embryo, enabled by culture on a highly oxygen permeable substrate, reduces the fraction of PSC within and the tumorigenic potential of differentiated populations. RESULTS: Differentiation and/or extended culture at low pO2 reduced measured pluripotency markers by up to four orders of magnitude for mouse PSCs (mPSCs). Combination with cell sorting increased the reduction to as much as six orders of magnitude. Upon implantation into immunocompromised mice, mPSCs differentiated at low pO2 either did not form tumors or formed tumors at a slower rate than at high pO2. CONCLUSIONS: Low pO2 culture alone or in combination with other methods is a potentially straightforward method that could be applied to future cell therapy protocols to minimize the possibility of tumor formation.
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PURPOSE OF REVIEW: To summarize recent reports on the effects of low oxygen on the undifferentiated phenotype and differentiation of embryonic stem cells (ESCs). RECENT FINDINGS: The oxygen level to which ESCs are exposed is an important environmental parameter. Under conditions maintaining the undifferentiated phenotype, low oxygen reduces spontaneous differentiation of human ESCs but reduces pluripotency gene expression in mouse ESCs, although reports are conflicting. Differentiation under low oxygen increases generation of neurons, cardiomyocytes, hematopoietic progenitors, endothelial cells, and chondrocytes. Many of the effects of low oxygen have been attributed to action by hypoxia inducible factor-1alpha (HIF-1alpha). The oxygen level in the gas phase (pO2gas) is often different than that experienced by the cells (pO2cell) and is unrecognized by investigators, which makes interpretation of the literature difficult. This difference increases with high cell densities, high cellular oxygen consumption rates, and large medium heights. The problem can be addressed by use of oxygen-permeable culture dishes and by estimation of pO2cell with mathematical models. SUMMARY: Low oxygen influences aspects of ESC pluripotency and differentiation. A better understanding of its effects and mechanism along with better estimation and control of pO2cell is important for applying low oxygen culture to regenerative medicine applications.