Abstract
Moderate hypoxic preconditioning of adipose-derived stem cells (ASCs) enhances properties such as proliferation and secretion of growth factors, representing a valuable strategy to increase the efficiency of cell-based therapies. In a wide variety of cells potassium (K+) channels are key elements involved in the cellular responses to hypoxia, suggesting that ASCs cultured under low oxygen conditions may display altered electrophysiological properties. Here, the effects of moderate hypoxic culture on proliferation, whole-cell currents, and ion channel expression were investigated using human ASCs cultured at 5% and 20% oxygen. Although cell proliferation was greatly enhanced, the dose-dependent growth inhibition by the K+ channel blocker tetraethylammonium (TEA) was not significantly affected by hypoxia. Under both normoxic and hypoxic conditions, ASCs displayed outward K+ currents composed by Ca2+-activated, delayed rectifier, and transient components. Hypoxic culture reduced the slope of the current-voltage curves and caused a negative shift in the voltage activation threshold of the whole-cell currents. However, the TEA-mediated shift of voltage activation threshold was not affected by hypoxia. Semiquantitative real-time RT-PCR revealed that expression of genes encoding for various ion channels subunits related to oxygen sensing and proliferation remained unchanged after hypoxic culture. In conclusion, outward currents are influenced by moderate hypoxia in ASCs through a mechanism that is not likely the result of modulation of TEA-sensitive K+ channels.
Highlights
Within the field of regenerative medicine, a multitude of clinical trials using autologous stem cell transplantation are currently under way [1]
By comparing adipose-derived stem cells (ASCs) cultured at 1%, 5%, and 21% oxygen, we demonstrated that the exposure to oxygen levels of 1% is optimal for promotion of the pro-angiogenic properties of ASC in terms of secretion of vascular endothelial growth factor (VEGF-1), whereas culture at 5% oxygen yields faster proliferation [4,5]
As shown by the TEA-mediated growth inhibition assay, K+ channels underlying the main components of the outward current (IKDR and IKCa) play a key role on the proliferation of ASCs
Summary
Within the field of regenerative medicine, a multitude of clinical trials using autologous stem cell transplantation are currently under way [1]. The ongoing and proposed clinical trials include transplantation of recently harvested cells, and expansion, preconditioning and predifferentiation of cells prior to implantation. In this context, it is noteworthy that culture of ASCs in hypoxic conditions alters their properties, both in terms of differentiation, secretion of various growth factors, as well as proliferation (reviewed by Zachar et al.) [3]. By comparing ASCs cultured at 1%, 5%, and 21% oxygen, we demonstrated that the exposure to oxygen levels of 1% is optimal for promotion of the pro-angiogenic properties of ASC in terms of secretion of vascular endothelial growth factor (VEGF-1), whereas culture at 5% oxygen yields faster proliferation [4,5]. The beneficial effect of moderate hypoxia on ASC proliferation without loss of multipotentiality has been demonstrated even for longer culture periods of almost two months [6]
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