In vitro hypoxic preconditioning of embryonic stem cells as a strategy of promoting cell survival and functional benefits after transplantation into the ischemic rat brain

Abstract

Hypoxic preconditioning (HP) and stem cell transplantation have been extensively studied as individual therapies for ischemic stroke. The present investigation is an initial effort to combine these methods to achieve increased therapeutic effects after brain ischemia. Sublethal in vitro hypoxia pretreatment significantly enhanced the tolerance of neurally-differentiating embryonic stem (ES) cells and primary bone marrow mesenchymal stem cells (BMSC) to apoptotic cell death (40–50% reduction in cell death and caspase-3 activation). The HP protective effects on cultured cells lasted for at least 6 days. HP increased secretion of erythropoietin (EPO) and upregulated expression of bcl-2, hypoxia-inducible factor (HIF-1α), erythropoietin receptor (EPOR), neurofilament (NF), and synaptophysin in ES cell-derived neural progenitor cells (ES-NPCs). The HP cytoprotective effect was diminished by blocking EPOR, while pretreatment of ES-NPCs with recombinant human EPO mimicked the HP effect. HP-primed ES-NPCs survived better 3 days after transplantation into the ischemic brain (30–40% reduction in cell death and caspase-3 activation). Finally, transplanted HP-primed ES-NPCs exhibited extensive neuronal differentiation in the ischemic brain, accelerated and enhanced recovery of sensorimotor function when compared to transplantation of non-HP-treated ES-NPCs. The cell-priming strategy aimed to promote transplanted cell survival and their tissue repair capability provides a simple yet effective way of optimizing cell transplantation therapy.