Hypoxia driven effects on regulating of stem progenitor cell proliferation and differentiation have been shown in a number of in vitro systems, such as rat mesencephalic cell cultures, where hypoxia promoted neuronal differentiation and hypoxia inducible factor 1 a overexpression Tipifarnib order lead to similar results as hypoxia. Contrary to these previously mentioned stu dies in primary mouse neural stem cells, cell death was increased even though proliferation and differentiation were improved. Murine neural progenitor cells that were exposed to hypoxia prior to engraft ment into a rat brain displayed a better survival than those without hypoxic preconditioning. Studer et al. reported an increased number of differentiated neu ronal cells and showed trophic and proliferative effects of lowered oxygen levels on rat neural precursors.

Accordingly, in vivo, global and focal ischemia increases the proliferation and neuronal differentiation of neural stem cells in the sub ventricular zone and in the sub granular zone of the dentate gyrus. HIF 1a is one of the major key factors involved in the response to hypoxia and mediates a variety of cellular responses to hypoxia. In hypoxic conditions HIF 1a is stabilized and induces several cellular responses such as the acti vation of numerous target genes e. g. erythropoietin, glycolytic enzymes, BMP, Notch and prosurvival genes which are described to be involved in the regulation of the neuronal progenitor production with an increased neurogenesis as a part of an intrinsic hypoxia response in mice.

In our study we were interested in the effect of hypoxia on the GSK-3 neuronal dif ferentiation of human NPCs. Furthermore as EPO sig naling is hypoxia inducible, we tested whether or not EPO can mimic the effects of hypoxia under normoxic conditions. Therefore we investigated the differentiation potential of human NPCs expanded and differentiated in different oxygen concentrations and the involvement of EPO in this differentiation process. As EPO is known to mimic the effects of hypoxia our main objective was to demonstrate the differential effects of EPO in normoxic conditions and to illustrate that EPO causes subtle changes, but does not completely mimic hypoxia as suggested by major publications. Moreover, we demonstrated a complex network of reactions of human NPC towards hypoxia and propose a mechanism of action within this model. Results In our study we used the human immortalized neural pro genitor cell line ReNcell VM. This cell line possesses the potential to differentiate into functional neuronal cells, expressing markers like bIII tubulin and tyrosine hydroxylase. Furthermore the cell line is characterised by a fast proliferation and a rapid onset of differentiation upon the withdrawl of growth factors.