Oxygen tension modulates beta-globin switching in embryoid bodies

Little is known about the factors influencing the hemoglobin switch in vert ebrates during develop ment. Inasmuch as the mammalian conceptus is exposed to changing oxygen tensions in utero, we examined the effect of different oxygen concentrations on beta-globin switching. We used an in vitro model o f mouse embryogenesis based on the differentiation of blastocyst-derived em bryonic stem cells to embryoid bodies (EBs). Cultivation of EBs at increasi ng oxygen concentrations (starting at 1% O-2) did not influence the tempora l expression pattern of embryonic (beta H1) globin compared to the normoxic controls (20% O-2). In contrast, when compared to normoxically grown EBs, expression of fetal/adult (beta(maj)) globin in EBs cultured at varying oxy gen concentrations was delayed by about 2 days and persisted throughout dif ferentiation. Quantitation of hemoglobin in EBs using a 2,7-diaminofluorene -based colorimetric assay revealed the appearence of hemoglobin in two wave s, an early and a late one. This observation was verified by spectrophotome tric analysis of hemoglobin within single EBs. These two waves might reflec t the switch of erythropoiesis from yolk sac to fetal liver. Reduced oxygen ation is known to activate the hypoxia-inducible factor-1 (HIF-1), which in turn specifically induces expression of a variety of genes among them eryt hropoietin (EPO). Although EBs increased EPO expression upon hypoxic exposu re, the altered beta-globin appearance was not related to EPO levels as det ermined in EBs overexpressing EPO. Since mRNA from both mouse HIF-1 alpha i soforms was detected in all EBs tested at different differentiation stages, we propose that HIV-1 modulates beta-globin expression during development.