Effect of oxygen on in vitro differentiation of mouse embryonic stem cells

The monolayer culture of embryonic stem (ES) cells and embryoid body (EB) formation were carried out under various oxygen tensions (i.e., 5% O2, 20% O2, and 40% O2). The ES cells cultured in 5% O2 and 20% O2 exhibited decreased specific alkaline phosphatase (AP) activity with culture time, whereas the ES cells cultured in 40% O2 retained their specific AP activity until culture day 3. The ES cells in 40% O2 maintained an octamer-binding transcription factor 4 (Oct-4; a marker of pluripotent undifferentiated ES cells) gene expression level higher than that at other oxygen tensions. The EB formed in 40% O2 maintained an Oct-4 gene expression level higher than that at other oxygen tensions. The generation of cardiomyocytes and the decline in Oct-4 gene expression level were earliest in the EB formed in 20% O2. It was suggested that the culture condition under a high oxygen tension (40% O2) retards the differentiation of ES cells and a normal oxygen tension (20% O2) allows spontaneous cell differentiation.