Oxygen Tension Alters the Effects of Cytokines on the Megakaryocyte, Erythrocyte, and Granulocyte Lineages

Many investigators believe that cytokines have exclusive control over proliferation and differentiation in stroma-free hematopoietic cultures. Although cytokines are indeed necessary, other culture parameters such as oxygen (O2) tension can greatly influence both hematopoietic cell proliferation and differentiation. We investigated the effects of cytokine combinations and O2 tension on the expansion of megakaryocyte (size and number of CD41a+ cells and number of colony-forming units-megakaryocyte [CFU-Mk]), granulocyte (CD15+ cells and CFU-granulocyte monocyte [CFU-GM]), and erythrocyte (hemoglobin+ cells and burst-forming units-erythrocyte [BFU-E]) lineages. Peripheral blood CD34+ cells were cultured in serum-free medium with interleukin (IL)-3, stem cell factor (SCF), and various combinations of thrombopoietin (TPO), erythropoietin (EPO), and/or granulocyte-colony stimulating factor (G-CSF). The effects of TPO, EPO, and G-CSF on Mks, erythrocytes, and granulocytes, respectively, were dependent on the O2 tension. Thrombopoietin-containing cultures under a gas phase of 20%) O2 tension produced 1.4- to 2.2-fold more Mks than those under 5% O2. The increase in Mk size with TPO was also much greater under 20% O2. Similarly, 2.1- to 2.4-fold more hemoglobin-containing cells were produced in EPO-containing cultures under 20% vs. 5% O2. In contrast, approximately twice as many CD15+ cells were produced in G-CSF-containing cultures under 5% vs. 20%) O2. The numbers of CFU-Mk in TPO-containing and CFU-GM in G-CSF-containing cultures were larger under 5% O2. Although the O2 tension had no effect on BFU-E production in EPO-containing cultures, BFU-E production under 5% O2 in cultures without EPO was equal to that in EPO-containing cultures. Our data also suggest that TPO, EPO, and G-CSF elicit stimulatory cross-lineage effects in the presence of IL-3 and SCF, and that these effects, too, are often dependent on O2 tension.