Oxygen tension has significant effects on human megakaryocyte maturation

Megakaryocytes (Mks) mature adjacent to bone marrow sinuses and release platelets within the sinusoidal space or in lung capillaries. Since the sites for platelet release have higher levels of oxygen tension (pO2) compared to the core of the bone marrow where stem and progenitor cells reside, we investigated whether pO2 influences Mk maturation. Mks were generated from CD34+ cells under 5% and 20% O2 with TPO, IL-3, and FL (n = 10). At day 15, CD41+ cell expansion in 20% and 5% O2 cultures was 85-fold and 31-fold respectively. 20% Oimage cultures also had higher levels of polyploid (≥8N) (8-fold higher) and proplatelet-displaying (5-fold higher) Mks. At day 21, 20% O2 cultures had a 5-fold higher number of apoptotic Mks. In contrast, the total number of CFU-Mks and the proportion of large-colony (≥20 Mks)-forming CFU-Mks were higher under 5% O2. Change from 5% to 20% O2 (n = 4) increased Mk differentiation compared to 5% O2. Change from 20% to 5% O2 (n = 3) reduced the level of Mk maturation and apoptosis compared to 20% O2. Cultures initiated with CD41+ cells showed that pO2 effects on Mks were direct (n = 3). RT-PCR indicated Mks under 20% O2 had a higher level of NF-E2 mRNA. These culture data may provide insights into in vivo Mk maturation. Decreased Mk differentiation at 5% O2 may explain observations that hypoxia causes thrombocytopenia in animals. Mk maturation and apoptosis followed similar kinetics under different pO2 environments supporting the hypothesis that these two processes are closely linked. The differential effects of pO2 on Mk progenitors versus mature Mks have implications beyond their physiological relevance. Using pO2 as a control parameter in ex vivo cultures one can (1) extend the level of Mk progenitor expansion (5% O2), or (2) generate a high level of non-apoptotic Mks for reinfusion into patients (5% to 20% O2).