Effect of culture PO2 on macrophage (RAW 264.7) nitric oxide production

Macrophages are commonly cultured at a Po-2 of 149 Torr, but tissue macrophages in vivo live in an environment of much lower oxygen tension. Despite the many potential mechanisms for changes in oxygen tension to influence nitric oxide (NO) synthesis, there have been few reports investigating the effect of Po-2 on macrophage NO production. With the use of a culture chamber designed to rigorously control oxygen tension, we investigated the effects of culture Po-2 on macrophage NO production, inducible nitric oxide synthase (iNOS) activity, iNOS protein, and tumor necrosis factor production. NO production and iNOS activity were linearly related in the range of 39.4 to 677 Torr, but not in the range of 1.03 to 39.4 Torr. Therefore, results obtained in vitro for the high oxygen tensions commonly used in cell culture were quantitatively and qualitatively different from results obtained in cells cultured at the lower oxygen tensions that more accurately reflect the in vivo environment. The influence of oxygen tension on NO production has implications for cell culture methodology and for the relationship between microcirculatory dysfunction and inflammatory responses in rodent models of sepsis.