Oxygen tension regulates the in vitro maturation of GM-CSF expanded murine bone marrow dendritic cells by modulating class II MHC expression

Conventional culture conditions for GM-CSF expanded murine bone marrow derived dendritic cells (BMDCs) uses ambient (hyperoxic) oxygen pressure (20% v/v, 152 Torr) and medium supplemented with the thiol 2-mercaptoethanol (2-Me). Given the redox activities of O₂ and 2-Me, the effects of 2%, 5%, 10%, and 20% v/v O₂ atmospheres and omitting 2-Me from the medium were tested upon the generation of GM-CSF expanded BMDCs. DC yield, phenotype and function were compared to BMDCs grown using conventional conditions. All cultures yielded DC subsets with CD11c⁺ MHC II^NEG, CD11c⁺ MHC II^INT, CD11c⁺ MHC II^HI expression phenotypes, classed as precursor, immature, and mature DCs (IDC, MDC). Low O₂ tensions generated significantly fewer precursor DCs, and more IDCs and MDCs. Cytometer sorted precursor DCs expressed surface class II MHC after transfer to low, but not high O₂ atmospheres. Expression of myeloid markers was similar between BMDC cultures generated in 5% O ₂ or conventional conditions, and MDCs from low O₂ cultures had the morphology typical of mature myeloid DCs. IDCs and MDCs from low O₂ and conventional culture conditions were similarly potent allostimulatory APCs. The O₂ tension (but not 2-Me addition) in vitro significantly influences overall DC subset frequencies and yield, and governs DC maturation by regulating the surface class II MHC expression of GM-CSF expanded BMDC cultures.