Hyperoxia Inhibits the Growth of Mouse Forebrain Oligodendrocyte Progenitors but Promotes Their Differentiation

Oligodendrocytes facilitate saltatory conduction and they support both neuronal and axonal survival; thus, when they are diseased or damaged, neurological problems ensue. Oligodendrocyte progenitors (OPCs) have been successfully isolated from rodents using magnetic beads, immunopanning and differential adhesion. Whereas rat OPCs are easy to propagate in vitro, expanding mouse OPCs in vitro has not been straightforward, which is problematic given that mice have been used to generate the majority of genetically engineered disease models. In this study, we developed and characterized a reproducible, straightforward method to prepare large numbers of nearly homogeneous cultures of mouse OPCs. Using the McCarthy and de Vellis mechanical separation method, we isolated OPCs from a mixture of glial cells and plated them onto fibronectin-coated tissue culture plates in a biochemically defined medium supplemented with fibroblast growth factor-2 (FGF-2) and medium conditioned by B104 neuroblastoma cells. However, when maintained in a standard tissue culture incubator, they proliferated very slowly and ∼ 20% died. By contrast, when they were maintained in an incubator with 2% oxygen they proliferated more rapidly and could be expanded for multiple passages with high viability. After three passages, greater than 99% of these OPCs expressed markers of early OPCs. In medium containing only FGF-2 they progressed to late-stage OPCs. When a medium supplemented with T3 was provided, a large subset of OPCs differentiated into O4+/MBP+ oligodendrocytes with sheet-like membranes. However, between 3 and 6 passages, less than 20% of the OPCs differentiated into MBP+ oligodendrocytes and this was associated with a loss of Sox-10. These studies reveal significant differences between mouse and rat OPCs and a role for oxygen tension in mouse OPC proliferation and differentiation.