In vitro model of rat perinatal hypoxia-ischemia. How temporal oxygen-glucose deprivation affects maturation of oligodendrocyte progenitor cells
INTRODUCTION: The frequent reason of brain damage in newborns is perinatal hypoxia-ischemia. It may cause the central nervous system hypomyelination, which often results in the long-term neurobehavioral disabilities. Oligodendrocytes, responsible for myelination, differentiate from oligodendrocyte progenitor cells (OPCs) in developing brain. AIM(S): In our study we investigated how oligodendrocyte progenitor cells respond to oxygen-glucose deprivation (OGD) in the in vitro model of neonatal asphyxia. METHOD(S): OPCs were obtained from rat primary glial cultures. 24 h after seeding, the cells were exposed to short OGD procedure. Its effect on the cell proliferation was measured by immunolabeling the dividing cells for Ki67 marker and BrdU incorporation. Cell viability was evaluated with AlamarBlue testing. Impact of OGD on OPC differentiation was assessed by making microscopic examination of immunocytochemically labeled cells with antibodies against NG2 for OPCs, GalC for immature oligodendrocytes, MBP for myelinating cells. Expression of characteristic myelin proteins was additionally verified by ELISA analysis. RESULTS: The obtained data revealed that OGD stimulated proliferation of oligodendrocyte progenitor cells, which was shown in 2% higher number of Ki67‑positive cells 1 day after OGD and 17% increase in the amount of BrdU- positive cells versus control 3 days after injury. Results of AlamarBlue assay also indicated the twofold higher viability of OGD‑affected cells versus control cells 6 days after injury. Microscopic analysis, confirmed by quantitative ELISA measurement, indicated a significant inhibition of oligodendrocyte maturation after injury. CONCLUSIONS: Hypoxic‑ischemic insult, reflected in vi‑ tro by OGD procedure, alters OPCs differentiation process causing the CNS hypomyelination. In our in vitro studies we showed that this may be associated with the enhanced proliferation of oligodendrocyte progenitors as a compensative mechanism for the injury and the disrupted maturation process. FINANCIAL SUPPORT: Supported by NCN grant 2014/15/B/NZ4/01875.