Abstract: Germinal matrix hemorrhage in preterm neonates often leads to white matter injury, contributing to long-term neurodevelopmental impairments. As resident brain immune cells, microglia play a complex role in injury response, including inflammation and repair. Although colony-stimulating factor 1 receptor inhibitors such as PLX5622 enable the selective depletion of microglia, their therapeutic potential in neonatal germinal matrix hemorrhage remains underexplored. Here, we used a collagenase-induced germinal matrix hemorrhage model in postnatal day 5 mice, and intraperitoneally administered PLX5622 72 hours post–germinal matrix hemorrhage to achieve targeted, temporary microglial depletion during the peak injury response. We then assessed the effects of this delayed intervention on oligodendrocyte lineage cell maturation, white matter integrity, and neurobehavioral outcomes. Additionally, RNA sequencing data from a germinal matrix hemorrhage rat model were analyzed using weighted gene co-expression network analysis to identify the critical phases for interventions. RNA sequencing data revealed a critical period in which key synaptic functions declined while immune responses intensified post–germinal matrix hemorrhage, thus pinpointing the critical response phases for potential interventions. Delayed PLX5622 treatment effectively depleted activated microglia, protecting against white matter injury and enhancing oligodendrocyte lineage cell maturation and myelination in subcortical white matter regions. Moreover, magnetic resonance imaging analysis revealed reduced brain lesion volumes in treated mice. Behaviorally, PLX5622-treated mice exhibited significant improvements in motor coordination and reduced hyperactivity compared with vehicle-treated germinal matrix hemorrhage model mice. These findings suggest that, when timed to avoid interference with initial oligodendrocyte lineage cell proliferation, targeted microglial depletion with PLX5622 significantly mitigates white matter damage and improves neurobehavioral outcomes in neonatal germinal matrix hemorrhage. The present study highlights the therapeutic potential of selectively modulating microglial reactivity to support neurodevelopment in preterm infants with brain injury.

Key words: colony-stimulating factor 1 receptor, germinal matrix hemorrhage, microglia, myelination, neonatal brain, oligodendrocyte lineage cell, PLX5622, white matter injury