Abstract
Neonatal hypoxic-ischemic encephalopathy (HIE) is a severe neurological condition associated with high rates of mortality or long-term disability. Despite its clinical significance, the detailed cellular mechanisms underlying HIE remain unclear. Single-cell RNA sequencing (scRNA-seq) has emerged as a powerful tool for investigating cellular heterogeneity across development, aging, and disease processes. However, no scRNA-seq studies have yet addressed neonatal HIE. In this study, we employed scRNA-seq to examine cellular heterogeneity during neonatal HIE. We analyzed a total of 87 580 high-quality brain cells to identify transcriptional changes associated with HIE. In the hyperacute phase, we observed astrocytes in response to tumor necrosis factors, involvement of microglia in phagocytosis, Stat3-mediated ischemic responses in oligodendrocyte precursor cells, and an increase in senescent lymphatic endothelial cells. In the acute phase, astrocytes were activated and involved in gliogenesis, while microglia proliferated. Neuroblasts were affected by metal ions, and oligodendrocytes decreased. In the subacute phase, astrocytes involved in inflammation and antigen presentation, while inflammatory microglia highly expressing MHC II were induced by the IL27 and type I interferon pathways and expanded. Additionally, peripheral immune cells played vital roles in HIE. Specifically, neutrophils infiltrated and expanded throughout all phases post-HIE. Spp1(high) macrophages, T cells, and plasmacytoid dendritic cells increased during the acute and subacute phases, and B cells expanded during the subacute phase. This study offers deep insights into the molecular alterations of key cell types following HIE, elucidating the pathological processes involved. These findings have significant implications for developing effective clinical strategies for managing HIE.