Gas6 restores microglial efferocytosis and limits neuroinflammation in neonatal hypoxic-ischemic encephalopathy by activating MerTK and the PI3K–Rac1 pathway

AIM: Hypoxic-ischemic encephalopathy is a major cause of neonatal disability and mortality. Its core pathology involves extensive neuronal apoptosis and persistent inflammatory responses. Microglia play a crucial role in maintaining brain homeostasis and promoting injury repair by recognizing and clearing apoptotic neurons. However, the regulatory mechanisms underlying this process remain unclear. METHOD: This study employed a co-culture model of apoptotic neurons, phagocytic function assays, cytokine analysis, transcriptome sequencing, Gas6 gene knockout and rescue experiments, combined with a mouse model of hypoxic-ischemic brain injury, to elucidate the role of microglia in the phagocytic process and the regulatory function of Gas6. RESULT: Injured neurons induced an early phase of pro-inflammatory activation and enhanced phagocytic capacity in microglia, followed by a shift towards an anti-inflammatory function. Transcriptome analysis suggested that co-culture with injured neurons activated pathways such as PI3K-AKT and NF-κB in microglia, concomitant with a significant upregulation of Gas6. Furthermore, we found that Gas6 deficiency significantly reduced the phosphorylation level of TAM receptors, leading to impaired downstream PI3K/AKT activation and a marked decrease in Rac1-GTP, thereby suppressing cytoskeletal rearrangement and phagocytic function. In parallel, Gas6-deficient microglia exhibited a sustained pro-inflammatory response, with both their efferocytic capacity and ability to regulate inflammation being significantly compromised. In vivo experiments showed that Gas6-KO mice displayed more severe neurological deficits, increased neuronal apoptosis, and stronger inflammatory responses after HIE. Supplementation with exogenous Gas6 elevated TAM receptor phosphorylation and the PI3K/AKT-Rac1 signaling pathway, partially restoring the phagocytic capacity of microglia. CONCLUSION: This study demonstrates the important role of the Gas6-TAM-PI3K/AKT-Rac1 signaling axis in modulating microglial efferocytic function and inflammatory state transition. It provides a potential therapeutic strategy for improving HIE prognosis by targeting the regulation of microglial phagocytosis.