ALKBH5 Demethylates the m6A Modification of SOCS3 in Microglia/Macrophages and Alleviates Neuroinflammation after Brain Injury

Microglia/macrophage_induced neuroinflammation plays a crucial role in the progression of traumatic brain injury (TBI). However, the involvement of N6_methyladenosine (m6A) RNA modifications in this process remains elusive. Single_cell RNA sequencing (scRNA_seq) and m6A RNA immunoprecipitation sequencing (MeRIP_seq) across multiple time points post_injury revealed a strong correlation between m6A modifications and genes enriched in microglia/macrophage. Furthermore, the m6A demethylase ALKBH5 was identified as a key regulator of dynamic m6A patterns at the injury site. ALKBH5 suppression in microglia/macrophage exacerbated neuroinflammation in vitro and worsened neurological deficits in controlled cortical impact (CCI) models. MeRIP_qPCR and RNA pull_down assays revealed SOCS3 was a downstream target of ALKBH5_mediated m6A demethylation. This demethylation stabilized Socs3 mRNA and enhanced its protein expression, which in turn suppressed neuroinflammation via inhibiting the JAK2_STAT3 pathway. Conversely, SOCS3 depletion impaired functional recovery after injury. These findings unveiled a critical ALKBH5_m6A_SOCS3 regulatory axis that mitigated microglia/macrophage_driven neuroinflammation after TBI, underscoring its potential as a therapeutic intervention target for TBI progression. Cortical tissues from mouse brains in the sham group and at 1, 3, 5 and 7 days following traumatic brain injury were minced and enzymatically dissociated for m6A sequencing.