RPL14 lysine lactylation orchestrates microglial inflammation and neuronal injury following ischemic stroke

Ischemic stroke causes extensive neuroinflammation and neuronal injury, but the molecular mechanisms linking metabolic stress to microglial dysfunction remain incompletely understood. Lysine lactylation, an analysis of post-translational modification connecting metabolic reprogramming to transcriptional regulation, has an unexplored role in post-stroke microglial dysfunction. Using time-resolved transcriptomics and single-cell RNA sequencing of brains from a middle cerebral artery occlusion (MCAO)-induced mouse model, we identified ribosomal protein L14 (RPL14) as a lactylation-associated driver of ischemic injury. Its causal role was validated via adeno-associated virus-short hairpin RNA (AAV-shRNA)-mediated silencing, site-specific mutagenesis, and p300/CBP lactyltransferase inhibition (C646) in vivo and in vitro . RPL14, a key metabolic-immune hub expressed mainly in microglia and monocytes, coordinates pro-inflammatory transforming growth factor β (TGFβ) and C-C motif chemokine ligand (CCL) signaling. Ischemic stress-induced lactate promotes RPL14 lactylation at K36/K39, a modification that acts as a molecular switch driving pro-inflammatory cytokine production and microglial apoptosis. Silencing RPL14 or blocking its lactylation with C646 significantly reduced cerebral infarction, attenuated neurological deficits, and suppressed neuronal apoptosis; mutating K36/K39 abolished lactate-induced inflammation. Our findings indicate that RPL14 lysine lactylation drives post-ischemic neuroinflammation and neuronal injury, and targeting this process may be a promising therapeutic strategy for stroke.