Mechanisms of methyltransferase-like 3-mediated microglial pyroptosis in sepsis-associated encephalopathy

Sepsis-associated encephalopathy (SAE) is a common and serious neurological complication of sepsis. This study aimed to investigate the mechanism of methyltransferase-like 3 (METTL3) in SAE-induced microglial pyroptosis and to identify new therapeutic targets for SAE treatment.

A SAE cell model was established using lipopolysaccharide (LPS)-treated BV-2 cells. The expression of interleukin-1β, interleukin-18, cleaved caspase-1, gasdermin D (GSDMD)-N, NOD-like receptor protein 3 (NLRP3), transforming growth factor beta receptor 3 (TGFBR3), and METTL3 was detected by. METTL3 was silenced in LPS-treated BV-2 cells to validate the role of METTL3 in microglial pyroptosis. Total N6-methyladenosine (m6A) content was measured. The binding of primary miRNA (pri-miR)-101-3p to DGCR8 and the m6A level of pri-miR-101-3p were analyzed by methylated RNA immunoprecipitation-qPCR. The expression of pri-miR-101-3p and miR-101-3p was measured by reverse transcription quantitative PCR. The downstream targets of miR-101-3p were predicted by databases, and the binding relationship between miR-101-3p and TGFBR3 was verified. Rescue experiments were performed to verify the role of METTL3/miR-101-3p/TGFBR3 axis in microglial pyroptosis.

LPS treatment decreased cell viability and promoted interleukin-1β, interleukin-18, METTL3, cleaved caspase-1, GSDMD-N, and NLRP3. Silencing METTL3 inhibited microglial pyroptosis. Mechanistically, METTL3 promoted the binding of pri-miR-101-3p to DGCR8 through m6A modification and increased mature miR-101-3p expression. miR-101-3p targeted TGFBR3 and inhibited TGFBR3 expression. miR-101-3p overexpression or TGFBR3 downregulation partially reversed the inhibitory effect of silencing METTL3 on LPS-induced microglial pyroptosis.

METTL3 is upregulated in SAE, enhances miR-101-3p expression through m6A modification, and inhibits TGFBR3 expression, finally leading to microglial pyroptosis in SAE.