Histone lactylation is associated with METTL3-dependent LCN2 m6A modification and astrocyte activation after intracerebral hemorrhage

Background

Intracerebral hemorrhage (ICH) is a major cause of secondary brain injury (SBI), which results in severe neurological deficits and poor clinical outcomes. Elevated serum lactate levels have been associated with unfavorable outcome in ICH patients. However, the role of lactate in ICH-induced SBI remain poorly understood.

Method

An autologous blood injection mouse model of ICH and lactate-treated C8D1A cells were employed as the in vivo and in vitro models, respectively. The establishment of ICH model was validated by behavior tests, and brain injury was assessed by H&E and Nissel staining. qRT-PCR, Western blot and IHC analysis were used to detect the expression of key molecules. Immunofluorescent (IF) staining was employed to evaluate astrocyte activation. Pro-inflammatory cytokine release was monitored by ELISA assay. The interaction between H3K18la and METTL3 was assessed by ChIP assay, and the association between METTL3 and LCN2 mRNA was assessed by RNA immunoprecipitation (RIP) assay.

Results

The levels of lactate, METTL3 and LCN2 are elevated in ICH model in mice. The inhibition of lactate decreased METTL3 expression and alleviated ICH-induced SBI. Mechanistically, histone H3K18 lactylation was associated with the upregulated levels of METTL3 and m⁶A in mouse brains. METTL3 regulated the m⁶A modification of LCN2 and upregulated its expression. In ICH mice, silencing of LCN2 inhibited A1 astrocyte activation. Histone lactylation-modulated LCN2 m⁶A modification is involved in astrocyte activation and the regulation of SBI in ICH mice.

Conclusion

These results suggested a mechanism whereby histone lactylation is implicated in the activation of A1 astrocytes through METTL3-mediated LCN2 m⁶A modification.