AGE- AND SEX-DEPENDENT DYNAMICS OF H4K20 METHYLATION ACROSS BRAIN CELL TYPES IN THE APP<SUP>NL-G-F</SUP> MOUSE MODEL OF ALZHEIMER'S DISEASE

Epigenetic dysregulation contributes to Alzheimer's disease (AD) pathogenesis, yet the temporal dynamics of these changes and their sex specificity across brain cell types remain unclear. H4K20 methylation –associated with DNA damage repair, chromatin compaction, and transcriptional regulation– may modulate the expression of genes relevant to AD. We investigated H4K20 methylation states (un-, mono-, di-, tri-methylation and acetylation), their writers (Kmt5a/b/c), erasers (Lsd1, Phf8, Rad23A/B) in the APP^NL-G-F knock-in mouse model (NLGF). Using our EpiFlow multiparametric spectral flow cytometry platform, we simultaneously quantified these epigenetic markers at single-cell resolution across six brain cortico-hippocampal cell populations: excitatory and inhibitory neurons, astrocytes, OPCs, mature oligodendrocytes, and microglia. Male and female wild-type (WT) and NLGF mice were analysed at 2, 5, and 10 months of age, following disease progression. We identified alterations with striking sex- and age-dependent patterns. Females showed predominant early changes at 2 and 5 months, while males exhibited mainly late-stage changes at 10 months old. NLGF effect peaks at 5 months across cell types, showing the largest shifts. Microglia displayed the highest vulnerability in both sexes, representing the cell type with the most significant epigenetic changes occurring in AD. The early female response, involving coordinated upregulation of epigenetic enzymes, may represent an initial aberrant epigenetic mechanism that could favour the increased susceptibility to AD observed in women. These findings reveal sexually dimorphic temporal trajectories of epigenetic dysregulation in AD, with females showing early responses and males exhibiting late alterations, suggesting sex-specific therapeutic windows for epigenetic interventions.