Environmental enrichment modulates, but does not normalize, aging-associated immune transcriptional programs after...

Aging remodels hippocampal transcriptional architecture by upregulating neuroinflammatory gene programmes and suppressing neuronal and metabolic networks, increasing ischemic injury susceptibility. Environmental enrichment (EE), a non-pharmacological intervention known to promote brain plasticity, has not previously been assessed at a systems level against aging- and stroke-associated co-expression programmes. Weighted gene co-expression network analysis (WGCNA) was applied to hippocampal RNA-seq data (GSE302188), yielding 45 modules, 24 of which were significantly age-associated (FDR < 0.05). Ten Age-UP modules were enriched for neuroinflammatory and immune activation pathways, nine Age-DOWN modules showed heterogeneous profiles including ribosomal RNA processing, and nominal enrichment for synaptic organisation pathways. Gene set enrichment analysis against an independent stroke dataset (GSE137482) identified 29 stroke-engaged modules (FDR < 0.05-0.25), 18 of which were Age-Neutral, indicating largely distinct aging and stroke transcriptional signatures. Cell-type deconvolution using MuSiC confirmed that age-associated module trajectories reflect coordinated within-cell transcriptional reprogramming rather than shifts in hippocampal cellular composition. In young adult mice (GSE95740; 3 months), EE did not preferentially reverse the concordantly dysregulated aging-stroke modules. To evaluate EE effects on established aging programmes, module eigengenes from an independent aged dataset (PRJEB58981;17 months) were projected onto the aging reference trajectory. EE partially opposed age-suppressed co-expression programmes but did not suppress the neuroinflammatory (turquoise) or interferon (sienna3) modules most strongly implicated in the aging-stroke overlap. Collectively, EE engaged hippocampal co-expression architecture principally through plasticity- and glial-related mechanisms. These findings identify complement and interferon signalling as priority targets for adjunctive pharmacological strategies in aged stroke-vulnerable populations. Adequately powered studies comparing young and aged animals across EE, stroke, and combined EE-plus-stroke conditions are required to determine EE's potential to suppress the concordant aging-stroke immune substrate represents a fundamental biological boundary or a tractable target in age-matched experimental designs.