Ageing-associated inflammation ("inflammaging") is characterised by chronic microglial priming or overactivity that elevates neurodegeneration risk. Microglia also accumulate autofluorescent lipofuscin as they age-aggregates of oxidised lipids, misfolded proteins and metals that generate reactive oxygen species. Characterising age-related changes in brain autofluorescence could provide insight into brain ageing dynamics and could identify associations with potential biomarkers in accessible tissues such as blood. We hypothesised that the ageing-related increase in microglial autofluorescent material produces a spectral signature that makes ageing autofluorescence distinct from that in young brains. To test this, we examined autofluorescence profiles in the hippocampus and arcuate nucleus (ARC) of young (3 months) and aged (21 months) female C57BL/6 mice. We found that ageing significantly increased the volume of the autofluorescent material in microglia, including driving an abundance of extra-large aggregates. In the ARC, larger aggregates-present only in aged brains-exhibited stronger far-red emission than the autofluorescence seen in young brains. Fluorescence lifetime imaging analysis indicated specific ageing-related molecular changes in brain autofluorescence. Lipidomic analysis from the plasma of these mice indicated 22 lipids that were differentially concentrated with age and one (sphingomyelin 32:2) that was significantly associated with the ageing brain spectral profile after controlling for multiple comparisons and validating in a separate cohort. These findings demonstrate that autofluorescent material aggregates with age in microglia and that brain autofluorescence undergoes a spectral red-shift associated with ageing and lipid alterations. Brain autofluorescence may thus reflect brain ageing with potential translational relevance for non-invasive monitoring.
