Spatially Resolved Free Fatty Acid Profiling Reveals Region- and Age-Dependent Remodeling of Alzheimer's Disease...

Free fatty acids (FFAs) are bioactive mediators of inflammation, energy metabolism, and membrane remodeling, yet their spatial organization within the Alzheimer's disease (AD) brain and at individual amyloid-β (Aβ) plaques has remained inaccessible. We developed a novel, chemically tailored MALDI workflow that enables simultaneous, spatially resolved detection of nearly 30 FFAs alongside over 100 complex lipid species within the same tissue section. Applying this approach to a transgenic AD mouse model across brain regions and disease stages, and combining it with single-plaque microenvironment analysis (SPMA) that treats each plaque as an individual analytical object, we uncover two previously inaccessible dimensions of plaque-associated lipid biology. FFA distributions form highly structured spatial compartments reflecting regional cytoarchitecture, with distinct enrichment of saturated, monounsaturated, and polyunsaturated species across cortical layers. Within Aβ plaques, nearly 75% of detected FFAs are significantly remodeled, with reciprocal enrichment of short saturated and highly unsaturated species alongside depletion of long-chain monounsaturated FFAs. This pattern is consistent with concurrent disruption of ELOVL-mediated elongation and FADS-mediated desaturation, including opposing enrichment of pro-inflammatory arachidonic acid and pro-resolving docosahexaenoic acid. Machine learning of single-plaque profiles reveals that FFA composition alone classifies plaque age with high accuracy, demonstrating that lipid remodeling continues after Aβ peptide composition has stabilized. Together, these findings establish spatial FFA profiling as a new analytical dimension in neurodegeneration research, revealing that Aβ plaques are dynamic lipid-metabolic microenvironments that continue to remodel long after Aβ deposition has stabilized.