IDENTIFICATION AND BIOLOGICAL CHARACTERIZATION OF EXTRACELLULAR VESICLE PROTEINS FOR PROPAGATION AND SPREAD OF TAU

Accumulation of hyper-phosphorylated tau aggregates are pathological hallmark of Alzheimer's disease (AD). Extracellular vesicles (EVs) derived from AD derived microglia contribute to tau propagation as previously shown in a mouse model of human disease. The data sets, derived from the entorhinal cortex to the dentate gyrus, support a role for EVs in tau-associated pathobiology. A quantitative proteomics profile was made from EVs isolated from cortical grey matter of AD (n=20) and control (CTRL) (n=18) brain tissues. Here, we have identified differentially expressed proteins between these brain tissue cohorts using predictive machine learning. Expressed EV proteins were validated by ELISA and by biological function then linked to AD progression. Differentially expressed EV proteins, recovered from AD and CTRL brain samples, were observed by quantitative proteomics employing machine learning. The candidate proteins were selected then validated from an independent cohort by a commercially available ELISA. We evaluated the effect of the identified proteins on microglial EV uptake. AD brain-derived EVs were labeled with PKH26 and preincubated with recombinant target proteins then tested with murine microglia for uptake measurements. Annexin V (ANXA5), neurosecretory protein VGF, and neuronal membrane glycoprotein M6-a (GPM6A) were differentially expressed in AD EVs. Each of the proteins were confirmed with 88% accuracy by machine learning analyses. There was significant difference in protein levels of ANXA5 (p = 4.72E-06, fold change = 2.10), VGF (p = 7.49E-07, fold change = 0.23), GPM6A (p = 5.00E-04, fold change = 1.56) between AD and CTRL brain-derived EVs. Increased protein level of ANXA5 in AD compared CTRL EVs (p = 4.80E-03) was confirmed by ELISA. Moreover, the ANXA5 and VGF recombinant proteins inhibited EV microglial uptake. ANXA5 and VGF distinguished AD from CTRL EVs. Preincubation of AD brain-derived EVs with ANXA5 or VGF inhibited EV microglial uptake. These data suggest an ANXA5 and VGF-dependent spread of pathogenic tau protein in the brain, and these molecules can be a potential therapeutic target for AD.