Procoagulant Extracellular Vesicles Increase Neuronal Tau expression, Metabolism and Processing Through Tissue Factor and Protease Activated Receptor 2

Neuro-inflammation is implicated in the onset of neuropathologies and can be promoted by stroke, trauma, toxins or infections. Brain tissue is rich in Tissue factor (TF) which is also released within cerebrospinal fluid as extracellular vesicles (EV). TF is an inflammatory protein which is increased during chronic conditions, and initiates blood coagulation and promotes tissue repair. This study examined the influence of TF on the expression, phosphorylation, aggregation and degradation of Tau protein in differentiated human cells SH-SY5Y and HCN-2, and rat neuronal cells. Studies were performed using vesicles containing TF or recombinant TF supplemented with factor VIIa (fVIIa) and also in the presence of various reagents and antibodies. Treatment of the differentiated cells with TF or TF-EV, upregulated the expression of Tau mRNA and protein, and was enhanced on repeated treatment. Incubation of cells with TF-fVIIa increased Tau expression and resulted in significant phosphorylation at Thr181, and was less at Ser202. Inhibition of the protease activity of TF-fVIIa, or blocking PAR2 activation on cells using SAM11 antibody, reduced Tau phosphorylation at Thr181. Examination of the Tau protein at intervals post-treatment indicated that Thr181 phosphorylation was present in bands of approximately 50 and 30-35 kDa while phosphorylation of Ser202 was associated with a 43 kDa band. Exposure of the cells to TF alone was sufficient to induce PKC-dependent phosphorylation of Tau. Prolonged treatment of differentiated SH-SY5Y cells with TF, resulted in higher staining with Amytracker dye. Finally, controlled digestion of recombinant full-length Tau with TF-fVIIa resulted in a smaller fragment. In conclusion, our data presents potential mechanisms by which TF influences Tau metabolism in neurons, being both beneficial in terms of clearance and regeneration, and having detrimental outcomes including aggregation.