Alpha-2-macroglobulin prevents platelet aggregation induced by Brain-Derived Neurotrophic Factor.

The brain-derived neurotrophic factor (BDNF) has been recently shown to have activating effects in isolated platelets. However, BDNF circulates in plasma and a mechanism to preclude constant activation of platelets appears necessary. Hence, we investigated the mechanism regulating BDNF bioavailability in blood. Protein-protein interactions were predicted by molecular docking and validated through immunoprecipitation. Platelet aggregation was assessed using light transmission aggregometry with washed platelets in response to classical agonists or BDNF, in the absence or presence of alpha-2-macroglobulin (αM), and in platelet-rich plasma. BDNF signaling was assessed with phospho-blots. As little as 25% autologous plasma was sufficient to completely abolish platelet aggregation in response to BDNF. Docking predicted two forms of BDNF binding to native or activated αM, parallel and perpendicular arrangements, and the model suggested that the BDNF-αM complex cannot bind to the high-affinity BDNF receptor, tropomyosin receptor kinase B (TrkB). Experimentally, native and activated αM formed stable complexes with BDNF preventing BDNF-induced TrkB activation and signal transduction. Both native and activated αM inhibited BDNF induced-platelet aggregation in a concentration-dependent manner with comparable half-maximal inhibitory concentrations (IC≈ 125-150 nM). Our study implicates αM as a physiological regulator of BDNF bioavailability, and as inhibitor of BDNF-induced platelet activation in blood.