Abstract Body: Background: Activated platelets exhibit metabolic plasticity, characterized by a shift from combined reliance on mitochondrial oxidative phosphorylation (OXPHOS) and aerobic glycolysis (the Warburg effect) towards a predominant dependence on aerobic glycolysis. This metabolic reprogramming helps meet the heightened ATP demands of energy-intensive activation processes, including aggregation, integrin αIIbβ3 activation, and granule secretion. Aerobic glycolysis, driven by lactate dehydrogenase (LDH)-mediated conversion of pyruvate to lactate under oxygen-rich conditions, enables rapid ATP production. This context-dependent metabolic reprogramming in activated platelets provides a unique avenue that can be targeted to attenuate platelet activation and thereby reduce susceptibility to thrombosis.
Aim: Determine whether pharmacological targeting of LDH using oxamate inhibits aerobic glycolysis in platelets, thereby reducing platelet activation.
Methods: Platelet aggregation and dense granule secretion in platelet-rich plasma (PRP) were assessed by aggregometry, while integrin αIIbβ3 activation and α-granule secretion were analyzed by flow cytometry. Outside-in signaling was examined by clot retraction. Glycolytic proton efflux rate (glycoPER) and ATP production were measured in washed platelets using Seahorse XFe.
Results: Oxamate treatment significantly reduced platelet aggregation in response to GPVI and GPCR agonists in both human and murine PRP. Agonist-induced integrin αIIbβ3 activation and secretion (from alpha and dense granules) was reduced in oxamate-treated platelets. Clot retraction was inhibited by oxamate, suggesting impaired outside-in signaling. Additionally, oxamate significantly reduced collagen-stimulated total tyrosine phosphorylation, suggesting reduced GPVI signaling. The antiplatelet effects of oxamate were mechanistically associated with reduced glycoPER and glycolytic ATP production, suggesting reduced aerobic glycolysis.
Conclusions: LDH inhibition by oxamate impaired platelet function by reducing aerobic glycolysis and glycolytic ATP production. As a translational potential, we are currently evaluating the effects of oxamate on ex vivo thrombosis in human whole blood using a microfluidic flow assay, as well as its in vivo effects on thrombosis and hemostasis using a FeCl₃-induced carotid artery thrombosis and tail clip-bleeding assay respectively.