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Updated: May 21, 2025

Preparation of Washed Human Platelets for Quantitative Metabolic Flux Studies
Published on: January 10, 2025
BAD-Glucokinase Axis Regulates Platelet Activation and Thrombosis
Mengnan Yang1, Shuang Chen1, Qing Li1
1Jiangsu Institute of Hematology, Cyrus Tang Medical Institute, The First Affiliated Hospital and Collaborative Innovation Center of Hematology, Suzhou Medical College, Soochow University, Key Laboratory of Thrombosis and Hemostasis, Ministry of Health, National Clinical Research Center for Hematological Diseases, Suzhou, China.
Background:
BAD (Bcl2-associated death promoter), a member of the Bcl2 proapoptotic family, promotes cell apoptosis by sequestering the prosurvival proteins Bcl-XL and Bcl2 from the proapoptotic proteins BAK (Bcl2 homologous antagonist/killer) and BAX (Bcl2-associated X protein) in nucleated cells. BAD is also expressed in platelets, playing a role in regulating platelet lifespan, apoptosis, and clearance. However, whether BAD regulates platelet activation and arterial thrombosis remains unclear.
Methods:
The role of BAD in platelet activation and arterial thrombosis was investigated using BAD-deficient mice (Bad-/-), in vitro functional studies, and arterial thrombosis models. The regulatory effect of BAD on platelet energy metabolism was detected using a Seahorse Extracellular Flux Analyzer. The regulatory effect of BAD on glucokinase was investigated by coimmunoprecipitation and activity measurement. The glucokinase heterozygous knockout mice (Gck+/-) and activator were used to study its role in platelet activation.
Results:
BAD-deficient mice (Bad-/-) and wild-type mice transfused with Bad-/- platelets displayed prolonged tail bleeding and arterial occlusion times. Bad-/- platelets exhibited decreased aggregation in response to stimulations by proteinase-activated receptor 4-activating peptide, thrombin, and U46619. Furthermore, BAD ablation suppressed platelet integrin αIIbβ3 activation, granule secretion, and clot retraction induced by these agonists. Mechanistically, BAD interacted with glucokinase, and BAD deficiency resulted in decreased platelet glucokinase activity, mitochondrial oxidative phosphorylation, and mitochondrial ATP production. The partial loss of glucokinase (Gck+/-) phenocopied platelet function defects caused by BAD deficiency, and a glucokinase activator rescued the impaired mitochondrial ATP production and function of Bad-/- platelets. Additionally, the glucokinase activator enhanced human platelet activation.
Conclusions:
Our findings demonstrate the critical role of the BAD-glucokinase axis in platelet activation and thrombosis, suggesting a potential target for antithrombotic therapy.
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