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α-Ketoglutarate Inhibits Thrombosis and Inflammation by Prolyl Hydroxylase-2 Mediated Inactivation of Phospho-Akt
Nishith M Shrimali1, Sakshi Agarwal1, Simrandeep Kaur1
1Regional Centre for Biotechnology; National Capital Region Biotech Science Cluster, Faridabad, India.
Background:
Phospho-Akt1 (pAkt1) undergoes prolyl hydroxylation at Pro125 and Pro313 by the prolyl hydroxylase-2 (PHD2) in a reaction decarboxylating α-ketoglutarate (αKG). We investigated whether the αKG supplementation could inhibit Akt-mediated activation of platelets and monocytes, in vitro as well as in vivo, by augmenting PHD2 activity.
Methods:
We treated platelets or monocytes isolated from healthy individuals with αKG in presence of agonists in vitro and assessed the signalling molecules including pAkt1. We supplemented mice with dietary αKG and estimated the functional responses of platelets and monocytes ex vivo. Further, we investigated the impact of dietary αKG on inflammation and thrombosis in lungs of mice either treated with thrombosis-inducing agent carrageenan or infected with SARS-CoV-2.
Findings:
Octyl αKG supplementation to platelets promoted PHD2 activity through elevated intracellular αKG to succinate ratio, and reduced aggregation in vitro by suppressing pAkt1(Thr308). Augmented PHD2 activity was confirmed by increased hydroxylated-proline and enhanced binding of PHD2 to pAkt in αKG-treated platelets. Contrastingly, inhibitors of PHD2 significantly increased pAkt1 in platelets. Octyl-αKG followed similar mechanism in monocytes to inhibit cytokine secretion in vitro. Our data also describe a suppressed pAkt1 and reduced activation of platelets and leukocytes ex vivo from mice supplemented with dietary αKG, unaccompanied by alteration in their number. Dietary αKG significantly reduced clot formation and leukocyte accumulation in various organs including lungs of mice treated with thrombosis-inducing agent carrageenan. Importantly, in SARS-CoV-2 infected hamsters, we observed a significant rescue effect of dietary αKG on inflamed lungs with significantly reduced leukocyte accumulation, clot formation and viral load alongside down-modulation of pAkt in the lung of the infected animals.
Interpretation:
Our study suggests that dietary αKG supplementation prevents Akt-driven maladies such as thrombosis and inflammation and rescues pathology of COVID19-infected lungs.
Funding:
Study was funded by the Department of Biotechnology (DBT), Govt. of India (grants: BT/PR22881 and BT/PR22985); and the Science and Engineering Research Board, Govt. of India (CRG/000092).
Insights
Alpha-ketoglutarate (αKG) supplementation enhances prolyl hydroxylase-2 (PHD2) activity, inhibiting Akt-mediated platelet and monocyte activation. Dietary αKG reduces inflammation and thrombosis, offering a potential therapeutic strategy for conditions like COVID-19.
Area of Science:
- Biochemistry
- Molecular Biology
- Immunology
Background:
- Phospho-Akt1 (pAkt1) is hydroxylated by prolyl hydroxylase-2 (PHD2), a reaction dependent on alpha-ketoglutarate (αKG).
- Akt signaling pathways are implicated in platelet and monocyte activation.
Purpose of the Study:
- To investigate if αKG supplementation can inhibit Akt-mediated activation of platelets and monocytes by augmenting PHD2 activity.
- To evaluate the therapeutic potential of αKG in inflammatory and thrombotic conditions.
Main Methods:
- In vitro studies involved treating isolated platelets and monocytes with αKG and agonists.
- In vivo studies included dietary αKG supplementation in mice, followed by ex vivo analysis of immune cell function.
- Inflammation and thrombosis models (carrageenan, SARS-CoV-2 infection) were used to assess αKG's effects in mice and hamsters.
Main Results:
- αKG supplementation increased PHD2 activity, suppressed pAkt1, and reduced platelet aggregation in vitro.
- Dietary αKG reduced ex vivo platelet and leukocyte activation, clot formation, and leukocyte accumulation in mice.
- In SARS-CoV-2 infected hamsters, αKG treatment reduced lung inflammation, thrombosis, and viral load, with decreased pAkt levels.
Conclusions:
- Dietary αKG supplementation effectively inhibits Akt-driven thrombosis and inflammation.
- αKG demonstrates a rescue effect in COVID-19-induced lung pathology.
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