Cardioprotective GLP-1 metabolite prevents ischemic cardiac injury by inhibiting mitochondrial trifunctional protein-α
M. Ahsan Siraj, Dhanwantee Mundil, Sanja Beca, Abdul Momen, Eric A. Shikatani, Talat Afroze, Xuetao Sun, Ying Liu, Siavash Ghaffari, Warren Lee, Michael B. Wheeler, Gordon Keller, Peter Backx, Mansoor Husain
M. Ahsan Siraj, Dhanwantee Mundil, Sanja Beca, Abdul Momen, Eric A. Shikatani, Talat Afroze, Xuetao Sun, Ying Liu, Siavash Ghaffari, Warren Lee, Michael B. Wheeler, Gordon Keller, Peter Backx, Mansoor Husain
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Research Article Cardiology Metabolism

Cardioprotective GLP-1 metabolite prevents ischemic cardiac injury by inhibiting mitochondrial trifunctional protein-α

Abstract

Mechanisms mediating the cardioprotective actions of glucagon-like peptide 1 (GLP-1) were unknown. Here, we show in both ex vivo and in vivo models of ischemic injury that treatment with GLP-1(28–36), a neutral endopeptidase–generated (NEP-generated) metabolite of GLP-1, was as cardioprotective as GLP-1 and was abolished by scrambling its amino acid sequence. GLP-1(28–36) enters human coronary artery endothelial cells (caECs) through macropinocytosis and acts directly on mouse and human coronary artery smooth muscle cells (caSMCs) and caECs, resulting in soluble adenylyl cyclase Adcy10–dependent (sAC-dependent) increases in cAMP, activation of protein kinase A, and cytoprotection from oxidative injury. GLP-1(28–36) modulates sAC by increasing intracellular ATP levels, with accompanying cAMP accumulation lost in sAC–/– cells. We identify mitochondrial trifunctional protein-α (MTPα) as a binding partner of GLP-1(28–36) and demonstrate that the ability of GLP-1(28–36) to shift substrate utilization from oxygen-consuming fatty acid metabolism toward oxygen-sparing glycolysis and glucose oxidation and to increase cAMP levels is dependent on MTPα. NEP inhibition with sacubitril blunted the ability of GLP-1 to increase cAMP levels in coronary vascular cells in vitro. GLP-1(28–36) is a small peptide that targets novel molecular (MTPα and sAC) and cellular (caSMC and caEC) mechanisms in myocardial ischemic injury.

Authors

M. Ahsan Siraj, Dhanwantee Mundil, Sanja Beca, Abdul Momen, Eric A. Shikatani, Talat Afroze, Xuetao Sun, Ying Liu, Siavash Ghaffari, Warren Lee, Michael B. Wheeler, Gordon Keller, Peter Backx, Mansoor Husain

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Figure 6

Pretreatment with GLP-1(28–36) increases glycolysis and glucose oxidation, and decreases FAO in coronary vascular cells.

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Pretreatment with GLP-1(28–36) increases glycolysis and glucose oxidatio...
(A) Seahorse XFe24 extracellular flux analyzer traces of a glycolysis stress test in caSMCs measured as the ECAR by sequential injection of 20 mM glucose, 1 μM oligomycin, and 100 mM 2-deoxy-d-glucose (2-DG). (B) Quantification of glycolysis, glycolytic capacity, and glycolytic reserves in caSMCs pretreated with 100 nM GLP-1(28–36) or scrambled control for 20 minutes. (C) Seahorse XFe24 traces of glucose oxidation, measured as the OCR by sequential injection of 20 mM glucose, 1 μM oligomycin, 1 μM FCCP [carbonyl cyanide-4-(trifluoromethoxy)phenylhydrazone], 1 μM rotenone, and 2 μM antimycin A into caSMCs cultured in normal (5.5 mmol/L) glucose media. (D) Quantification of glucose oxidation measured in caSMCs that were pretreated with 100 nM GLP-1(28–36) or scrambled control for 20 minutes. (E) Seahorse XFe24 traces of exogenous palmitic acid oxidation measured as the OCR following sequential injection into caSMCs of 200 μM palmitic acid conjugated to BSA or BSA control, 1 μM oligomycin, 1 μM FCCP, 1 μM rotenone, and 2 μM antimycin A. (F) Quantification of exogenous palmitic acid oxidation measured in caSMCs that were pretreated with 100 nM GLP-1(28–36) or scrambled control for 20 minutes. (G) Seahorse XFe24 traces from a glycolysis stress test in caECs, measured as described above. (H) Quantification of glycolysis, glycolytic capacity, and glycolytic reserves in caECs pretreated with 100 nM GLP-1(28–36) or scrambled control for 20 minutes. (I) Quantification of glucose oxidation in caECs pretreated with 100 nM GLP-1(28–36) or scrambled control for 20 minutes. n = 3/treatment in all Seahorse assays, each in triplicate. Data represent the mean ± SEM. *P < 0.01, **P < 0.001, and ***P < 0.0001, by 1-way ANOVA with Bonferroni’s post hoc test.