Activation of AMPK via inhibition of PAR2 limits myocardial ischemia-reperfusion injury

Background/Introduction Protease activated receptor 2 (PAR2) is a G protein-coupled receptor reported to regulate vascular contractility and heart failure. However, its role in mediating acute myocardial infarction is currently unclear. AMP-activated protein kinase (AMPK) is an enzyme that maintains cellular energy balance, and its activation during myocardial infarction plays a key role in preventing cardiomyocyte death and limiting post-ischemic cardiac damage. Purpose This study is the first to explore the relationship between PAR2 and AMPK in cardiomyocytes during ischemia, hoping to open up new therapeutic strategies for the treatment of post-ischemic cardiac injury. Methods In this study, we used PAR2 genetic knockout mice, Langendorff heart perfusion system and isolated primary adult cardiomyocytes, combined with molecular and cellular techniques, to investigate a novel signaling pathway by which PKA/CaMKKβ in regulating AMPK activity during hypoxia and ischemia. Results PAR2 is highly expressed in the heart. PAR2 deficiency significantly improved cardiac functional recovery and limited infarct size following acute global ischemia, which were associated with increased AMPK activation. Interestingly, knockdown of PAR2 in cardiomyocytes also improved AMPK and limited cell death following hypoxia, suggesting that cardiomyocyte PAR2 may be key to regulate cardiac ischemia-reperfusion injury. We also found that PAR2 activation stimulated PKA, leading to an inhibitory phosphorylation of the AMPK upstream kinase, CaMKKβ at Ser495. In addition, PAR2 activation promoted the binding of CaMKKβ to 14-3-3, a protein that maintains CaMKKβ inactivation. Thus, blocking PAR2 activation by GB83 upregulated AMPK by attenuating inhibitory phosphorylation of CaMKKβ and dissociating the binding between CaMKKβ and 14-3-3 during hypoxia. Conclusion Inhibition of cardiac PAR2 during acute ischemia can upregulate AMPK activity and cell survival through the PKA/CaMKKβ signaling pathway. Thus, the development of cardiac PAR2 as a pharmaceutical target will have important clinical significance for the future therapy of myocardial infarction.Graphic abstract