N-arachidonoylphenolamine alleviates ischaemia/reperfusion-induced cardiomyocyte necroptosis by restoring proteasomal activity.

Necroptosis and apoptosis contribute to the pathogenesis of myocardial ischaemia/reperfusion (I/R) injury and subsequent heart failure. N-arachidonoylphenolamine (AM404) is a paracetamol lipid metabolite that has pleiotropic activity to modulate the endocannabinoid system. However, the protective role of AM404 in modulating I/R-mediated myocardial damage and the underlying mechanism remain largely unknown. A murine I/R model was generated by occlusion of the left anterior descending artery. AM404 (20 mg/kg) was injected intraperitoneally into mice at 2 and 24 h before the I/R operation. Our data revealed that AM404 administration to mice greatly ameliorated I/R-triggered impairment of myocardial performance and reduced infarct area, myocyte apoptosis, oxidative stress and inflammatory response accompanied by the reduction of receptor interacting protein kinase (RIPK)1/3- mixed lineage kinase domain-like (MLKL)-mediated necroptosis and upregulation of the immunosubunits (ß2i and ß5i). In contrast, administration of epoxomicin (a proteasome inhibitor) dramatically abolished AM404-dependent protection against myocardial I/R damage. Mechanistically, AM404 treatment increases ß5i expression, which interacts with Pellino-1 (Peli1), an E3 ligase, to form a complex with RIPK1/3, thereby promoting their degradation, which leads to inhibition of cardiomyocyte necroptosis in the I/R heart. In conclusion, these findings demonstrate that AM404 could prevent cardiac I/R damage and may be a promising drug for the treatment of ischaemic heart disease.

The mechanosensitive Piezo1 channel exacerbates myocardial ischaemia/reperfusion injury by activating caspase-8-mediated PANoptosis.

PANoptosis is a newly discovered type of cell death characterized by pyroptosis, apoptosis and/or necroptosis and has been implicated in the inflammatory response. Piezo1 is a mechanosensitive ion channel that plays important roles in physiological development and various diseases. However, whether cardiomyocytes undergo PANoptosis during myocardial ischaemia/reperfusion (I/R) injury and the role of Piezo1 in this process remain largely unexplored. In this study, our results revealed that the expression levels of the main components of the PANoptosome, including caspase-8, caspase-3, NLRP3, caspase-1, GSDMD, RIPK1, RIPK3 and MLKL, were significantly upregulated in I/R heart tissues over time, indicating the occurrence of PANoptosis in I/R hearts. Accordingly, Piezo1 expression was significantly upregulated in I/R-injured hearts and hypoxia/reoxygenation (H/R)-treated cardiomyocytes. In contrast, pharmacological inhibition of Piezo1 by the inhibitor GsMTx4 in mice markedly attenuated the I/R-mediated decline in cardiac contractile function and increases in infarct size, apoptosis, oxidative stress and inflammation accompanied by the inhibition of PANoptosis-related mediators in I/R hearts. Consistently, the effects of Piezo1 on calcium influx and PANoptosis were further verified by GsMTx4 and Piezo1 activator Yoda1 in H/R-treated cardiomyocytes in vitro. Moreover, caspase-8 rather than calcium influx was required for H/R-induced PANoptosis in vitro. Mechanistically, Piezo1 interacts with caspase-8, a key initial activator of the PANoptosome complex, which subsequently activates cardiomyocyte PANoptosis, leading to cardiac dysfunction. In summary, these data suggest that Piezo1 is a new cardiac mechanosensor that promotes cardiac I/R injury possibly through the caspase-8-mediated activation of cardiomyocyte PANoptosis and highlight that Piezo1 may represent a new target for treating ischaemic heart disease.