Abstract 359: Autosis is Triggered by Ischemia/Reperfusion in the Heart

Abstract

Although autophagy generally acts as a protective function in cardiomyocytes (CMs), uncontrolled or excessive activation of autophagy can be detrimental to CMs under some conditions. However, whether autophagy mediates cell death for CMs remains largely unknown. Recently, it has been shown in HeLa cells that excessive activation of autophagy induces cell death with characteristic morphological and biochemical features, and this form of cell death is termed autosis. Here we investigated whether death of CMs is induced by autosis in the presence of TAT-Beclin1, an autophagy-inducing peptide, and ischemia/reperfusion. TAT-Beclin1 treatment dose-dependently increased the level of LC3-II and decreased that of p62. TAT-Beclin1-treated CMs showed increases in trypan-blue-positive cells suggesting that autophagy-related cell death is induced by TAT-Beclin1. TAT-Beclin1-treated CMs showed the typical morphological features of autosis, including increased autophagic vacuoles and empty vacuoles, perinuclear spaces with cytoplasmic materials, and ballooning under electron microscopic observation. TAT-Beclin1-induced autosis in CMs was not inhibited by inhibitors of apoptosis, necrosis, or lysosomal degradation. Although autosis is known to be inhibited by cardiac glycosides, since cardiomyocyte Na + -K + -ATPase in rodents is relatively resistant to cardiac glycosides, we generated adenovirus harboring shRNA-α1 subunit of Na + -K + -ATPase. We found that downregulation of the α1 subunit of Na + -K + -ATPase inhibits TAT-Beclin1-induced autosis in CMs. We also found that ischemia/reperfusion induces autosis in the mouse heart, as evidenced by the presence of characteristic morphological features of autosis, including the presence of autophagosomes, empty vacuoles, and perinuclear space. Lastly, in order to elucidate the molecular mechanism mediating autosis in the heart, we conducted unbiased screenings of 150 kinase inhibitors and identified several classes of kinase inhibitors suppressing TAT-Beclin1-induced autosis in CMs. These results indicate that autosis occurs in the heart during ischemia/reperfusion injury. Our findings may provide novel therapeutic targets for treatment of cardiac cell death during ischemia/reperfusion.