CaMKII/calpain interaction mediates ischemia/reperfusion injury in isolated rat hearts

Hong-Ting Lu,Jia-Kun Tang,Feng Gao,Jun Ren,Ren-Qian Feng,Jing-Jun Zhou

doi:10.1038/s41419-020-2605-y

Hong-Ting Lu, Jia-Kun Tang + Show 4 more

Open Access

https://doi.org/10.1038/s41419-020-2605-y

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Abstract

Previous studies indicated that Ca2+/calmodulin-dependent kinase II (CaMKII), a kinase involved in the modulation of ryanodine receptor activity, activates Ca2+-regulated protease μ-calpain to promote myocardial ischemia/reperfusion injury. This study was performed to explore the underlying mechanisms in CaMKII-induced calpain activation to better understand heart injury. To examine the Ca2+ paradox and ischemia/reperfusion injury, isolated rat hearts were subjected to a Ca2+-free solution for 3 min, or left coronary artery occlusion for 40 min, prior to restoration of normal perfusion. Blockade of trans-sarcoplasmic reticulum Ca2+ flux using ryanodine and thapsigargin failed to prevent Ca2+ paradox-induced heart injury. In contrast, the Ca2+ paradox increased CaMKII auto-phosphorylation at Thr287, while the CaMKII inhibitor KN-62 and the Na+/Ca2+ exchanger inhibitor KB-R7943 alleviated heart injury and calpain activity. Intriguingly, the binding of μ-calpain large subunit calpain-1 (CAPN1) to phospho-CaMKII was blunted by both inhibitors. Thus, a Ca2+ leak via the ryanodine receptor is not an essential element in CaMKII-elicited calpain activation. In hearts receiving vector injection, ischemia/reperfusion caused elevated calpain activity and α-fodrin degradation, along with membrane integrity damage, similar to the effects noted in control hearts. Importantly, all these alterations were diminished with delivery of adeno-associated virus expressing mutant CaMKIIδC T287A. Ischemia/reperfusion increased CaMKII auto-phosphorylation and binding of CAPN1 to phospho-CaMKII, and facilitated the translocation of phospho-CaMKII and CAPN1 to the plasma membrane, all of which were reversed by injecting CaMKII mutant. Furthermore, the relocation capacity and the interaction of CaMKII with CAPN1 appeared to be dependent upon CaMKII autophosphorylation, as its mutant delivery increased the level of CaMKII, but did not increase membrane content of CaMKII and CAPN1, or their interactions. Together, CaMKII/calpain interaction represents a new avenue for mediating myocardial ischemia/reperfusion injury, and CaMKII likely serves as both a kinase and a carrier, thereby promoting calpain membrane translocation and activation.

Highlights

Myocardial ischemia/reperfusion injury remains an unresolved medical problem and imposes a heavy healthcare burden[1]
The calmodulin-dependent protein kinase II (CaMKII)/ryanodine receptor cascade is considered the final instigator of intracellular Ca2+ overload-induced injury[8,11,12]
We previously demonstrated that CaMKII activates calpain, another culprit factor involved in ischemia/reperfusion injury[10]

Summary

Introduction

Myocardial ischemia/reperfusion injury remains an unresolved medical problem and imposes a heavy healthcare burden[1]. Ischemia/reperfusion injury induces cell death, arrhythmia, and cardiac dysfunction[1]. Upon external Ca2+ stimulation, ryanodine receptors release Ca2+ from the SR to provoke systolic cytosolic Ca2+ peaks and maintain cardiomyocyte contractility[6]. Aberrant intracellular Ca2+ may leak through the ryanodine receptor to provoke cell death in pathological conditions such as myocardial ischemia/ reperfusion injury[6,7,8]. Ryanodine receptor activity is tightly regulated by Ca2+/calmodulin-dependent protein kinase II (CaMKII), a serine/threonine kinase containing catalytic, autoinhibitory, and Ca2+/calmodulin binding domains. Once Ca2+/calmodulin bound, CaMKII undergoes autophosphorylation at the Thr[287] residue in the autoinhibitory domain and remains persistently activated even after dissociation from Ca2+/calmodulin.

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