Bcl-2–associated athanogene 3 protects the heart from ischemia/reperfusion injury

Introduction Bcl-2–associated athanogene 3 (BAG3) is a 575–amino acid protein that is highly conserved in nature (1). First identified by its ability to bind Bcl-2, BAG3 is expressed most abundantly in the heart and skeletal muscle and in many cancers. BAG3 serves as a cochaperone with members of the heat shock family of proteins to regulate protein quality control, interacts with Bcl-2 to inhibit apoptosis, and maintains the structural integrity of the sarcomere by linking filamen with the Z-disc through binding with the actin capping protein β-1 (CapZβ1) (2). Recent studies have demonstrated that BAG3 plays a pivotal role in maintaining cardiac homeostasis: (a) homozygous deletion of BAG3 in mice led to severe LV dysfunction, myofibril disorganization, and death by 4 weeks of age (3); (b) a single allele mutation in children was associated with progressive limb and axial mus-cle weakness, severe respiratory insufficiency, and cardiomyopathy (4, 5), whereas deletions in BAG3 have been associated with heart failure with reduced ejection fraction (HFrEF) independent of peripheral muscle weakness or neurologic complications (6, 7); (c) BAG3 levels were reduced in mice and pigs with HFrEF secondary to a LAD occlusion and in patients with end-stage HFrEF (6); and (d) knockdown of BAG3 in neonatal myocytes led to myofibrillar disarray when the cells were stretched (8). However, in adult myo-cytes, BAG3 localized at the sarcolemma and t-tubules, where it modulates myocyte contraction and action potential duration through specific interaction with the β1-adrenergic receptor and L-type Ca 2+ channel (9). Furthermore, overexpression of BAG3 using an adeno-associated virus serotype 9 (rAAV9-BAG3) restored left ventricular (LV) function in mice with diminished LV function secondary to a myocardial infarction (10). Bcl-2–associated athanogene 3 (BAG3) is an evolutionarily conserved protein expressed at high levels in the heart and the vasculature and in many cancers. While altered BAG3 expression has been associated with cardiac dysfunction, its role in ischemia/reperfusion (I/R) is unknown. To test the hypothesis that BAG3 protects the heart from reperfusion injury, in vivo cardiac function was measured in hearts infected with either recombinant adeno-associated virus serotype 9–expressing (rAAV9-expressing) BAG3 or GFP and subjected to I/R. To elucidate molecular mechanisms by which BAG3 protects against I/R injury, neonatal mouse ventricular cardiomyocytes (NMVCs) in which BAG3 levels were modified by adenovirus expressing (Ad-expressing) BAG3 or siBAG3 were exposed to hypoxia/reoxygenation (H/R). H/R significantly reduced NMVC BAG3 levels, which were associated with enhanced expression of apoptosis markers, decreased expression of autophagy markers, and reduced autophagy flux. The deleterious effects of H/R on apoptosis and autophagy were recapitulated by knockdown of BAG3 with Ad-siBAG3 and were rescued by Ad-BAG3. In vivo, treatment of mice with rAAV9-BAG3 prior to I/R significantly decreased infarct size and improved left ventricular function when compared with mice receiving rAAV9-GFP and improved markers of autophagy and apoptosis. These findings suggest that BAG3 may provide a therapeutic target in patients undergoing reperfusion after myocardial infarction.