EphrinA1-Fc Attenuates Ventricular Remodeling and Dysfunction in Chronically Nonreperfused WT but not EphA2-R-M mice.

BACKGROUND: EphrinA1-Fc abolishes acute I/R injury and attenuates nonreperfused cardiac injury 4 days after permanent occlusion in mice. The goal of this study was to assess the capacity of a single intramyocardial administration of ephrinA1-Fc at the time of coronary artery ligation, to determine the degree to which early salvage effects translate to reduced adverse remodeling after 4 weeks of nonreperfused myocardial infarction (MI) in wild-type B6 and EphA2-R-M (EphA2 receptor null) mice. METHODS: At 4 weeks post-MI, echocardiography, histologic and immunohistochemical analyses of B6 mouse hearts were performed. Primary mouse cardiac fibroblasts (FBs) isolated from B6 mice cultured in the presence of low and high dose ephrinA1-Fc, both with and without pro-fibrotic TGF-ß stimulation and Western blots, were probed for relative expression of remodeling proteins MMP-2, MMP-9 and TIMP-1, in addition to DDR2 and (p)SMAD2/3/totalSMAD2/3. RESULTS: EphrinA1-Fc preserved a significant degree of contractile function, decreased adverse left ventricular remodeling, attenuated excessive compensatory hypertrophy, and decreased interstitial fibrosis in wild-type (WT) B6 mouse hearts. In contrast, most of these parameters were poorer in ephrinA1-Fc-treated EphA2-R-M mice. Of note, fibrosis was proportionately decreased, implying that other EphA receptor(s) are more important in regulating the pro-fibrotic response. Primary FBs showed disparate alteration of MMP-2, MMP-9 and TIMP-1, as well as DDR2 and p-SMAD2/3/totalSMAD2/3, which indicates that matrix remodeling and cardiac fibrosis in the injured heart are influenced by ephrinA1-Fc. CONCLUSION: This study demonstrates the capacity of a single administration of ephrinA1-Fc at the onset of injury to attenuate long-term nonreperfused post-MI ventricular remodeling that results in progressive heart failure, and the important role of EphA2 in mitigating the deleterious effects.

Intracardiac administration of ephrinA1-Fc preserves mitochondrial bioenergetics during acute ischemia/reperfusion injury.

AIMS: Intracardiac injection of recombinant EphrinA1-Fc immediately following coronary artery ligation in mice reduces infarct size in both reperfused and non-reperfused myocardium, but the cellular alterations behind this phenomenon remain unknown. MAIN METHODS: Herein, 10 wk-old B6129SF2/J male mice were exposed to acute ischemia/reperfusion (30minI/24hrsR) injury immediately followed by intracardiac injection of either EphrinA1-Fc or IgG-Fc. After 24 h of reperfusion, sections of the infarct margin in the left ventricle were imaged via transmission electron microscopy, and mitochondrial function was assessed in both permeabilized fibers and isolated mitochondria, to examine mitochondrial structure, function, and energetics in the early stages of repair. KEY FINDINGS: At a structural level, EphrinA1-Fc administration prevented the I/R-induced loss of sarcomere alignment and mitochondrial organization along the Z disks, as well as disorganization of the cristae and loss of inter-mitochondrial junctions. With respect to bioenergetics, loss of respiratory function induced by I/R was prevented by EphrinA1-Fc. Preservation of cardiac bioenergetics was not due to changes in mitochondrial JH2O2 emitting potential, membrane potential, ADP affinity, efficiency of ATP production, or activity of the main dehydrogenase enzymes, suggesting that EphrinA1-Fc indirectly maintains respiratory function via preservation of the mitochondrial network. Moreover, these protective effects were lost in isolated mitochondria, further emphasizing the importance of the intact cardiomyocyte ultrastructure in mitochondrial energetics. SIGNIFICANCE: Collectively, these data suggest that intracardiac injection of EphrinA1-Fc protects cardiac function by preserving cardiomyocyte structure and mitochondrial bioenergetics, thus emerging as a potential therapeutic strategy in I/R injury.

EphrinA1-Fc attenuates myocardial ischemia/reperfusion injury in mice.

EphrinA1, a membrane-bound receptor tyrosine kinase ligand expressed in healthy cardiomyocytes, is lost in injured cells following myocardial infarction. Previously, we have reported that a single intramyocardial injection of chimeric ephrinA1-Fc at the time of ischemia reduced injury in the nonreperfused myocardium by 50% at 4 days post-MI by reducing apoptosis and inflammatory cell infiltration. In a clinically relevant model of acute ischemia (30min)/reperfusion (24hr or 4 days) injury, we now demonstrate that ephrinA1-Fc reduces infarct size by 46% and completely preserves cardiac function (ejection fraction, fractional shortening, and chamber dimensions) in the short-term (24hrs post-MI) as well as long-term (4 days). At 24 hours post-MI, diminished serum inflammatory cell chemoattractants in ephrinA1-Fc-treated mice reduces recruitment of neutrophils and leukocytes into the myocardium. Differences in relative expression levels of EphA-Rs are described in the context of their putative role in mediating cardioprotection. Validation by Western blotting of selected targets from mass spectrometry analyses of pooled samples of left ventricular tissue homogenates from mice that underwent 30min ischemia and 24hr of reperfusion (I/R) indicates that ephrinA1-Fc administration alters several regulators of signaling pathways that attenuate apoptosis, promote autophagy, and shift from FA metabolism in favor of increased glycolysis to optimize anaerobic ATP production. Taken together, reduced injury is due a combination of adaptive metabolic reprogramming, improved cell survival, and decreased inflammatory cell recruitment, suggesting that ephrinA1-Fc enhances the capacity of the heart to withstand an ischemic insult.