Background <p>Myocardial ischemia can lead to myocardial injury. Eukaryotic elongation factor 1A2 (eEF1A2) serves as a required translational elongation factor specific to cardiac function. Yet, the action of eEF1A2 in myocardial injury is still unexplored largely.</p> Objective <p>To investigate the expression of eEF1A2 in myocardial cell injury and its potential effect and mechanism on myocardial injury in ischemic cardiomyopathy.</p> Methods <p>We used qRT-PCR to quantify eEF1A2’s relative expression, immunofluorescence and western blotting to acquire related proteins’ contents. Cell vitality and apoptotic rate were tested with the CCK-8 assay, lactate dehydrogenase (LDH) test and flow cytometry method separately. We also tested the contents of oxidative stress factors: reactive oxygen species (ROS), malondialdehyde (MDA), and glutathione peroxidase (GSH-Px). In-vivo experiments were made to analyze the intervention of excess eEF1A2 in myocardial injury.</p> Results <p>Silencing eEF1A2 enhanced hypoxia’s inhibitory impact on the vitality of H9c2 cells and stimulatory influence on the cells’ apoptosis and oxidative stress; in contrast, overexpressing eEF1A2 exhibited reverse outcomes. Overexpression of eEF1A2 alleviated hypoxia-induced suppression of the phosphorylation of AMP-activated protein kinase (AMPK) and promotional effect on the phosphorylation of IκBα and p65. This phenomenon was reversed with the addition of Compound C. Compound C reversed the influence of eEF1A2 on cells vitality, apoptosis, and oxidative stress injury. In addition, eEF1A2 overexpression reduced the expression of creatine phosphokinase (CPK) and LDH, alleviated myocardial injury and fibrosis, diminished myocardial injury’s impeding impact on Ki-67 and AMPK’s phosphorylation and stimulatory impacts on cell apoptosis and on the phosphorylation of Collagen I, Collagen III, IκBα, and p65.</p> Conclusion <p>Via the AMPK/NF-κB signal pathway, eEF1A2 is capable of ameliorating myocardial injury in ischemic cardiomyopathy. This finding positions eEF1A2 as a viable target for both diagnosis and molecular therapy of myocardial injury.</p> Graphical abstract <p>Through the AMPK/NF-κB signaling pathway, eEF1A2 improves the cell viability of cardiomyocytes, reduces apoptosis and oxidative stress, and thus alleviates myocardial injury.</p> <p></p> Highlights <p><OrderedList> <ListItem> <ItemNumber>1.</ItemNumber> <ItemContent> <p>eEF1A2 level in hypoxic H9c2 cells is significantly decreased.</p> </ItemContent> </ListItem> <ListItem> <ItemNumber>2.</ItemNumber> <ItemContent> <p>eEF1A2 mediates the vitality, apoptosis, and oxidative stress response of hypoxic H9c2 cells.</p> </ItemContent> </ListItem> <ListItem> <ItemNumber>3.</ItemNumber> <ItemContent> <p>eEF1A2, through the AMPK/NF-κB signal pathway, ameliorates myocardial injury in ischemic cardiomyopathy.</p> </ItemContent> </ListItem> </OrderedList></p>