Abstract Body (Do not enter title and authors here): Background:
Delayed reperfusion (DR) in myocardial ischemia contributes to significant cardiac injury, characterized by mitochondrial dysfunction, oxidative stress, and cardiomyocyte death. Mitochondrial respiratory complexes are essential for maintaining cardiac energy metabolism during DR. EN1, known to support mitochondrial function in neurons, has recently been suggested to regulate mitochondrial complexes. However, its role in cardiomyocytes and the underlying molecular mechanisms remain poorly defined.
Methods:
We evaluated EN1 expression in murine models of myocardial DR injury and hypoxia/reoxygenation (H/R)-treated primary cardiomyocytes. The functional role of EN1 was examined using cardiomyocyte-specific En1 knockout and overexpression mouse models. Myocardial injury, cardiac function, and mitochondrial respiratory complex expression were assessed. To elucidate the molecular mechanism, we conducted co-immunoprecipitation to identify protein interactions, and employed PGC-1α inhibition and TFAM knockdown to dissect downstream signaling.
Results:
EN1 was significantly upregulated in both mouse myocardium post-DR and cardiomyocytes post-H/R, with nuclear localization. Specific knockout of En1 in cardiomyocytes markedly aggravated DR-induced myocardial infarction, elevated cardiomyocyte apoptosis, and impaired left ventricular systolic function. Loss of EN1 disrupted mitochondrial structure and led to reduction in the expression of key mitochondrial respiratory complex subunits (including UQCRC2, SDHB, and NDUFB8) at both the transcript and protein levels. In contrast, cardiomyocyte-specific EN1 overexpression reduced myocardial infarct size, preserved mitochondrial respiratory complex integrity, and restored cardiac function after DR injury. Mechanistically, EN1 interacted with PGC-1α and promoted the transcription of nuclear-encoded respiratory complex genes, while simultaneously upregulating TFAM to enhance mitochondrial DNA transcription. Inhibition of PGC-1α or silencing of TFAM impaired these effects, confirming that EN1 mediates mitochondrial and cardioprotective effects through the PGC-1α/TFAM pathway.
Conclusions:
EN1 plays a crucial role in maintaining mitochondrial respiratory function and conferring resistance to DR-induced cardiac injury through the PGC-1α/TFAM signaling axis. These findings identify EN1 as a promising molecular target for improving mitochondrial function and cardiac outcomes in myocardial DR injury.