Abstract Body (Do not enter title and authors here): Background: Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is a key glycolytic enzyme traditionally considered a housekeeping protein. Recent studies, however, suggest that GAPDH plays a role in cellular processes such as apoptosis and more importantly, in mitochondrial (MITO) function through regulation of mitophagy, a process of selective degradation and clearance of damaged mitochondria by lysosomes. We previously showed that accumulation of damaged mitochondria is a characteristic feature of constituent cardiomyocytes of the failing left ventricular (LV) myocardium. This abnormality points to dysregulation of mitophagy in heart failure (HF) that can lead to overall MITO dysfunction in HF with subsequent energy deprivation.
Purpose: In this study, we tested the hypothesis that GAPDH protein levels are selectively reduced in MITO fractions isolated from LV tissue of dogs with chronic HF compared to healthy normal (NL) dogs.
Methods: Studies were performed using LV tissue from 7 healthy NL dogs and 7 HF dogs (LV ejection fraction≤35%) produced by multiple intracoronary microembolizations. GAPDH (37 kDa) protein levels were measured in lithium dodecyl sulfate (LDS) lysate of 1) LV homogenate, 2) cytosolic fraction, 3) MITO fractions and 4) sarcolemmal fractions by Western blotting coupled with chemiluminescence using a canine-specific commercially available antibody. Band intensities were expressed in densitometric units (du).
Results: Results are shown in the table. A significant reduction in GAPDH levels was observed in the MITO fractions of HF dogs compared to NL dogs (p< 0.05). In contrast, GAPDH levels in the homogenate, cytosol, and sarcolemma fractions showed no significant differences between HF and NL groups.
Conclusion: GAPDH protein levels are selectively reduced in mitochondria of LV myocardium of dogs with chronic HF. This observation is consistent with abnormal mitophagy in HF evidenced by accumulation of damaged mitochondria in cardiomyocytes. Given the importance of mitochondria in heart function, targeting MITO dysfunction remains a promising therapeutic approach for the treatment of HF.