Abstract 287: Altered Mitochondrial Flash Activity and mPTP Opening in the Aged Heart Are Reversed by Elamipretide Treatment

Abstract

Rationale: The mitochondrial theory of aging pinpoints mitochondria as the major reactive oxygen species (ROS) production site and target of oxidative stress during aging. Mitochondrial flash (mitoflash) is recently discovered excitable event that is a manifestation of ROS production combined with pH change and coupled with mitochondrial permeability transition pore (mPTP) opening. Our preliminary data shown that 8-week SS-31 peptide (elamipretide) protects the functional performance of the aged mouse heart. This study aims to investigate mitoflash activity and mPTP opening in the aged heart, especially whether and how SS-31 protects the heart function involving modulation of these activities. Methods and Results: By using confocal microscopy imaging on the isolated rat cardiomyocytes with overexpression of mitochondrial targeted cpYFP, we found mitoflash activity in the cells from old (26 mo) was higher than that in young (5 mo) rat cells (2.2 ± 0.2 in old vs 1.3 ± 0.2 in young, /1000μm 2 /100s, n=27-64, p<0.05). SS-31 10 μM treatment normalized the mitoflash activity to the young level. Similarly, by photon-excitation induced mPTP opening, we found that aging increased the mPTP opening and 1 μM SS-31 stabilized the mPTP. Furthermore, by using Seahorse Assay on intact primary isolated cardiomyocytes, we found mitochondrial basal respiration in cells from old mice was higher than that in young mouse cells, however, the maximal respiration rate was without significant difference. The increased basal respiration was attributed to a higher proton leak in old cardiomyocytes (131 ± 14 in 24 month vs 94 ± 11 in young, pmol/min/800cells, n=17-35, p<0.05). Interestingly, acute (2 hrs) in vitro treatment of the old cardiomyocytes with SS-31 100 nm decreased proton leak. Moreover, SS-31 (100 nM) decreased the superoxide production as measured by ratio of MitoSOX Red to MitoTrackerGreen. Conclusion: These results indicate that SS-31 directly protects cardiac aging through rapid rejuvenation of mitochondrial respiration in cardiomyocytes, and in particular, by reducing proton leak, mitochondrial flash activity, and decreasing the mPTP opening. This study helps to uncover the mechanism of the cardiac aging and the protective effect from SS-31 treatment.