Effect of the activation of mitochondrial biogenesis by nicotinamide mononocleotide on cardioprotection induced by melatonin in myocardial ischemia-reperfusion injury of aged male rats

Abstract

Introduction: Ischemic heart diseases are the major reasons for disability and mortality in elderly. In this study, the effect of combined therapy of NMN and melatonin on cardiac function, myocardial infarct size, mitochondrial function, oxidative stress and mitochondrial biogenesis and miR-499 changes as well as histopathological changes in aged rat heart with region IR injury have been investigated. Methods: Sixty aged Wistar rats were randomly allocated to five groups including sham, control, NMN-receiving, melatonin-receiving, and combination therapy (NMN+melatonin). NMN (100 mg/kg/day/i.p.) was administered for every other day for 28 days before IR. The rat’s heart are suspended in the Langendorff apparatus and, myocardial IR injury was induced by LAD coronary artery ligation (ischemia) for 30 minutes and then reopening of it (reperfusion) for 60 mimutes. In the groups receiving melatonin 5 min before the start of reperfusion, 50 μM melatonin was added to the Krebs solution and the hearts were reperfused with Krebs containing melatonin for 15 min. At the end of the isolated heart experiments, a series of isolated hearts were used to assess mryocardial infarct size by using evans blue and TTC dye staining, and in the other series of hearts were used for mitochondrial isolation and mitochondrial function studies (production of ROS and mitochondrial membrane potential), oxidative stress, and expression of mitochondrial biogenesis genes and miR-499 also histopathological changes. Results: Melatonin and NMN had cardioprotective effects in old rats. They improved cardiac function (p<0.001), significantly reduced creatine kinase and infarct size compared to control group. Also, there was a significant increase in antioxidant enzyme activity in treated groups in comparison with control group (p<0.001). Moreover, pretreatment with NMN increased the cardioprotection by melatonin. Mitochondrial ROS production was decreased in the treated groups compared to the control group (p <0.001). There was an increase in mitochondrial membrane depolarization (p<0.001) in the control group in comparison to the sham group. Melatonin (p<0.01) and NMN (p<0.001) treatment prevented mitochondrial membrane depolarization. The effects of combination therapy on reduction of mitochondrial ROS and oxidative status and improvement of mitochondrial membrane potential as well as mitochondrial biogenesis were greater than those of alone treatments. In addition, melatonin and NMN increased the expression of mitochondrial biogenesis genes and miR-499 expression (p<0.001). Also combination treatment improved the damaged tissue following IR injury.