Oxidative stress and function of putative cardio-protective compounds in isolated cardiomyocytes

Abstract

The objectives of the studies conducted for this thesis were to further develop our understanding of the effect of homocysteine (Hcy) as a risk factor for cardiovascular disease (CVD) and the cardioprotective roles of different putative protective compounds in freshly isolated cardiomyocytes under mild and toxic oxidative conditions. We examined the effects of acute homocysteinemia in middle-aged (36 weeks) rat hearts on the susceptibility of freshly isolated cardiomyocytes to oxidative stress under normal and oxidative stress conditions and the effects of putative protective compounds. We also measured ROS production, [Ca⁺⁺]i and contractility in young adult rat cardiomyocytes of both sexes, isolated with or without taurine, using fluorescence microscopy. Finally, serum Hcy and glutathione concentrations were measured using liquid chromatography mass spectrometry (LCMS) in young adult and in middle-aged Wistar rats of both sexes. The results showed that the percentage of rod-shaped cells and the viability of isolated cells decreased following acute exposure to DL-Hcy and oxidative stress. However, we show that putative protective compounds supplement (e.g., L-carnitine, L-glutamate + L-carnitine in combination or taurine) can counteract free radical accumulation and protect against oxidative damage. This is the first study to demonstrate the effects of perfusing freshly isolated heart cells with a combination of L-glutamate and L-carnitine. Our findings also show that 1% L-methionine supplementation significantly increased serum Hcy levels in young adult and middle-aged male rats and had no significant effects in female rats. Also, a small increase in serum glutathione (GSH) concentration was observed in female rats with no significant changes in male. Our study provides new evidence that ROS production is lower in female cardiomyocytes than in males in both control and methionine groups under acute oxidative stress. Interestingly, our findings show an increase in [Ca⁺⁺]i in female compared to male cardiomyocytes under acute oxidative stress in both control and methionine groups which might explain the higher contractility of the female cardiomyocytes under these conditions. We also show that taurine treatment is more effective in mild oxidative stress in female control and methionine groups compared to male groups. Similar protective effects of taurine are reported during acute oxidative stress following exposure to chronic ROS induced by methionine in both male and female cardiomyocytes. In conclusion, these results suggest that acute exposure to DL-Hcy and oxidative stress affect cell morphology and viability and highlight the role of putative protective compounds against oxidative stress. Also our results provide new evidence to show that 1% L-methionine supplementation reduces ROS production, increases [Ca⁺⁺]i and increases contraction in female compared to male cardiomyocytes under acute oxidative stress in both control and methionine groups and reveals the protective effect of taurine.