Time-resolved fluorescence spectroscopy investigation of the effect of 4-hydroxynonenal on endogenous NAD(P)H in living cardiac myocytes.

Lipid peroxidation is a major biochemical consequence of the oxidative deterioration of polyunsaturated lipids in cell membranes and causes damage to membrane integrity and loss of protein function. 4-hydroxy-2-nonenal (HNE), one of the most reactive products of n-6 polyunsaturated fatty acid peroxidation of membrane phospholipids, has been shown to be capable of affecting both nicotinamide adenine dinucleotide (phosphate) reduced [NAD(P)H] as well as NADH production. However, the understanding of its effects in living cardiac cells is still lacking. Our goal was to therefore investigate HNE effects on NAD(P)H noninvasively in living cardiomyocytes. Spectrally resolved lifetime detection of endogenous fluorescence, an innovative noninvasive technique, was employed. Individual fluorescence components were resolved by spectral linear unmixing approach. Gathered results revealed that HNE reduced the amplitude of both resolved NAD(P)H components in a concentration-dependent manner. In addition, HNE increased flavoprotein fluorescence and responsiveness of the NAD(P)H component ratio to glutathione reductase (GR) inhibitor. HNE also increased the percentage of oxidized nucleotides and decreased maximal NADH production. Presented data indicate that HNE provoked an important cell oxidation by acting on NAD(P)H regulating systems in cardiomyocytes. Understanding the precise role of oxidative processes and their products in living cells is crucial for finding new noninvasive tools for biomedical diagnostics of pathophysiological states.

Metabolic remodelling of cardiac myocytes during pregnancy: the role of mineralocorticoids.

BACKGROUND: Pregnancy is associated with significant cardiac adaptations. The regulatory mechanisms involved in functional cardiac adaptations during pregnancy are still largely unknown. In pathologic conditions, mineralocorticoids have been shown to mediate structural as well as functional remodelling of the heart. However, their role in cardiac physiological conditions is not completely understood. Here, we examined cardiac cell metabolic remodelling in the late stages of rat pregnancy, as well as mineralocorticoid involvement in this regulation. METHODS: We have applied rapid video imaging, echocardiography, patch clamp technique, confocal microscopy, and time-resolved fluorescence spectroscopy. RESULTS: Our results revealed that cardiac cells undergo metabolic remodelling in pregnancy. Inhibition of mineralocorticoid receptors during pregnancy elicited functional alterations in cardiac cells: blood levels of energy substrates, particularly lactate, were decreased. As a consequence, the cardiomyocyte contractile response to these substrates was blunted, without modifications of L-type calcium current density. Interestingly, this response was associated with changes in the mitochondrial metabolic state, which correlated with modifications of bound reduced nicotinamide adenine dinucleotide (phosphate) NAD(P)H levels. We also noted that mineralocorticoid receptor inhibition prevented pregnancy-induced decrease in transient outward potassium current. CONCLUSIONS: This study demonstrates that in pregnancy, mineralocorticoids contribute to functional adaptations of cardiac myocytes. By regulating energy substrate levels, in particular lactate, in the plasma and metabolic state in the cells, mineralocorticoids affect the contractility responsiveness to these substrates. In the future, understanding cardiac adaptations during pregnancy will help us to comprehend their pathophysiological alterations.