Cardiac proteome analysis of a novel genetic mouse model of hypertension: the ET+/+eNOS-/-mice
Zusammenfassung der Projektergebnisse
The vasoconstrictive hormone Endothelin‐1 (ET‐1) and the vasodilator nitric oxide (NO) belong to a complex and tightly adjusted network and control cardiac function. The clinical relevance of this delicate interplay has been acknowledged because of its implication in many cardiovascular diseases, such as pulmonary arterial hypertension, systemic hypertension, and coronary artery disease. However, the underlying molecular mechanisms remain to be fully clarified. ET‐1 transgenic (ET+/+) mice develop pulmonary and cardiac fibrosis but remain normotensive. The functional antagonist of ET‐1, NO, was assumed to counteract the ET‐1 effect on blood pressure in ET+/+ mice. To test this hypothesis in vivo, ET+/+ mice were crossbred with eNOS knockout (eNOS‐/‐) mice. Similar to the eNOS‐/‐ model, the ET+/+eNOS‐/‐ mice develop high blood pressure compared to wild type (WT) and ET+/+ animals. To the contrary to our hypothesis, at the age of nine months, the eNOS‐/‐, but not ET+/+eNOS‐/‐ mice, are characterized by diastolic dysfunction. These findings suggested that transgenic overexpression of ET‐1 on an eNOS‐/‐ background could be beneficial for diastolic function. In this study, it is shown that cardiac ET‐1 gene expression was elevated in both eNOS‐/‐ and ET+/+eNOS‐ /‐ mice compared to WT at the age of nine months with no significant difference between these groups. This suggests that the restored left ventricular function between eNOS‐/‐ and ET+/+eNOS‐/‐ animals are due to adaptive mechanisms triggered by the chronic overexpression of ET‐1. In line with this, eNOS‐/‐ animals developed cardiac hypertrophy at the age of nine months, whereas ET+/+eNOS‐/‐ mice showed enlarged cardiomyocytes at the age of three months that were not detectable in animals nine months of age. Histological analysis showed that cardiac arterioles were dilated in both ET+/+ and ET+/+eNOS‐/‐ mice compared to WT and eNOS‐/‐ mice. By enhancing blood flow, this could be beneficial for cardiac functions. In order to dissect the causal molecular changes underlying this phenomenon, the cardiac proteome of the different genotypes were compared to WT at the age of three months using two‐dimensional electrophoresis coupled to mass spectrometry. This study revealed that transgenic overexpression of ET‐ 1, with or without eNOS, led to a higher abundance of proteins regulating oxidative stress indicating that, in contrast to eNOS‐/‐ animals, ET+/+ and ET+/+eNOS‐/‐ mice developed molecular mechanisms limiting oxidative damages. Moreover, diastolic dysfunction observed in eNOS‐/‐ mice may be explained by the differential abundance of proteins involved in the contractile machinery. Overexpression of ET‐1 may have benefited the cardiac function in eNOS‐/‐ mice by restoring these changes. Finally, this study indicated that a shift from fatty acid to glucose metabolism, considered as cardioprotective, may have occurred to a greater extent in crossbred animals than in eNOS‐/‐ mice. Despite the lack of specificity of commercial antibodies, we could in part confirm and analyze in more details the results of the proteomics study by classic and 2‐dimensional Western Blot. Taken together, this study showed that transgenic overexpression of ET‐1 in mice can have beneficial effects on cardiac function, even in the absence of eNOS, by modulating various systems (oxidative stress, contractile machinery, and energy metabolism). The clinical relevance of these findings should be confirmed by analyzing the impact of a pharmacological interference with these systems on cardiac function.
Projektbezogene Publikationen (Auswahl)
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Endothelin-1 overexpression restores left ventricular function in eNOS knockout mice
N. Vignon-Zellweger, K. Relle, E. Kienlen, M. Alter, P. Seider, J. Sharkovska, S. Heiden, P. Kalk, K. Schwab, B. Albrecht-Küpper, J.-P. Stasch, F. Theuring, B. Hocher
