Proteome Analysis Suggests That Mitochondrial Dysfunction in Stressed Endothelial Cells Is Reversed by a Soy Extract and Isolated Isoflavones Dagmar Fuchs,† Barbara Dirscherl,† Joyce H. Schroot,‡ Hannelore Daniel,† and Uwe Wenzel*,§ Department of Food and Nutrition, Molecular Nutrition Unit, Technical University of Munich, Am Forum 5, D-85350 Freising, Germany, Food Technology Centre, part of Wageningen University and Research Centre, Bornsesteeg 59, 6708 PD Wageningen, The Netherlands, Molecular Nutrition Research, Interdisciplinary Research Center, Justus-Liebig-University of Giessen, Heinrich-Buff-Ring 26-32, D-35392 Giessen, Germany Received October 17, 2006
Apoptosis is a driving force in atherosclerosis development. A soy extract or a combination of the soy isoflavones genistein and daidzein inhibited apoptosis induced by oxidized LDL in endothelial cells. Proteome analysis revealed that the LDL-induced alterations of numerous proteins were reversed by the extract and the genistein/daidzein mixture but only three protein entities, all functionally linked to mitochondrial dysfunction, were regulated in common by both treatments. Keywords: proteomics • atherosclerosis • ox-LDL • apoptosis • reactive oxygen species
Introduction Epidemiological studies suggest that the consumption of soycontaining foods has the ability to prevent or to slow-down the development of cardiovascular disease.1,2 Systematic reviews have assessed the effects of soy isoflavones on lipid levels and concluded that a diet supplemented with soy protein isolates containing isoflavones reduce LDL cholesterol levels.3 In contrast, purified soy isoflavones or soy protein preparation not containing isoflavones failed to show statistically significant effects.3 It therefore remains unclear which active components of soy mediate the cardioprotective actions in vivo. Studies in vitro revealed that isolated isoflavones interfere with a variety of mechanisms that play a role in atherosclerosis development.4-7 One crucial mechanism involved in atherosclerosis is endothelial apoptosis, since it leads to an increased exposure of phosphatidylserine groups on the outer leaflet of the plasma membrane that in the presence of coagulation factors V and VII, promotes thrombin generation.8 In addition, membrane vesicles and blebs from apoptotic endothelial cells contain oxidized phospholipids that induce monocyte adhesion to the endothelium.9 We have previously shown that isoflavones by altering the steady-state levels of a variety of cellular proteins are able to inhibit apoptosis when induced by the endothelial stressors homocysteine or oxidized LDL (ox-LDL).10-12 Since the effects of isoflavones in vivo appear to depend on the presence of an intact soy matrix and other soy compounds than * To whom correspondence should be addressed. Uwe Wenzel, Molecular Nutrition Research, Interdisciplinary Research Center, Justus-LiebigUniversity of Giessen, Heinrich-Buff-Ring 26-32, D-35392 Giessen, Germany. Phone, +49 (0) 641/99-39220; fax, +49 (0) 641/99-39229; e-mail,
[email protected]. † Technical University of Munich. ‡ Wageningen University and Research Centre. § Justus-Liebig-University of Giessen.
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Journal of Proteome Research 2007, 6, 2132-2142
Published on Web 05/16/2007
isoflavones could contribute to the effects as well,3 we assessed in the present study if and to which extent a soy extract in comparison to a genistein and daidzein mixture can affect apoptosis in endothelial cells in culture when stressed with 5 µg/mL ox-LDL. The soy extract employed was cleaved by β-glucosidase in vitro to deliver 2.5 µM genistein and 1.0 µM of daidzein. These concentrations of isoflavone aglycones are commonly found in plasma of humans on a high intake of soy products as in Japan, whereas on a Western-type diet, plasma concentrations are usually
