The role of oxysterols in control of endothelial stiffness

Abstract

Endothelial dysfunction is a key step in atherosclerosis development. Our recent studies suggested that oxLDL-induced increase in endothelial stiffness plays a major role in dyslipidemia-induced endothelial dysfunction. In this study, we identify oxysterols, as the major component of oxLDL, responsible for the increase in endothelial stiffness. Using Atomic Force Microscopy to measure endothelial elastic modulus, we show that endothelial stiffness increases with progressive oxidation of LDL and that the two lipid fractions that contribute to endothelial stiffening are oxysterols and oxidized phosphatidylcholines, with oxysterols having the dominant effect. Furthermore, endothelial elastic modulus increases as a linear function of oxysterol content of oxLDL. Specific oxysterols, however, have differential effects on endothelial stiffness with 7-ketocholesterol and 7α-hydroxycholesterol, the two major oxysterols in oxLDL, having the strongest effects. 27-hydroxycholesterol, found in atherosclerotic lesions, also induces endothelial stiffening. For all oxysterols, endothelial stiffening is reversible by enriching the cells with cholesterol. oxLDL-induced stiffening is accompanied by incorporation of oxysterols into endothelial cells. We find significant accumulation of three oxysterols, 7α-hydroxycholesterol, 7β-hydroxycholesterol, and 7-ketocholesterol, in mouse aortas of dyslipidemic ApoE−/− mice at the early stage of atherosclerosis. Remarkably, these are the same oxysterols we have identified to induce endothelial stiffening. Endothelial dysfunction is a key step in atherosclerosis development. Our recent studies suggested that oxLDL-induced increase in endothelial stiffness plays a major role in dyslipidemia-induced endothelial dysfunction. In this study, we identify oxysterols, as the major component of oxLDL, responsible for the increase in endothelial stiffness. Using Atomic Force Microscopy to measure endothelial elastic modulus, we show that endothelial stiffness increases with progressive oxidation of LDL and that the two lipid fractions that contribute to endothelial stiffening are oxysterols and oxidized phosphatidylcholines, with oxysterols having the dominant effect. Furthermore, endothelial elastic modulus increases as a linear function of oxysterol content of oxLDL. Specific oxysterols, however, have differential effects on endothelial stiffness with 7-ketocholesterol and 7α-hydroxycholesterol, the two major oxysterols in oxLDL, having the strongest effects. 27-hydroxycholesterol, found in atherosclerotic lesions, also induces endothelial stiffening. For all oxysterols, endothelial stiffening is reversible by enriching the cells with cholesterol. oxLDL-induced stiffening is accompanied by incorporation of oxysterols into endothelial cells. We find significant accumulation of three oxysterols, 7α-hydroxycholesterol, 7β-hydroxycholesterol, and 7-ketocholesterol, in mouse aortas of dyslipidemic ApoE−/− mice at the early stage of atherosclerosis. Remarkably, these are the same oxysterols we have identified to induce endothelial stiffening. Multiple studies have shown that oxidative modifications of LDL (oxLDL) are a major factor in the development of atherosclerosis (1.Witztum J.L. Steinberg D. Role of oxidized low density lipoprotein in atherogenesis.J. Clin. Invest. 1991; 88: 1785-1792Crossref PubMed Scopus (2472) Google Scholar, 2.Witztum J.L. Steinberg D. The oxidative modification hypothesis of atherosclerosis: does it hold for humans?.Trends Cardiovasc. Med. 2001; 11: 93-102Crossref PubMed Scopus (387) Google Scholar, 3.Stocker R. Keaney Jr, J.F. Role of oxidative modifications in atherosclerosis.Physiol. Rev. 2004; 84: 1381-1478Crossref PubMed Scopus (2105) Google Scholar, 4.Libby P. Inflammation in atherosclerosis.Nature. 2002; 420: 868-874Crossref PubMed Scopus (6940) Google Scholar, 5.Paoletti R. Gotto A.M. Hajjar D.P. Inflammation in atherosclerosis and implications for therapy.Circulation. 2004; 109: III-20-III-26Crossref Google Scholar–6.Levitan I. Volkov S. Subbaiah P.V. Oxidized LDL: diversity, patterns of recognition and pathophysiology.Antioxidants & Redox Signaling. 2010; 13: 39-75Crossref PubMed Scopus (313) Google Scholar). The level of oxLDL increases dramatically with hypercholesterolemia in animal models of atherosclerosis (7.Hodis H.N. Kramsch D.M. Avogaro P. Bittolo-Bon G. Cazzolato G. Hwang J. Peterson H. Sevanian A. Biochemical and cytotoxic characteristics of an in vivo circulating oxidized low density lipoprotein (LDL.J. Lipid Res. 1994; 35: 669-677Abstract Full Text PDF PubMed Google Scholar, 8.Holvoet P. Theilmeier G. Shivalkar B. Flameng W. Collen D. LDL hypercholesterolemia is associated with accumulation of oxidized LDL, atherosclerotic plaque growth, and compensatory vessel enlargement in coronary arteries of miniature pigs.Arterioscler. Thromb. Vasc. Biol. 1998; 18: 415-422Crossref PubMed Scopus (96) Google Scholar), and in humans (9.Cazzolato G. Avogaro P. Bittolo-Bon G. Characterization of a more electronegatively charged LDL subfraction by ion exchange HPLC.Free Radic. Biol. Med. 1991; 11: 247-253Crossref PubMed Scopus (99) Google Scholar, 10.van Tits L.J. van Himbergen T.M. Lemmers H.L. de Graaf J. Stalenhoef A.F. Proportion of oxidized ldl relative to plasma apolipoprotein b does not change during statin therapy in patients with heterozygous familial hypercholesterolemia.Atherosclerosis. 2006; 185: 307-312Abstract Full Text Full Text PDF PubMed Scopus (41) Google Scholar), it is found in atherosclerotic lesions (11.Ylä-Herttuala S. Palinski W. Rosenfeld M.E. Parthasarathy S. Carew T.E. Butler S. Witztum J.L. Steinberg D. Evidence for the presence of oxidatively modified low density lipoprotein in atherosclerotic lesions of rabbit and man.J. Clin. Invest. 1989; 84: 1086-1095Crossref PubMed Google Scholar). It is also well established that dyslipidemia-induced dysfunction of vascular endothelial cells is a critical step in the early stage of atherosclerosis (e.g., Refs. 12.Shimokawa H. Primary endothelial dysfunction: atherosclerosis.J. Mol. Cell. Cardiol. 1999; 31: 23-37Abstract Full Text PDF PubMed Scopus (363) Google Scholar, 13.Kinlay S. Libby P. Ganz P. Endothelial function and coronary artery disease.Curr. Opin. Lipidol. 2001; 12: 383-389Crossref PubMed Scopus (307) Google Scholar–14.Sima A.V. Stancu C.S. Simionescu M. Vascular endothelium in atherosclerosis.Cell Tissue Res. 2009; 335: 191-203Crossref PubMed Scopus (187) Google Scholar) and a strong predictor of cardiovascular disease (CVD) development (15.Gonzalez M.A. Selwyn A.P. Endothelial function, inflammation, and prognosis in cardiovascular disease.Am. J. Med. 2003; 115: 99S-106SAbstract Full Text Full Text PDF PubMed Scopus (216) Google Scholar, 16.Heitzer T. Schlinzig T. Krohn K. Meinertz T. Munzel T. Endothelial dysfunction, oxidative stress, and risk of cardiovascular events in patients with coronary artery disease.Circulation. 2001; 104: 2673-2678Crossref PubMed Scopus (1653) Google Scholar–17.Schächinger V. Britten M.B. Zeiher A.M. Prognostic impact of coronary vasodilator dysfunction on adverse long-term outcome of coronary disease.Circulation. 2000; 101: 1899-1906Crossref PubMed Scopus (2344) Google Scholar). The mechanisms, however, of dyslipidemia-induced endothelial dysfunction are still poorly understood. Our recent studies have shown that exposure to oxLDL in vitro or to plasma dyslipidemia in vivo significantly increases the stiffness of aortic endothelial cells, which in turn is associated with an increase in endothelial contractility, enhanced angiogenic potential, and sensitivity to shear stress (18.Byfield F.J. Tikku S. Rothblat G.H. Gooch I. increases endothelial and Lipid Res. 2006; Full Text Full Text PDF PubMed Scopus Google Scholar, I. Gooch Subbaiah P.V. M. I. oxLDL-induced in lipid of is with endothelial stiffness and J. 2010; PubMed Scopus Google F.J. I. oxLDL of aortic endothelial J. Cell. PubMed Scopus Google Scholar). Furthermore, studies that dyslipidemia-induced endothelial stiffening is not by by of lipid of in endothelial cells (18.Byfield F.J. Tikku S. Rothblat G.H. Gooch I. increases endothelial and Lipid Res. 2006; Full Text Full Text PDF PubMed Scopus Google Scholar, I. Gooch Subbaiah P.V. M. I. oxLDL-induced in lipid of is with endothelial stiffness and J. 2010; PubMed Scopus Google Scholar, I. of oxLDL on PubMed Google Scholar). with these oxLDL-induced endothelial stiffness by enriching the cells with oxLDL on the content of endothelial I. Gooch Subbaiah P.V. M. I. oxLDL-induced in lipid of is with endothelial stiffness and J. 2010; PubMed Scopus Google Scholar). studies to the hypothesis that oxLDL induces endothelial dysfunction by oxysterols into the plasma in the of and endothelial stiffening. The of this study, to the impact of oxysterols on endothelial stiffness. in by of oxidation and of R. Keaney Jr, J.F. Role of oxidative modifications in atherosclerosis.Physiol. Rev. 2004; 84: 1381-1478Crossref PubMed Scopus (2105) Google Scholar, I. Volkov S. Subbaiah P.V. Oxidized LDL: diversity, patterns of recognition and pathophysiology.Antioxidants & Redox Signaling. 2010; 13: 39-75Crossref PubMed Scopus (313) Google Scholar, H. J. H. G. The role of lipid and in oxidative modification of Radic. Biol. Med. 13: PubMed Scopus Google Scholar, H. M. T. modified LDL is an oxidized LDL with oxidized of 2003; PubMed Scopus Google Scholar). LDL is oxidized a oxidized which oxidized to oxidized LDL, which is in oxysterols, as well in as and R. Keaney Jr, J.F. Role of oxidative modifications in atherosclerosis.Physiol. Rev. 2004; 84: 1381-1478Crossref PubMed Scopus (2105) Google Scholar, I. Volkov S. Subbaiah P.V. Oxidized LDL: diversity, patterns of recognition and pathophysiology.Antioxidants & Redox Signaling. 2010; 13: 39-75Crossref PubMed Scopus (313) Google Scholar, H. M. T. modified LDL is an oxidized LDL with oxidized of 2003; PubMed Scopus Google Scholar). the in oxLDL, oxysterols have shown to at all of atherosclerosis and an role in plaque G. B. S. G. oxidation in the vascular to Med. 2009; PubMed Scopus Google Scholar, W. and 1999; Full Text Full Text PDF PubMed Scopus Google Scholar). In the early stage of oxysterols are in of endothelial as endothelial V. G. A. P. of of oxidized oxidized in are of PubMed Scopus Google Scholar), G. B. B. V. P. G. of in by Redox 2006; PubMed Scopus Google Scholar), and of the S. B. endothelial dysfunction in 11: PubMed Scopus (41) Google Scholar, G. M. S. P. of of in and vascular endothelial cells by PubMed Scopus Google Scholar). In the stage of an increase in the of oxysterols shown to contribute to of vascular cells G. M. S. P. of of in and vascular endothelial cells by PubMed Scopus Google Scholar, of Opin. Lipidol. 2001; 12: PubMed Scopus Google Scholar) and of M. I. M. of of in to of atherosclerotic 1999; PubMed Scopus Google Scholar, S. B. P. G. G. oxysterols induce of in cells the of atherosclerotic Radic. Biol. Med. PubMed Scopus Google Scholar), which to of the In this study, we that oxysterols a major role in the increase in endothelial stiffness and identify oxysterols responsible for this effect. aortic endothelial cells and in and in a at with and with as I. Rothblat G.H. content of in endothelial 2000; 115: PubMed Scopus Google Scholar). 7-ketocholesterol, 7α-hydroxycholesterol, 7β-hydroxycholesterol, and plasma in the LDL by in the density to The is three of and at to to the to the to LDL with a of The oxidation at for and and by The content of in LDL and oxLDL by as with of of LDL In LDL oxidized by the with of at as by of low density lipoprotein to an oxidized and cytotoxic Lipid Res. 1991; Full Text PDF PubMed Google Scholar). The oxidation of LDL by the of by at a that in the studies of low density lipoprotein to an oxidized and cytotoxic Lipid Res. 1991; Full Text PDF PubMed Google Scholar, P.V. K. function of in Biol. 1999; Full Text Full Text PDF PubMed Scopus Google Scholar). by the and of lipid and J. PubMed Scopus Google Scholar) and on a with the of of and on a on the same in for the with a and the to The to of lipid and in of by the lipid The of by content The of J. PubMed Google Scholar, P.V. D. R. of in on the function of 2009; PubMed Scopus Google Scholar). by For LDL and oxLDL as an of the by with of at for The of by a with and as W. and during of low density lipoprotein and by Lipid Res. Full Text PDF PubMed Google Scholar). In a of The for at a of hold of and at for The and and with LDL or oxLDL of cells three with and and The by the and and the of to the For the to cells. and mice at of and on the to and and to a The by the and and the of by as in the to the for the and of by the of The in with the established by the and of the of at The elastic modulus of endothelial cells with a as I. Gooch Subbaiah P.V. M. I. oxLDL-induced in lipid of is with endothelial stiffness and J. 2010; PubMed Scopus Google Scholar, I. M. of during of J. Full Text Full Text PDF PubMed Scopus Google Scholar). with as The the the and the and at three at the and of cells for The by the at of the as it and the The the at a of a of which to or of the The and to the is the is is the to is the of the and is the elastic The to the a The are as of the elastic for a is in elastic modulus to to this not contribute to the in all on cells and cells to on the same and on the same of endothelial cells as in studies F.J. H. Rothblat G.H. I. increases stiffness of aortic endothelial J. 2004; Full Text Full Text PDF PubMed Scopus Google Scholar, F.J. I. Evidence for the role of stiffness in of 2006; PubMed Scopus Google Scholar). the with a and with for the with to a by a a with of LDL endothelial LDL oxidized with for of and to or oxidized LDL, as in studies W. and during of low density lipoprotein and by Lipid Res. Full Text PDF PubMed Google Scholar, H. D.P. Gotto Jr, A.M. Lipid and in oxidation of low density Lipid Res. 31: Full Text PDF PubMed Google Scholar, S. A. of oxysterol during in vitro oxidation of low density PubMed Scopus Google Scholar). The of oxidation by the of a of LDL oxidation J. of PubMed Scopus Google Scholar). we show that progressive LDL oxidation in increase in stiffness and increase in endothelial stiffness is the in the of the elastic in cells to oxLDL with cells to LDL as well as the increase in the elastic for cells to oxidized LDL a significant increase in endothelial elastic modulus is cells are to oxLDL with a of of which is as oxidized LDL of oxidative modification of low density lipoprotein the of to coronary artery PubMed Scopus Google Scholar, Witztum J.L. that and modified Thromb. Vasc. Biol. PubMed Scopus Google Scholar). an increase in LDL oxidation to the oxidation to in stiffening effect. in stiffness with LDL oxidation increase in endothelial stiffness is also cells are to LDL oxidized with of LDL oxidation that in oxidized LDL of low density lipoprotein to an oxidized and cytotoxic Lipid Res. 1991; Full Text PDF PubMed Google Scholar, P.V. K. function of in Biol. 1999; Full Text Full Text PDF PubMed Scopus Google Scholar, Witztum J.L. that and modified Thromb. Vasc. Biol. PubMed Scopus Google Scholar). In the endothelial stiffness by progressive into the an that in studies (18.Byfield F.J. Tikku S. Rothblat G.H. Gooch I. increases endothelial and Lipid Res. 2006; Full Text Full Text PDF PubMed Scopus Google Scholar, F.J. H. Rothblat G.H. I. increases stiffness of aortic endothelial J. 2004; Full Text Full Text PDF PubMed Scopus Google Scholar). increase in endothelial stiffness is the of LDL oxidation It is that the lipid of LDL and oxLDL into major of and of lipid of PubMed Scopus Google Scholar). lipid of the with the fractions by to the of the of PubMed Google Scholar). we this to which fractions induce the stiffening effect. In this of we oxidized LDL as stiffening these that the lipid of oxLDL is to and to and The of in as in and the effects of the lipid fractions on endothelial endothelial elastic modulus in cells to lipid for for of the fractions The fractions of lipid LDL as We show that the on endothelial stiffness cells to the oxidized by the of cells with elastic as in the of the In endothelial stiffening also in cells to In exposure of and fractions on endothelial stiffness to studies W. and during of low density lipoprotein and by Lipid Res. Full Text PDF PubMed Google Scholar), major oxysterols are identified in oxidized LDL of by the and and 7-ketocholesterol The oxysterol is 7β-hydroxycholesterol, by 7-ketocholesterol, with and oxysterols and The of oxysterols increase with progressive LDL oxidation with the significant increase in oxidized LDL In of oxysterols is accompanied with a in oxLDL increase in endothelial stiffness is with the of oxysterols which oxysterols induce an increase in cells to of of the oxysterols identified Our show that oxysterols found in oxLDL have differential effects on endothelial stiffness. 7-ketocholesterol and significantly endothelial stiffness a that not stiffening effects in cells to and the in the of oxysterols to induce endothelial stiffening associated with the in to into endothelial cells, we the level of incorporation of 7-ketocholesterol, an oxysterol that the strongest on endothelial with an oxysterol that effect. Our that in the of the incorporation of the two oxysterols into the cells In we the effects of two more oxysterols, and two oxysterols not significant of oxLDL found in atherosclerotic lesions W. and 1999; Full Text Full Text PDF PubMed Scopus Google Scholar, I. B. and for a role of this in of 1994; PubMed Scopus Google Scholar, M. J. V. I. of in atherosclerotic PubMed Scopus Google Scholar). is in atherosclerotic lesions and cells W. and 1999; Full Text Full Text PDF PubMed Scopus Google Scholar, I. B. and for a role of this in of 1994; PubMed Scopus Google Scholar, M. J. V. I. of in atherosclerotic PubMed Scopus Google Scholar). We show that exposure to in a significant increase of endothelial exposure to We have that oxLDL-induced increase in endothelial stiffness by the cells to a to the I. Gooch Subbaiah P.V. M. I. oxLDL-induced in lipid of is with endothelial stiffness and J. 2010; PubMed Scopus Google Scholar). to the hypothesis that the cells to the exchange and oxysterols into the and the cells the stiffening effect. In this study, we this hypothesis by the cells to of 7-ketocholesterol, an oxysterol that identified in as having a on endothelial and to the cells to 7-ketocholesterol in incorporation into the and this by the exposure of the cells to we the cells that to 7-ketocholesterol with also the stiffening of the We that this is the an increase in the elastic modulus in cells by the exposure to Endothelial stiffening by also by the impact of on endothelial we to the exposure to oxysterols the of oxysterols on endothelial stiffness. this cells to a that in enriching the cells with and to with the exposure to 7-ketocholesterol in a significant increase in endothelial elastic modulus, the and the of this effect. stiffening cells to 7-ketocholesterol that an increase in have a endothelial stiffening. we the of significant of oxidized is into the endothelial by the of oxysterols in of endothelial cells to oxLDL or to LDL as a Endothelial cells to to of oxidized LDL or to LDL for cells with to and for the of lipid The of oxysterols in the lipid by the that exposure to oxLDL in a significant increase in the level of 7-ketocholesterol in lipid of endothelial cells, as is by the of 7-ketocholesterol level in cells with oxLDL with cells with LDL also increase in the of and 7β-hydroxycholesterol, these increases not The in the of the three oxysterols the that exposure to oxLDL in a significant increase in 7-ketocholesterol not in and with studies (18.Byfield F.J. Tikku S. Rothblat G.H. Gooch I. increases endothelial and Lipid Res. 2006; Full Text Full Text PDF PubMed Scopus Google Scholar), we found change in the level of in cells to oxLDL. we the dyslipidemia in vivo also in the accumulation of oxysterols in the vascular of mouse aortas at the early stage of atherosclerosis development in ApoE−/− a for atherosclerosis I. Volkov S. Subbaiah P.V. Oxidized LDL: diversity, patterns of recognition and pathophysiology.Antioxidants & Redox Signaling. 2010; 13: 39-75Crossref PubMed Scopus (313) Google Scholar, J.L. R. mice lesions of all of atherosclerosis the Thromb. 1994; PubMed Google Scholar, J.L. models of PubMed Scopus Google Scholar). studies have shown that ApoE−/− mice as a for the early stage of atherosclerosis these mice are development of atherosclerosis is significantly with lesions cells and lesions J.L. R. mice lesions of all of atherosclerosis the Thromb. 1994; PubMed Google Scholar). In study, the of oxysterols in aortas to ApoE−/− mice a and mice of the same as The relative of of the oxysterols, as well as to the level of are shown in Our that in ApoE−/− is a significant increase in the two oxysterols that identified in this to induce endothelial 7-ketocholesterol and also increase in the of oxysterols the level of does not change and relative to as shown in however, that in to change in oxysterol in mice in the of plasma and of studies that endothelial major in endothelial function F.J. I. oxLDL of aortic endothelial J. Cell. PubMed Scopus Google Scholar, F.J. I. Evidence for the role of stiffness in of 2006; PubMed Scopus Google Scholar, C.S. M. S. of and PubMed Scopus Google Scholar, M. K. M. K. by for endothelial cells to shear 2000; PubMed Scopus Google Scholar, Gooch The relative of endothelial and stiffness in Res. 2004; PubMed Scopus Google H. H. de M. vascular endothelium and 104: PubMed Scopus Google Scholar). The that endothelial and however, are still poorly understood. Our recent studies have shown that endothelial elastic modulus is by oxidative modifications of LDL I. Gooch Subbaiah P.V. M. I. oxLDL-induced in lipid of is with endothelial stiffness and J. 2010; PubMed Scopus Google Scholar), a major lipoprotein associated with the development of atherosclerosis (1.Witztum J.L. Steinberg D. Role of oxidized low density lipoprotein in atherogenesis.J. Clin. Invest. 1991; 88: 1785-1792Crossref PubMed Scopus (2472) Google Scholar, 2.Witztum J.L. Steinberg D. The oxidative modification hypothesis of atherosclerosis: does it hold for humans?.Trends Cardiovasc. Med. 2001; 11: 93-102Crossref PubMed Scopus (387) Google Scholar, 3.Stocker R. Keaney Jr, J.F. Role of oxidative modifications in atherosclerosis.Physiol. Rev. 2004; 84: 1381-1478Crossref PubMed Scopus (2105) Google Scholar, 4.Libby P. Inflammation in atherosclerosis.Nature. 2002; 420: 868-874Crossref PubMed Scopus (6940) Google Scholar, 5.Paoletti R. Gotto A.M. Hajjar D.P. Inflammation in atherosclerosis and implications for therapy.Circulation. 2004; 109: III-20-III-26Crossref Google Scholar–6.Levitan I. Volkov S. Subbaiah P.V. Oxidized LDL: diversity, patterns of recognition and pathophysiology.Antioxidants & Redox Signaling. 2010; 13: 39-75Crossref PubMed Scopus (313) Google Scholar). oxidation of LDL is a that an of with and the on the of LDL The key are the critical that are responsible for oxLDL-induced The of to the impact of the of LDL oxidation on endothelial stiffness and to identify oxLDL that are critical for an increase in endothelial elastic the of oxLDL-induced effects the that the of LDL oxidation significantly or which in turn have major impact on effects (1.Witztum J.L. Steinberg D. Role of oxidized low density lipoprotein in atherogenesis.J. Clin. Invest. 1991; 88: 1785-1792Crossref PubMed Scopus (2472) Google Scholar, I. Volkov S. Subbaiah P.V. Oxidized LDL: diversity, patterns of recognition and pathophysiology.Antioxidants & Redox Signaling. 2010; 13: 39-75Crossref PubMed Scopus (313) Google Scholar, I. S. The role of oxidized low density lipoprotein in atherogenesis.J. PubMed Google Scholar). Our that stiffening is in to oxidized LDL, and the increases as the oxidation oxidized LDL that is by to to LDL and of is associated with the early of atherosclerosis H. M. T. modified LDL is an oxidized LDL with oxidized of 2003; PubMed Scopus Google Scholar). It also shown to induce in endothelial and Sevanian A. S. B. M. A.M. modified low density lipoprotein endothelial Clin. Invest. PubMed Scopus Google Scholar). the oxidative stress LDL and cytotoxic effects on endothelial cells and Oxidized and in atherosclerosis. for Cardiovasc. Med. 2001; 11: PubMed Scopus Google Scholar, H. A. M. H. H. T. S. of to the cytotoxic of oxidized low density Lipid Res. 1998; Full Text Full Text PDF PubMed Google Scholar). however, have of the effects of and oxidized LDL on the same Witztum J.L. in in to oxidized LDL and role of and Biol. Cell. 2003; PubMed Scopus Google Scholar) that and oxidized LDL in with a for oxidized In to study, show that endothelial stiffening increases with progressive LDL that oxLDL-induced endothelial stiffening at early and of disease The of the oxLDL that we found to responsible for oxLDL-induced endothelial stiffening to of oxidized oxysterols and with oxysterols having the dominant effect. Furthermore, we found a an increase in endothelial stiffness and the of oxysterols in oxLDL. are with oxidized LDL having more impact on endothelial stiffness with oxidized LDL as major oxidized H. M. T. modified LDL is an oxidized LDL with oxidized of 2003; PubMed Scopus Google Scholar), oxidized LDL and oxysterols W. and during of low density lipoprotein and by Lipid Res. Full Text PDF PubMed Google Scholar, H. D.P. Gotto Jr, A.M. Lipid and in oxidation of low density Lipid Res. 31: Full Text PDF PubMed Google Scholar). Furthermore, the of oxysterols, the stiffening is not by oxidized with the oxidized at having the effects. The same oxysterol shown to in atherosclerotic lesions W. and during of low density lipoprotein and by Lipid Res. Full Text PDF PubMed Google Scholar) and of oxidized in to on in Cardiovasc. Full Text Full Text PDF PubMed Scopus Google Scholar). We also show that endothelial stiffening is by which shown to the oxysterol in atherosclerotic lesions W. and 1999; Full Text Full Text PDF PubMed Scopus Google Scholar). that endothelial stiffening an factor in endothelial dysfunction in In of the we that exposure to oxLDL in a in lipid of and an increase in of the that is and I. Gooch Subbaiah P.V. M. I. oxLDL-induced in lipid of is with endothelial stiffness and J. 2010; PubMed Scopus Google Scholar). Furthermore, a in lipid of the in cells to 7-ketocholesterol I. Gooch Subbaiah P.V. M. I. oxLDL-induced in lipid of is with endothelial stiffness and J. 2010; PubMed Scopus Google Scholar). 7-ketocholesterol shown to with the major of a of of with 2001; Full Text Full Text PDF PubMed Scopus Google Scholar). the of these we that oxLDL-induced endothelial stiffening not the incorporation of oxysterols into the endothelial plasma In the study, we this by three oxysterols that induce endothelial the of endothelial stiffening by oxysterols with and that exposure to oxLDL in significant incorporation of oxysterols, 7-ketocholesterol, into the endothelial Our show that oxysterols have differential effects on endothelial stiffness that this to the in the of these oxysterols to into the is that to in oxysterols have differential effects on the lipid of the and in to the of however, are to this We also show that enriching the cells with to the exposure to oxysterols endothelial cells the of oxysterols on endothelial stiffness. that enriching endothelial cells with by on endothelial that the of stiffening the of to a of the two effects. We that the of to the incorporation of into the incorporation of oxysterols into endothelial endothelial and is to to endothelial function, the level of and In of the presence of oxysterols in studies have shown that oxysterols are found in vascular of ApoE−/− mice with as well as in atherosclerotic lesions W. and 1999; Full Text Full Text PDF PubMed Scopus Google Scholar, J. M. S. T. A. S. of oxysterols in atherosclerotic lesions and plasma Radic. Res. 2001; PubMed Scopus Google Scholar). Our studies show that a significant accumulation of oxysterols found in aortic of ApoE−/− mice a at a early stage of disease of lesions are J.L. R. mice lesions of all of atherosclerosis the Thromb. 1994; PubMed Google Scholar). we show oxysterol accumulation in ApoE−/− change in in ApoE−/− The and to the level of in the plasma it to the in oxysterol accumulation and at the early of atherosclerosis a significant studies have shown that are significantly more to the development of atherosclerosis T.M. and vascular in atherosclerotic the of 2010; PubMed Scopus Google Scholar, M.E. in the cardiovascular of Rev. Cardiol. 2009; PubMed Scopus Google Scholar). We that accumulation of the oxysterols in the vascular in stiffening of endothelial cells and that this plays an role in the of the The for in the with aortic endothelial oxidized LDL oxidized LDL oxidized