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(Circulation. 2003;107:1520.)
© 2003 American Heart Association, Inc.

Basic Science Reports

L-4F, an Apolipoprotein A-1 Mimetic, Restores Nitric Oxide and Superoxide Anion Balance in Low-Density Lipoprotein-Treated Endothelial Cells

Zhijun Ou, MD; Jingsong Ou, MD, PhD^*; Allan W. Ackerman, MS; Keith T. Oldham, MD; Kirkwood A. Pritchard, Jr, PhD^*

From the Department of Surgery (Z.O., J.O., A.W.A., K.T.O., K.A.P.), the Division of Pediatric Surgery and Pharmacology & Toxicology (K.A.P.), and the Cardiovascular Center (Z.O., J.O., K.T.O., K.A.P.) and Free Radical Research Center (Z.O., J.O., K.T.O., K.A.P.), Medical College of Wisconsin, Children’s Hospital of Wisconsin, Milwaukee.

Correspondence to Kirkwood A. Pritchard Jr, PhD, Medical College of Wisconsin, 8701 Watertown Plank Rd, Milwaukee, WI 53226. E-mail kpritch{at}mcw.edu

	Abstract

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Background— Low-density lipoprotein (LDL) impairs endothelial cell function by uncoupling endothelial nitric oxide synthase (eNOS) activity, which allows superoxide anion (O₂^·-) to be generated rather than nitric oxide (·NO). Recent reports indicate that apolipoprotein (apo) A-1 mimetics inhibit the development of atherosclerotic lesions in LDL receptor-null mice. Here we hypothesize that L-4F, an apoA-1 mimetic that inhibits atherosclerosis induced by hypercholesterolemia, protects endothelial cell function by preventing LDL from uncoupling eNOS activity.

Methods and Results— Bovine aortic endothelial cells were incubated with LDL±L-4F, and changes in A23187-stimulated ·NO and O₂^·- generation were determined by ozone chemiluminescence and superoxide dismutase-inhibitable ferricytochrome c reduction, respectively. Western analysis of eNOS immunoprecipitates was used to determine effects of LDL and L-4F on heat shock protein 90 (hsp90) interactions with eNOS. LDL decreased ·NO production and increased eNOS-dependent O₂^·- generation. Pretreatment of LDL with L-4F increased ·NO and decreased O₂^·- generation. By itself, L-4F had no effect on O₂^·- but did increase ·NO generation. Stimulation of endothelial cells incubated with LDL decreased the association of hsp90 with eNOS. Pretreatment of LDL with L-4F prevented a decrease in hsp90 association with eNOS and often enhanced association on stimulation.

Conclusions— These data demonstrate that L-4F protects endothelial cell function by preventing LDL from uncoupling eNOS activity. L-4F allows endothelial cell to maintain coupled eNOS activity to generate ·NO even in the face of atherogenic concentrations of LDL.

Key Words: apolipoproteins • cells, endothelial • lipoproteins • nitric oxide • nitric oxide synthase

	Introduction

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Endothelial cell dysfunction is hypothesized to play a causal role in the premature development of atherosclerosis.^1,2 Previously, our laboratory and others have demonstrated that low-density lipoprotein (LDL) induces endothelial cell dysfunction by increasing superoxide anion (O₂^·-) generation.^3–6 As the balance of nitric oxide (·NO) and O₂^·- has major implications for vascular function, not only with respect to atherosclerosis,⁷ but also vasodilation,⁸ it is essential that we investigate alternative mechanisms for protecting endothelial cell function in the face of atherogenic challenges.

Several clinical trials suggest that increasing high-density lipoprotein (HDL) improves vascular function to inhibit atherosclerosis.^9–11 HDL appears to protect vascular function by a number of mechanisms. In vitro, HDL inhibits LDL oxidation^12–14 and LDL-induced monocyte chemotactic activity.^13,14 Intravenous infusion of HDL rapidly increases forearm blood flow providing proof that HDL improves endothelial- and endothelial nitric oxide synthase (eNOS)-dependent vascular function.¹⁵ Ironically, hypercholesterolemia and inflammation inhibit HDL function with respect to LDL oxidation or monocyte chemotactic activity.^16,17 Such findings support the concept that it is the failure of HDL to perform its critical duties that allows atherogenic mechanisms induced by LDL to proceed unchecked.

Experimental evidence indicates that HDL function can be enhanced also by apoA-1 mimetics. Intraperitoneal injection of an apoA-1 mimetic (5F) and parental administration of another apoA-1 mimetic (D-4F) enhance the ability of HDL to inhibit LDL oxidation and protect mice from diet-induced atherosclerosis.^14,18 D-4F restores antiinflammatory properties of HDL that are lost during influenza infection.¹⁷ On the basis of these reports, we reasoned that L-4F, another apoA-1 mimetic, might protect endothelial cell function from the atherogenic effects of LDL.

Here we investigate whether L-4F protects against LDL-induced increases in endothelial cell O₂^·- generation. We find that L-4F inhibits LDL-induced increases in stimulated O₂^·- production and restores coupled eNOS activity to increase ·NO production, at least in part by restoring and increasing heat shock protein 90 (hsp90) interactions with eNOS.

	Methods

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Synthesis of L-4F
L-4F (Ac-DWFKAFYDKVAEKFKEAFNH₂) was synthesized by the Protein & Nucleic Acid Share Facility of the Medical College of Wisconsin in Milwaukee. Purity (typically >98%) and structure were assessed by GC/MS.

Endothelial Cells
Bovine aortic endothelial cells (BAECs) were purchased from VEC Technologies (Rensselaer, NY). BAECs were initially cultured and expanded in MCDB-131C from VEC Technologies (passage 0 to 1) and gradually conditioned over 4 days so that they could be expanded and maintained in RPMI 1640 media containing 20% FBS (SH30070, HyClone).

LDL
LDL (1.019 to 1.063 g/mL) was isolated by sequential density ultracentrifugation with density adjustments made by the addition of solid KBr as described.⁵ For the incubations here, LDL was used at 6.2 mmol/L (240 mg/dL), and pretreatment of LDL was made by adding L-4F (10 µg/mL, final concentration) to the media for 30 minutes before incubating with endothelial cell cultures.

Stimulated Endothelial Cell O₂^·- Generation
Superoxide dismutase (SOD)-inhibitable ferricytochrome c reduction based on molar extinction coefficient (=21 000 mol/L^-1 · cm^-1) was used to calculate stimulated endothelial cell O₂^·- generation. After experimental treatments with LDL (6.2 mmol/L [240 mg/dL]) and/or L-4F (10 µg/mL) for 24 hours, endothelial cell cultures were washed 3 times with Hanks Balanced Salts Solution (HBSS) and stimulated in HBSS containing A23187 (5 µmol/L). To quantify eNOS-dependent O₂^·- generation, cultures were incubated with L-nitroargininemethylester (L-NAME, 1 mmol/L) 30 minutes before and during incubation with ferricytochrome c (50 µmol/L). Measurements of ferricytochrome c reduction were performed in triplicate ±SOD (1000 U/mL) and cell proteins determined in duplicate as described.¹⁹

Endothelial Cell ·NO Generation
Endothelial cell nitrite + nitrate production was determined by ozone chemiluminescence with VCl₃ as described.¹⁹ After the endothelial cell cultures were incubated with L-4F (10 µg/mL) for either 30 minutes or 24 hours, they were washed (3x) with HBSS and then incubated with HBSS containing L-arginine (25 µmol/L, basal activity) or with HBSS containing L-arginine (25 µmol/L) and A23187 (5 µmol/L) (stimulated activity) for 30 minutes. Each experiment was performed in triplicate; ·NO measurements were made in duplicate, and cell proteins determined in duplicate as described.¹⁹

Western Analysis
Hsp90 interactions with eNOS and phosphorylation of eNOS (S1179) were determined as described.^4,5 Briefly, after experimental treatments, cultures were washed with HBSS and cells lysed in modified RIPA buffer.²⁰ eNOS was immunoprecipitated with H32 (SH-258, BioMol) 1-µg antibody per 100-µg cell lysate (300 to 500 µg total) as described.^4,5 Immunoblots were visualized by enhanced chemiluminescence with Immunostar reagents (Biorad).

Statistical Analysis
Data were analyzed by Student’s t test for experiments with 2 groups and ANOVA with a Newman-Keul’s test as a post-hoc test for experiments with more than 2 groups for determining levels of significance. Minimum levels of significance were set at P<0.05.

	Results

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Incubation of BAEC cultures to L-4F (30 minutes) enhanced stimulated ·NO generation (Figure 1A). Incubation of cultures with L-4F (24 hours) enhanced basal and stimulated ·NO generation (Figure 1B) without altering stimulated O₂^·- generation (Figure 1C). Short-term incubation with LDL (6.2 mmol/L, [240 mg cholesterol/dL], 24 h) attenuated basal and stimulated ·NO generation (Figure 2). Pretreatment of LDL with L-4F (10 µg/mL, 30 minutes) before incubation with the cultures, however, enhanced basal and stimulated ·NO generation (Figure 3A) and attenuated stimulated O₂^·- generation (Figure 3B). Previous studies showed that short-term incubation (24 hours) of isolated arterial segments with LDL increased endothelial O₂^·- generation by a predominantly eNOS-dependent mechanism.⁴ L-NAME inhibition of stimulated O₂^·- generation in LDL-treated cultures confirms that LDL uncouples eNOS activity (Figure 3B). Pretreatment of LDL with L-4F markedly decreased stimulated O₂^·- generation (Figure 3B). The observation that L-NAME did not further decrease O₂^·- in LDL+L-4F cultures is consistent with the idea that eNOS activity in the LDL+L-4F cultures is coupled.

View larger version (21K): [in this window] [in a new window]   Figure 1. L-4F enhances stimulated ·NO generation. A, Acute exposure of BAEC culture with L-4F (10 µg/mL, 30 minutes) increases A23187 (5 µmol/L, 30 minutes)-stimulated ·NO generation. B, Exposure of BAEC cultures with L-4F (10 µg/mL, 24 hours) increases basal and A23187 (5 µmol/L, 30 minutes)-stimulated ·NO generation. C, Under similar conditions as in B, L-4F had no effect on A23187-stimulated O2·- generation and L-NAME (1 mmol/L) increased stimulated O2·- generation indicating that eNOS activity, in BAEC cultures treated with L-4F, is coupled. *P<0.05; **P<0.01; n=4 to 6.

View larger version (18K): [in this window] [in a new window]   Figure 2. LDL inhibits endothelial cell ·NO generation. BAEC cultures incubated with LDL (6.2 mmol/L [240 mg/dL], 24 hours) generate less nitrite + nitrate than control cultures under basal and A23187-stimulated conditions [5 µmol/L, 30 minutes]. **P<0.01, n=5 to 8.

View larger version (26K): [in this window] [in a new window]   Figure 3. L-4F improves coupled eNOS activity. A, L-4F (10 µg/mL, 24 h) increases ·NO generation in LDL-treated cultures under basal and stimulated conditions. B, As in A, L-4F decreased LDL-induced increases in stimulated O2·- generation. L-NAME decreased stimulated O2·- generation in LDL-treated cultures. L-NAME did not further decrease stimulated O2·- generation in cultures incubated with LDL that was pretreated with L-4F. These data demonstrate that LDL uncouples stimulated eNOS activity that is prevented by pretreating LDL with L-4F. **P<0.01, n=5 to 6.

Numerous reports indicate that an increase in the association of hsp90 with eNOS plays an important role in increasing ·NO generation.^19–23 L-4F had modest effects on the levels of phospho-eNOS (S1179) on eNOS isolated by immunoprecipitation from control cultures under basal and stimulated conditions (Figure 4, top panel, lane 2 versus lane 1 and lane 4 versus lane 3, respectively). L-4F appears to increase hsp90 association with eNOS under basal conditions (Figure 4, bottom panel, lane 2 versus lane 1). Stimulation markedly increases hsp90 association with eNOS (Figure 4, bottom panel, lane 3 versus lane 1), which was slightly reduced in cultures incubated with L-4F (Figure 4, bottom panel, lane 4 versus lane 3).

View larger version (37K): [in this window] [in a new window]   Figure 4. L-4F increases hsp90 interactions with eNOS in LDL-treated BAEC cultures. Representative images of Western analysis for phospho-eNOS (S1179) and hsp90 association with eNOS (isolated by immunoprecipitation) show that L-4F restores hsp90-dependent signal transduction mechanisms in LDL-treated BAEC cultures. In control cultures, low levels of hsp90 are associated with eNOS (lane 1). A23187 stimulation markedly increases in hsp90 association with eNOS (lane 3). Under basal conditions, LDL slightly increases hsp90 interactions with eNOS (lane 5 versus lane 1); however, on stimulation, LDL decreases hsp90 association with eNOS (lane 7 versus lane 3). Pretreatment of the LDL with L-4F restores hsp90 interactions with eNOS when LDL-treated cultures are stimulated (lane 8 versus lane 3). The bar graphs represent mean±SEM of changes in band densities of autoradiograms relative to band densities of the control (lane 1) set at 1.0. C, Control BAEC cultures (n= 10) and LDL cultures (n=6).

Previously, we reported that under basal conditions, long-term LDL exposure (4 days) increases uncoupled eNOS activity by increasing phospho-eNOS (S1179) levels on eNOS and decreasing hsp90 interactions on stimulation.⁵ The salient features of the blots in Figure 4 are that A23187 stimulation increases phospho-eNOS (S1179) and hsp90 association with eNOS (lane 3 versus lane 1); that LDL decreases hsp90 association with eNOS in A23187-stimulated cultures (bottom panel, lane 7 versus lane 3) without inhibiting phosphorylation (top panel, lane 7 versus lane 3); and that pretreatment of LDL with L-4F restores hsp90 interactions with eNOS (bottom panel, lane 8 versus lane 3) in addition to slightly increasing phosphorylation of eNOS (top panel, lane 8 versus lane 3). These data show that stimulation of LDL-treated cultures decreased hsp90 interactions with eNOS compared with A23187-stimulated control cultures, confirming previous findings.⁴ The marked increase in hsp90 association with eNOS in the LDL + L-4F-treated cultures suggest that L-4F helps endothelial cells maintain coupled eNOS activity by enhancing hsp90 interactions in the face of an LDL-induced atherogenic challenge.

	Discussion

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In this report we show that L-4F increases endothelial cell ·NO generation under basal and stimulated conditions and decreases LDL-induced increases in endothelial cell O₂^·- generation. Western analysis of eNOS immunoprecipitates reveals that one of the mechanisms by which this apoA-1 mimetic increases ·NO generation is by restoring hsp90 interactions with eNOS in LDL-treated endothelial cells. Stimulation of LDL-treated endothelial cell cultures decreased hsp90 association with eNOS compared with stimulated controls (Figure 4, lane 7 versus lane 3), confirming previous reports.⁴ However, when LDL was pretreated with L-4F and then incubated with cultures, hsp90 interactions with eNOS were maintained under basal conditions and enhanced under stimulated conditions. Thus, L-4F protects endothelial cell function against LDL, at least in part, by increasing hsp90 interactions with eNOS, an essential step in the mechanisms by which endothelial cells generate ·NO.^19–26 In spite of the fact that L-4F increases ·NO and decreases O₂^·- generation in LDL-treated cultures, the mechanisms by which this apoA-1 mimetic restores eNOS function remain unclear. For example, when control cultures are stimulated with A23187, L-4F slightly decreases the association of hsp90 with eNOS, yet increases stimulated ·NO generation. Such discrepancies suggest that alternative mechanisms are governing eNOS activity and function beyond hsp90. As L-4F was designed to mimic apoA-1 and increase HDL function, it is possible that this mimetic may be activating eNOS by the scavenger receptor, a ceramide-dependent and calcium-independent mechanism.²⁷ Regardless, these data demonstrate that L-4F is highly effective at protecting endothelial cell function against LDL-induced shifts in ·NO and O₂^·- balance.

One of the earliest events in vascular disease is a loss in ·NO activity. Decreases in ·NO activity appear to develop before structural changes in the vessel wall.²⁸ On the basis that ·NO is a chain-breaking antioxidant that inhibits LDL oxidation,²⁹ platelet aggregation,³⁰ and smooth muscle cell proliferation,³¹ it is easy to understand how a loss in ·NO activity increases atherosclerosis. If the underlying atherogenic mechanisms cannot be corrected, then logically providing ·NO may help prevent vascular disease. Indeed, this approach is effective based on recent reports showing that ·NO donors inhibited atherosclerosis in hypercholesterolemic mice³² and decreased inflammation of the gastrointestinal tract in a murine model of inflammatory bowel disease.³³

However, if the underlying defect in endothelial cell function could be corrected, then the atherogenic effects of LDL should be eliminated or at least minimized. If LDL induces endothelial cell dysfunction by increasing O₂^·- generation, which inactivates ·NO^3–5 and HDL improves endothelial- and eNOS-dependent forearm blood flow,¹⁵ then L-4F, an apoA-1 mimetic, should prevent LDL-induced increases in endothelial cell O₂^·- generation. Our findings that L-4F preserves ·NO balance by preventing LDL-induced increases in uncoupled eNOS activity are consistent with the notion that HDL function is essential for protecting endothelial cell function¹⁵ and that apoA-1 mimetics can be used to enhance HDL function to increase atheroprotection, as was shown earlier.^14,18,34 Although L-4F prevented LDL from uncoupling eNOS, we also observed that it markedly increased ·NO production in control cultures after just 30 minutes incubation, with no other intervention (Figure 1A). Mechanistically, we find this effect of L-4F on eNOS ·NO generation in control cultures to be as exciting as the observation that L-4F restores ·NO balance in LDL-treated endothelial cell cultures, in that it suggests that even under the best culture conditions, negative regulators of eNOS ·NO generation are present. This idea is reinforced by findings that L-4F increased ·NO but had no effect on O₂^·- generation in control cultures in contrast to the effects of the mimetic on LDL-treated cultures in which L-4F increased ·NO and decreased O₂^·- generation.

The mechanisms by which L-4F improves endothelial cell ·NO generation remain unclear at this time. L-4F inhibits LDL oxidation, confirming previous reports with D-4F^14,18 (Data Supplement Figure). Yet, in vitro, its ability to inhibit LDL oxidation is modest compared with the antioxidant properties of probucol or BHT.³⁵ Such differences suggest that although oxidative stress induces endothelial cell dysfunction, L-4F is doing more than simply inhibiting LDL oxidation. Possibly, L-4F partitions or removes proinflammatory oxidized lipids³⁴ such that they are no longer available to negatively regulate eNOS activity. Future studies are required to determine whether such mechanisms are involved in how L-4F increases coupled eNOS activity to shift the balance of ·NO and O₂^·- generation toward ·NO.

	Acknowledgments

 
This study was supported in part by the Marie Z. Uihlein Endowed Chair Award, the Children’s Hospital Foundation (Milwaukee, Wis) (K.T.O.), and the National Heart, Lung, and Blood Institute (HL 61417 and HL 71214, K.A.P.).

	Footnotes

 
These authors contributed equally to the experimental design of the study.

This article originally appeared Online on March 17, 2003 (Circulation. 2003;107:r48–r52).

The online-only Data Supplement Figure is available at http://www.circulationaha.org.

Received January 22, 2003; accepted February 3, 2003.

	References

Top Abstract Introduction Methods Results Discussion References

 
1. Cohen RA, Zitnay KM, Haudenschild CC, et al. Loss of selective endothelial cell vasoactive functions caused by hypercholesterolemia in pig coronary arteries. Circ Res. 1988; 63: 903–910.[Abstract/Free Full Text]

2. Munro JM, Cotran RS. The pathogenesis of atherosclerosis: atherogenesis and inflammation. Lab Invest. 1988; 58: 249–261.[Medline] [Order article via Infotrieve]

3. Pritchard Jr KA, Groszek L, Smalley DM, et al. Low density lipoprotein increases endothelial cell nitric oxide synthase generation of superoxide anion. Circ Res. 1995; 77: 510–518.[Abstract/Free Full Text]

4. Stepp DW, Ou J, Ackerman AW, et al. Native LDL and minimally oxidized LDL differentially regulate superoxide anion in vascular endothelium in situ. Am J Physiol Heart Circulation. 2002; 283: H750–H759.[Abstract/Free Full Text]

5. Pritchard KA Jr, Ackerman AW, Ou J, et al. Native low-density lipoprotein induces endothelial nitric oxide synthase dysfunction: role of heat shock protein 90 and caveolin-1. Free Radic Biol Med. 2002; 33: 52–62.[CrossRef][Medline] [Order article via Infotrieve]

6. Vergnani L, Hatrik S, Ricci F, et al. Effect of native and oxidized low-density lipoprotein on endothelial nitric oxide and superoxide production: key role of L-arginine availability. Circulation. 2000; 101: 1261–1266.[Abstract/Free Full Text]

7. White CR, Darley-Usmar V, Berrington WR, et al. Circulating plasma xanthine oxidase contributes to vascular dysfunction in hypercholesterolemic rabbits. Proc Natl Acad Sci U S A. 1996; 93: 8745–8749.[Abstract/Free Full Text]

8. Miller FJ Jr, Gutterman DD, Rios CD, et al. Superoxide production in vascular smooth muscle contributes to oxidative stress and impaired relaxation in atherosclerosis. Circ Res. 1998; 82: 1298–1305.[Abstract/Free Full Text]

9. Ericsson CG, Nilsson J, Grip L, et al. Effect of bezafibrate treatment over five years on coronary plaques causing 20% to 50% diameter narrowing: the Bezafibrate Coronary Atherosclerosis Intervention Trial (BECAIT). Am J Cardiol. 1997; 80: 1125–1129.[CrossRef][Medline] [Order article via Infotrieve]

10. Frick MH, Syvanne M, Nieminen MS, et al. Prevention of the angiographic progression of coronary and vein-graft atherosclerosis by gemfibrozil after coronary bypass surgery in men with low levels of HDL cholesterol: the Lopid Coronary Angiography Trial (LOCAT) Study Group. Circulation. 1997; 96: 2137–2143.[Abstract/Free Full Text]

11. Secondary prevention by raising HDL cholesterol and reducing triglycerides in patients with coronary artery disease: the Bezafibrate Infarction Prevention (BIP) study. Circulation. 2000; 102: 21–27.[Abstract/Free Full Text]

12. Parthasarathy S, Barnett J, Fong LG. High density lipoprotein inhibits the oxidative modification of low density lipoprotein. Biochim Biophys Acta. 1990; 1044: 275–283.[Medline] [Order article via Infotrieve]

13. Navab M, Berliner JA, Subbanagounder G, et al. HDL and the inflammatory response induced by LDL-derived oxidized phospholipids. Arterioscler Thromb Vasc Biol. 2001; 21: 481–488.[Abstract/Free Full Text]

14. Garber DW, Datta G, Chaddha M, et al. A new synthetic class A amphipathic peptide analogue protects mice from diet-induced atherosclerosis. J Lipid Res. 2001; 42: 545–552.[Abstract/Free Full Text]

15. Spieker LE, Sudano I, Hurlimann D, et al. High-density lipoprotein restores endothelial function in hypercholesterolemic men. Circulation. 2002; 105: 1399–1402.[Abstract/Free Full Text]

16. Van Lenten BJ, Hama SY, de Beer FC, et al. Anti-inflammatory HDL becomes pro-inflammatory during the acute phase response: loss of protective effect of HDL against LDL oxidation in aortic wall cell cocultures. J Clin Invest. 1995; 96: 2758–2767.[Medline] [Order article via Infotrieve]

17. Van Lenten BJ, Wagner AC, Anantharamaiah GM, et al. Influenza infection promotes macrophage traffic into arteries of mice that is prevented by D-4F, an apolipoprotein A-I mimetic peptide. Circulation. 2002; 106: 1127–1132.[Abstract/Free Full Text]

18. Navab M, Anantharamaiah GM, Hama S, et al. Oral administration of an Apo A-I mimetic Peptide synthesized from D-amino acids dramatically reduces atherosclerosis in mice independent of plasma cholesterol. Circulation. 2002; 105: 290–292.[Abstract/Free Full Text]

19. Ou J, Ou Z, Ackerman AW, et al. Inhibition of heat shock protein 90 (hsp90) in proliferating endothelial cells uncouples endothelial nitric oxide synthase activity. Free Radic Biol Med. 2003; 34: 269–276.[CrossRef][Medline] [Order article via Infotrieve]

20. Garcia-Cardena G, Fan R, Shah V, et al. Dynamic activation of endothelial nitric oxide synthase by Hsp90. Nature. 1998; 392: 821–824.[CrossRef][Medline] [Order article via Infotrieve]

21. Feron O, Dessy C, Desager J.-P., et al. Hydroxy-methylglutaryl-coenzyme a reductase inhibition promotes endothelial nitric oxide synthase activation through a decrease in caveolin abundance. Circulation. 2001; 103: 113–118.[Abstract/Free Full Text]

22. Fontana J, Fulton D, Chen Y, et al. Domain mapping studies reveal that the M domain of hsp90 serves as a molecular scaffold to regulate Akt-dependent phosphorylation of endothelial nitric oxide synthase and NO release. Circ Res. 2002; 90: 866–873.[Abstract/Free Full Text]

23. Shi Y, Baker JE, Zhang C, et al. Chronic hypoxia increases endothelial nitric oxide synthase generation of nitric oxide by increasing heat shock protein 90 association and serine phosphorylation. Circ Res. 2002; 91: 300–306.[Abstract/Free Full Text]

24. Russell KS, Haynes MP, Caulin-Glaser T, et al. Estrogen stimulates heat shock protein 90 binding to endothelial nitric oxide synthase in human vascular endothelial cells: effects on calcium sensitivity and NO release. J Biol Chem. 2000; 275: 5026–5030.[Abstract/Free Full Text]

25. Bucci M, Roviezzo F, Cicala C, et al. Geldanamycin, an inhibitor of heat shock protein 90 (Hsp90) mediated signal transduction has anti-inflammatory effects and interacts with glucocorticoid receptor in vivo [In Process Citation]. Br J Pharmacol. 2000; 131: 13–16.[CrossRef][Medline] [Order article via Infotrieve]

26. Brouet A, Sonveaux P, Dessy C, et al. Hsp90 ensures the transition from the early Ca2 +-dependent to the late phosphorylation-dependent activation of the endothelial nitric-oxide synthase in vascular endothelial growth factor-exposed endothelial cells. J Biol Chem. 2001; 276: 32663–32669.[Abstract/Free Full Text]

27. Li XA, Titlow WB, Jackson BA, et al. High-density lipoprotein binding to scavenger receptor, Class B, type I activates endothelial nitric-oxide synthase in a ceramide-dependent manner. J Biol Chem. 2002; 277: 11058–11063.[Abstract/Free Full Text]

28. Tsao PS, McEvoy LM, Drexler H, et al. Enhanced endothelial adhesiveness in hypercholesterolemia is attenuated by L-arginine. Circulation. 1994; 89: 2176–2182.[Abstract/Free Full Text]

29. Goss SP, Hogg N, Kalyanaraman B. The effect of nitric oxide release rates on the oxidation of human low-density lipoprotein. J Biol Chem. 1997; 272: 21647–21653.[Abstract/Free Full Text]

30. Radomski MW, Palmer RM, Moncada S. Comparative pharmacology of endothelium-derived relaxing factor, nitric oxide and prostacyclin in platelets. Br J Pharmacol. 1987; 92: 181–187.[Medline] [Order article via Infotrieve]

31. Garg UC, Hassid A. Nitric oxide-generating vasodilators and 8-bromo-cyclic guanosine monophosphate inhibit mitogenesis and proliferation of cultured rat vascular smooth muscle cells. J Clin Invest. 1989; 83: 1774–1777.[Medline] [Order article via Infotrieve]

32. Napoli C, Ackah E, De Nigris F, et al. Chronic treatment with nitric oxide-releasing aspirin reduces plasma low-density lipoprotein oxidation and oxidative stress, arterial oxidation-specific epitopes, and atherogenesis in hypercholesterolemic mice. Proc Natl Acad Sci U S A. 2002; 99: 12467–12470.[Abstract/Free Full Text]

33. Fiorucci S, Antonelli E, Distrutti E, et al. NCX-1015, a nitric-oxide derivative of prednisolone, enhances regulatory T cells in the lamina propria and protects against 2,4,6-trinitrobenzene sulfonic acid-induced colitis in mice. Proc Natl Acad Sci U S A. 2002; 99: 15770–15775.[Abstract/Free Full Text]

34. Navab M, Hama SY, Ready ST, et al. Oxidized lipids as mediators of coronary heart disease. Curr Opin Lipidol. 2002; 13: 363–372.[CrossRef][Medline] [Order article via Infotrieve]

35. Kalyanaraman B, Darley-Usmar VM, Wood J, et al. Synergistic interaction between the probucol phenoxyl radical and ascorbic acid in inhibiting the oxidation of low-density lipoprotein. J Biol Chem. 1992; 267: 6789–6795.[Abstract/Free Full Text]

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				S. J. Peterson, D. Husney, A. L. Kruger, R. Olszanecki, F. Ricci, L. F. Rodella, A. Stacchiotti, R. Rezzani, J. A. McClung, W. S. Aronow, et al. Long-Term Treatment with the Apolipoprotein A1 Mimetic Peptide Increases Antioxidants and Vascular Repair in Type I Diabetic Rats J. Pharmacol. Exp. Ther., August 1, 2007; 322(2): 514 - 520. [Abstract] [Full Text] [PDF]

				N. M. Gharavi, P. S. Gargalovic, I. Chang, J. A. Araujo, M. J. Clark, W. L. Szeto, A. D. Watson, A. J. Lusis, and J. A. Berliner High-Density Lipoprotein Modulates Oxidized Phospholipid Signaling in Human Endothelial Cells From Proinflammatory to Anti-inflammatory Arterioscler. Thromb. Vasc. Biol., June 1, 2007; 27(6): 1346 - 1353. [Abstract] [Full Text] [PDF]

				A. Kontush and M. J. Chapman Functionally Defective High-Density Lipoprotein: A New Therapeutic Target at the Crossroads of Dyslipidemia, Inflammation, and Atherosclerosis Pharmacol. Rev., September 1, 2006; 58(3): 342 - 374. [Abstract] [Full Text] [PDF]

				C. Mineo, H. Deguchi, J. H. Griffin, and P. W. Shaul Endothelial and Antithrombotic Actions of HDL Circ. Res., June 9, 2006; 98(11): 1352 - 1364. [Abstract] [Full Text] [PDF]

				M. J Jackson Reactive oxygen species and redox-regulation of skeletal muscle adaptations to exercise Phil Trans R Soc B, December 29, 2005; 360(1464): 2285 - 2291. [Abstract] [Full Text] [PDF]

				M. Navab, G.M. Anantharamaiah, and A. M. Fogelman An Apolipoprotein A-I Mimetic Works Best in the Presence of Apolipoprotein A-I Circ. Res., November 25, 2005; 97(11): 1085 - 1086. [Full Text] [PDF]

				J. Ou, J. Wang, H. Xu, Z. Ou, M. G. Sorci-Thomas, D. W. Jones, P. Signorino, J. C. Densmore, S. Kaul, K. T. Oldham, et al. Effects of D-4F on Vasodilation and Vessel Wall Thickness in Hypercholesterolemic LDL Receptor-Null and LDL Receptor/Apolipoprotein A-I Double-Knockout Mice on Western Diet Circ. Res., November 25, 2005; 97(11): 1190 - 1197. [Abstract] [Full Text] [PDF]

				M. Navab, G.M. Anantharamaiah, S. Hama, G. Hough, S. T. Reddy, J. S. Frank, D. W. Garber, S. Handattu, and A. M. Fogelman D-4F and Statins Synergize to Render HDL Antiinflammatory in Mice and Monkeys and Cause Lesion Regression in Old Apolipoprotein E-Null Mice Arterioscler. Thromb. Vasc. Biol., July 1, 2005; 25(7): 1426 - 1432. [Abstract] [Full Text] [PDF]

				A. L. Kruger, S. Peterson, S. Turkseven, P. M. Kaminski, F. F. Zhang, S. Quan, M. S. Wolin, and N. G. Abraham D-4F Induces Heme Oxygenase-1 and Extracellular Superoxide Dismutase, Decreases Endothelial Cell Sloughing, and Improves Vascular Reactivity in Rat Model of Diabetes Circulation, June 14, 2005; 111(23): 3126 - 3134. [Abstract] [Full Text] [PDF]

				H. Gupta, C. R. White, S. Handattu, D. W. Garber, G. Datta, M. Chaddha, L. Dai, S. H. Gianturco, W. A. Bradley, and G.M. Anantharamaiah Apolipoprotein E Mimetic Peptide Dramatically Lowers Plasma Cholesterol and Restores Endothelial Function in Watanabe Heritable Hyperlipidemic Rabbits Circulation, June 14, 2005; 111(23): 3112 - 3118. [Abstract] [Full Text] [PDF]

				M. Toporsian, R. Gros, M. G. Kabir, S. Vera, K. Govindaraju, D. H. Eidelman, M. Husain, and M. Letarte A Role for Endoglin in Coupling eNOS Activity and Regulating Vascular Tone Revealed in Hereditary Hemorrhagic Telangiectasia Circ. Res., April 1, 2005; 96(6): 684 - 692. [Abstract] [Full Text] [PDF]

				R. M. Epand, R. F. Epand, B. G. Sayer, G. Datta, M. Chaddha, and G. M. Anantharamaiah Two Homologous Apolipoprotein AI Mimetic Peptides: RELATIONSHIP BETWEEN MEMBRANE INTERACTIONS AND BIOLOGICAL ACTIVITY J. Biol. Chem., December 3, 2004; 279(49): 51404 - 51414. [Abstract] [Full Text] [PDF]

				X. Li, K.-Y. Chyu, J. R. F. Neto, J. Yano, N. Nathwani, C. Ferreira, P. C. Dimayuga, B. Cercek, S. Kaul, and P. K. Shah Differential Effects of Apolipoprotein A-I-Mimetic Peptide on Evolving and Established Atherosclerosis in Apolipoprotein E-Null Mice Circulation, September 21, 2004; 110(12): 1701 - 1705. [Abstract] [Full Text] [PDF]

				M. Navab, G.M. Anantharamaiah, S. T. Reddy, S. Hama, G. Hough, V. R. Grijalva, A. C. Wagner, J. S. Frank, G. Datta, D. Garber, et al. Oral D-4F Causes Formation of Pre-{beta} High-Density Lipoprotein and Improves High-Density Lipoprotein-Mediated Cholesterol Efflux and Reverse Cholesterol Transport From Macrophages in Apolipoprotein E-Null Mice Circulation, June 29, 2004; 109(25): 3215 - 3220. [Abstract] [Full Text] [PDF]

				M. Navab, G. M. Ananthramaiah, S. T. Reddy, B. J. Van Lenten, B. J. Ansell, G. C. Fonarow, K. Vahabzadeh, S. Hama, G. Hough, N. Kamranpour, et al. Thematic review series: The Pathogenesis of Atherosclerosis The oxidation hypothesis of atherogenesis: the role of oxidized phospholipids and HDL J. Lipid Res., June 1, 2004; 45(6): 993 - 1007. [Abstract] [Full Text] [PDF]

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