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(Circulation Research. 2001;89:1155.)
© 2001 American Heart Association, Inc.

Molecular Medicine

Transforming Growth Factor-ß₁ Modulates Oxidatively Modified LDL–Induced Expression of Adhesion Molecules

Role of LOX-1

Hongjiang Chen, Dayuan Li, Tom Saldeen, Jawahar L. Mehta

From the Departments of Internal Medicine and Physiology (H.C., D.L., J.L.M.), University of Arkansas for Medical Sciences and Central Arkansas Veterans Healthcare System, Little Rock, Arkansas; and the Department of Surgical Sciences (T.S.), University of Uppsala, Sweden.

Correspondence to Jawahar L. Mehta, MD, PhD, Division of Cardiovascular Medicine, University of Arkansas for Medical Sciences, 4301 West Markham, Mail Slot 532, Little Rock, AR 72205-7199. E-mail MehtaJL{at}uams.edu

	Abstract

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Oxidatively modified LDL (ox-LDL) activates a lectin-like receptor, LOX-1, which results in the expression of adhesion molecules on endothelial surface. We investigated the regulation of the expression of transforming growth factor-ß₁ (TGF-ß₁) and its receptors by ox-LDL and the functional significance of this interaction with regard to adhesion molecule expression in human coronary artery endothelial cells (HCAECs). Ox-LDL, in a time- and concentration-dependent manner, upregulated the expression of all 3 subtypes (1, 2, and 3 [including endoglin]) of TGF-ß₁ receptors and decreased active TGF-ß₁ synthesis (all P<0.05 versus control and native-LDL–treated cells). Treatment of HCAECs with a monoclonal antibody to LOX-1 attenuated ox-LDL–mediated upregulation of TGF-ß₁ receptors and decrease in TGF-ß₁ synthesis (P<0.05 versus ox-LDL alone). Ox-LDL also enhanced the expression of P-selectin and ICAM-1 as well as monocyte adhesion to HCAECs (P<0.05 versus control untreated cells). Pretreatment with recombinant TGF-ß₁ attenuated the enhanced expression of adhesion molecules and monocyte adhesion to HCAECs (P<0.05 versus ox-LDL alone). Effects of recombinant TGF-ß₁ were blocked by antibody to TGF-ß₁ receptor type 2, but not by antibody to endoglin. Thus ox-LDL, via activation of LOX-1, increases the expression of TGF-ß₁ receptors and decreases TGF-ß₁ synthesis in HCAECs. Recombinant TGF-ß₁, by binding to TGF-ß₁ type 2 receptors, modulates ox-LDL–mediated expression of adhesion molecules and monocyte adhesion to HCAECs.

Key Words: adhesion molecules • endothelial cells • oxidatively modified LDL • transforming growth factor-ß₁

	Introduction

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Oxidatively modified LDL (ox-LDL) is a critical pathogenic factor in atherosclerosis. Uptake of LDL, especially its oxidized form, by endothelial cells is considered an important step in the initiation and progression of atherosclerosis.¹ LOX-1, a membrane protein that belongs structurally to C-type selectin family, is a specific receptor that facilitates uptake of ox-LDL by endothelial cells.^2,3 Ox-LDL binding to LOX-1 in endothelial cells induces a cascade of events, such as upregulation of angiotensin-II type 1 receptor expression, release of reactive oxygen species, reduction in nitric oxide synthesis, and activation of redox-sensitive transcription factor NF-B.^4–6 Studies by several investigators have shown that ox-LDL, via upregulation of LOX-1, induces monocyte adhesion and enhances expression of adhesion molecules, such as P-selectin, vascular cell adhesion molecule (VCAM-1), and intracellular adhesion molecule (ICAM-1), as well as monocyte chemotactic protein (MCP-1) in endothelial cells.^7–9 Expression of adhesion molecules allows the binding of inflammatory cells to the activated endothelium.^10–12 Growth factors, such as transforming growth factor-ß₁ (TGF-ß₁), have been shown to modulate the expression of adhesion molecules.^13–16

TGF-ß₁, a member of the family of growth factors, is a multifunctional polypeptide.¹⁷ It manifests its biological effects by binding to three distinct TGF-ß₁ receptors on cell surface (type 1, 2 and 3 [including endoglin]).^18–20 There is evolving evidence that TGF-ß₁ regulates proliferation and differentiation of cells, remodeling of blood vessels and myocardium after injury, angiogenesis, inflammation, release of cytokines, and lipid metabolism.^{10,15–18,21–23} In keeping with the potential role that TGF-ß₁ may play in adhesion molecule expression and possibly atherogenesis, Koglin et al²⁴ showed more extensive atherosclerosis in TGF-ß₁–knockout mice. Grainger et al,²⁵ and later Tashiro et al,²⁶ showed reduced plasma TGF-ß₁ levels in patients with atherosclerosis. Other investigators have shown alteration in the expression of TGF-ß₁ receptors in atherosclerosis.^27,28 Recently, Minami et al²⁹ and Draude et al³⁰ reported that LOX-1 expression in endothelial cells is upregulated in response to TGF-ß₁.

The present study was designed to investigate the regulation of TGF-ß₁ synthesis and expression of its receptors on human coronary artery endothelial cells (HCAECs) by ox-LDL. We focused this work on endothelial cells because this is the first cell line to become dysfunctional in atherosclerosis.

	Materials and Methods

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Cell Culture
The initial batch of HCAECs was purchased from Clonetics Corp. The endothelial cells were pure, based on morphology and staining for factor VIII and acetylated LDL. These cells were 100% negative for -actin expression. The endothelial cell growth medium consisted of 500 mL basal medium, 5 ng human recombinant epidermal growth factor, 5 mg hydrocortisone, 25 mg gentamycin, 25 µg amphotericin B, 6 mg bovine brain extract, and 25 mL FBS.^3,4 Fifth passage HCAECs were used in this study.

To explore the role of ox-LDL on the expression of TGF-ß₁ and its receptors, HCAECs were incubated with native-LDL (n-LDL) or ox-LDL in different concentrations (10, 20, and 40 µg/mL) for different times (1, 6, 12, and 24 hours). Parallel groups of cells were pretreated with a blocking antibody to LOX-1 (10 µg/mL) (gift of T. Sawamura, Osaka, Japan). Culture media were collected for measuring active TGF-ß₁ levels, and the cells were harvested for determination of the expression of TGF-ß₁ receptors. The concentrations of ox-LDL were chosen based in previous studies.^2–4,8

To examine the functional significance of TGF-ß₁ in ox-LDL–induced expression of adhesion molecules, cultured HCAECs were pretreated with recombinant TGF-ß₁ (2 ng/mL) (Sigma) prior to incubation with ox-LDL (40 µg/mL), and then adhesion of monocytes to endothelial cells quantified. Parallel groups of cells were pretreated with antibody to TGF-ß₁ receptor type 2 (10 µg/mL) or antibody to endoglin (10 µg/mL) (both from Santa Cruz Biotechnology). The specificity of these antibodies was 100%.

Preparation of Lipoproteins
Ox-LDL was prepared as described earlier.^3,4 In brief, human n-LDL was oxidized by exposure to CuSO₄ (5 µmol/L free Cu²⁺ concentration) in PBS at 37° for 24 hours. The thiobarbituric acid reactive substance content of ox-LDL was 18.2±0.28 (versus 0.56±0.16 nmol per 100 µg protein in the n-LDL preparation, P<0.01). LDL and ox-LDL were kept in 50 mmol/L Tris-HCl, 0.15 mol/L NaCl, and 2 mmol/L EDTA at pH 7.4 and were used within 10 days of preparation.

TGF-ß₁ Assay by ELISA
TGF-ß₁ level in the culture media was measured by ELISA (Promega).³¹

Isolation and Adhesion of Human Monocytes
The technique for isolation of human peripheral monocytes and for quantitation of their adhesion to HCAECs has been described earlier.⁸ Isolated cell preparation was 94% to 98% pure and the cells had intact function.⁸

¹²⁵I-TGF-ß₁ Binding and Cross-Linking
TGF-ß₁ receptors were studied by chemically cross-linking ¹²⁵I-TGF-ß₁ to the cell surface followed by SDS-PAGE and autoradiography. Briefly, 1x10⁵ HCAECs were kept in regular culture medium for 24 hours and then treated with n-LDL, ox-LDL, or antibody to LOX-1. Cells were incubated in fresh serum-free medium for 1 hour before applying ¹²⁵I-TGF-ß₁ (1 ng/mL/well) in binding medium (with 0.05% gelatin) at 4°C with 50-fold excess unlabeled TGF-ß₁. After 3 hours of incubation, DSS was added to the well at a final concentration of 1 mmol/L for 15 minutes. The cells were washed with binding medium and scraped into detachment buffer (10 mmol/L Tris-HCl, pH 7.4, 1 mmol/L EDTA, 10% glycerol, 0.3 mmol/L PMSF) and centrifuged at 12 000g for 2 minutes. The cells were resuspended in SDS sample buffer with ß-mercaptoethanol for 30 minutes at 37°C, centrifuged, and the dissolved proteins were separated on a 8% SDS-PAGE gel. The dried gel was then exposed to film.^28,32

Determination of P-Selectin and ICAM-1 Protein and mRNA
The methodology for protein and mRNA determination has been described earlier.^3,4 For protein assay (Western analysis), the primary polyclonal antibodies to P-selectin and ICAM-1 were obtained from Santa Cruz Biotechnology. The primer pairs specific to P-selectin were as follows: forward, 5'-AATTTCTCCCAACTCAGTG-3'; reverse, 5'-CGAACTGTGTCAGCTCCTCTC-3'. The ICAM-1 primers were as follows: forward, 5'-GTGTCTCCTTCTTTGACACT-3'; reverse, 5'-ATGACATGCTTGAGCCAGG-3'.

Data Analysis
Data are presented as mean±SEM. Statistical significance was determined in multiple comparisons among independent groups of data in which ANOVA and the Student-Newman-Keuls test indicated the presence of significant differences. A value of P<0.05 was considered statistically significant.

	Results

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Ox-LDL and Active TGF-ß₁ Synthesis and TGF-ß₁ Receptor Expression
Active TGF-ß₁ levels were lower in the supernatants of HCAECs treated with ox-LDL compared with those treated with n-LDL. The decrease in active TGF-ß₁ levels in response to ox-LDL occurred in a time- and concentration-dependent manner (n=5, P<0.05) (Figure 1).

View larger version (33K): [in this window] [in a new window]   Figure 1. Effect of oxidized-LDL (ox-LDL) on active TGF-ß1 levels in human coronary artery endothelial cells (HCAECs). Incubation of HCAECs with ox-LDL, but not native-LDL (n-LDL), decreased active TGF-ß1 levels in the HCAEC culture medium. The decrease in TGF-ß1 levels in response to ox-LDL occurred in a time- and concentration-dependent manner. Importantly, pretreatment of HCAECs with antibody to LOX-1 prior to incubation with ox-LDL attenuated the decrease in TGF-ß1 levels induced by ox-LDL (40 µg/mL). Data from 5 separate experiments are shown in mean±SEM.

By ¹²⁵I-TGF-ß₁ binding and cross-linking, we observed that incubation of HCAECs with ox-LDL, but not n-LDL, augmented expression of all three TGF-ß₁ receptor types in the HCAECs. Increase in the expression of TGF-ß₁ receptors in response to ox-LDL occurred in a time- and concentration-dependent manner. A representative example of increased TGF-ß₁ receptor expression is shown in Figure 2, and densitometric analysis from 3 separate experiments is shown in Figure 3. It is noteworthy that the expression of type 2 and type 3 receptors was similar and more marked than the expression of type 1 receptor. The increase in TGF-ß₁ receptor expression correlated with the decrease in active TGF-ß₁ levels.

View larger version (61K): [in this window] [in a new window]   Figure 2. Expression of TGF-ß1 receptors in HCAECs by binding and cross-linking with 125I-TGF-ß1. Incubation of HCAECs with ox-LDL, but not n-LDL, augmented expression of TGFß1 receptors in HCAECs; the increase occurred in a time- and concentration-dependent manner. The expression of type 2 and 3 receptors was similar and prominent. Data are representative of 3 independent experiments.

View larger version (24K): [in this window] [in a new window]   Figure 3. Role of LOX-1 in the expression of TGF-ß1 receptors on HCAECs treated with ox-LDL. Incubation of HCAECs with ox-LDL (40 µg/mL) increased the expression of all TGF-ß1 receptor types, especially types 2 and 3. Pretreatment of HCAECs with antibody to LOX-1 attenuated ox-LDL enhanced expression of TGF-ß1 receptors, suggesting that the effects of ox-LDL are mediated via activation of LOX-1. Densitometric data are from 3 different binding and cross-linking experiments and are shown in mean±SEM.

Treatment of HCAECs with LOX-1 antibody attenuated the decrease in TGF-ß₁ levels induced by ox-LDL (P<0.05 versus ox-LDL alone) (Figure 1). Treatment of HCAECs with antibody to LOX-1 prior to incubation with ox-LDL also attenuated the enhanced expression of TGF-ß₁ receptors induced by ox-LDL (Figure 3). These observations suggest that the effects of ox-LDL are mediated via activation of LOX-1 receptor.

TGF-ß₁ and Its Receptors and Expression of Adhesion Molecules
Ox-LDL, but not n-LDL, markedly increased the expression of P-selectin and ICAM-1 (protein and mRNA) in HCAECs. This is in accordance with previous observations of upregulation of expression of adhesion molecules on HCAECs exposed to ox-LDL.⁹ Pretreatment of cells with recombinant TGF-ß₁ markedly reduced adhesion molecule expression in HCAECs induced by ox-LDL. The effect of TGF-ß₁ was blocked by antibody to TGF-ß₁ type 2 receptor but not by antibody to endoglin (Figure 4).

View larger version (35K): [in this window] [in a new window]   Figure 4. Effect of TGF-ß1 on the expression of ICAM-1 and P-selectin by Western blot and RT-PCR. The left panel is representative experiment showing expression of ICAM-1 and P-selectin (protein and mRNA). The right panel is the result of densitometric analysis from 4 different experiments. (1) Control; (2) n-LDL; (3) ox-LDL (40 µg/mL); (4) TGF-ß1 (2 ng/mL) alone; (5) TGF-ß1 (2 ng/mL)+ox-LDL (40 µg/mL); (6) antibody to TGF-ß1 receptor type 2 (10 µg/mL)+TGF-ß1 (2 ng/mL)+ox-LDL (40 µg/mL); and (7) antibody to TGF-ß1 receptor type 3 (endoglin, 10 µg/mL)+TGF-ß1 (2 ng/mL)+ox-LDL (40 µg/mL). Ox-LDL, not n-LDL, markedly increased the expression of ICAM-1 and P-selectin in HCAECs. Pretreatment of cells with recombinant TGF-ß1 reduced adhesion molecule expression in HCAECs treatment with ox-LDL. The effect of TGF-ß1 was blocked by antibody to TGF-ß1 type 2 receptor but not by antibody to endoglin. Data from 4 separate experiments shown in mean±SEM.

TGF-ß₁ and Monocyte Adhesion Induced by Ox-LDL
To evaluate the functional significance of adhesion molecule expression in response to ox-LDL, we quantitated adhesion of monocytes to HCAECs. As described earlier,⁹ ox-LDL markedly increased the adhesion of monocytes to HCAECs (n=5 in each group, P<0.01 versus control), whereas n-LDL had no effect. Importantly, pretreatment of cells with recombinant TGF-ß₁ did not affect the adhesion of monocytes to HCAECs cultured under normal conditions. However, it reduced monocyte adhesion to HCAECs treated with ox-LDL (P<0.05 versus ox-LDL alone). Further, the enhanced monocyte adhesion was blocked by antibody to TGF-ß₁ type 2 receptor but not by antibody to endoglin (Figure 5).

View larger version (31K): [in this window] [in a new window]   Figure 5. Effect of TGF-ß1 on monocyte adhesion induced by ox-LDL. Ox-LDL, but not n-LDL, markedly increased the adhesion of monocytes to HCAECs. Pretreatment of cells with recombinant TGF-ß1 did not affect the adhesion of monocytes to HCAECs cultured under normal conditions; however, it significantly reduced the increase in monocyte adhesion induced by ox-LDL. The enhanced monocyte adhesion in response to ox-LDL was blocked by antibody to TGF-ß1 type 2 receptor but not by antibody to endoglin. Data are shown from 5 separate experiments in mean±SEM.

	Discussion

Top Abstract Introduction Materials and Methods Results Discussion References

 
Role of growth factors in atherosclerosis is an area of great interest. Growth factors regulate angiogenesis, smooth muscle cell growth, differentiation, and inflammatory reactions. For example, insulin and insulin-like growth factors have clearly been shown to promote atherogenesis.³³ Recently, vascular endothelial growth factor, a potent angiogenic factor, has also been shown to promote atherogenesis in experimental animal models.^34,35

Role of TGF-ß₁ in atherosclerosis has been debated. Chen et al³⁶ showed increase in TGF-ß₁ expression in the proliferating lesions of cholesterol-fed rabbits. On the other hand, Grainger et al³⁷ have reported a decrease in the expression of TGF-ß₁ in the transgenic apolipoprotein(a) mice that develop severe atherosclerosis. Later studies by Grainger et al²⁵ and Tashiro et al²⁶ also reported low levels of active TGF-ß₁ in the serum of patients with severe atherosclerosis. In keeping with these reports,^25,26 Blann et al²⁷ described an increase in the plasma levels of TGF-ß₁ type 2 receptor in the plasma of patients with atherosclerosis.

We attempted to clarify the interrelationship between ox-LDL and TGF-ß₁ in HCAECs. We found that treatment of HCAECs with ox-LDL, but not n-LDL, reduced TGF-ß₁ synthesis in a time- and concentration-dependent manner. Concurrently, there was an upregulation of the expression of all 3 TGF-ß₁ receptor types (both mRNA and protein). These observations suggest that ox-LDL reduces TGF-ß₁ synthesis with a feedback upregulation of TGF-ß₁ receptors. The decrease in active TGF-ß₁ levels in the supernates of HCAECs treated with ox-LDL is consistent with the observations of Grainger et al^25,37 and Tashiro et al.²⁶ Ox-LDL–mediated upregulation of TGF-ß₁ receptors is also consistent with the description of increased plasma levels of TGF-ß₁ receptors in atherosclerosis.²⁷ Because a specific antibody to LOX-1 blocked the modulation of TGF-ß₁ and its receptors in response to ox-LDL, we conclude that the effect of ox-LDL is mediated by activation of LOX-1.

The reduction in TGF-ß₁ synthesis in atherosclerosis may also involve ox-LDL–mediated inhibition of plasminogen and, hence, plasmin synthesis. Decrease in plasmin has been shown to reduce expression and activation of TGF-ß₁.^38,39 Ox-LDL has also been shown to upregulate angiotensin II type 1 receptors in atherosclerosis.⁴ Angiotensin II type 1 receptor expression, like ox-LDL, decreases TGF-ß₁ synthesis and increases the expression of its receptors,⁴⁰ and may contribute to alterations in TGF-ß₁ and its receptors.

TGF-ß₁ has a multitude of effects on lipid metabolism.^41,42 It inhibits proliferation of smooth muscle cells and accelerates their differentiation.²² Atherosclerosis is considered an inflammatory process, and the endothelial cells from atherosclerotic regions generally exhibit increased expression of adhesion molecules.^9,13,14 There is strong evidence that TGF-ß₁ can inhibit expression of these adhesion molecules and the adhesion of monocytes to the vessel wall.¹⁵ In cats subjected to coronary artery occlusion and reperfusion, TGF-ß₁ decreased inflammatory reaction and limited reperfusion injury.⁴³ Blockade of TGF-ß₁ gene expression accelerates atherosclerosis in mice.²⁴ In the present study, we found that ox-LDL decreased active TGF-ß₁ levels in the supernates of HCAECs and concurrently increased expression of P-selectin and ICAM-1 and monocyte adhesion to endothelial cells. The importance of TGF-ß₁ in the expression of adhesion molecules became evident from experiments in which recombinant TGF-ß₁ attenuated ox-LDL–induced expression of adhesion molecules and subsequent monocyte adhesion. These observations suggest that reduction of TGF-ß₁ synthesis may, at least in part, be the mechanism by which ox-LDL induces expression of adhesion molecules and facilitates attachment of monocytes to the activated endothelium.

Although TGF-ß₁ is thought to participate in several pathophysiologic processes, its signaling pathway is still not completely understood. It has been suggested that TGF-ß₁ functions by binding first to type 3 receptors, which then facilitate binding to type 2 receptors, and finally transferring the binding signal to type 1 receptors.¹⁸ Others have postulated that TGF-ß₁ exerts its function by binding directly to type 2 receptors, which transfer the signal to type 1 receptors. Endoglin can only modulate this binding signal, but it does not directly participate in signal transduction.¹⁹ Recent studies indicate that endoglin is associated with growth of smooth muscle cells, fibroblasts and cancer cells.^44–46 The present study shows that the effects of recombinant TGF-ß₁ (inhibition of ox-LDL–mediated expression of adhesion molecules and monocyte adhesion to HCAECs) are blocked by antibody to type 2 receptor, but not by antibody to endoglin. These observations indicate that the effect of TGF-ß₁ on the expression of adhesion molecules is mediated by activation of type 2 receptor and has no direct relationship with the activation of endoglin.

In conclusion, the present study in HCAECs suggests that (1) ox-LDL decreases active TGF-ß₁ synthesis and increases the expression of all TGF-ß₁ receptor types; (2) activation of LOX-1 is a key element in this process; (3) ox-LDL–mediated expression of adhesion molecules is related to a reduction in TGF-ß₁ synthesis; and (4) activation of TGF-ß₁ type 2 receptor is a initial step in the modulatory effect of TGF-ß₁ on ox-LDL–mediated inflammatory reaction in the vessel wall.

	Acknowledgments

 
This study has been supported by a Merit Review grant from the Department of Veterans Affairs (J.L.M.), a contract with the Department of Defense (J.L.M.), funds from the Swedish Medical Research Council (T.S.), an American Heart Association Scientist Development Grant (D.Y.L.), the Wrenette Worthen Williamson Cardiology Research Endowment (J.L.M.), and the Howard and Elsie Stebbins Chair in Cardiology (J.L.M.).

Received July 18, 2001; revision received October 12, 2001; accepted October 12, 2001.

	References

Top Abstract Introduction Materials and Methods Results Discussion References

 
1. Steinberg D. Oxidative modification of LDL and atherogenesis. Circulation. 1997; 95: 1062–1071.

2. Sawamura T, Kume N, Aoyama T, Moriwaki H, Hoshikawa H, Aiba Y, Tanaka T, Miwa S, Katsura Y, Kita T, Masaki T. An endothelial receptor for oxidized low-density lipoprotein. Nature. 1997; 386: 73–77.

3. Li D, Mehta JL. Upregulation of endothelial receptor for oxidized LDL (LOX-1) by oxidized LDL and implications in apoptosis of human coronary artery endothelial cells: evidence from use of antisense LOX-1 mRNA and chemical inhibitors. Arterioscler Thromb Vasc Biol. 2000; 20: 1116–1122.

4. Li D, Saldeen T, Romeo F, Mehta JL. Oxidized LDL upregulates angiotensin II type 1 receptor expression in cultured human coronary artery endothelial cells: the potential role of transcription factor NF-B. Circulation. 2000; 102: 1970–1976.

5. Cominacini L, Rigoni A, Pasini AF, Garbin U, Davoli A, Campagnola M, Pastorino AM, Lo Cascio V, Sawamura T. The binding of oxidized low density lipoprotein (ox-LDL) to ox-LDL receptor-1 reduces the intracellular concentration of nitric oxide in endothelial cells through an increased production of superoxide. J Biol Chem. 2001; 276: 13750–13755.

6. Cominacini L, Pasini AF, Garbin U, Davoli A, Tosetti ML, Campagnola M, Rigoni A, Pastorino AM, Lo Cascio V, Sawamura T. Oxidized low density lipoprotein (ox-LDL) binding to ox-LDL receptor-1 in endothelial cells induces the activation of NF- B through an increased production of intracellular reactive oxygen species. J Biol Chem. 2000; 275: 12633–12638.

7. Takei A, Huang Y, Lopes-Virella MF. Expression of adhesion molecules by human endothelial cells exposed to oxidized low density lipoprotein: influences of degree of oxidation and location of oxidized LDL. Atherosclerosis. 2001; 154: 79–86.

8. Li D, Mehta JL. Antisense to LOX-1 inhibits oxidized LDL-mediated upregulation of monocyte chemoattractant protein-1 and monocyte adhesion to human coronary artery endothelial cells. Circulation. 2000; 101: 2889–2895.

9. Li D, Chen H, Mhatre A, Romeo F, Saldeen T, Mehta JL. Statins inhibit ox-LDL-induced expression of adhesion molecules and monocyte adhesion to human coronary endothelial cells: role of mitogen-activated protein kinase and NF-B. Circulation. 2000; 102: II-251.Abstract.

10. Ross R. Atherosclerosis: an inflammatory disease. N Engl J Med. 1999; 340: 115–126.

11. Dong ZM, Chapman SM, Brown AA, Frenette PS, Hynes RO, Wagner DD. The combined role of P- and E-selectins in atherosclerosis. J Clin Invest. 1998; 102: 145–152.

12. Ramos CL, Huo Y, Jung U, Ghosh S, Manka DR, Sarembock IJ, Ley K. Direct demonstration of P-selectin- and VCAM-1–dependent mononuclear cell rolling in early atherosclerotic lesions of apolipoprotein E-deficient mice. Circ Res. 1999; 84: 1237–1244.

13. Meager A. Cytokine regulation of cellular adhesion molecule expression in inflammation. Cytokine Growth Factor Rev. 1999; 10: 27–39.

14. Yeh CH, Peng HC, Huang TF. Cytokines modulate integrin _vß₃-mediated human endothelial cell adhesion and calcium signaling. Exp Cell Res. 1999; 251: 57–66.

15. Koyanagi M, Egashira K, Kubo-Inoue M Usui M, Kitamoto S, Tomita H, Shimokawa H, Takeshita A. Role of transforming growth factor-ß1 in cardiovascular inflammatory changes induced by chronic inhibition of nitric oxide synthesis. Hypertension. 2000; 35: 86–90.

16. Park SK, Yang WS, Lee SK, Ahn H, Park JS, Hwang O, Lee JD. TGF-ß₁ down-regulates inflammatory cytokine-induced VCAM-1 expression in cultured human glomerular endothelial cells. Nephrol Dial Transplant. 2000; 15: 596–604.

17. Moses HL, Yang EY, Pietenpol JA. TGF-ß stimulation and inhibition of cell proliferation: new mechanistic insights. Cell. 1990; 63: 245–247.

18. McCaffrey TA. TGF-ßs and TGF-ß receptors in atherosclerosis. Cytokine Growth Factor Rev. 2000; 11: 103–114.

19. Blobe GC, Schiemann WP, Lodish HF. Role of transforming growth factor-ß in human disease. N Engl J Med. 2000; 342: 1350–1358.

20. Massague J, TGF-ß signal transduction. Annu Rev Biochem. 1998; 67: 753–791.

21. August P, Leventhal B, Suthanthiran M. Hypertension-induced organ damage in African Americans: transforming growth factor-ß₁ excess as a mechanism for increased prevalence. Curr Hypertens Rep. 2000; 2: 184–191.

22. Mii S, Ware JA, Kent KC. Transforming growth factor-ß inhibits human vascular smooth muscle cell growth and migration. Surgery. 1993; 114: 464–470.

23. Park HJ, Galper JB. 3-Hydroxy-3-methylglutaryl CoA reductase inhibitors up-regulate transforming growth factor-ß signaling in cultured heart cells via inhibition of geranylgeranylation of RhoA GTPase. Proc Natl Acad Sci U S A. 1999; 96: 11525–11530.

24. Koglin J, Glysing-Jensen T, Raisanen-Sokolowski A, Russell ME. Immune sources of transforming growth factor-ß₁ reduce transplant arteriosclerosis: insight derived from a knockout mouse model. Circ Res. 1998; 83: 652–660.

25. Grainger DJ, Kemp PR, Metcalfe JC, Liu AC, Lawn RM, Williams NR, Grace AA, Schofield PM, Chauhan A. The serum concentration of active transforming growth factor-ß is severely depressed in advanced atherosclerosis. Nat Med. 1995; 1: 74–79.

26. Tashiro H, Shimokawa H, Yamamoto K, Momohara M, Tada H, Takeshita A. Altered plasma levels of cytokines in patients with ischemic heart disease. Cor Artery Dis. 1997; 8: 143–147.

27. Blann AD, Wang JM, Wilson PB, Kumar S. Serum levels of the TGF-ß receptor are increased in atherosclerosis. Atherosclerosis. 1996; 120: 221–226.

28. McCaffrey TA, Consigli S, Du B, Falcone DJ, Sanborn TA, Spokojny AM, Bush HL Jr. Increased type II/type I TGF-ß receptor ratio in cells derived from human atherosclerotic lesions: conversion from an antiproliferative to profibrotic response to TGF-ß1. J Clin Invest. 1995; 96: 2667–2675.

29. Minami M, Kume N, Kataoka H, Morimoto M, Hayashida K, Sawamura T, Masaki T, Kita T. Transforming growth factor-ß₁ increases the expression of lectin-like oxidized low-density lipoprotein receptor-1. Biochem Biophys Res Commun. 2000; 272: 357–361.

30. Draude G, Lorenz RL. TGF-ß₁ downregulates CD36 and scavenger receptor A but upregulates LOX-1 in human macrophages. Am J Physiol Heart Circ Physiol. 2000; 278: 1042–1048.

31. Mehta JL, Yang BC, Strates BS, Mehta P. Role of TGF-ß1 in platelet-mediated cardioprotection during ischemia-reperfusion in isolated rat hearts. Growth Factors. 1999; 16: 179–190.

32. Massague J, Kelly B. Internalization of transforming growth factor-ß and its receptor in BALB/c 3T3 fibroblasts. J Cell Physiol. 1986; 128: 216–222.

33. Bayes-Genis A, Conover CA, Schwartz RS. The insulin-like growth factor axis: a review of atherosclerosis and restenosis. Circ Res. 2000; 86: 125–130.

34. Celletti FL, Hilfiker PR, Ghafouri P, Dake MD. Effect of human recombinant vascular endothelial growth factor 165 on progression of atherosclerotic plaque. J Am Coll Cardiol. 2001; 37: 2126–2130.

35. Celletti FL, Waugh JM, Amabile PG, Brendolan A, Hilfiker PR, Dake MD. Vascular endothelial growth factor enhances atherosclerotic plaque progression. Nat Med. 2001; 7: 425–429.

36. Chen YL, Wu HW, Jiang MJ. Transforming growth factor-ß₁ gene and protein expression associated with atherogenesis of cholesterol-fed rabbits. Histol Histopathol. 2000; 15: 421–428.

37. Grainger DJ, Kemp PR, Liu AC, Lawn RM, Metcalfe JC. Activation of transforming growth factor-ß s inhibited in transgenic apolipoprotein(a) mice. Nature. 1994; 370: 460–462.

38. Pedrozo HA Schwartz Z, Robinson M, Gomes R, Dean DD, Bonewald LF, Boyan BD. Potential mechanisms for the plasmin-mediated release and activation of latent transforming growth factor-ß1 from the extracellular matrix of growth plate chondrocytes. Endocrinology. 1999; 140: 5806–5816.

39. Lawn RM, Pearle AD, Kunz LL, Rubin EM, Reckless J, Metcalfe JC, Grainger DJ. Feedback mechanism of focal vascular lesion formation in transgenic apolipoprotein(a) mice. J Biol Chem. 1996; 271: 31367–31371.

40. Li D, Chen H, Mehta JL. Angiotensin-II via AT1 receptor activation upregulates endoglin expression in human coronary artery endothelial cells. J Am Coll Cardiol. 2001; 37: 294A.Abstract.

41. Brand C, Cherradi N, Defaye G, Chinn A, Chambaz EM, Feige JJ, Bailly S. Transforming growth factor-ß1 decreases cholesterol supply to mitochondria via repression of steroidogenic acute regulatory protein expression. J Biol Chem. 1998; 273: 6410–6416.

42. Zuckerman SH, Panousis C, Evans G. TGF-ß reduced binding of high-density lipoproteins in murine macrophages and macrophage-derived foam cells. Atherosclerosis. 2001; 155: 79–85.

43. Lefer AM, Ma XL, Weyrich AS, Scalia R. Mechanism of the cardioprotective effect of transforming growth factor ß₁ in feline myocardial ischemia and reperfusion. Proc Natl Acad Sci U S A. 1993; 90: 1018–1022.

44. Ma X, Labinaz M, Goldstein J, Miller H, Keon WJ, Letarte M, O’Brien E. Endoglin is overexpressed after arterial injury and is required for transforming growth factor-ß–induced inhibition of smooth muscle cell migration. Arterioscler Thromb Vasc Biol. 2000; 20: 2546–2552.

45. Miller DW, Graulich W, Karges B, Stahl S, Ernst M, Ramaswamy A, Sedlacek HH, Muller R, Adamkiewicz J. Elevated expression of endoglin, a component of the TGF-ß-receptor complex, correlates with proliferation of tumor endothelial cells. Int J Cancer. 1999; 81: 568–572.

46. Guerrero-Esteo M, Lastres P, Letamendia A, Perez-Alvarez MJ, Langa C, Lopez LA, Fabra A, Garcia-Pardo A, Vera S, Letarte M, Bernabeu C. Endoglin overexpression modulates cellular morphology, migration, and adhesion of mouse fibroblasts. Eur J Cell Biol. 1999; 78: 614–623.

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