| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
|
|
|
|||||||
|
|||||||||
Institut National de la Santé et de la Recherche Médicale U. 689, Cardiovascular Research Center Lariboisiere, and University Paris 7, Paris, France
ABSTRACT I. INTRODUCTION A. Historical Perspective B. Atherosclerosis as an Immunoinflammatory Disease II. THE ATHEROSCLEROTIC CYTOKINE NETWORK A. Cytokine Families B. Cytokine-Associated Signaling Pathways 1. NF-kappaB 2. JNK/AP-1 3. JAK/STAT 4. Smads 5. TLR/Myd88 signaling pathways III. INDUCERS OF CYTOKINE PRODUCTION IN ATHEROSCLEROSIS A. Initial Trigger(s) B. Secondary Triggers 1. HSP 2. Immune complexes 3. Infectious agents 4. Defective clearance of apoptotic cells 5. Cellular microparticles 6. MMPs 7. Inflammasome 8. Oxygen radicals 9. Angiotensin II 10. AGEs 11. Proinflammatory cytokines 12. TLR endogenous agents 13. Mechanical factors 14. Adipokines 15. Platelet products and coagulation factors/others IV. CYTOKINES AND CYTOKINE RECEPTORS IN HUMAN ATHEROSCLEROTIC PLAQUES A. Cytokine Expression in Plaques B. Cellular Sources of Cytokines 1. Vascular cells 2. Leukocytes 3. Platelets 4. Mast cells C. Biological Effects of Cytokines 1. Effects on endothelial permeability 2. Activation of adhesion molecule and chemokine expression 3. Modulation of scavenger receptor expression and lipid metabolism 4. Effect of SMC migration/proliferation 5. Modulation of extracellular matrix remodeling 6. Mobilization of vascular progenitor cells 7. Regulation of neovessel formation/promotion of intraplaque neovascularization 8. Induction of apoptosis 9. Modulation of procoagulant activity and fibrinolysis 10. Regulation of immune response V. CYTOKINE AND CYTOKINE RECEPTOR-ASSOCIATED MODULATION OF PLAQUE DEVELOPMENT AND STABILITY A. Proinflammatory Cytokines 1. TNF-alpha 2. IL-1 3. IL-2 4. IL-6 5. IL-12/IL-18/IFN-gamma 6. CD40/CD40L 7. Osteopontin 8. MIF B. Anti-inflammatory Cytokines 1. IL-10 2. IL-4/IL-13 3. TGF-beta C. Chemokines/Chemokine Receptors 1. MCP1/CCR2 2. Fractalkine/CX3CR1 3. IL-8/CXCR2 4. RANTES/CCR5 5. MIF D. Hematopoietic Factors/M-CSF E. Platelet-Derived Factors VI. CYTOKINES AND ADAPTIVE IMMUNITY IN ATHEROSCLEROSIS A. Role of T/B Cells in Atherosclerosis B. Cytokines and Pathogenic Immune Response in Atherosclerosis 1. Cytokines and DC maturation 2. Cytokines and Th1 differentiation 3. Cytokines and Th1/Th2 paradigm C. Immunological Tolerance and Regulatory T Cells 1. Development and function of natural regulatory T cells 2. Cytokines and regulatory T cells 3. Regulatory T cells in atherosclerosis VII. CYTOKINES AND CARDIOVASCULAR RISK A. TNF-alpha B. IL-2 C. IL-6 D. IL-7 E. IL-8 F. IL-18 G. sCD40L H. IL-10 I. M-CSF VIII. THERAPEUTIC POTENTIAL A. Use of Anticytokines B. Targetting Downstream Inflammasome C. Targetting the JAK/STAT Pathway D. Activation of the Natural Anti-inflammatory Intracellular Pathway (SOCS) E. Stimulation of Treg Cells F. Stimulation of Macrophage Emigration From Atherosclerotic Lesions IX. CONCLUSION GRANTS ACKNOWLEDGMENTS REFERENCES
 |
ABSTRACT |
|---|
 Top NextReferences |
|---|
play a critical role. The purpose of this review is to bring together the current information concerning the role of cytokines in the development, progression, and complications of atherosclerosis. Specific emphasis is placed on the contribution of pro- and anti-inflammatory cytokines to pathogenic (innate and adaptive) and regulatory immunity in the context of atherosclerosis. Based on our current knowledge of the role of cytokines in atherosclerosis, we propose some novel therapeutic strategies to combat this disease. In addition, we discuss the potential of circulating cytokine levels as biomarkers of coronary artery disease. |
I. INTRODUCTION |
|---|
|
TopPreviousNextReferences |
|---|
The earliest visible lesion in the development of atherosclerosis is the fatty streak. This comprises an area of intimal thickening composed of macrophages distended by lipid droplets (known as foam cells), lymphocytes, and smooth muscle cells. The American Heart Association (AHA) Committee on Vascular Lesions provided a classification of human atherosclerotic lesions which correlate the histological lesion types, from type I to type VI, with corresponding clinical syndromes (648, 649). This classification should not be understood as an orderly, linear pattern of plaque progression (704). Plaques develop as a result of the accumulation of low-density lipoproteins (LDL) in the subendothelial space, followed by the diapedesis of leukocytes and formation of foam cells, proliferation of smooth muscle cells, and production of connective tissue. The landmark work of Seymour Glagov showed that the arterial wall can remodel itself in response to plaque growth by increasing its external diameter to accommodate the plaque without narrowing of the lumen (234). Thrombosis is the ultimate stage in the disease process that is responsible for clinically observable adverse events implicating coronary, cerebrovascular, and peripheral vascular beds (394). Studies indicate that in patients with atherothrombotic disease plaque formation is likely to be widespread throughout the vasculature, often affecting more than one vascular bed (93).
Even though atherosclerosis is reaching epidemic proportions nowadays, it is not in any way a disease specific to the modern times; it was already present in antiquity. Sir Marc Ruffer was able to identify in 1911 degenerative arterial changes suggestive of atherosclerosis in the left subclavian artery from an Egyptian mummy (583). Later on, paleopathologist A. T. Sandison, using modern technical methods for tissue fixation, confirmed that Egyptian mummies had histological evidence of atherosclerosis with lipid deposits, reduplication of the internal elastic lamina, and medial calcification in arteries (593).
Atherosclerosis is nowadays recognized as a chronic inflammatory disease of large arteries (235, 265, 395, 417, 578). Remarkably, the very first description of the cause of angina pectoris referred to inflammation. Yet, the belief in this notion was subjected to peaks and troughs from early dates up to recent times.
According to the historian J. O. Leibowitz (381), the Italian surgeon and anatomist Antonio Scarpa (1752–1832) was the first to present an anatomopathological description of arterial wall degeneration in full detail. In his 1804 monograph on aneurysms, Scarpa opposed the view that a dilatation of the aorta was the intrinsic cause of an aneurysm leading to rupture. He emphasizes that "... especially the internal coat is subject, from slow internal cause, to an ulcerated and steatomatous disorganization, as well as to a squamous and earthy rigidity and brittleness," introducing the concept of an underlying metabolic disorder in the process of atherosclerosis, rather than the theory of inflammation that already prevailed at that time, the expression "heart abscess" being frequently used to describe heart pathology (reviewed in Ref. 381).
The term atheroma, derived from Greek and meaning "porridge," was first proposed by Albrecht von Haller in 1755 to designate the degenerative process observed in the intima of arteries. London surgeon Joseph Hodgson (1788–1869) published in 1815 his Treatise on the Diseases of Arteries and Veins in which he claimed that inflammation was the underlying cause of atheromatous arteries. But thereafter, most of pathologists of the 19th century following Carl Rokitanski (1804–1878) abandoned the view that inflammation was an etiological factor and considered that atherosclerosis was a degenerative process, with intimal proliferation of connective tissue and calcification, best described by the term arteriosclerosis proposed in 1833 by French pathologist Jean Lobstein (1777–1835). However, German pathologist Rudolf Virchow (1821–1902), a leading authority of his day in pathology and the greatest contributor to the notion of thrombosis, considered atheroma as a chronic inflammatory disease of the intima, that he called "chronic endarteritis deformans". In his opinion, the accumulation of lipids was a late manifestation of atheroma (701). Finally, the Leipzig pathologist Marchand in 1904 first used the term atherosclerosis, which since has been widely adopted, instead of arteriosclerosis, to designate the degenerative process of the intimal layer of the arteries.
Until the beginning of the 20th century, the theories put forward to explain the pathogenesis of atherosclerosis remained purely descriptive and were based on the anatomical observation of human atherosclerotic vessels. A first revolution in the mechanistic assessment of atherosclerosis was initiated in 1908 when the Russian scientist Alexander Ignatowski showed that experimental atherosclerosis could be induced in rabbits by feeding them a diet of milk and egg yolk (301). Soon thereafter, in 1913, N. Anitschkov and S. Chalatov reproduced experimental atherosclerosis by adding pure cholesterol to rabbit food (21). This gave rise to the lipid theory of atherosclerosis that predominated for most of the 20th century. The next significant leap only came during the 1970s when Brown and Goldstein showed that the LDL receptor that they had discovered, a cell surface protein that binds LDL and removes them from blood (reviewed in Ref.88), is not involved in macrophage foam-cell formation and proposed that a macrophage receptor that recognized acetylated LDL plays a key role in this process (237). Subsequently, during the 1980s, the central role of oxidized LDL (oxLDL) in the pathogenesis of atherosclerosis was exposed by Daniel Steinberg and his group (650), and a number of scavenger receptors mediating their uptake by macrophages were identified (reviewed in Ref. 387). The model of the Watanabe heritable hyperlipidemic (WHHL) rabbit, introduced in 1980 (726) was particularly useful in establishing the role of oxLDL in atherogenesis. A second revolution occurred at the beginning of the 1990s when mouse models of atherosclerosis, apolipoprotein E (apoE)- and LDL receptor (LDLr)-deficient mice, were derived by homologous recombination techniques (304, 306, 543, 784). In contrast to the previous models, mice lacking functional apoE or LDLr genes were shown to develop widely distributed arterial lesions that progress from foam cell-rich fatty streaks to fibro-proliferative plaques with lipid/necrotic cores, typical of the spectrum of human lesions (305, 487, 564). The possibility of abolishing the expression of a single gene of interest, or of overexpressing it, in these mouse models opened a new era of atherosclerosis research at a mechanistic level.
B. Atherosclerosis as an Immunoinflammatory Disease
A ripple in the lipid theory appeared in the mid 1970s, when Russel Ross developed his popular "response to injury" hypothesis of atherogenesis, postulating that atherosclerotic lesions arise as a result of focal injury to the arterial endothelium, followed by adherence and aggregation of platelets (580). During the resulting release reaction, platelet-derived growth factor (PDGF) is secreted from the platelets and promotes the proliferative response of smooth muscle cells (SMC). Uncontrolled exuberant SMC proliferation was believed to eventually cause artery occlusion. SMC were considered at that time to be the main promoter of atherosclerotic lesion formation. Instead, it has since been clearly established that SMC proliferation in the plaque is rather modest, and actually tends to be beneficial since it contributes to plaque stabilization (158, 731). In addition, the endothelium actually remains morphologically intact during the development of atherosclerosis (197, 578), although it is activated and directly involved in the immunoinflammatory response. Poole and Florey (547) were the first to observe that soon after initiation of cholesterol feeding in rabbits, monocytes adhere to the endothelium and migrate through the yet intact endothelial monolayer. Michael Gimbrone first proposed the concept of endothelial dysfunction that acknowledged the central role of the normal endothelium in protecting against atherosclerosis while hypothesizing that its cellular functions were altered, "activated" in the disease (232). Ross revisited his "response to injury" theory in 1986 (579) considering that "subtle endothelial injury" was the primum movens in atherosclerosis, and published in 1999 in the New England Journal of Medicine a remarkable review entitled: "Atherosclerosis: a chronic inflammatory disease" (578). The view that atherosclerosis is indeed a chronic inflammatory disease initiated by monocyte/lymphocyte adhesion to activated endothelial cells (EC) is now widely accepted and substantiated by experimental and clinical observations. Several excellent reviews have been published on the theme of atherosclerosis and inflammation since the founding Ross review (52, 235, 265, 395, 417, 578).
Instrumental in the change of opinion regarding the role of inflammation and immunity, rather than SMC proliferation, in the pathogenesis of atherosclerosis was the precise identification of the cell components of human atherosclerotic plaques using modern immunohistochemical techniques by Göran Hansson and colleagues (316). Histologically, the lipid-laden foam cells of the fatty streak, which characterizes the plaque at an early stage, are derived from macrophages. In time, the lipid/necrotic core is covered with fibrous tissue composed mainly of 
-actin positive SMC, and thus forms the fibrolipid plaque. Rather large amounts of T lymphocytes, 
20%, are found as well, surrounding the plaque and in the fibrous cap, pointing to a role of immunity in atherosclerosis (268, 316).
Also determinant in the understanding of the pathogenesis of atherosclerosis were the works by the pathologists Michael Davies (158, 159) and Erling Falk (198), later confirmed and extended by the group of Renu Virmani (704), in their quest for the causes of acute coronary syndromes. Their works emphasized that coronary atherosclerotic plaques exist under two major phenotypes: 1) stable plaques, characterized for the most part by a thick fibrous cap isolating a relatively small lipid core from the lumen, which are associated with a very low risk of thromboembolic complications; and 2) unstable (or vulnerable) plaques, most of which are characterized by a large lipid core covered by a thin fibrous cap prone to rupture and thrombus formation, and which are thought to be associated with a higher risk for thromboembolic complications (218). Analysis of culprit atherosclerotic lesions in patients with acute myocardial infarction revealed that inflammation is crucially determinant in precipitating plaque rupture and some forms of superficial plaque erosion (157, 353, 690).
Virmani uncovered another mechanism of coronary thrombosis occurring in unruptured noninflammatory plaques, described as plaque erosion (199, 703). Eroded plaques differ from ruptured plaques in that they have a base rich in proteoglycans and SMCs. These lesions are more often seen in younger individuals and women, they are associated with less luminal narrowing and less calcification, and they are less likely to have foci of macrophages and T cells compared with ruptured plaque (199). We recently provided experimental evidence that endothelial apoptosis might be a major determinant of plaque erosion (182, 679).
Inflammation, which "is a complex set of interactions among soluble factors and cells that can arise in any tissue in response to traumatic, infectious, postischemic, toxic or autoimmune injury" (493) appears to be involved at all stages of atherosclerosis. It is implicated in the formation of early fatty streaks, when the endothelium is activated and expresses chemokines, including monocyte chemotactic protein (MCP)-1 and interleukin (IL)-8, and adhesion molecules, including intercellular adhesion molecule (ICAM)-1, vascular adhesion molecule (VCAM)-1, E- and P-selectin, leading to monocyte/lymphocyte recruitment and infiltration into the subendothelium (265). It also acts at the onset of adverse clinical vascular events, when activated cells within the plaque secrete matrix proteases that degrade extracellular matrix proteins and fragilize the fibrous cap, leading to rupture and thrombus formation (399). Cells involved in the atherosclerotic process include vascular (endothelial and smooth muscle) cells, monocytes/macrophages, lymphocytes (T, B, NKT), dendritic cells, and mast cells. They secrete or are stimulated by soluble factors including peptides, glycoproteins, proteases, and a set of cytokines.
The purpose of this review is to bring together the current information concerning the role of cytokines in the development, progression, and complications of atherosclerosis. Specific emphasis is placed on the contribution of pro- and anti-inflammatory cytokines, in modulating innate, adaptive, and regulatory immunity in the context of atherosclerosis. In addition, we discuss the potential of the circulating cytokine levels as biomarkers of (coronary) artery disease. Finally, we propose some novel therapeutic strategies targeting the cytokine network to combat atherosclerosis.
|
II. THE ATHEROSCLEROTIC CYTOKINE NETWORK |
|---|
|
TopPreviousNextReferences |
|---|
Stanley Cohen introduced for the first time the word cytokine in 1974 (132, 133). Until then the term lymphokine, proposed by Dudley Dumonde in 1969, had been used to designate lymphocyte-derived factors and more generally proteins secreted from a variety of cell sources, affecting the growth or function of many types of cells, collectively (181). At the second International Lymphokine Workshop held in 1979, the name interleukin was proposed to characterize proteins with "the ability to act as communication signals between different populations of leukocytes" (473). Later on in 1989, Balkwill and Burke (33) defined cytokine as "one term for a group of protein cell regulators, variously called lymphokines, monokines, interleukins, interferons (we should add "chemokines"), which are produced by a wide variety of cells in the body, play an important role in many physiological responses, are involved in the pathophysiology of a range of diseases, and have therapeutic potential."
Nowadays, the cytokines consist of more than 50 secreted factors involved in intercellular communication, which regulate fundamental biological processes including body growth, lactation, adiposity, and hematopoiesis (77). Cytokines are clustered into several classes: interleukins (33 have been identified to date), tumor necrosis factors (TNF), interferons (IFN), colony stimulating factors (CSF), transforming growth factors (TGF), and chemokines. They are especially important for regulating inflammatory and immune responses and have crucial functions in controlling both innate and adaptive immunity. The predominant actors in adaptive immunity, helper-T (Th) cells, have been categorized on the basis of the pattern of cytokines that they can secrete, resulting in either a cell-mediated immune response (Th1) associated with IL-2 and IFN-
secretion, or a humoral immune response (Th2), associated with IL-4, IL-5, IL-10, and IL-13 secretion.
Cytokines are categorized according to the structural homology of their receptors as class I or class II cytokines (77, 369) (Table 1). Most ILs, CSFs, and IFNs belong to one of these two classes of cytokines, which mediate their effects through the Janus kinase-signal transducers and activators of transcription (JAK-STAT) pathway. Three other major cytokine families encompass the IL-1 family (including IL-1, IL-1, IL-1ra, and IL-18), TNF family, and TGF- superfamily (Table 1). IL-1 and TNF family members activate the nuclear factor-
B (NF-B) and mitogen-activated protein (MAP) kinase signaling pathways, while TGF- superfamily members activate signaling proteins of the Smad family.
|
superfamily is composed of many multifunctional cytokines including TGF-1–2-3, activins, inhibins, anti-Müllerian hormone (AMH), bone morphogenetic proteins (BMPs), and myostatin (540). TGF- family members are secreted as inactive complexes bound to the latency-associated peptide (LAP), a protein derived from the NH2-terminal region of the TGF- gene product. The LAP forms covalent bonds with the latent TGF- binding proteins (LTBPs), high-molecular-weight proteins of which four different isoforms exist (571). The resulting large latent complexes are sequestered within the extracellular matrix. Proteases in the extracellular matrix can digest LTBP, dissociating LAP from TGF-. Cytokines share a number of specific features.
and IFN-, MCP-1 and IL-8) markedly amplifies their activity. This also holds true when a cytokine induces the expression of (an)other cytokine receptor(s). -chain, the IL-6 family cytokines share the gp130 subunit, and the three IFN-
isoforms utilize a heterodimeric receptor composed of its specific receptor subunit IFN-R (or IL-28R) and the subunit IL-10R2 of the IL-10R, also shared with IL-10 and the IL-10-related cytokines, IL-22 and IL-26. One must admit that many of these properties are also shared by growth factors. However, one difference is that the production of growth factors, including PDGF, epidermal growth factor (EGF), fibroblast growth factor (FGF), and vascular endothelial growth factor (VEGF), tends to be constitutive and is not as tightly regulated as that of cytokines. Also, the target cells of growth factors are mainly nonimmune.
Cytokines are often classified according to their pro- (TNF, IL-1, IL-12, IL-18, IFN-) or anti-inflammatory (IL-4, IL-10, IL-13, TGF-) activities. In light of the data obtained from experimental and clinical studies, described below, regarding the pathophysiological role of cytokines in atherosclerosis, we propose to cluster cytokines as pro- or antiatherogenic (Table 2).
|
The NF-B pathway is one of the main signaling pathways activated in response to proinflammatory cytokines, including TNF-, IL-1, and IL-18, as well as following activation of the Toll-like receptors (TLR) by the pattern recognition of pathogen-associated molecular patterns (PAMPs). Activation of this pathway plays a central role in inflammation through the regulation of genes encoding pro-inflammatory cytokines, adhesion molecules, chemokines, growth factors, and inducible enzymes such as cyclooxygenase-2 (COX2) and inducible nitric oxide synthase (iNOS) (166). NF-B is a dimeric transcription factor formed by the hetero- or homodimerization of proteins of the Rel family, including p50 and p65. In its inactive form NF-B is bound to inhibitor of B (I-B/) in the cytoplasm. Proinflammatory cytokines and pathogens act through distinct signaling pathways that converge on the activation of an IB kinase (IKK) complex containing two kinases IKK1/IKK and IKK2/IKK, and the regulatory protein NEMO (NF-B essential modifier, also named IKK) (762); IKK activation initiates IB/ phosphorylation at specific NH2-terminal serine residues (782). Phosphorylated IB is then ubiquitinated, leading to its degradation by the 26S proteasome. This releases NF-B dimers from the cytoplasmic NF-B-IB complex, allowing them to translocate to the nucleus (Fig. 1). Once in the nucleus, NF-B binds to B enhancer elements on specific genes promoting transcription. Targets genes of NF-B include IB, the synthesis of which ensures that NF-B is transiently activated. This negative-feedback regulation gives rise to oscillations in NF-B translocation (496).
|
B is a redox-sensitive transcription factor, and the intracellular redox status of the cell is extremely important in the regulation of NF-B activity (reviewed in Ref. 311). Antioxidants, such as aspirin, N-acetylcysteine (NAC), and flavonoids can therefore inhibit the activation of NF-B. A number of natural constitutive or inducible pathways inhibiting NF-B activity also exist (see review in Ref. 669). For example, A1 and A20, two cytoprotective genes, are induced in response to inflammatory stimuli to protect EC from exaggerated activation and from undergoing apoptosis even when NF-B is blocked (139). A20 terminates NF-B activation by promoting reaccumulation of IB through its interaction with proteins involved in TNF- signaling upstream of IB degradation (375). Consequently, A20-deficient mice fail to terminate TNF-induced NF-B activity, having a persistently active IKK complex that prevents reaccumulation of IB protein, are hypersensitive to TNF-, and suffer from severe inflammation. The inducible form of the heme oxygenase (HO-1) is another example of endogenous anti-inflammatory pathway induced in response to inflammatory stimuli. HO-1 can be upregulated in human EC by TNF and IL-1 (674), and HO-1 possesses potent antiapoptotic and anti-inflammatory properties (742). HO-1 deficiency in humans is associated with the presence of severe and persistent endothelial damage (761). The anti-inflammatory properties of HO-1 seem to be related to an inhibitory action on P- and E-selectin expression on EC (688).
Activated NF-B has been identified in SMC, macrophages, and EC of human atherosclerotic lesions (78, 82, 480). Enhanced endothelial activation of NF-B has been shown to occur in LDLr-deficient mice very early on following a high-fat diet, in regions of the proximal aorta with high probability for atherosclerotic lesion development (262). Furthermore, supershift analysis in cells isolated from human carotid atherosclerotic plaques, composed in majority of macrophages and SMC, demonstrate that activated NF-B consists of p65, c-Rel, and p50, but not relB or p52 subunits (480). NF-B activation in these cells controls the expression of proinflammatory cytokines TNF-; IL-6 and IL-8; matrix metalloproteinases (MMP)-1, -3, and -9; and tissue factor (TF), as shown by their selective inhibition following blockade of the NF-B pathway by overexpression of IB or dominant-negative IKK-2 (480). Interestingly, in this study NF-B inactivation did not affect the expression of the anti-inflammatory cytokine IL-10 or the matrix metalloproteinase inhibitor TIMP-1.
The actual in vivo role of the NF-B pathway has recently been addressed in experimental models of atherosclerosis. Kanters et al. (324), using LDLr-deficient mice with a cell-specific deletion of IKK2 preventing NF-B activation in macrophages, unexpectedly found increased atherosclerotic lesion formation and inflammation in these animals. This result was associated with a significant reduction in the anti-inflammatory and antiatherogenic cytokine IL-10, suggesting that a certain level of NF-B activation is required to modulate the inflammatory reaction and counteract proatherogenic responses (Fig. 2). This finding is in favor of a central role for NF-B in the induction of "protective" antiapoptotic and anti-inflammatory genes, critical to the resolution of the inflammatory process (374). However, the detrimental effect of NF-B inhibition in atherogenesis is likely to depend on how NF-B activity is inhibited. In a subsequent study, Kanters et al. (323) examined the effects of hematopoietic NF-B1 (the p50 subunit of NF-B) deficiency in the development of atherosclerotic lesions, transplanting bone marrow from mice deficient in NF-B1 into irradiated LDLr–/– mice. Instead of promoting the formation of larger inflammatory lesions, as was the case with specific IKK2 deficiency in macrophages, hematopoietic NF-B1 deficiency was associated with a significant decrease in lesion size, despite enhanced accumulation of T and B lymphocytes within the lesions. This could be explained, at least in part, by the observation that in contrast to IKK2 deficiency, NF-B1 deficiency did not alter the inflammatory balance in favor of a proatherogenic phenotype. Despite increased TNF- expression by NF-B1-deficient macrophages, other major proatherogenic molecules such as MCP-1 were downregulated, whereas critical antiatherogenic factors such as IL-10 were significantly upregulated. Decreased MCP-1 production and increased IL-10 expression may have contributed to the limitation of plaque size despite enhanced accumulation of T cells. Another plausible explanation for reduced lesion development in NF-B1-deficient animals could be a defect in the uptake of oxLDL by macrophages, as characteristic foam cells were absent in NF-B1-deficient lesions. Moreover, both scavenger receptor class A (SR-A) expression and uptake of oxLDL were significantly reduced in NF-B1-deficient macrophages stimulated ex vivo with lipopolysaccharide (LPS), although in vivo relevance of this in vitro effect remains to be determined. In summary, NF-B appears to be at the crossroads of the inflammatory response in atherosclerosis, fine-tuning the response of the vessel wall to injury (Fig. 2).
|
AP-1 (activator protein-1) is a transcription factor consisting of homodimers or heterodimers of Fos (c-Fos, FosB, Fra-1 and Fra2), Jun (c-Jun, JunB, JunD), or ATF subunits which recognize either 12-O-tetradecanoylphorbol-13-acetate (TPA) response elements or cAMP response elements (CRE) (626). Jun proteins can homodimerize, but Fos proteins can only form stable dimers with Jun. Phosphorylation of c-Jun by c-Jun NH2-terminal kinases (JNKs) results in enhanced transcriptional activity of complexes containing AP-1 dimers (734).
JNK belongs to the family of stress-activated protein kinases that also includes the p38 protein kinases. Three highly related but distinct gene products, JNK1, JNK2, and JNK3, can be expressed as a total of 10 isoforms as a result of variable mRNA splicing (259). JNK1 and JNK2 show a broad tissue distribution, whereas JNK3 is expressed predominantly in neurons but also in cardiac smooth muscle and the testes (770). Targeted deletion of the genes coding for JNK1 or JNK2 results in abnormal thymocyte selection (588) and loss of T-lymphocyte differentiation and effector function (179). JNK3 knockout mice show resistance to neuronal apoptosis, directly implicating JNK in at least some specific instances of programmed cell death (678, 768).
JNK phosphorylation is mediated by two MAPK kinases (MAPKKs), MAP2K4 (or MKK4) and MAP2K7 (or MKK7), that they can cooperatively activate JNK (Fig. 1). Gene disruption studies in mice demonstrate that both MAP2K4 and MAP2K7 are required for full activation of JNK by environmental stressors and that MKK7 is essential for JNK activation by TNF (677).
Many proinflammatory genes, including those encoding TNF-, IL-2, IL-6, E-selectin, ICAM-1, VCAM-1, MCP-1, COX2, and MMPs-1, -9, -12, and -13 (500), are regulated by the JNK pathway, through interaction of AP-1 with other cis-acting sequences in their promoters and with certain transcription factors that bind to these sequences (Fig. 2).
A recent study showed that atherosclerotic lesions were significantly reduced in JNK2-deficient apoE–/– mice, but not in JNK1-deficient apoE–/– mice, compared with apoE–/– mice (568). JNK2 expression in leukocytes, rather than in vascular cells, appeared to be responsible for this effect. Indeed, transplantation of apoE–/– JNK2–/– bone marrow into apoE–/– mice reduced atherosclerosis to an extent similar to that of apoE–/– JNK2–/– mice transplanted with apoE–/– JNK2–/– bone marrow, whereas apoE–/–JNK2–/– mice transplantedwith apoE–/– bone marrow showed atherosclerotic lesions equivalent to those of apoE–/– mice transplanted with apoE–/– bone marrow (568).
The class I and II cytokines induce homodimerization and activation of their cognate receptors, resulting in the activation of associated JAK kinases (JAK1, JAK2, JAK3, and Tyk2) (Table 1) (520a). The activated JAKs phosphorylate the receptor cytoplasmic domains, which creates docking sites for SH2-containing signaling proteins. Among the tyrosine phosphorylated substrates are members of the STAT family of proteins (Table 1) (520a). Receptor engagement and tyrosine phosphorylation activate the cytosolic inactive STATs, resulting in their nuclear translocation and gene activation. This pathway was originally found to be activated by IFNs, but a number of cytokines, growth factors, and hormonal factors also activate JAK and/or STAT proteins (Fig. 1). In particular, IL-6 binds to the IL-6 receptor -chain and gp130, which activate JAK1 and STAT3. IFN- utilizes JAK1 and JAK2, and usually activates STAT1. It is noteworthy that the anti-inflammatory cytokine IL-10 also activates JAK and/or STAT proteins (reviewed in Ref. 481). The IL-10/IL-10R interaction activates JAK1 and Tyk2, which are associated with the IL-10R1 and IL-10R2, respectively.
STAT3 can be activated by a number of cytokines, especially those of the IL-6 family, mediating the expression of several acute-phase response genes. Yet, STAT3 appears to play a critical negative role in controlling inflammation, as shown in mice with STAT3 deletion in specific cell types, including keratinocytes (594), T cells (666), macrophages/neutrophils (664), cardiomyocytes (309), or endothelial cells (322), STAT3 deficiency being embryonically lethal. STAT3-deficient T cells show severely impaired IL-6-induced cell proliferation, due to the lack of IL-6-mediated prevention of T-cell apoptosis (666). STAT3 deletion in mice within the macrophage/neutrophil lineage results in chronic inflammation and pathological colitis with age, due to the enhancement of the Th1 response by blockade of IL-10 signaling (664). Removal of STAT3 from hematopoietic progenitors also results in increased proinflammatory cytokine production, inflammatory bowel disease, and an expanded macrophage population (732). Interestingly, STAT3-deficient macrophages and neutrophils show increased production of inflammatory cytokines in response to LPS, which cannot be reduced by IL-10 (664). STAT3 activation by IL-10 is therefore central for anti-inflammatory responses in macrophages and neutrophils (Fig. 2). It is noteworthy that mice with conditional STAT3 deletion in endothelium also show exaggerated inflammation and leukocyte infiltration in multiple organs upon LPS challenge (322). An endothelium-derived soluble factor that is dependent on STAT3 is likely to control IFN- production during LPS-induced inflammation (322).
In terms of immunoregulation, STAT4 and STAT6 are crucially important for the differentiation of Th cells. IL-4 activates STAT6 and promotes the differentiation of Th2 cells (634). Conversely, IL-12 activates STAT4 and drives the differentiation of naive T cells into Th1 cells that produce IFN- (325). In atherosclerosis, the Th cell response is of the Th1 type, characterized by abundant secretion of IFN- (264). Yet, Th2 profile does not necessarily offer protection against atherosclerosis and might even be proatherogenic (see sect. VIB3). Therefore, targeting STAT4 and STAT6 could be of use in the treatment of atherosclerosis. Interestingly, statins, which are believed to exert beneficial effects in cardiovascular disease beyond cholesterol lowering (350), have been reported to inhibit Th1-mediated disease and to block activation of STAT4 (386, 492, 778) and induction of major histocompatibilty complex (MHC)-II expression by IFN- (366). Other drugs, including rapamycin and lisofylline, have also been reported to block STAT4 activation (127, 771). Interestingly, a recent study showed that rapamycin reduces atherosclerosis in apoE–/– mice, with concomitant decreased expression of IL-12p40, IFN- and IL-10 mRNA, and enhanced expression of TGF-1 (190). Pentoxifylline, a methylxanthine derivative of lisofylline, has been reported to have protective effects against atherosclerosis in apoE–/– mice, associated with a reduced Th1 polarization of Th lymphocytes (373).
Cytokine signaling by the JAK/STAT pathway is regulated, in part, by a family of endogenous JAK kinase inhibitor proteins termed suppressors of cytokine signaling (SOCS) (748). The SOCS family consists of eight members [SOCS-1 to SOCS-7 and cytokine-inducible SH2 proteins (CIS)] all sharing a central SH2 domain and a COOH-terminal SOCS box. Both SOCS1 and SOCS3 inhibit JAK tyrosine kinase activity; SOCS1 directly binds to the activation loop of JAKs through the SH2 domain, while SOCS3 binds to cytokine receptors (Fig. 2). SOCS1 regulates INF signaling, and deficiency leads to lethal disease, which is characterized by exaggerated effects of IFN-. Interestingly, mice lacking both SOCS-1 and IFN-, though saved from the lethal perinatal syndrome observed in SOCS-1-deficient mice, develop a variety of chronic infections or inflammatory lesions as adults (466). In contrast, SOCS2 regulates growth hormone, and SOCS-knockout mice show gigantism. SOCS3 is preferentially expressed in Th2 cells and plays an important role in regulating the onset and maintenance of Th2-mediated allergic immune disease (619).
Very little is known regarding the role of SOCS in atherosclerosis. It has been reported that SOCS-1 inhibits IFN--induced CD40 expression in macrophages by blocking IFN--mediated STAT-1 activation, and in so doing suppressing IFN--induced TNF- secretion and subsequent NF-B activation (733). Inasmuch as the CD40/CD40L pathway actively participates in plaque development and progression (reviewed in Ref. 605), mimics or inducers of SOCS1 might be useful to attenuate the effects of IFN- in the context of atherosclerosis.
TGF--triggered signals are transduced by proteins belonging to the Smad (for vertebrate homologs of Sma and Mad) family. Smads serve as substrates for TGF- receptors type I and II, in which the cytoplasmic domain possesses serine/threonine kinase activity (453). The type I receptor recognizes and phosphorylates Smad2 and Smad3, which associates with Smad4, forming complexes that participate in DNA binding and recruitment of transcription factors (Fig. 1). Smad3 may also have antagonistic properties, as it plays a critical role in TGF--dependent repression of vascular inflammation by inhibiting AP-1 activity (Fig. 2) (200, 201). In addition to these agonistic Smads, inhibitory Smads (I-Smad) such as Smad6 and Smad7, which associate with activated receptors and interfere with Smad2 and Smad3 binding, are present. Expression of Smad7 is induced by IFN- as a negative regulator of the TGF-/Smad pathway (684). Recent advances in the study of atherosclerosis point to an important role of TGF- signaling in the protection against excessive plaque inflammation, loss of collagen content, and induction of regulatory immunity (see below and reviews in Refs. 243, 444, 445). Immunohistochemistry and RT-PCR anlaysis of human plaques reveal Smad2, Smad3, and Smad4 expression in macrophages of fibrofatty lesions and in SMC of fibrous caps (320). We also detected phosphorylated Smad2 in the aortic sinus of apoE–/– mice, indicative of TGF- activity in atherosclerotic lesions (438).
5. TLR/Myd88 signaling pathways
At least 10 TLRs (TLR1–10) recognize different PAMPs associated with different classes of pathogens (review in Refs. 303, 665). For example, TLR4 recognizes LPS, which is unique to Gram-negative bacteria, and TLR2 recognizes peptidoglycan found in Gram-positive bacteria. TLR9 recognizes unmethylated CpG motifs, which are abundant in prokaryotic genomes and virus DNA. TLR3 recognizes double-stranded RNA (dsRNA) produced during viral infections. TLRs are characterized by a 150-amino acid intracytoplasmic domain named TIR (Toll/IL-1R/R), which they share with members of the IL-1 receptor (IL-1R) family and plant disease resistance (R) genes, and by an extracellular domain composed of NH2-terminal leucine-rich repeats (LRRs) flanked by characteristic cysteine clusters on the COOH-terminal (CF motif) or NH2-terminal (NF motif) side of the LRRs. Upon stimulation, TLRs and the IL-1R family activate the transcription factors NF-B and AP-1, leading to production of proinflammatory cytokines. TIR domains play a critical role in TLR signaling. They allow homophilic interactions with the cytoplasmic factor MyD88 that also contains a TIR domain (Fig. 1). MyD88, which is recruited to the receptors after stimulation, contains an NH2-terminal death domain that enables it to bind the death domain-containing serine-threonine kinases of the IL-1R-associated kinases (IRAK) family (reviewed in Ref. 312). As a result, MyD88 functions as an adaptor between receptors of the TLR or IL-1R families and downstream signaling kinases. Following association of MyD88 with IRAK-4, IRAK-4 is autophosphorylated, dissociates from the receptor complex, and interacts with TNF-receptor-associated factor (TRAF)-6. Once activated, TRAF6 activates a heterodimer composed of two ubiquitination proteins called Uev1A and Ubc13, which triggers its association with the MAP3 kinase TAK1 (716). From TAK1, two signaling pathways diverge; one ultimately leads to NF-B activation and the other to MAP kinase activation. Studies using MyD88-deficient mice showed that this factor is essential for the NF-B-dependent induction of TNF- and IL-6 in response to TLR agonists (331). Interestingly, analysis of MyD88 mutant mice unexpectedly pointed to the existence of a MyD88-independent pathway downstream of some TLRs. Indeed, TLR4- or TLR3-mediated activation of NF-B and AP-1 by LPS and dsRNA, respectively, was not abolished but only delayed in MyD88-deficient mice (13, 331).
Recently, enhanced expression of TLR4 was detected in murine (apoE–/– mice) and human carotid and coronary atherosclerotic plaques (186, 760). TLR1 and TLR2 expression has also been found in human carotid (186) but not in coronary plaques (760). Human epidemiological data demonstrate that an Asp299Gly TLR4 polymorphism, which attenuates receptor signaling, is associated with a decreased risk of atherosclerosis and acute coronary events (18, 64, 335). Functional TLR4 expression has also been correlated with the development of aortic intimal hyperplasia in a mouse model of artery injury (699), and TLR4 activation by LPS increases atherosclerotic plaque formation in the apoE3*Leiden atherosclerotic mouse model (287).
Interestingly, TLR4 appears to be involved in several aspects of the inflammatory response even in the absence of infection, by recognizing endogenous ligands produced during inflammation. Extracellular matrix components, including the type III repeat extra domain A of fibronectin, low-molecular-weight oligosaccharides of hyaluronic acid, and polysaccharide fragments of heparan sulfate, provoke immunostimulatory responses similar to those induced by LPS, via TLR4 (315, 517, 673). Moreover, fibrinogen (642) and minimally modified LDL (mmLDL) (471) are able to induce the production of chemokines and cytokines from macrophages through recognition by TLR4. Together, these recent findings indicate that TLR4 may exert LPS-independent atherogenic activities (468, 469). Two lines of evidence support this hypothesis: 1) oxLDL enhances TLR4 expression in macrophages (760), and 2) TLR4 or its intracellular adaptor protein, MyD88, reduces atherosclerosis in uninfected apoE-deficient mice, concomitant with a marked reduction in macrophage infiltration and MCP-1 expression in the atherosclerotic lesions (54), and decreased circulating levels of IL-12 and MCP-1 (470).
Remarkably, CD4+CD25+ Treg cells selectively express TLR4–5-7–8 (110). This is of particular importance given the role that Treg cells play in atherosclerosis, as we recently reported (8, 437; see sect. VIC).
|
III. INDUCERS OF CYTOKINE PRODUCTION IN ATHEROSCLEROSIS |
|---|
|
TopPreviousNextReferences |
|---|
According to the classical view of inflammation, cytokines are produced by cells of the innate immune system (monocytes, neutrophils, NKT cells) in response to microbial infection, toxic reagents, trauma, antibodies, or immune complexes (493). In the host, TLRs and intracellular proteins (NOD1 and NOD2, for "nucleotide-binding oligomerization domain") act as sensors of the conserved molecular motifs present on a wide range of different microbes, the PAMPs. Hence, cytokines are secondary mediators of inflammation and not the primary triggers. An etiologic role for infectious agents in atherosclerosis, especially Chlamydia pneumoniae and cytomegalovirus (CMV), has been repeatedly evoked (396) since the first seroepidemiologic evidence of an association of the chlamydia TWAR strain with acute myocardial infarction and chronic coronary disease was reported in 1988 (589). However, the most recent clinical trials, including Weekly Intervention with Zithromax for Atherosclerosis and its Related Disorders (WIZARD) (511), Azithromycin in Acute Coronary Syndrome (AZACS) (114), Antibiotic Therapy After Acute Myocardial Infarction (ANTIBIO) (781), Pravastatin or Atorvastatin Evaluation and Infection Therapy (PROVE-IT) (108), and Azithromycin and Coronary Events Study (ACES) (250), assessing the potential benefits of antibiotic therapy with the goal of targeting Chlamydia pneumoniae showed no effect of treatment in patients with CAD. Moreover, experimental studies showed that infection is not necessary for initiation or progression of atherosclerosis in apoE-deficient mice. Atherosclerosis develops identically in germ-free animals and in animals raised with ambient levels of microbial challenge (749). One must therefore conclude that pathogens do not serve as etiologic agents for atherosclerosis, even though one cannot rule out a role in disease exacerbation. Several reports indicate that inoculation of atherosclerosis-prone mice with high doses of C. pneumoniae fosters atherosclerosis (292, 475). Yet, the atherogenic effect of C. pneumoniae requires elevated serum cholesterol levels (292).
Atherosclerosis clearly does not develop in any animal model without a significant level of plasma cholesterol, and the dominant role of cholesterol is also well established in humans. While hypertension, diabetes, and smoking are factors that dramatically increase the risk of atherosclerosis, it is not rare to have clinically significant atherosclerosis in the absence of these risk factors. In contrast, below a certain level of cholesterol (150 mg/dl), atherosclerosis is practically absent in human populations (106), and risk gradually increases with increased plasma cholesterol levels (647). Moreover, primary and secondary clinical trials have established the efficacy of lowering cholesterol with statins for prevention of cardiovascular disease (256, 694). It is therefore tempting to hypothesize that the primary trigger of cytokine release in atherosclerosis has a link with cholesterol. Atherogenic cholesterol exists mainly in the form of LDL, which are the main culprit in CAD. In fact, several lines of evidence support the hypothesis that oxidized lipids, including oxLDL, are the most likely triggering factors for cytokine production.
Quantitative analysis of atherosclerosis in fetal aorta showed that fatty streaks are already present at this early stage of life, lesions being more abundant in fetus from hypercholesterolemic mothers than from normocholesterolemic mothers (489). Interestingly, qualitative analysis of lesions depicted similar distribution of native LDL, oxLDL, and macrophages in lesions of offspring from both hypercholesterolemic and normocholesterolemic mothers. The presence of macrophages alone, without native LDL or oxLDL, or their association with native LDL, was almost never observed, and most of the lesions contained both oxLDL and macrophages. A few lesions with native LDL or oxLDL without macrophages were also present. This seminal study allows us to describe the exact chronology of events leading to fatty streak formation in humans, starting with native LDL uptake by the arterial intima, followed by LDL oxidation and, finally, monocyte recruitment after endothelial activation by oxLDL.
C3H mice, which do not develop atherosclerotic lesions either when fed an atherogenic diet or when crossed with the atherosclerosis prone apoE–/– mice, do not respond to in vivo administration of oxLDL, in contrast to C57BL/6 mice (391). Their EC are not activated in the presence of oxLDL, whereas cells from C57BL/6 mice express M-CSF, MCP-1, and VCAM-1 in the same conditions. Yet, C3H EC respond perfectly well to activation by the proinflammatory cytokines IL-1 and TNF- (628, 629).
oxLDL behaves as a potent inflammatory agent. In vivo administration of oxidized LDL to C57BL/6 mice causes rapid induction of circulating M-CSF and upregulation of genes encoding JE (the murine analog of MCP-1) as well as other inflammatory proteins in various tissues (392). OxLDL stimulates the expression of adhesion molecules on EC (337). OxLDL has chemoattractant activity on monocytes, promotes their differentiation into macrophages, but inhibits their mobility (555, 556). Binding of oxLDL to CD36 triggers the release of proinflammatory cytokines in macrophages (310). In addition, incubating human blood mononuclear cells with oxLDL results in T-lymphocyte activation, as assessed by increased expression of IL-2 receptors and HLA-DR antigens on T lymphocytes (215).
Oxidation of LDL generates many "neo-self determinants" that induce an active immune response (288) and may challenge the regulatory pathways responsible for immune homeostasis. Both humoral and cellular immune responses can profoundly affect atherosclerotic development and progression (268).
The amount of lipid retained in macrophages depends on unregulated uptake of oxidized lipoproteins by scavenger receptors, as first identified by Brown and Goldstein (87), counterbalanced by degradation and efflux.
Altogether these findings point to a role of oxLDL as a very early trigger of vascular inflammation. LDL accumulation and modification in the subendothelium trigger monocyte and lymphocyte recruitment. Thereafter, activated macrophages and lymphocytes secrete abundant amounts of cytokines that in turn can activate EC, SMC, and macrophages/lymphocytes to foster cytokine production, leading to a self-perpetuating inflammatory process that becomes less dependent on the presence of oxLDL. This might explain why oxLDL, while instrumental in triggering the early atherosclerotic events, are less critical in upholding the inflammatory environment. This might also explain in part the efficiency of antioxidant therapies in the prevention of atherosclerosis when these therapies are administered at the very beginning of the atherosclerotic process in animal models, but their failure to do so in most secondary or primary prevention clinical trials in humans (reviewed in Ref. 743), where treatment is administered at later stages of the disease when secondary inflammatory mediators become as important as the initial oxidative-related stimulus. It is noteworthy that atherosclerotic plaques do not regress, or regress very slowly, in cholesterol-fed rabbits following short-term withdrawal of cholesterol feeding and normalization of cholesterol plasma levels (2, 155). It is only after a prolonged cholesterol withdrawal period that decrease in plaque size, together with reduced vascular inflammation and plaque stabilization, is observed (7, 347, 697). In humans, aggressive lipid lowering treatment using statins has been shown to be very effective in limiting plaque development and reducing plaque progression (142, 143, 505, 506, 777). The cytokine network may thus serve as a final common proinflammatory pathway regardless of the initiating event and provides a supplemental therapeutic target, especially in late stages of the disease.
Several oxidized lipids and/or phospholipids are lipid bioactive mediators and may serve as primary triggers of the atherosclerotic process. Bioactive lipids have been identified in the atherosclerotic plaque, including the potent inflammatory mediator platelet activating factor (PAF), PAF-like lipids, oxidized phospholipids (oxPL), and lysophosphatidylcholine (lysoPC) (494). Like oxLDL, PAF induces TNF- production by monocytes (71, 213) and MHC class II dependent IFN- secretion by T lymphocytes (213). Oxidized phospholipids upregulate tissue factor expression in EC (62), as well as in SMC (146). Similarly, lysoPC can enhance IFN- secretion and gene expression in human T lymphocytes (503) and stimulate the production of IL-1 in macrophages (407). It also stimulates ICAM-1 and VCAM-1 expression (362, 793) and induces the release of IL-6 and IL-8 (662) in EC and MCP-1 in EC (663) and SMC (575).
Lipid oxidation products such as lysoPC, 4-hydroxy-2-nonenal (4HNE), and oxysterols are contained in oxLDL (662). Oxidized 1-palmitoyl-2-arachidonyl-sn-glycero-3-phosphorylcholine (OxPAPC), which is present in minimally modified LDL, is a PAF-like lipid that is found in atherosclerotic plaques. OxPAPC, but not native PAPC, is able to stimulate EC to bind monocytes and to secrete MCP-1, IL-8, and growth-related oncogene (GRO)- (376, 565, 773; see review in Ref. 382). Individual lipids identified in OxPAPC include 1-palmitoyl-2-(5-oxovaleroyl)-sn-glycero-3-phosphorylcholine (POVPC), 1-palmitoyl-2-glutaroyl-sn-glycero-3-phosphorylcholine (PGPC), and epoxy-isoprostane-PC (727, 728). Oxidized 1-palmitoyl-2-linoleoyl-sn-glycero-3-phosphorylcholine (PLPC) also promotes monocyte-endothelial interactions (4). Moreover, epoxyisoprostane and epoxycyclopentenone phospholipids have been identified in OxPAPC that induce MCP-1 and IL-8 in EC (653). Oxidized phospholipids can upregulate tissue factor both in EC (62) and in SMC (146).
Interestingly, it has been shown that OxPAPC inhibits the binding of LPS to LPS-binding protein and CD14, which are required for presenting LPS to TLR-4 (61). It is therefore likely that upon acute bacterial inflammation oxidized phospholipids exert anti-inflammatory properties by inhibiting NF-B pathway, while under conditions of chronic inflammation, the pro-inflammatory activity of lipid oxidation products becomes more pathologically relevant (382).
Eicosanoids are well-known lipid mediators of inflammation. They comprise a variety of compounds [prostaglandins, thromboxanes, leukotrienes (LT), hydroxyand epoxy-fatty acids, lipoxins, and isoprostanes] that are derived from arachidonic acid. Leukotrienes are a class of eicosanoids that are derived through the action of the 5-lipoxygenase (5-LO). 5-LO is pivotal for the generation of both proinflammatory (LTB4 and LTC4) and anti-inflammatory (lipoxins) mediators. However, in contrast to their inhibitory effects on PMN and eosinophils, lipoxins are potent stimuli for peripheral blood monocytes, stimulating monocyte chemotaxis and adherence (428). Recent biologic and genetic findings implicate the 5-LO pathway in atherosclerosis (162, 279, 461, 552). Mehrabian et al. (461) reported that heterozygotes for the 5-LO gene on the LDLr–/– background had considerably reduced aortic lesions, compared with the advanced lesions observed in 5-LO+/+LDLr–/– mice, despite equivalent hypercholesterolemia. 5-LO pathway also promotes pathogenesis of hyperlipidemia-dependent aortic aneurysm (787). Furthermore, clinical findings showed that variant alleles of 5-LO genes were associated with a significant increase of carotid intima thickness (184). Most recently, a significant association was drawn between the gene encoding 5-LO activating protein (FLAP) and myocardial infarction by<
