Atherosclerosis is a disease of the vessel wall involving lipid accumulation, chronic inflammation, cell death, and thrombosis that causes heart disease and stroke. Although elevated cholesterol levels are a recognized risk factor for atherosclerosis, a growing number of studies suggest that oxidized phospholipids may also play an important role in this condition.1,2 Phospholipids, essential components of lipoproteins and cell membranes, are composed of fatty acids bound to a glycerol backbone containing a polar head group. They are susceptible to free-radical or enzymatic oxidation by myeloperoxidase, lipoxygenase, and other enzymes that are present in the vessel wall. The addition of oxygen to the polyunsaturated fatty acids produces prostaglandin-like molecules, some of which then decompose and fragment to form additional bioactive molecules.
Oxidized phospholipids accumulate under conditions of oxidative stress during viral infections and in inflammatory conditions such as rheumatoid arthritis and atherosclerosis; they are also generated in apoptotic and necrotic cells.(1,2) Oxidized, but not native, phospholipids can interact with specific receptors that mediate atherogenesis. In addition, oxidized phospholipids contain reactive groups that can bind covalently to proteins, forming lipid–protein adducts. These modified proteins become dysfunctional, which can contribute to atherosclerosis. Phospholipid oxidation elicits an immune response by creating new epitopes that are recognized by antibodies of innate immunity, such as E06.(3) Thus, oxidized phospholipids are fundamentally distinct from unoxidized phospholipids in their ability to interact with cells, proteins, and the immune system in order to promote atherogenesis.
In vivo studies in human tissue have demonstrated the accumulation of oxidized phospholipids in the vessel wall at all stages of atherosclerosis, from early fatty streaks (in infants of mothers with hyperlipidemia) to advanced complex lesions, suggesting that these lipids may contribute to all stages of atherogenesis. In vitro studies and studies in which oxidized phospholipids were injected into animals have demonstrated that specific oxidized phospholipids can mediate many atherogenic processes — from the earliest entry of monocytes into the vessel wall to thrombus formation

A Model of the Roles of Oxidized Phospholipids in the Development of Atherosclerosis.
Low-density lipoprotein (LDL) moves into the subendothelial space and becomes oxidized (Panel A). Inflammatory cells are recruited into the vessel wall, take up the oxidized LDL through scavenger receptors, and become foam cells (Panel B). The cell membranes of apoptotic cells continue to recruit inflammatory cells into the vessel wall (Panel C). Oxidized phospholipids also promote thrombosis through the modulation of thrombotic factors (Panel D). Modified Lp(a) lipoprotein, which accumulates in atherosclerotic lesions, can be detected at higher levels in the blood after angioplasty with the use of E06, an antibody that recognizes oxidized phospholipids (Panel E).

Oxidized phospholipids activate the endothelium to bind monocytes (but not neutrophils) and cause the endothelial cells and smooth-muscle cells to produce the potent monocyte chemoattractant protein 1 and the differentiation factor macrophage colony-stimulating factor. (Oxidized phospholipids can also induce responses that protect cells from oxidative stress and inhibit some acute, neutrophil-mediated inflammatory responses.(2))
In vivo, the presence of monocyte-binding molecules and chemotactic factors causes monocytes to migrate into the subendothelial space and to differentiate into macrophages. These macrophages can then release additional reactive oxygen species, further oxidizing low-density lipoprotein to a form that is recognized by scavenger receptors on macrophages and on smooth-muscle cells; this uptake results in the formation of foam cells. Oxidized phospholipids, either free or adducted to apolipoprotein B-100, are recognized by the CD36 scavenger receptor.(4) Furthermore, these phospholipids bind to C-reactive protein and could promote foam-cell formation through the Fcg receptor.
As atherogenesis progresses (in response to cytokines produced by activated endothelial cells and macrophages), smooth-muscle cells proliferate, enter the intima, and form foam cells. Specific oxidized phospholipids at low concentrations stimulate the proliferation of smooth-muscle cells. Ultimately, the foam cells die by necrosis or apoptosis, and a necrotic core is formed. At higher concentrations, oxidized phospholipids have been shown to regulate smooth-muscle apoptosis by increasing the level of ceramide and facilitating the release of cytochrome c from mitochondria. All the while, the inflammation continues, with further entry of monocytes and lymphocytes into the vessel wall. This continuing entry may be facilitated by oxidized phospholipids that are present in the membranes of apoptotic and necrotic cells.
Ultimately, the plaque may rupture or erode, causing a thrombus to form. Key enzymes in the coagulation pathway are also targets of oxidized phospholipids, which increase the expression of tissue factor in endothelial cells, while decreasing the expression of thrombomodulin and the activity of tissue-factor–pathway inhibitor. Platelet activation is also stimulated by oxidized phospholipids. Thus, oxidized phospholipids have proatherogenic effects on all vascular-wall cells.
Although many of the studies cited above were performed in vitro, there is growing evidence that oxidized phospholipids have a role in atherogenesis in vivo. Knocking out or inhibiting receptors that recognize oxidized phospholipids (including the platelet-activating–factor [PAF] receptor, CD36, and toll-like receptors 2 and 4) leads to a decrease in experimental atherosclerosis. Knocking out 12/15 lipoxygenase, an enzyme that oxidizes polyunsaturated fatty acids, also results in decreased atherosclerosis. Levels of myeloperoxidase, another oxidative enzyme, are correlated with the risk of coronary artery disease. High-density lipoprotein (HDL) has been shown to play a protective role in atherogenesis and alters the metabolism of oxidized phospholipids. HDL contains proteins (such as apolipoprotein A-I [apo A-I]) and enzymes (such as lecithin–cholesterol acyltransferase, paraoxonase, and PAF–acetylhydrolase) that can prevent the formation of oxidized phospholipids or destroy them once they have formed. Apo A-I transfers the phospholipids to HDL for destruction. Knocking out paraoxonase or PAF–acetylhydrolase increases atherosclerosis.
The same enzymes associated with HDL that destroy oxidized phospholipids are also inhibited by them, creating a balance so that in the absence of continued inflammation, HDL maintains enough functioning apo A-I and enzyme activity to be antiinflammatory. During an acute-phase response (e.g., after surgery) or during a chronic response (e.g., a chronic systemic inflammation such as atherosclerosis), the balance can shift, and HDL can become proinflammatory. In animal models of atherosclerosis, the balance has been shifted back by the transgenic or adenovirus-mediated expression of high concentrations of apo A-I or the exogenous administration of apo A-I or apo A-I–mimetic peptides.(5)
The study by Tsimikas et al., reported in this issue of the Journal (pages 46–57), demonstrated a correlation between the levels of oxidized phospholipids in the blood and levels of Lp(a) lipoprotein. The investigators also determined that increased levels of Lp(a) lipoprotein and oxidized phospholipids, in particles containing apolipoprotein B-100, correlated with the risk of coronary artery disease and that combined hypercholesterolemia plus increased levels of either oxidized phospholipids or Lp(a) lipoprotein greatly increased the odds of coronary artery disease. Thus, this study is the first to establish a causal connection between the levels of oxidized phospholipids and the risk of coronary artery disease.
In summary, phospholipids are ubiquitous molecules that are important to the structural integrity of cells and lipoproteins. When oxidized, however, they can promote inflammation, are taken up by scavenger receptors on macrophages, and are recognized by the innate immune system. Studies suggest that proteins and enzymes that remove or destroy oxidized phospholipids prevent atherosclerosis and that proteins and enzymes that produce or retain oxidized phospholipids promote atherosclerosis. Thus, oxidized phospholipids may be a diagnostic marker of coronary artery disease or may represent a potential target for therapeutic intervention.

Source Information: Dr. Berliner is a professor of medicine and pathology and Dr. Watson is an assistant professor of medicine at the David Geffen School of Medicine, University of California, Los Angeles.

References

1) Berliner JA, Subbanagounder G, Leitinger N, Watson AD, Vora D. Evidence for a role of phospholipid oxidation products in atherogenesis. Trends Cardiovasc Med 2001;11:142-147.
2) Leitinger N. Oxidized phospholipids as modulators of inflammation in atherosclerosis. Curr Opin Lipidol 2003;14:421-430.
3) Binder DJ. The role of natural antibodies in atherosclerosis. J Lipid Res (in press).
4) Podrez EA. Identification of a novel family of oxidized phospholipids that serve as ligands for the macrophage scavenger receptor CD 36. J Biol Chem 2002;277:38503-38516.
5) Navab M, Ananthramaiah GM, Reddy ST, et al. The oxidation hypothesis of atherogenesis: the role of oxidized phospholipids and HDL. J Lipid Res 2004;45:993-1007.