Chapter 1

Atherosclerotic Biology and Epidemiology of Disease

Epidemiology of Cardiovascular Disease

Atherosclerosis, with its complications, is the leading cause of mortality and morbidity in the developed
world. In the United States, a snapshot of the population reveals that 60 million adults currently suffer from
atherosclerotic cardiovascular disease, which accounts for 42% of all deaths annually, at a cost to the nation
of $128 billion. Fortunately, despite this catastrophic burden of disease, much evidence has emerged over the
last decade suggesting that the progression of atherosclerosis can be slowed or even reversed in many people
with appropriate lifestyle and drug interventions.
The origin of the current epidemic of cardiovascular disease can be traced back to the time of
industrialization in the 1700s. The three factors largely responsible for this were an increase in the use of
tobacco products, reduced physical activity, and the adoption of a diet high in fat, calories, and cholesterol.
This rising tide of cardiovascular disease continued into the twentieth century, but began to recede when data
from the Framingham study identified a number of modifiable risk factors for cardiovascular disease,
including cigarette smoking, hypertension, and hypercholesterolemia (1).
The number of deaths per 100,000 attributable to cardiovascular disease peaked in the Western world in 1964
to 1965, since which time there has been a gradual decline in death rates (Fig. 1.1) (2). The age-adjusted
coronary heart disease (CHD) mortality in the United States dropped by more than 40% and cerebrovascular
disease mortality by more than 50%, with the greatest reductions being seen among whites and men. This
reduction has occurred despite a quadrupling of the proportion of the population older than 65 years of age
and has been due to a number of factors, particularly major health promotion campaigns aimed at reducing
the prevalence of Framingham risk factors. Indeed, there has been a substantial change in the prevalence of
population cardiovascular risk factors over the last 30 years (Table 1.1). The war is not won, however, and
the decline in the death rate from cardiovascular disease slowed in the 1990s (Fig. 1.2). This is likely owing
to a large increase in the prevalence of both obesity and type 2 diabetes mellitus, as well as a resurgence of
cigarette smoking in some sectors of society (3). Female death rates from cardiovascular disease overtook
male death rates in 1984 and have shown a smaller decline over the last 30 years (4). The consequences of
atherosclerosis are also beginning to be felt in less well-developed regions of the globe (5), with death from
atherosclerotic cardiovascular disease set to replace infection as the leading cause of death in the Third World
in the near future. This phenomenon is further illustrated by the increase in CHD mortality in countries of
Eastern and Central Europe (most notably countries of the former Soviet Union). For example, in the
Ukraine the age standardized death rate in the year 2000 was just over 800 per 100,000 people representing
an increase of over 60% when compared to 1990 (6).
A further note of caution should also be struck. Western countries are experiencing a dramatic increase in the
prevalence of heart failure. In the United States, almost 5 million people carry a diagnosis of heart failure (7),
thus singling it out as an emerging epidemic (8). However, the determinants of this epidemic have yet to be
fully elucidated, with some epidemiologic studies pointing toward hypertension as the driving factor (9) and
others suggesting CHD as the predominant cause (10).

Biology of Atherosclerosis

Traditionally, atherosclerosis has been viewed as a degenerative disease, affecting predominantly older
people, slowly progressing over many years, and eventually leading to symptoms through mechanical effects
of blood flow. The perceived insidious and relentless nature of its development has meant that
a somewhat pessimistic view of the potential to modify its progression by medical therapy has held sway.
There has been little emphasis on the diagnosis and treatment of high-risk asymptomatic patients. Disease
management has instead been dominated by interventional revascularization approaches, targeting the largest
and most visible or symptomatic lesions with coronary angioplasty or bypass surgery.

Recently, for several reasons, this defeatist view of the pathogenesis and progression of atherosclerosis has
begun to change. First, careful descriptive studies of the underlying pathology of atherosclerosis have
revealed that atherosclerotic plaques differ in their cellular composition and that the cell types predominating
in the plaque can determine the risk of fatal clinical events. A high degree of plaque inflammation is
particularly dangerous. Second, recent epidemiologic work has identified many new potentially modifiable
risk factors for atherosclerosis, above and beyond those highlighted as a result of the Framingham study (11).
The third and most important reason is because several large-scale clinical trials have reported that drugs in
particular, the HMG-CoA reductase inhibitors (statins) are able to reduce the number of clinical events in
patients with established atherosclerosis and do so without necessarily affecting the size of atherosclerotic
plaques. These three strands of evidence have shown that, rather than being an irreversibly progressive
disease, atherosclerosis is a dynamic, inflammatory process that may be amenable to medical therapy.
Understanding the cellular and molecular interactions that determine the development and progression of
atherosclerosis brings with it opportunities to develop novel therapeutic agents targeting key molecular and
cellular interactions in its etiology. In addition, the recognition that the clinical consequences of
atherosclerosis depend almost entirely on plaque composition argues for a new approach to diagnosis, with
less emphasis placed on the degree of lumen narrowing and more interest in the cellular composition of the
plaque.
TABLE 1.1 Temporal Changes in Coronary Risk Factors
Cigarette smoking 1960 Men, 55%; women, 33%

1990 Men, 30%; women, 27%

Undiagnosed hypertension 1960 52%
1980 29%
Mean serum cholesterol 1960 225mg/dL
1990 208 mg/dL
Diabetes mellitus 1970 2.6%
1990 9.1%
Sedentary lifestyle 1970 41%
1985 27%
Obesity 1960 25%
1990 38%

Normal Artery

The healthy artery consists of three histologically distinct layers. Innermost and surrounding the lumen is the
tunica intima, which comprises a single layer of endothelial cells in close proximity to the internal elastic
lamina. The tunica media surrounds the internal elastic lamina, and its composition varies depending on the
type of artery. The tunica media of the smallest arterial vessels, arterioles, comprises a single layer of
vascular smooth muscle cells (VSMCs). Small arteries have a similar structure but with a thicker layer of
medial VSMCs. Arterioles and small arteries are termed resistance vessels because they contribute vascular
resistance and, hence, directly affect blood pressure. At the opposite end of the spectrum are large elastic or
conduit arteries, named for the high proportion of elastin in the tunica media. The tunica media of all arteries
is contained within a connective tissue layer that contains blood vessels and nerves and that is known as the
tunica adventitia. In normal arteries, the vessel lumen diameter can be altered by contraction and relaxation
of the medial VSMCs in response to a variety of systemic and locally released signals.

Atherosclerotic Vessel

Atherosclerosis is primarily a disease affecting the intimal layer of elastic arteries. For reasons that remain
largely unknown, some arterial beds appear more prone than others. Coronary, carotid, cerebral, and renal
arteries and the aorta are most often involved. The arteries supplying the lower limbs are also vulnerable to
disease. Interestingly, the internal mammary artery is almost always spared, making it an invaluable vessel
for coronary bypass surgery.
Atherosclerotic lesions develop over many years and pass through several overlapping stages. Histologically,
the earliest lesion is a subendothelial accumulation of lipid-laden macrophage foam cells and associated T
lymphocytes known as a fatty streak. Fatty streaks are asymptomatic and nonstenotic. Postmortem
examinations have shown that they are present in the aorta at the end of the first decade of life, are present in
the coronary arteries by the second, and begin to appear in the cerebral circulation by the third decade. With
time, the lesion progresses and the core of the early plaque becomes necrotic, containing cellular debris,
crystalline cholesterol, and inflammatory cells, particularly macrophage foam cells. This necrotic core
becomes bounded on its luminal aspect by an endothelialized fibrous cap, consisting of VSMCs embedded in
an extensive collagenous extracellular matrix. Inflammatory cells are also present in the fibrous cap,
concentrated particularly in the shoulder regions, where T cells, mast cells, and especially macrophages have
a tendency to accumulate. Advanced lesions may become increasingly complex, showing evidence of
calcification, ulceration, new microvessel formation, and rupture or erosion (12). Microvessels within the
plaque may play important roles in the formation of macrophage-rich vulnerable atheroma by providing an
extended surface area of activated endothelial cells to hasten recruitment of further inflammatory cells as
well as by promotion of intraplaque hemorrhage (13).

Thus, the composition of atherosclerotic plaques is variable, dynamic and complex, and it is the interaction
between the various cell types within a plaque that determines the progression, complications, and outcome
of the disease.

Cellular Roles in Atherogenesis

Endothelial Cells

The endothelium plays a central role in maintaining vascular health by virtue of its vital anti-inflammatory
and anticoagulant properties. Many of these characteristics are mediated by the nitric oxide (NO) molecule.
This molecule was discovered in the 1980s, having been isolated from lipopolysaccharide-primed
macrophages (14). NO is synthesized by endothelial cells under the control of the enzyme endothelial NO
synthase (NOS) and has a number of anti-atherogenic properties. First, it acts as a powerful inhibitor of
platelet aggregation on endothelial cells. Second, it can reduce inflammatory cell recruitment into the intima
by abrogating the expression of genes involved in this process, such as those encoding intercellular adhesion
molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), P-selectin, and monocyte
chemoattractant protein-1 (MCP-1) (15,16,17). There is some evidence that NO may also reduce lipid entry
into the arterial intima (18). NO is also a potent anti-inflammatory molecule and, depending on
concentration, may be a scavenger or a producer of potentially destructive oxygen free radicals, such as
peroxynitrite (19,20,21). The earliest detectable manifestation of atherosclerosis is a decrease in the
bioavailability of NO in response to pharmacologic or hemodynamic stimuli (22). This may occur for two
reasons. Either there may be decreased manufacture of NO because of endothelial cell dysfunction, or
increased NO breakdown may take place. There is evidence that both mechanisms may be important in
different situations (23). Many atherosclerosis risk factors can lead to impaired endothelial function and
reduced NO bioavailability. For example, hyperlipidemic patients have reduced NO-dependent
vasodilatation, which is reversed when patients are treated with lipid-lowering medication (24). Diabetics
also have impaired endothelial function, occurring primarily as a result of impaired NO production. There is,
however, some evidence to suggest that increased oxidative stress leading to enhanced NO breakdown may
also be a factor in early endothelial dysfunction (25). Similarly, other risk factors for atherosclerosis, such as
hypertension and cigarette smoking, are associated with reduced NO bioavailability (26,27). In cigarette
smokers, endothelial impairment is thought to be caused by enhanced NO degradation by oxygen-derived
free-radical agents such as the superoxide ion. There are also other consequences of an increased reactivity
between NO and superoxide species. The product of their interaction, ONOO (peroxynitrite), is a powerful
oxidizing agent and can reach high concentrations in atherosclerotic lesions. This may result in cellular
oxidative injury.
Another consequence of endothelial cell dysfunction that occurs in early atherosclerosis is the expression of
surface-bound selectins and adhesion molecules, including P-selectin, ICAM-1, and VCAM-1. These
molecules attract and capture circulating inflammatory cells and facilitate their migration into the
subendothelial space (22). Normal endothelial cells do not express these molecules, but their appearance may
be induced by abnormal arterial shear stress, subendothelial oxidized lipid, and, in diabetic patients, the
presence of advanced glycosylation products in the arterial wall. The importance of selectins and adhesion
molecules in the development of atherosclerosis is demonstrated in experiments using mice, which lack their
expression. These animals develop smaller lesions with a lower lipid content and fewer inflammatory cells
than control mice when fed a lipid-rich diet (28). Animal models have reinforced the importance of
inflammatory cell recruitment to the pathogenesis of atherosclerosis, but because inflammatory cells are
never seen in the intima in the absence of lipid, the results suggest that subendothelial lipid accumulation is
also necessary for the development of atherosclerosis.
The tendency for atherosclerosis to occur preferentially in particular sites may be explained by subtle
variations in endothelial function. This is probably caused by variations in local blood flow patterns,
especially conditions of low flow, which can influence expression of a number of endothelial cell genes,
including those encoding ICAM-1 and endothelial NOS (29,30). In addition to flow speed, flow type can
have a direct effect on cell morphology. In areas of laminar flow (atheroprotective flow), endothelial cells
tend to have an ellipsoid shape, contrasting with the situation found at vessel branch points and curves,
where turbulent flow (atherogenic flow) induces a conformational change toward polygonal-shaped cells
(31). Such cells have an increased permeability to low-density lipoprotein (LDL) cholesterol and may
promote lesion formation (32).
These data are consistent with the idea that the primary event in atherogenesis is endothelial dysfunction.
The endothelium can be damaged by a variety of means, leading to dysfunction and, by unknown
mechanisms, subsequent subendothelial lipid accumulation. In this situation, the normal homeostatic features
of the endothelium break down; it becomes more adhesive to inflammatory cells and platelets, it loses its
anticoagulant properties, and there is reduced bioavailability of NO. Importantly, endothelial function is
improved by drugs that have been shown to substantially reduce death from vascular disease, including
statins and angiotensin-converting enzyme inhibitors (33,34).

Inflammatory Cells

LDL from the circulation is able to diffuse passively through the tight junctions that bind neighboring
endothelial cells. The rate of passive diffusion is increased when circulating levels of LDL are elevated. In
addition, other lipid fractions may be important in atherosclerosis. Lipoprotein(a) has the same basic
molecular structure as LDL, with an additional apolipoprotein(a) element attached by a disulfide bridge. It
has been shown to be highly atherogenic (35), accumulate in the arterial wall in a manner similar to LDL
(36), impair vessel fibrinolysis (37), and stimulate smooth muscle cell proliferation (38). The accumulation
of subendothelial lipids, particularly when at least partly oxidized, is thought to stimulate the local
inflammatory reaction that initiates and maintains activation of overlying endothelial cells. The activated
cells express a variety of selectins and adhesion molecules and also produce a number of chemokines in
particular, MCP-1, whose expression is upregulated by the presence of oxidized LDL in the subendothelial
space (39). Interestingly, the protective effect of high-density lipoprotein (HDL) against atherosclerotic
vascular disease may be partly explained by its ability to block endothelial cell expression of adhesion
molecules (40,41). Chemokines are proinflammatory cytokines responsible for chemoattraction, migration,
and subsequent activation of leukocytes. Mice lacking the MCP-1 gene develop smaller atherosclerotic
lesions than normal animals (42). The first stage of inflammatory cell recruitment to the intima is the
initiation of rolling of monocytes and T cells along the endothelial cell layer. This phenomenon is mediated
by the selectin molecules, which selectively bind ligands found on these inflammatory cells. The subsequent
firm adhesion to and migration of leukocytes through the endothelial cell layer depends on the endothelial
expression of adhesion molecules such as ICAM-1 and VCAM-1 and their binding to appropriate receptors
on inflammatory cells. Once present in the intima, monocytes differentiate into macrophages under the
influence of chemokines such as macrophage colony-stimulating factor. Such molecules also stimulate the
expression of the scavenger receptors that allow macrophages to ingest oxidized lipids and to develop into
macrophage foam cells, the predominant cell in an early atherosclerotic lesion. The formation of scavenger
receptors is also regulated by peroxisome proliferator-activated receptor-γ (PPAR-γ a nuclear transcription
factor expressed at high levels in foam cells) (43). PPAR-γ agonists (glitazones), which are used to treat
patients with type 2 diabetes, have been shown to have many anti-atherogenic effects, including increasing
production of NO (44), decreased endothelial inflammatory cell recruitment and reduced vascular endothelial
growth factor (VEGF) expression (45). Also, PPAR-γ agonists can reduce the lipid content of plaques by
enhancing reverse cholesterol transport from plaque to liver. Positive results with these drugs in patients with
type 2 diabetes are emerging. As well as reducing matrix metalloproteinase (MMP) 9 levels, glitazones also
significantly ameliorated C-reactive protein (CRP) and CD40 ligand levels, as well as causing direct plaque
regression in a rabbit atheroma model (46). Clearly their use in large clinical trials in patients without
diabetes as anti-atheroma drugs is awaited with interest.
In early atherosclerosis at least, the macrophage can be thought of as performing a predominantly beneficial
role as a neutralizer of potentially harmful oxidized lipid components in the vessel wall. However,
macrophage foam cells also synthesize a variety of proinflammatory cytokines and growth factors that
contribute both beneficially and detrimentally to the evolution of the plaque. Some of these factors are
chemoattractant (osteopontin) and growth-enhancing (platelet-derived growth factor) for VSMCs (12,47).
Under the influence of these cytokines, VSMCs migrate from the media to the intima, where they adopt a
synthetic phenotype, well-suited to matrix production and protective fibrous cap formation.
However, activated macrophages have a high rate of apoptosis. Once dead, they release their lipid content,
which becomes part of the core of the plaque, thereby contributing to its enlargement. The apoptotic cells
also contain high concentrations of tissue factor, which may invoke thrombosis if exposed to circulating
platelets (48). Interestingly, the selective glycoprotein 2b3a receptor antagonist abciximab has been shown to
have an effect on the levels of tissue factor found in monocytes. In an in vitro study by Steiner (49), the drug
attenuated both the amount of tissue factor and its RNA levels. As tissue factor is a potent instigator of the
clotting cascade, this role may explain part of the protective effect of abciximab on the microcirculation of
patients with acute coronary syndromes (50).
It is now generally recognized that the pathologic progression and consequences of atherosclerotic lesions
are determined by dynamic interactions between inflammatory cells recruited in response to subendothelial
lipid accumulation, and the local reparative wound healing response of surrounding VSMCs.

Vascular Smooth Muscle Cells

VSMCs reside mostly in the media of healthy adult arteries, where their role is to regulate vascular tone.
Thus, medial VSMCs contain large amounts of contractile proteins, including myosin, α-actin, and
tropomyosin. Continued expression of this contractile phenotype is maintained by the influence of
extracellular proteins in the media, which act via integrins in the VSMC membrane. In atherosclerosis,
however, the cells become influenced by cytokines produced by activated macrophages and endothelial cells.
Under these influences, VSMCs migrate to the intima and undergo a phenotypic change characterized by a
reduction in content of contractile proteins and a large increase in the number of synthetic organelles. This
migration of VSMCs from the media to the intima, and the consequent change from a contractile to a
synthetic phenotype, was previously thought to be a crucial step in the development of atherosclerosis in the
modified response to injury hypothesis discussed previously. More recently, it has been recognized that
intimal VSMCs in atherosclerotic plaques bear a remarkable similarity to VSMCs found in the early
developing blood vessels (51), suggesting that intimal VSMCs may be performing a beneficial, reparative
role rather than a destructive one in atherosclerosis. VSMCs are well-equipped for this action. First, they can
express the proteinases that they require to break free from the medial basement membrane and allow them
to migrate to the site of inflammation or injury in response to chemokines. Second, they can produce various
growth factors, including VEGF and platelet-derived growth factor, that act in an autocrine loop to facilitate
their proliferation at the site of injury. Finally, and most important, they produce large quantities of matrix
proteins, in particular glycosaminoglycans, elastin, and collagen isoforms 1 and 3, necessary to repair the
vessel and form a fibrous cap over the lipid-rich core of the lesion. This fibrous cap separates the highly
thrombogenic lipid-rich plaque core from circulating platelets and the proteins of the coagulation cascade
and also confers structural stability to the atherosclerotic lesion. And because the VSMC is the only cell
capable of synthesizing this cap, it follows that VSMCs play a pivotal role in maintaining plaque stability
and protecting against the potentially fatal thrombotic consequences of atherosclerosis (52).
Cellular Interactions and Lesion Stability

Generally, early atherosclerosis progresses without symptoms until a lesion declares itself in one of two
ways. As discussed, macrophage foam cells may undergo apoptosis, especially in the presence of high
concentrations of oxidized LDL. Their cellular remnants then become part of an enlarging lipid-rich core.
Plaque size thus increases, and there may be a consequent reduction in vessel lumen area. At times of
increased demand, such as exercise, this may be sufficient to cause ischemic symptoms such as angina. More
hazardous is if the plaque presents with disruption of the fibrous cap, leading to exposure of the
thrombogenic lipid core. This is likely to result in subsequent platelet accumulation and activation, fibrin
deposition, and intravascular thrombosis. Depending on factors such as collateral blood supply, extent of
arterial thrombus, and local fibrinolytic activity, the end result may be arterial occlusion and downstream
necrosis.
By studying the pathology of ruptured plaques, several characteristics have been identified that seem to be
predictive of the risk of rupture in individual lesions (53). Plaques that are vulnerable to rupture tend to have
thin fibrous caps (<65 µm) with a high ratio of inflammatory cells to VSMCs and contain a lipid core that
occupies more than 50% of the volume of the plaque. Of these, the most important is the cellular
composition of the fibrous cap. Plaques containing a heavy inflammatory cell infiltrate and relatively few
VSMCs have the highest risk of rupture (54). Interestingly, in the coronary tree, such high-risk plaques are
usually located in the proximal portions of the main arteries (55). Recently, evidence of the importance of
intraplaque hemorrhage from microvessels in plaque destabilization has emerged. It is thought that the
leaking vessels initiate platelet and red cell phagocytosis by plaque macrophages, causing them to become
activated and tipping the balance toward plaque rupture (56).
Inflammatory cells in plaques act to promote plaque rupture by a number of synergistic mechanisms. First,
activated T cells produce proinflammatory cytokines, typified by IFN-γ, that directly inhibit VSMC
proliferation (57) and almost completely shut down collagen synthesis (58,59). Thus, VSMCs in the vicinity
of activated T cells in plaques are poorly able to lay down or repair extracellular matrix. Second,
macrophage-derived inflammatory cytokines, in particular interleukin-1β and tumor necrosis factor-α along
with IFN-γ from T cells, are synergistically cytotoxic for VSMCs, causing depletion in cell number by
apoptosis (60). These cytokines are found at high levels in vulnerable plaques (61). Third, activated
macrophages can induce VSMC apoptosis by direct cell cell contact (62). Finally, and probably most
important, macrophages secrete a variety of matrix metalloproteinases that degrade the matrix components of
the fibrous cap by proteolytic cleavage of its protein components (52). The production of matrix
metalloproteinases is upregulated by inflammatory mediators such as tumor necrosis factor-α. As well as
being under threat from such an array of insults, VSMCs themselves within the fibrous cap of a mature
plaque have a reduced ability to proliferate (63,64) and an enhanced susceptibility to apoptosis (65). Thus,
inflammatory cells can destroy the fabric of the fibrous cap, and resident VSMCs are poorly equipped to
compensate, particularly in the presence of inhibitory inflammatory cytokines. It is important to note that all
of these features can be present in small, hemodynamically insignificant plaques that are clinically silent and
angiographically invisible. Thus, plaque composition is far more important than plaque size in determining
outcome.

Inflammatory Markers in Atherosclerosis

The cell biology of plaque development and subsequent rupture illustrates that atherosclerosis is
fundamentally an inflammatory condition. Confirmation of this inflammatory basis has come from several
studies of different patient populations that have all demonstrated a correlation between levels of markers of
systemic inflammation, principally CRP, and risk of a clinical event owing to plaque rupture
(66,67,68,69,70). However, unlike in other systemic inflammatory conditions, such as rheumatoid arthritis,
levels of CRP in atherosclerosis are characteristically not elevated above the conventional normal range, and
a correlation between CRP level and coronary events was demonstrated only after development of a highly
sensitive assay for CRP that was capable of measuring levels below the lower limit of detection of
conventional assays. The risk of clinical events associated with an elevated CRP seems to be independent of
the presence of other Framingham risk factors for atherosclerosis. Elevated CRP also predicts near-term
plaque rupture events as well those up to 20 years in the future, suggesting that inflammation is important in
both early and late atherosclerosis (71). Additionally, CRP level accurately indicates the likelihood of sudden
cardiac death, a condition usually associated with multiple atherosclerotic plaque ruptures (72). However,
despite initial enthusiasm, large meta-analyses have suggested that the relative risk of a cardiovascular event
is increased by only approximately 1.5 times in those people with a baseline elevated CRP (above 3 mg/dL)
(73,74). With such a modest predictive value, it may be that the routine measurement of CRP alone in
asymptomatic patients is not yet justified for accurate disease prediction.
Similar, although less compelling, correlations with clinical events have also been published for other
markers of inflammation, including soluble ICAM-1, VCAM-1, P-selectin, and interleukin-6 (the primary
driver of CRP production) (75,76,77,78). Results of these studies have been interpreted by some as indicating
that atherosclerosis arises as a consequence of a systemic inflammatory process (e.g., chronic infection) and
by others that it reflects the inflammatory processes of atherosclerosis itself. However, there is accumulating
evidence in favor of the latter interpretation.

Two Forms of Plaque Disruption: Fibrous Cap Rupture and Endothelial Erosion

Atherosclerotic plaques become life threatening when they initiate clot formation in the vessel lumen and
disturb blood flow. This can occur in two different ways. Either there can be fibrous cap rupture, with
consequent exposure of the thrombogenic extracellular matrix of the cap and the tissue factor rich lipid core
to circulating blood, or less commonly, there is erosion of the endothelial cells covering the fibrous cap, also
potentially leading to the formation of a platelet-rich thrombus. Endothelial erosion probably accounts for
approximately 30% of acute coronary syndromes overall and seems particularly common in women (79).
Both forms of plaque disruption invariably lead to local platelet accumulation and activation. This may result
in triggering of the clotting cascade, thrombus formation, and, if extensive, complete vessel occlusion.
Platelet-rich thrombus contains chemokines and mitogens, in particular platelet-derived growth factor and
thrombin that induce migration and proliferation of VSMCs from the arterial media to the plaque and
transforming growth factor-β that contributes to healing of the disrupted lesion (80). Platelets also express
CD40 on their cell membrane, which causes local endothelial cell activation, resulting in the recruitment of
more inflammatory cells to the lesion and perpetuating the cycle of inflammation, rupture, and thrombosis.
However, fibrous cap rupture or erosion does not invariably lead to vessel occlusion. Up to 70% of plaques
causing high-grade stenosis contain histologic evidence of previous subclinical plaque rupture with
subsequent repair (81). This is particularly likely to occur if high blood flow through the vessel prevents the
accumulation of a large occlusive thrombus. Thus, nonocclusive plaque rupture induces formation of a new
fibrous cap over the organizing thrombus, which restabilizes the lesion but at the expense of increasing its
size. Because this occurs suddenly, there is little opportunity for adaptive remodeling of the artery, and the
healed lesion may now impede flow sufficiently to produce ischemic symptoms. This explains why patients
who have previously had normal exercise tolerance may suddenly develop symptoms of stable angina
pectoris. It also follows that if lesions can grow as a consequence of repeated episodes of silent rupture and
repair, an inhibition of plaque rupture rate will reduce progression of atherosclerosis. Therefore,
atheromatous plaques may become larger by two methods. The first is a gradual increase in size as a
consequence of macrophage foam cell accumulation and incorporation of apoptotic cells into an enlarging
necrotic lipid-laden plaque core. The second is a stepwise increase in size because of repeated, often silent
episodes of plaque rupture or erosion with subsequent VSMC-driven repair.

Balance of Atherosclerosis: Therapeutic Implications

Atherosclerosis is a dynamic process in which the balance between the destructive influence of inflammatory
cells and the reactive, stabilizing effects of VSMCs determines outcome (Fig. 1.3). This balance can be
tipped toward plaque rupture by factors such as an atherogenic lipoprotein profile, high levels of lipid
oxidation, local free radical generation, and genetic variability in expression and activity of certain central
inflammatory molecules. For example, an association between plaque progression and a polymorphism in the
stromelysin-1 gene promoter has been described (82). Until recently, it was also thought that infectious
organisms might be involved in atherosclerosis, either as plaque initiators or as having some role in initiating
plaque rupture. Chlamydia pneumoniae is found in plaques, localizing at high concentrations within
macrophages, but is rarely found in normal arteries (83). Although these data imply a pathologic association
between the presence of chlamydia infection and atherosclerosis, neither a causative role nor an association
between serum markers of infection and ischemic heart disease has been established. Although animal work
has shown that healthy rabbits nasally inoculated with chlamydia develop extensive atherosclerosis (84), the
situation appears to be somewhat different in humans. Two large prospective studies and an extensive meta-
analysis of previous data failed to show any association between serum markers of infection with chlamydia
and incidence of or mortality from ischemic heart disease (85,86). These results effectively excluded a strong
association but allowed the possibility of a weaker link. This hypothesis has now been effectively rejected
after several negative trials of antibiotics in coronary artery disease (87,88,89).

The balance can be tipped toward plaque stability by a reduction in plaque inflammation or an increase in
VSMC-driven repair. Lipid reduction, by whatever means, reduces clinical events. Evidence that this may be
due to a plaque-stabilizing effect comes from animal studies that showed that statins reduced inflammatory
cell and increased VSMC content of plaques (90,91), changes that would be expected to enhance stability.
Dietary lipid lowering in rabbits also reduced the number of microvessels in the aortic intima, suggesting
another mechanism of favorably altering the biology of plaques (92).

More important, however, evidence from human clinical studies also points to a plaque-stabilizing effect of
statins. Despite angiographic studies showing that statins produce only a small, hemodynamically
insignificant reduction in lumen stenosis (93,94), more sensitive intravascular ultrasound studies have shown
beyond doubt that statins can halt lesion enlargement in the coronary arteries, with the most benefit being
seen with higher doses of the most potent drugs (95). Statins can also reduce new lesion formation, and,
importantly, the number of new vessel occlusions. These arise after a plaque ruptures, leading to an occlusive
thrombus in the context of a well-collateralized myocardial circulation. This seems to imply that statins
stabilize plaques by reducing rupture rate. This conclusion is supported by the results of all the large primary
and secondary prevention studies, which have demonstrated that statins (pravastatin, simvastatin, and
lovastatin) produce major reductions in events owing to plaque rupture, such as myocardial infarction and
stroke (34,96,97,98,99). Because statins have only a modest effect on plaque size but cause profound
reductions in the number of clinical events, these studies highlight the inadequacy of angiography for the
prediction of clinical events and suggest that statins have beneficial effects on plaque inflammation in
addition to, or as a result of, their lipid-lowering effects. Importantly, this notion is supported by the
observation that the reduction in clinical events due to statin therapy is accompanied by a parallel reduction
in CRP levels that is unlikely to be caused by effects of statins on nonatherosclerotic inflammation
(100,101). Also, in the first study of its kind, it has been shown that statins reduce inflammation and increase
plaque collagen content in human carotid artery atherosclerosis (102). However, the various statins do differ
in their anti-inflammatory effect; in the REVERSAL study, atorvastatin achieved a far greater reduction in
CRP than pravastatin (95); whether this has important clinical relevance is not yet known.

Statin drugs may help to stabilize plaques in a number of different ways. It is known that they can exert
direct effects on endothelial cell function, inflammatory cell number and activity, VSMC proliferation,
platelet aggregation, and thrombus formation (103,104,105,106,107). Evidence that non lipid-lowering
effects may be important in vivo comes from animal studies in which pravastatin caused beneficial changes
in plaque composition (but not size), even when lipid levels were maintained at pretreatment levels (91).
Additionally, in mice, simvastatin has direct anti-inflammatory effects comparable to those of indomethacin
(108). Recently, a newly recognized effect of statins as immune modulators has been described, whereby
major histocompatibility complex class II mediated T-cell activation is reduced by a variety of statins (109).
However, the matter of non lipid-lowering effects of statins is not yet proven beyond doubt: several of the
pleiotropic anti-inflammatory effects of statins (decreased expression of MMPs, and tissue factor) occur in
animals on a lipid-lowering diet alone, without exposure to drugs of any kind (110). In addition, the
administration of other forms of anti-inflammatory drugs to patients with atherosclerosis does not seem to
confer any clinical benefit and may do harm; the cyclooxygenase-2 (COX-2) class of drugs are a case in
point, causing a doubling of the rate of myocardial infarction in one study (111).

Restenosis

Restenosis describes the late loss of gain in lumen diameter achieved immediately after balloon dilatation of
an atherosclerotic plaque. For many years, it has been thought of as an undesirable response to vascular
injury. However, in effect, it represents an extreme form of plaque stabilization. Whether performed on a
stable or unstable plaque, balloon angioplasty causes endothelial disruption and often substantial damage to
the full thickness of the vessel wall. The initial thrombotic response that would otherwise lead to early vessel
occlusion is prevented by antiplatelet and antithrombotic therapy. There then follows a reparative response
driven by medial VSMCs and adventitial myofibroblasts. The former form a matrix-rich neointima over the
exposed plaque, whereas the latter produce a collagenous matrix in the adventitia. The net result is that the
adventitial reaction splints the vessel and prevents the positive remodeling that would normally allow
expansion of the vessel to accommodate the neointima. However, although this phenomenon may lead to
angiographic or clinical restenosis, much more important, it renders the lesion stable, making the likelihood
of a further plaque rupture at that site extremely remote. In effect, by stimulating a vigorous VSMC repair
response, balloon angioplasty tips the balance of atherosclerosis in favor of plaque stability. This
phenomenon undoubtedly underlies the success of angioplasty in the treatment of acute myocardial
infarction. Most of the adverse effects of the response to balloon angioplasty on remodeling can be countered
by deployment of a stent, particularly the drug-eluting variety, where significant restenosis is rarely
encountered. The drugs used to coat the stents are antiproliferative agents, and are highly effective at
eliminating restenosis (112). However, by impairing the synthetic ability of the VSMCs of the cap, there
have been reports of early thrombotic occlusions of treated arteries, although longer term analysis of the data
suggest that this is not frequent (113). Nevertheless, drug-eluting stents are likely to become universally used
in the catheter laboratory in the near future.

Controversies and Personal Perspectives

Many issues concerning the initiation and progression of atherosclerosis remain to be resolved. In particular,
controversy persists over the extent to which endothelial dysfunction precedes or is the consequence of
intimal lipid accumulation; the relative contributions of endothelial erosion and plaque rupture to clinical
events; and the extent to which statins achieve their plaque-stabilizing effects directly via lipid lowering or
by their so-called pleiotropic effects on the intercellular interactions that lead to plaque rupture. Integral to
this last issue is the outstanding question of what is the optimal level of lipid reduction. In other words, is
lower LDL always better?

Despite these controversies, it is certain that drug treatment will become increasingly prominent in the
management of patients with, and at high risk of developing, atherosclerosis. Improvements in drug design
will come from a number of complementary approaches. First, improvement will come by modifications of
existing molecules, based on understanding how currently available drugs such as statins and angiotensin-
converting enzyme inhibitors influence plaque progression. This will include evaluation of how other lipid-
modifying strategies, such as inhibiting cholesterol absorption in the gut and modifying the balance between
pro- and anti-atherogenic lipoproteins and triglycerides, might influence the atherosclerotic process. Second,
improvements will come by targeting molecular interactions known to be involved in atherogenesis. Likely
candidates include endothelial adhesion molecules, MMPs, inflammatory cytokines and their signaling
molecules, in particular, nuclear factor-κ B and its downstream transcriptional activators. Here the challenge
lies in identifying pathways or molecular species that are specific for atherosclerosis whose modification will
not compromise the normal inflammatory response to pathogens. This approach will include developing
regulators of VSMC behavior, such as modulators of transforming growth factor-β driven matrix production,
that may lead to enhanced maintenance of the fibrous cap. Another important example includes establishing
the role of drugs targeting peroxisome proliferator activated receptors in modifying inflammation and the
vascular consequences of the metabolic syndrome that links insulin resistance, diabetes, hypertension, and
dyslipidemia with premature atherosclerosis. The third approach is to use new technologies such as
proteomics to design new therapeutic molecules and gene array technologies to identify new molecular
targets in vascular disease. In addition, as a consequence of sequencing the human genome, a number of
orphan receptors have already been identified that might provide vascular-specific targets for novel therapies.
Finally, local drug delivery to high-risk plaques with drug-eluting stents has been proposed as a means of
reducing risk of rupture (plaque passivation) (115,116). This approach will need better methods of
identifying high-risk plaques, which will probably include invasive imaging data derived from IVUS and
thermography coupled with noninvasive methods such as high-resolution molecular magnetic resonance
imaging and possibly Fluorodeoxyglucose positron emission tomography (FDG-PET) (117,118).

The Future

It is almost inconceivable that advances in our understanding of the atherosclerotic disease process will not
lead to the development of new anti-atheroma drugs that will act synergistically with statins and angiotensin-
converting enzyme inhibitors. For example, a novel HDL-like molecule has recently been shown to reduce
atheroma burden when given by intravenous infusion over 5 weeks to a high-risk group of patients (114).
Furthermore, we predict that advances in genetics and diagnostics will combine with therapeutic advances to
produce substantial reductions in premature cardiovascular deaths. Thus, new gene polymorphisms and
mutations will be identified that confer increased likelihood either of developing atheroma or of experiencing
its consequences. This will lead, in turn, to better prescription of lifestyle modifications and better targeting
of current and new therapies for primary prevention of cardiovascular events. This approach will be led by
new diagnostic tests based on specific circulating markers of vascular inflammation and imaging of the
inflammatory process underlying plaque rupture that will allow better preclinical diagnosis of patients at
greatest risk of cardiovascular events and subsequent monitoring of plaque-modifying therapies.