Atherosclerosis Pathology: Definition, Etiology, Epidemiology

A study of coronary arteries of 63 hearts obtained from deceased fetuses, infants, children, and adolescents found that coronary intimal thickening begins in fetuses and progresses to atherosclerosis in the pediatric population and adolescents. ^[17]  The most common sites of intimal thickening were near bifurcation sites in the left anterior descending coronary artery (55.6%) and in areas free of bifurcation in the right coronary artery (75%). The extent of intimal thickening was significantly associated with older ages in this population. ^[17]

The early lesions consist of two nonatherosclerotic intimal lesions referred to as adaptive intimal thickening and intimal xanthoma ("fatty streak" in the AHA classification) (see the image below). Intimal xanthoma denotes a lesion rich in foamy macrophages without extracelullar lipid pools. Adaptive intimal thickening is present from birth and grow in areas of low shear stress, and are consist mainly of smooth muscle cells in a proteoglycan rich matrix.

Observations from experimental models and autopsy studies in young human subjects suggest that monocyte adherence to the endothelial surface and transmigration into the intima occur as the earliest events in the development of atherosclerotic lesions. Adaptive intimal thickening is characterized by retention of modified lipoproteins within the proteoglycan rich matrix in the intima. The initiation of adhesion increases the expression of selectins, which facilitates the rolling of monocytes, followed by firm attachment by endothelial integrins. Low density lipoprotein (LDL) oxidation, a critical step in atherosclerosis development, has been shown to occur through induction of lipoxoygenases, myeloperoxidases, inducible nitric oxide synthase and nicotinamide adenine dinucleotide phosphate (NADPH) oxidase. (See the image below.)

Pathologic Intimal Thickening

Pathologic intimal thickening (PIT) or type III lesions in the AHA classification are thought to represent the earliest of the progressive plaques. This lesion is primarily composed of smooth muscle cells near the lumen and matrix consisting chiefly of proteoglycan and type III collagen. Focal areas of accumulated lipid ("lipid pools") are found localized toward the abluminal medial wall as areas devoid of smooth muscle cells but rich in proteoglycans.

It is thought that lesions of PIT that show the presence of macrophages are at a more advanced stage, as demonstrated by Nakashima et al in their study of early coronary lesions progression near branch points. ^[18] This study demonstrated that PIT and intimal xanthoma occur simultaneously and are not truly separable lesions. A variable number of T lymphocytes are also observed at this stage, but a true necrotic core is absent. Areas of lipid pools may also contain free cholesterol appearing as cholesterol clefts on paraffin stained sections.

Although the precise origin of the "lipid pool" is debatable, studies suggest that the loss of smooth muscle cells (death by apoptosis) may be involved, as their remnant basement membranes can be visualized by periodic acid Schiff (PAS) staining and show microcalcification. In addition, the confirmation of lipids by oil red O staining is highly suggestive of a lipid retention process facilitated by select proteoglycans and oxidation, which may lead to activation of factors responsible for apoptosis.

Progression to Complex Atherosclerotic Lesions

The following generally describes the progression of atherosclerotic lesions. Also, see the image below.

• Fibroatheroma

• Thin cap fibroatheroma ("vulnerable plaque") and plaque rupture

• Necrotic core expansion and risk for plaque rupture

• Intraplaque hemorrhage and necrotic core expansion

• Healed plaque rupture (HPR)

• Erosion

  

  Atherosclerosis pathology. Simplified scheme for classifying atherosclerotic lesions modified from the current American Heart Association (AHA) recommendations. The boxed areas represent the seven categories of lesions. Dashed lines are used for two categories, because there is controversy over the role that each plays in the initial phase of lesion formation, and both "lesions" can exist without progressing to a fibrous cap atheroma (ie, AHA type IV lesion). The processes responsible for progression are listed between categories. Lines (solid and dotted; dotted lines represent the least-established processes) depict current concepts of how one category may progress to another, with the thickness of the line representing the strength of supportive evidence that the events occur. (Reproduced with permission fromVirmani R, Kolodgie FD, Burke AP, Farb A, Schwartz SM. Lessons from sudden coronary death: a comprehensive morphological classification scheme for atherosclerotic lesions. Arterioscler Thromb Vasc Biol. May 2000;20(5):1262-75.)

Fibroatheroma

The fibrous cap atheroma is the first of the advanced lesions of coronary atherosclerosis by the AHA classification scheme. Its defining feature is the presence of a lipid-rich necrotic core encapsulated by collagen rich fibrous tissue. The fibrous cap atheroma may result in significant luminal narrowing and is also prone to complications of surface disruption, thrombosis, and calcification. The origin and development of the core is fundamental to understanding the progression of coronary artery disease. The fibrous cap consists of collagen, smooth muscle cells, and proteoglycan with varying degrees of inflammatory cells—mostly macrophages and lymphocytes. The thickness of the fibrous cap distinguishes the fibroatheroma (relatively thick) from the thin fibrous cap atheroma (classic "vulnerable" plaque).

Recognition of early necrosis is identified by macrophage infiltration within lipid pools associated with a substantial increase in free cholesterol and breakdown of extracellular matrix, presumably by matrix metalloproteinase (MMP) activity. This, together with the death of macrophages in the setting of defective phagocytic clearance of apoptotic cells, is thought to contribute to the development of late plaque necrosis. Ultimately, the size of the necrotic core is a strong predictor of lesion vulnerability. ^{[18, 19]}

Thin cap fibroatheroma ("vulnerable plaque") and plaque rupture

The thin cap atheroma is thought to be a precursor lesion to plaque rupture and is characterized by a necrotic core overlaid by a thin, fibrous cap (65 µm or less), which is heavily infiltrated by macrophages and T lymphocytes. The density of macrophages at the site of rupture is typically very high, although in some cases macrophages may be sparse. However, because plaque rupture is responsible for 76% of fatal coronary events associated with thrombi in sudden coronary death patients, identification of the thin cap atheroma is critical. A common mechanism of disruption of the fibrous cap atheroma occurs via the thinning or weakening of the fibrous cap, resulting in fissures and ruptures.

Plaque rupture is defined as an area of fibrous cap disruption in which the overlying thrombus is in contact with the underlying necrotic core. The fibrous cap is composed of type I collagen with varying degrees of macrophages and lymphocytes and very few, if any, alpha-actin positive smooth muscle cells. The luminal thrombus is platelet rich at the rupture site. Plaque ruptures are most prevalent in the proximal coronary artery near branch points and are frequently found in the proximal left anterior descending coronary artery, followed by the right and left circumflex coronary arteries.

The causes of plaque rupture are poorly understood, but responsible factors include expression of factors that weaken the fibrous cap, such as MMPs, enzymes (eg, myeloperoxidases) produced by inflammatory cells, high shear regions, stress points, macrophage calcification, and iron deposition. Data are also beginning to emerge that demonstrate critical differences in gene expression between stable and unstable atherosclerotic plaques. ^[20] In one of these studies, differential expression of 18 genes associated with lesion instability included the metalloproteinase ADAMDEC1, retinoic acid receptor responser-1, cysteine protease legumain (a potential activator of MMPs) and cathepsins. ^[20]

More recently, in an autopsy study, investigators reported an association between B lymphocytes and macrophages in the perivascular adipose tissue (PvAT) with coronary atherosclerosis. ^[21] They concluded that the "density of CD20+ B lymphocytes and CD68+ macrophages in periplaque PvAT was increased with plaque size, and the CD68+ macrophages were greater near unstable atherosclerotic plaques than near stable lesions." Moreover, there was more intense inflammation in the periplaque PvAT than in the PvAT distal to the atherosclerotic plaques. ^[21]

Necrotic core expansion and risk for plaque rupture

Expansion of the necrotic core is an important pathogenic process contributing to plaque vulnerability. The presenting inflammatory stimuli for macrophage recruitment into lipid pools are poorly understood along with the respective signaling pathways for subsequent apoptotic cell death and necrosis. Evidence exists pointing toward the involvement of the endoplasmic reticulum (ER) stress pathway, or so-called unfolded protein response (UPR), as the primary mechanisms of macrophage cell death in plaques. This pathway promotes the death of macrophages—the resultant accumulation of dead macrophages coupled with defective phagocytic clearance has been cited as one of the principal factors causing necrotic core expansion.

Intraplaque hemorrhage and necrotic core expansion

Data from the authors' laboratory provide evidence that repeat intraplaque hemorrhage is a contributing factor to necrotic core expansion, as red blood cells are a rich source of free cholesterol, which is an important constituent of ruptured plaques. ^[22] The red blood cells are enriched with lipids constituting 40% of their weight and free cholesterol content within membranes exceeding all other cell types. The expression of glycophorin-A (a protein exclusive to red blood cell membranes) within the necrotic cores of advanced coronary atheroma is strongly positive, whereas its presence in plaques with early necrosis or pathologic intimal thickening remains absence or low.

Intraplaque hemorrhage likely occurs from leaky vasa vasorum that infiltrate the plaque as the lesion thickness increases. The authors have reported that microvessel density is increased in advanced plaques compared with early plaques. Microvessels in normal and atherosclerotic arteries are thin-walled, with compromised structural integrity characterized by poor endothelial junctions. ^[23] Therefore, intraplaque hemorrhage together with the death of macrophages in the setting of defective phagocytic clearance of apoptotic cells is thought to contribute to the development of necrotic core in advance stage plaques.

Healed plaque rupture

Morphologic studies suggest that plaque progression beyond 40-50% cross-sectional luminal narrowing occurs secondary to repeated ruptures. Ruptured lesions with healed repair sites, namely, healed plaque ruptures (HPRs) as shown by Mann and Davies, are easily detected microscopically by the identification of breaks in the fibrous cap with an overlying repair reaction consisting of proteoglycans and/or collagen, depending on the phase of healing. ^[24] Early-healed lesions are rich in proteoglycan, which are eventually replaced by type I collagen.

The prevalence of silent ruptures in the clinical population is unknown. Few angiographic studies have demonstrated plaque progression, and short-term studies have suggested that thrombosis is the likely cause. Mann and Davies showed that the frequency of healed plaque ruptures increases along with lumen narrowing. ^[24] In plaques with 0-20% diameter stenosis, the incidence of healed plaque ruptures was 16%; in lesions with 21-50% stenosis, the incidence was 19%; and in plaques with >50% narrowing, the incidence was 73%. ^[24]

The authors have shown a high frequency of healed plaque ruptures in the coronary arteries from patients dying suddenly with severe coronary disease. ^[25] The percentage of cross-sectional luminal narrowing was dependent on the number of healed repair sites.

Erosion

Although plaque rupture is the most common cause of coronary thrombosis, acute coronary syndromes may occur in the absence of rupture. As mentioned earlier, thrombi may occur as a result of 3 different events: plaque rupture, plaque erosion, or, rarely, a calcified nodule (see Etiology). Plaque erosion is characterized by absence of the endothelium at the site of erosion, with exposed intima composed of smooth muscle cells and proteoglycans, as well as typically minimal inflammation.

In a series of 20 patients who died with acute myocardial infarction, van der Wal et al found plaque ruptures in 60% of lesions with thrombi, whereas the remaining 40% showed "superficial erosion." ^[26] The term "erosion" was chosen because the luminal surface beneath the thrombus lacked endothelial cells. In these lesions, the thrombus was confined to the most luminal portion of the plaque, and there was an absence of ruptures following serial sectioning of these lesions.

In addition, the authors' laboratory studied nearly 100 cases of sudden coronary death and found that 60% of all thrombi could be attributed to plaque rupture and 40% to erosions. Morphologically, major differences exist in the cellular composition of ruptured versus erosion lesions. Unlike the prominent fibrous cap inflammation described in ruptures, eroded surfaces contain few macrophages (rupture 100% vs erosion 50%, P< 0.0001) and T lymphocytes (rupture 75% vs erosion 32%, P< 0.004). Cell activation, indicated by human lymphocyte antigen (HLA)-DR staining, was identified in macrophages and T cells in 89% of plaque ruptures and in 36% of plaque erosions (P = 0.0002). ^[27] The smooth muscle cells near the erosion site appeared "activated," often displaying bizarre shapes with hyperchromatic nuclei and prominent nucleoli. The incidence of calcification was also less common in erosion than in ruptures.

The authors have also shown that more than 85% of thrombi in erosions exhibited late stages of healing characterized by acute inflammatory cell lysis, invasion by smooth muscle cells and/or endothelial cells, or organized layers of smooth muscle cells and proteoglycans with varying degrees of platelet/fibrin layering, whereas in ruptures only one half of thrombi showed healing. ^[28] Postmortem coronary thrombi superimposed on eroded plaques have been shown to contain a higher density of myeloperoxidase-positive cells than those superimposed on ruptured plaques. ^[29] Also, circulation blood myeloperoxidase levels are elevated in patients with acute coronary sinus with erosion compared with those with rupture, suggesting that elevations in selective inflammatory biomarkers may reflect specific acute coronary events.

Probe electrospray ionization mass spectrometry appears to have the potential to detect new biomarkers of atherosclerosis (eg, cholesterol sulfate, phospholipid PE18:0/24:0), although current findings are from animal models (rabbits). ^[30]

Comparison of plaque rupture and plaque erosion

Both clinical and morphologic differences are widely apparent between plaque rupture and erosion. Beginning with age, patients with plaque rupture tend to be significantly older (53 ± 10 y) than those with erosion (44 ± 7 y) (P< 0.02). Survival is also a critical factor, because an estimation of fatal ruptures in the fifth decade of life is 17 per 100,000 per year compared with 6 per 100,000 for plaque erosion.

Although the relationship between risk factors and culprit plaques is similar between women and men, the proportion of women younger than the age of 50 years dying suddenly with plaque erosion is remarkably higher. Plaque burden expressed as the percentage of cross-sectional area stenosis excluding the thrombus is greater in plaque ruptures (78 ± 12%) than erosions (70 ± 11%) (P< 0.03), whereas eccentric plaques are more common in erosions. Unlike the prominent fibrous cap inflammation described in ruptures, eroded surfaces contain fewer macrophages and T lymphocytes. Taken together, eroded plaques tend to be eccentric lesions rich in smooth muscle cells and proteoglycans with very little inflammation or calcification.