Objectives: The purpose of this study was to investigate which histological changes associated with risk factors could contribute to the progression from the initial atherosclerotic lesions including fatty streaks to the advanced lesions.Methods: We examined the associations of histomorphometric findings in the determined anatomical sites of mid-thoracic aortas (TAs) and left anterior descending coronary arteries (LADs) with major risk factors for atherosclerosis, using a young autopsied series from the the Pathobiological Determinants of Atherosclerosis in Youth (PDAY) study. The histological classification by the American Heart Association was graded for 1013 TAs and 1009 LADs. Histometric study, including immunohistochemistry, was performed in type 2 lesions (fatty streaks) of TAs from 59 subjects and LADs from 45 ones.Results: For the progression from the initial lesions into the advanced atherosclerotic lesions, the most effective lipid profiles were low plasma HDL-C in TA and elevated serum non-HDL-C in LAD. This lipid profile of each artery correlated with number or density of intimal smooth muscle cell-derived foam cells, respectively. The serum concentration of non-HDL-C correlated with macrophage foam cells in TAs. Hypertension and hyperglycemia were associated with increase of intimal area and/or collagen content in both arteries, but not with either types of foam cell proliferation. Smoking correlated with increased collagen content in TAs.Conclusion: There were histologically different ways of progressing from fatty streaks to advanced atherosclerotic lesions depending on the risk factors. For the atherosclerosis progression from type 2 lesions to advanced lesions, increase in number of smooth muscle cell-derived foam cells could be an important indicator. (C) 2011 Elsevier Ireland Ltd. All rights reserved.

Objective— We investigated the influence of genetic variants (rare and common) in the gene encoding periostin ( POSTN ) on atherosclerosis as measured in arterial specimens from the Pathobiological Determinants of Atherosclerosis in Youth (PDAY) study. Methods and Results— A comprehensive survey of common POSTN variants (87 single-nucleotide polymorphisms [SNPs]) in PDAY subjects (n=2527) identified numerous SNPs associated with raised lesions in abdominal aorta and with fatty streaks in thoracic aorta. These SNPs belonged to a small number of correlation bins that spanned the entire locus. To examine effects of rare variants, we resequenced POSTN functional regions in PDAY cases with raised lesions (n=291) and controls with no raised lesions (n=294). However, we found no significant associations with case-control status for carriers of POSTN rare variants using the weighted-sum method for rare variant analysis. Conclusion— We identified common variants in POSTN that are associated with arterial lesions in young persons from the PDAY study. This finding strongly supports a role for periostin in atherogenesis, as suggested by recent proteomics analysis that found abundant expression of periostin in atherosclerotic lesions. Genetic variation may influence atherosclerosis via periostin's known involvement in multiple relevant pathways, including angiogenesis, vascular remodeling, and stimulation of migration and differentiation of vascular smooth muscle cells.

Heparin cofactor II (HCII) is a serine protease inhibitor (serpin) that has been shown to be a predictor of decreased atherosclerosis in the elderly and protective against atherosclerosis in mice. HCII inhibits thrombin in vitro and HCII-thrombin complexes have been detected in human plasma. Moreover, the mechanism of protection against atherosclerosis in mice was determined to be the inhibition of thrombin. Despite this evidence, the presence of HCII in human atherosclerotic tissue has not been reported. In this study, using samples of coronary arteries obtained from the Pathobiological Determinants of Atherosclerosis in Youth (PDAY) study, we explore the local relationship between HCII and (pro)thrombin in atherosclerosis. We found that HCII and (pro)thrombin are co-localized in the lipid-rich necrotic core of atheromas. A significant positive correlation between each protein and the severity of the atherosclerotic lesion was present. These results suggest that HCII is in a position to inhibit thrombin in atherosclerotic lesions where thrombin can exert a proatherogenic inflammatory response. However, these results should be tempered by the additional findings from this, and other studies, that indicate the presence of other plasma proteins (antithrombin, albumin, and alpha(1)-protease inhibitor) in the same localized region of the atheroma.

We have established a strong correlation between PARP activation and markers of oxidative stress in a mouse model of atherosclerosis. The present study presents evidences establishing an active role for PARP in human atherosclerosis. Human aortic atherosclerotic plaques (AHA grade 2 and 3) obtained from the PDAY study were subjected to immunostaining to detect iNOS (increased nitric oxide production), nitrotyrosine (nitration of proteins), 8‐oxo‐dG (oxidative DNA damage), PAR (PARP activation), active caspase‐3 (apoptotic cell death) and CD‐68 (macrophage/foam cells). These markers were selected to test the hypothesis that nitration of protein induced by increased nitric oxide production leads to DNA damage and PARP activation which ultimately participates in cell death and inflammation in atherosclerosis. PARP activation was localized within and in the vicinity of plaques suggesting active role of this enzyme in plaque dynamics. PARP activation correlated strongly with the presence of nitrotyrosine, iNOS and 8‐oxo‐dG as analyzed by double immunofluorescence. Moderate correlation was observed between PAR and active caspase immunoreactivity. Little correlation between PARP activation and CD‐68 was observed in this study. Overall the data suggest that PARP is activated in human atherosclerotic plaques at early stages and strongly correlates with markers of protein/DNA damage and cell death.

Atherosclerosis begins in childhood or adolescence and progresses during the young adult years to cause clinical coronary heart disease (CHD) in middle-aged and older individuals. This article reviews evidence regarding the association of the major established CHD risk factors with atherosclerosis in adolescents and young adults, with emphasis on the findings of the Pathobiological Determinants of Atherosclerosis in Youth (PDAY) study. Age, non-high-density lipoprotein cholesterol concentration, smoking, hypertension, obesity, and hyperglycemia are positively associated with atherosclerotic lesions, whereas female gender and high-density lipoprotein cholesterol concentration are negatively associated with lesions. The PDAY study developed a risk score that provides a weighted summary of the risk factor effects on advanced atherosclerosis. Although developed to predict advanced atherosclerotic lesions, the risk score is also associated with all stages of lesion severity, including the transition from normal tissue to the earliest anatomically detectable lesion. Application of the PDAY risk score to data from longitudinal studies of risk factors in young adults shows that early measurement of risk factors predicts atherosclerosis assessed noninvasively up to 15 years later, and that subsequent change in risk score during the 15-year interval also predicts atherosclerosis. These findings provide strong support for maintaining a low lifetime risk and for focusing on preventing atherosclerosis as the most effective way to prevent CHD. Risk-factor control beginning in the late teenage years will retard development of the earliest stage of atherosclerosis and its progression, and will reduce or delay occurrence of CHD.

Background: The histologically topographic comparisons on atherosclerosis progression among three anatomical sites, mid-thoracic and lower abdominal aorta and left anterior descending coronary artery (LAD) were performed using a young population (age 15-34 years) from the Pathobiological Determinants of Atherosclerosis in Youth (PDAY) study.Methods and results: The histological classification based on the American Heart Association grading scheme showed that in the thoracic aorta type 2 lesions (numerous macrophage foam cells with fine particles but no pools of extracellular lipid) appeared in the first 10-year age group, with no significant change in prevalence in the next 10 years. Lesions greater than type 2 were rarely seen in the thoracic aorta. Although type 2 lesions appeared later in the LAD than in the aorta, the lesions within the LAD progressed rapidly to more advanced lesions (types 4 and 5) or atheroma. Lesion development in the abdominal aorta was intermediate to lesion development in the thoracic aorta and the LAD.Conclusions: The most striking topographic difference on lesion progression among the three anatomical sites was the vulnerability of type 2 lesions to progress into advanced lesions. The histology study, including immunohistochemistry limited to the type 2 lesions suggested that lesion progression was related to the intimal thickness and the amount of collagen but not to the number of macrophage foam cells. (c) 2007 Elsevier Ireland Ltd. All rights reserved.

Background: Disease is sometimes best studied by examination of tissue obtained from autopsied individuals. Results derived from autopsied persons are considered biased because many factors influence the decision to perform an autopsy, and variables measured postmortem may be affected by changes immediately prior to death and emergency medical treatment. Methods: The Pathobiological Determinants of Atherosclerosis in Youth (PDAY) study measured coronary heart disease risk factors postmortem in autopsied young adults 23-34 years of age dying from external causes (accidents, homicides, suicides). The Coronary Artery Risk Development in Young Adults (CARDIA) study measured risk factors in living subjects of similar ages. Results: Within sex, race, and age groups, the differences in body mass index between PDAY and CARDIA were significant but small. The prevalences of hyperglycemia/diabetes within sex, race, and age groups were similar in PDAY and CARDIA; overall, blacks in PDAY, but not in CARDIA, had a higher prevalence than whites. Serum lipoprotein concentrations were slightly and significantly higher and significantly more variable in PDAY subjects than in CARDIA subjects; the greater variability was interpreted as due primarily to emergency medical treatment. Prevalences of smoking and hypertension were substantially higher in PDAY subjects. Conclusions: Although there were statistically significant differences, the overall similarity of the risk factors in the two studies supports the validity of investigating associations of risk factors measured postmortem with anatomically determined arterial lesions in autopsied young adults dying from external causes. The greater variability in postmortem serum measurements attenuates but does not obscure associations. (c) 2007 Elsevier Inc. All rights reserved.

A risk score formula to estimate the probability of advanced atherosclerosis using coronary heart disease (CHD) risk factors was developed for persons 15-34 years of age by the Pathobiological Determinants of Atherosclerosis in Youth (PDAY) study. We applied the PDAY risk score to autopsied individuals from the Community Pathology Study (CPS), a different population that included middle-aged as well as young subjects. The PDAY risk score was associated with extent of raised lesions in the coronary arteries of CPS cases 15-34 years of age. The PDAY risk score computed from only the modifiable risk factors was associated with extent of raised lesions in the coronary arteries of subjects 35-54 years of age. The association of the PDAY risk score with lesions in 15-34 year old CPS subjects validates the PDAY risk score. The associations in both younger (15-34 years) and older (35-54 years) subjects suggest a seamless progression of the effects of the modifiable risk factors on atherosclerosis from 15 to 54 years of age. These results support the proposal that early control of risk factors is likely to prevent or delay the onset of CHD.

OBJECTIVES. Atherosclerosis begins in childhood and progresses during adolescence and young adulthood. The Pathobiological Determinants of Atherosclerosis in Youth Study previously reported risk scores to estimate the probability of advanced atherosclerotic lesions in young individuals aged 15 to 34 years using the coronary heart disease risk factors (gender, age, serum lipoprotein concentrations, smoking, hypertension, obesity, and hyperglycemia). In this study we investigated the relation of these risk scores to the early atherosclerotic lesions. METHODS. We measured atherosclerotic lesions in the left anterior descending coronary artery, right coronary artery, and abdominal aorta and the coronary heart disease risk factors in persons 15 to 34 years of age who died as a result of external causes and were autopsied in forensic laboratories. RESULTS. Risk scores computed from the modifiable risk factors were associated with prevalence of microscopically demonstrable lesions of atherosclerosis (American Heart Association grade 1) in the left anterior descending coronary artery and with the extent of the earliest detectable gross lesion (fatty streaks) in the right coronary artery and abdominal aorta. Risk scores computed from the modifiable risk factors also were associated with prevalence of lesions of higher degrees of microscopic severity (intermediate as well as advanced) in the left anterior descending coronary artery and with extent of lesions of higher degrees of severity (intermediate and raised lesions) in the right coronary artery and abdominal aorta. CONCLUSIONS. Risk scores calculated from traditional coronary heart disease risk factors to identify individual young persons with high probability of having advanced atherosclerotic lesions also are associated with earlier atherosclerotic lesions, including the earliest anatomically demonstrable atherosclerotic lesion. These results support lifestyle modification in youth to prevent development of the initial lesions and the subsequent progression to advanced lesions and, thereafter, to prevent or delay coronary heart disease.

Transverse, white-streak 'wrinkles' in the aorta were first described as Querlinien (cross lines) or Wellenlinien (wave lines) in the German literature in the early 20th century. These rhythmic structures were previously thought to be artifacts of stretching and shrinkage of the aorta. Not until the 1970s was it proposed that the areas of rhythmic wrinkling (RW) might be part of the process of atherosclerosis. We analyzed 2,650 aortas from the Pathobiological Determinants of Atherosclerosis in Youth (PDAY) study for prevalence, extent, and topographical distribution of these areas of RW. Furthermore, we investigated the possible relationship of RW to atherosclerotic sudanophilic stained 'fatty streaks' and elevated intimal lesions called 'raised lesions' (RL). This study provides evidence that (1) the prevalence of RW is fairly high in the aorta and occurs in a specific distribution in both the thoracic and abdominal aorta; (2) RW seems to precede the development of RL, with RL occurring in the same topographical areas as RW; and (3) RW may be associated with the subsequent development of advanced atherosclerosis, particularly raised lesions.

Essential polyunsaturated fatty acids (PUFA) of the ω-3 family are believed to protect against cardiovascular disease. A rich source of ω-3 PUFA is found in fish and marine mammals (seal, walrus, whale), which are a large part of the traditional diet of Alaska Natives (Eskimo, American Indians, Aleuts), a group that has been reported to have a lower mortality rate from cardiovascular disease than non-Natives.

This paper reports the results of the largest autopsy-based comparative study of atherosclerotic lesions between young Japanese and Americans, aged 15–34 years and autopsied between 1987 and 1995, by analyzing the data from the Japanese second nation-wide study of atherosclerosis and Pathobiological Determinants of Atherosclerosis in Youth study in the USA. In the right coronary arteries, in Japanese, fatty streaks were well established in the second decade of life with very little increase in the remaining age groups up to age of 34 years. In contrast, in American subjects, the average percentage of surface involvement of fatty streaks and raised lesions proceeded steadily with age without an obvious plateau throughout the 20-year period. The extent and prevalence of the raised lesions was much greater in Americans than in Japanese in 25–29 and 30–34-year age groups. Moreover, the rate of progression of raised lesions was much more rapid in Americans. These findings are in keeping with the fact that coronary heart disease (CHD) death rates are much higher in the USA than in Japan. In light of data showing that the risk factor profiles for CHD have become very similar between the two countries, these differences need to be explained.

Background—Blacks have a high rate of sudden coronary death (SCD). We determined the rate of SCD in men and women 30 to 69 years of age in a 6-year period recorded at a state Medical Examiner’s Office. Methods and Results—In a subset of 327 whites and 130 blacks, hearts were systematically studied to determine the extent of coronary disease, presence and type of thrombus (acute rupture, acute erosion, stable plaque), and heart weight. These parameters were correlated with the presence of conventional risk factors. The estimated rate of SCD in blacks was similar to that in whites under the age of 40 years but increased compared with whites with advancing age, becoming 1.5 times the rate for whites in the 7th decade (95% of the increase in the 6th decade was due to sudden death with stable plaque). Among the autopsied group with severe coronary atherosclerosis, HDL cholesterol was higher and hypertension more prevalent in blacks, but there was no difference in the prevalence of healed infarcts, plaque burden, heart weight, acute thrombi, or rates of diabetes, cigarette smoking, and total cholesterol. Conclusions—When compared with a control autopsy group of 568 deaths, multivariate analysis showed a significant association in blacks between stable plaque and left ventricular hypertrophy (risk ratio, 7.6), type 1 diabetes (risk ratio, 3.6), hypertension (risk ratio, 3.5), elevated total cholesterol (risk ratio, 3.1) and type 2 diabetes (risk ratio, 2.9). Because these risk factors are associated with SCD in blacks, they may be important targets for reducing the disparately high rate of SCD in blacks as compared with whites.

Background — Elevations in serum C-reactive protein measured by high-sensitivity assay (hs-CRP) have been associated with unstable coronary syndromes. There have been no autopsy studies correlating hs-CRP to fatal coronary artery disease. Methods and Results — Postmortem sera from 302 autopsies of men and women without inflammatory conditions other than atherosclerosis were assayed for hs-CRP. There were 73 sudden deaths attributable to atherothrombi, 71 sudden coronary deaths with stable plaque, and 158 control cases (unnatural sudden deaths and noncardiac natural deaths without conditions known to elevate CRP). Atherothrombi were classified as plaque ruptures (n=55) and plaque erosion (n=18); plaque burden was estimated in each heart. Total cholesterol, high-density lipoprotein cholesterol, diabetes, smoking history, and body mass index were also determined. Immunohistochemical stains for CRP and numbers of thin cap atheromas per heart were quantitated in coronary deaths with hs-CRP in the highest and lowest quintiles. The median hs-CRP was 3.2 μg/mL in acute rupture, 2.9 μg/mL in plaque erosion, 2.5 μg/mL in stable plaque, and 1.4 μg/mL in controls. Mean log hs-CRP was higher in rupture ( P <0.0001), erosion ( P =0.005), and stable plaque ( P =0.0003) versus controls. By multivariate analysis, atherothrombi ( P =0.02), stable plaque ( P =0.003), and plaque burden ( P =0.03) were associated with log hs-CRP independent of age, sex, smoking, and body mass index. Mean staining intensity for CRP of macrophages and lipid core in plaques was significantly greater in cases with high hs-CRP than those with low CRP ( P =0.0001), as were mean numbers of thin cap atheromas ( P <0.0001). Conclusions — hs-CRP is significantly elevated in patients dying suddenly with severe coronary artery disease, both with and without acute coronary thrombosis, and correlates with immunohistochemical staining intensity and numbers of thin cap atheroma.

Background —Subclinical episodes of plaque disruption followed by healing are considered a mechanism of increased plaque burden. Detailed pathological studies of healed ruptures, however, are lacking. Methods and Results —We identified acute and healed ruptures from 142 men who died of sudden coronary death and performed morphometric measurements of plaque burden, luminal stenosis, and smooth muscle cell phenotype. Healed ruptures were found in 61% of hearts and were associated with healed myocardial infarction, increased heart weight, dyslipidemia, and diabetes. Multiple healed rupture sites with layering were frequently found in segments with acute and healed rupture; the percent area luminal narrowing increased with increased numbers of healed sites of previous rupture. The underlying percent luminal narrowing for acute ruptures (mean 79±15%) exceeded that for healed ruptures (mean 66±14%, P =0.0001), and the area within the internal elastic lamina was significantly less in healed ruptures than in acute ruptures, when segments were grouped by distance from the ostium. Healed ruptures favored the accumulation of immature smooth muscle cells at repair sites, with a cellular proliferation index of 0.40±0.09%, significantly higher than the index at the sites of rupture ( P =0.008). Conclusions —These data provide evidence that silent plaque rupture is a form of wound healing that results in increased percent stenosis. Healed ruptures occur in arteries with less cross-sectional area luminal narrowing than acute ruptures and are a frequent finding in men who die suddenly with severe coronary atherosclerosis.

Elevated levels of serum free fatty acids (FFA) may be the metabolic alteration in obesity that leads to insulin resistance (IR) and type 2 diabetes mellitus (DM). The obese Zucker rat (ZR) is a genetic model of juvenile-onset obesity and type 2 DM. Compared with its lean sibling, the obese ZR is hyperinsulinemic, hypertriglyceridemic, and, beginning at about 6 months, hyperglycemic. The obese ZR demonstrates also IR, hyperphagia, increased lipogenesis, adipocyte hypertrophy and hyperplasia, and increased serum FFA levels. This study was designed to determine if serum FFA levels in lean and obese ZRs correlate with metabolic parameters associated with altered energy metabolism and IR. We hypothesized that serum FFA levels correlate with such serum parameters such as insulin, glucose, triglyceride, and total cholesterol, as well as such tissue parameters as retroperitoneal, perirenal, and epididymal fat pad weights and liver total lipid content. Twenty lean and 20 obese ZR were age/weight matched. For 14 days each rat had ad libitum access to a single bowl diet that was 50% fat, 30% carbohydrate, and 20% protein. Body weights and caloric intakes were measured daily. After 14 days, all animals were fasted overnight and euthanized. Serum and tissue measurements were made and various parameters were correlated with FFA levels. Serum FFA levels were almost 2 times higher in the obese ZR (approximately 1 mmol/L) compared to the lean (approximately 0.6 mmol/L). Each variable measured was significantly (p < or = 0.05) greater in the obese ZR compared to the lean. There were significant correlations between serum FFA levels and certain variables when data from all ZR were plotted against serum and tissue parameters. However, within phenotypes, there were no significant correlations. Serum FFA levels predict serum and tissue parameters that accompany obesity and IR when comparing lean and obese rats. However, FFA do not predict such parameters within one phenotype.

Background Coronary artery disease in women appears 10 to 15 years later than in men. To test the hypothesis that the effects of estrogen may manifest themselves as histologic differences in coronary plaques, we examined the hearts of premenopausal and postmenopausal women who died suddenly from coronary artery disease.Methods We studied 51 cases of sudden coronary death and 47 deaths in women who died from noncoronary causes. Coronary deaths were classified on the basis of histologic features. The number of acute plaque ruptures, healed plaque ruptures, vulnerable plaques, and acute plaque erosions were compared between groups. Postmortem values of serum total cholesterol, HDL cholesterol, and thiocyanate were measured, and menopausal status was confirmed by calculating body mass index.Results Women older than 50 years of age were much more likely to have a ruptured plaque than were younger, premenopausal women. Plaque rupture was significantly associated with elevated total cholesterol level. In the 51 women who died of coronary disease, the mean number of vulnerable plaques increased significantly as women advanced into the postmenopausal years.Conclusions Our data suggest that estrogen has an anti-inflammatory effect on atherosclerotic plaques, resulting in plaque stabilization. Plaque erosion, the major substrate for thrombosis in premenopausal women, does not appear to be inhibited by estrogen. Because plaque progression may result both from repeated rupture and repeated erosion, a better understanding of the effect of estrogen on atherosclerosis may yield insights into the nature of coronary artery disease.

Insulin-resistant muscle tissue contains low proportions of arachidonic acid (AA), and increased proportions of muscle AA correlate with improved insulin sensitivity. Dehydroepiandrosterone (DHEA) and AA, like the thiazolidinedione drugs that decrease insulin resistance (IR), are peroxisome proliferators. Long-chain fatty acids (FA) have been named the "one true" endogenous ligand for activating the peroxisome proliferator-activator receptor (PPAR), and DHEA has been named a "good candidate" as a naturally occurring indirect activator of PPAR. This study was conducted to determine DHEA's effects on lipid profiles of skeletal and cardiac muscle in lean and obese Zucker rats (ZR), a model of IR, type 2 diabetes mellitus, and obesity. We hypothesize that DHEA may alter long-chain FA profiles in muscle tissue of obese rats such that they more closely resemble that of the lean. In our experiments, we employed a DHEA and a pair-fed (PF) group (n = 6) for 12 lean and 12 obese ZR. For 30 d, the diet of the two DHEA groups was supplemented with 0.6% DHEA; PF groups were given the average daily calories consumed by their corresponding treatment group. Hearts and gastrocnemius muscles were assayed for phospholipid (PL), free FA, and triglyceride (TG) FA profiles. The proportion of PL AA was significantly greater in both muscle types of lean compared to obese rats. Hearts from both DHEA groups had greater PL proportions of AA and less oleic (18:1) acid than their PF controls. Likewise, 18:1 proportions were significantly lower in the gastrocnemius; however, AA proportions were not significantly different. Similar phenotypic profile differences were observed in the TG fraction of both muscle types. There were no DHEA-related TG FA profile alterations.