Isotope kinetics of human skin cholesterol secretion.

Specific radioactivity (SA) time curves of plasma and skin surface cholesterol collected at several skin areas were recorded in 10 patients on formula diets after single intravenous injections of radioactive cholesterol. Earliest detectable radioactivity on skin surface was found in 4-6 days; depending on the skin site, SA's peaked in 13-75 days. SA's of free cholesterol were almost always higher than those of esterified cholesterol. The general forms of the SA time curves were in keeping with the idea that plasma cholesterol is carried to the skin surface in association with the epidermal and sebaceous cells, whereby (a) cholesterol synthesized de novo is mixed with derived from plasma and (b) appearances of plasma cholesterol on the skin surface is delayed by a time factor that corresponds to the movement of epidermal and sebaceous cells from the basal layer to the skin surface. The shorter mean transit times of plasma cholesterol on skin areas rich in sebaceous glands (22-24 days on the head) than on those poor in these glands (38 days on forearms and 72 days on feet) suggest that cholesterol passes faster through the sebaceous glands than through the epidermis, and faster through thin than thick epidermis. The fraction of skin surface cholesterol (f) that is derived from plasma cholesterol was estimated by three independent methods, and comparable results were obtained. Values of f were lower on skin areas rich in sebaceous glands (0.29-0.46 on forehead) than on areas poor in these glands (0.41-0.70 for forearms; 0.60 on feet) and lower for esterified (0.27-0.33) than for free (0.39-0.48) cholesterol. These data suggest that higher proportions of sebaceous gland and of esterified cholesterol, respectively, are synthesized de novo than epidermal and of free cholesterol. In two patients it was possible to calculate that f of total skin surface cholesterol was 0.49 and 0.37, respectively, and that the maximum amount of plasma cholesterol lost through the skin was 29 and 22 mg/day, respectively. Knowing the total daily excretion of total neutral and acidic steroids in feces in these patients, and assuming a total daily urinary steroid excretion 50 mg, we estimated that no more than 3.2% of total steroid excretion occurred via the skin.

Transfer of 15-lipoxygenase gene into rabbit iliac arteries results in the appearance of oxidation-specific lipid-protein adducts characteristic of oxidized low density lipoprotein.

Oxidized low density lipoprotein (LDL) possesses several atherogenic properties. The mechanisms by which LDL becomes oxidized in vivo remain unknown, but previous studies have suggested that 15-lipoxygenase may be one of the factors involved in the initiation of LDL oxidation in the arterial wall. 3 wk after a retrovirus-mediated 15-lipoxygenase gene transfer into iliac arteries of normocholesterolemic rabbits there was a threefold increase in 15-lipoxygenase activity but no signs of LDL oxidation. However, when animals were made moderately hypercholesterolemic by feeding a 0.13% cholesterol diet for 2-3 wk starting from day 4 after the gene transfer, oxidation-specific lipid-protein adducts characteristic of oxidized LDL were detected in 15-lipoxygenase-transduced arteries. Control experiments in which contralateral iliac arteries were transduced with beta-galactosidase-containing retroviruses showed only occasional signs of the presence of oxidation-specific adducts. The results support the hypothesis that products derived from the 15-lipoxygenase activity are involved in the induction of LDL oxidation within the arterial wall, provided that sufficient concentrations of lipoproteins are present in the artery.

Expression of alpha 2-macroglobulin receptor/low density lipoprotein receptor-related protein and scavenger receptor in human atherosclerotic lesions.

Macrophage- and smooth muscle cell (SMC)-derived foam cells are typical constituents of human atherosclerotic lesions. At least three receptor systems have been characterized that could be involved in the development of foam cells: alpha 2-macroglobulin receptor/LDL receptor-related protein (alpha 2 MR/LRP), scavenger receptor, and LDL receptor. We studied the expression of these receptors in human atherosclerotic lesions with in situ hybridization and immunocytochemistry. An abundant expression of alpha 2MR/LRP mRNA and protein was found in SMC and macrophages in both early and advanced lesions in human aortas. alpha 2MR/LRP was also present in SMC in normal aortas. Scavenger receptor mRNA and protein were expressed in lesion macrophages but no expression was found in lesion SMC. LDL receptor was absent from the lesion area but was expressed in some aortas in medial SMC located near the adventitial border. The results demonstrate that (a) alpha 2MR/LRP is, so far, the only lipoprotein receptor expressed in lesions SMC in vivo; (b) scavenger receptors are expressed only in lesion macrophages; and (c) both receptors may play important roles in the development of human atherosclerotic lesions.

Serum fatty acids and coronary heart disease in Finnish populations.

Finland has one of the highest coronary heart disease (CHD) rates in the whole world, and within the country, the rates are higher in men and eastern Finland than in women and western Finland, respectively. The differences are not wholly explained by the 3 classical risk factors. Several cross-sectional and longitudinal studies suggest that the intake of dietary linoleate (18:2), as reflected in the composition of serum and tissue fatty acids, has an inverse association with CHD, although there are also studies with negative results. Our own retrospective study failed to show any differences in the fatty acid composition of serum CE, TG or PL between men who had died of CHD and age- and risk factor-matched controls, but the negative finding may be due to changes in the fatty acids during storage. Among Finnish populations, most serum CE fatty acids had highly significant correlations with those in other serum lipid fractions, adipose tissue and platelets, and they had good "tracking" for up to 4 yr. Serum CE and TG 18:2, and total omega 6 polyunsaturated fatty acids (PUFA) had strong correlations with dietary 18:2 and PUFA, and can thus be used in the evaluation of population differences in the intake of PUFA. CE fatty acids were analyzed in a total of 2820 free-living 1- to 85-yr-old males and females. The percentages of serum fatty acids were age-dependent. West-Finnish populations had higher contents of 18:2 than those in eastern Finland, and middle-aged women had higher proportions of 18:2 than men. The content of 18:2 in adipose tissue and/or serum CE of middle-aged men was comparable to that in Scotland and lower than those in Italy and Sweden. These findings are compatible with the idea that 18:2 is a negative risk factor of CHD, but the evidence is only circumstantial. Possible mechanisms whereby 18:2 could affect CHD have been discussed. The contents of omega 3 fatty acids in CE were opposite to those of omega 6 fatty acids in that eastern Finnish and male populations had higher contents of 18:3 omega 3 and 20:5 omega 3 than western Finns and females, respectively. This finding is apparently due to substitution of vegetable fat for milk fat in the diet, resulting in a relative deficiency of the omega 3 fatty acids at the expense of 18:2. Experimental evidence for such a mechanism was obtained in an intervention study.(ABSTRACT TRUNCATED AT 400 WORDS)

Receptor-mediated binding and degradation of subfractions of human plasma low-density lipoprotein by cultured fibroblasts.

The receptor-mediated metabolism of human plasma low-density lipoprotein (LDL) subfractions was studied. LDL was isolated from healthy donors and further fractionated by density gradient ultracentrifugation into three subfractions: (I) d = 1.031-1.037, (II) d = 1.037-1.041 and (III) d = 1.041-1.047 g/ml, comprising 24 +/- 7%, 46 +/- 8% and 30 +/- 9% of the total LDL protein, respectively. As assessed by electron microscopy and gradient gel electrophoresis, the LDL particle size decreased and the relative protein content increased from fraction I towards fraction III. Fraction II had the highest (Kd 2.6 micrograms/ml) and fraction I the lowest (Kd 5.8 micrograms/ml) binding affinity to LDL receptors of human fibroblasts at 4 degrees C. The rate of receptor-mediated degradation of fraction II was also higher than that of the other two fractions at 37 degrees C. These results suggest that LDL subfractions have different rates of receptor-mediated catabolism depending on particle size or composition, and therefore their metabolic fate and atherogenic properties may also differ.

In vivo low density lipoprotein oxidation relates to coronary reactivity in young men.

OBJECTIVES: This study was undertaken to examine the relation of in vivo low density lipoprotein (LDL) oxidation and other lipid risk factors to coronary reactivity in normal subjects. BACKGROUND: Experimental studies have shown that oxidized LDL (ox-LDL) particles are injurious to the vascular wall by impairing its normal vasodilator function. METHODS: We used noninvasive positron emission tomographic (PET) imaging with intravenous dipyridamole to measure coronary flow reserve, a marker of coronary endothelial and smooth muscle function, in 30 healthy men (mean [+/-SD] age 34.4 +/- 3.2 years). As a marker of in vivo LDL oxidation, the autoantibody titer against ox-LDL was measured by the enzyme-linked immunosorbent assay method. RESULTS: Plasma levels of autoantibody titer against ox-LDL were inversely associated with coronary flow reserve (r = -0.42, p = 0.023). High LDL cholesterol levels (above median > 3.0 mmol/liter) were associated with a low coronary flow reserve only in subjects expressing simultaneously high levels of ox-LDL titer (above median). Subjects with simultaneously high levels of LDL cholesterol and ox-LDL titer had lower coronary flow reserve values than subjects in other groups (3.89 vs. > 5.0 in other groups, p = 0.066). CONCLUSIONS: These data provide evidence for the role of ox-LDL in affecting the coronary reactivity in vivo and support the concept that oxidative modification of LDL particles provides a mechanism by which high LDL concentrations exhibit injurious effects on the coronary vascular bed.

Relationship of angiotensin-converting enzyme gene polymorphism to carotid wall thickness in middle-aged men.

The insertion/deletion (I/D) polymorphism of the human angiotensin-converting enzyme (ACE) gene is a major determinant of circulating ACE levels. The D allele has been suggested to be a potent risk factor for coronary artery disease; however, the effect of the ACE gene on carotid atherosclerosis remains controversial. We therefore studied the relationship between the ACE gene I/D polymorphism and carotid artery intima-media thickness (IMT). A random sample of 300 men aged 50-59 years living in southern Finland were selected, and 233 agreed to participate (74%). Data were collected in 219 subjects. Quantitative B-mode ultrasonography was used to measure the maximum near and far wall IMT of right and left common, bifurcation, and internal carotid artery. The mean maximum IMT (overall mean) was calculated as the mean of 12 maximum IMTs at 12 standard sites. Patients with an IMT higher than 1.7 mm in at least one of 12 standard sites were assumed to have carotid atherosclerosis. The I/D polymorphism was determined by polymerase chain reaction. Overestimation of the frequency of the DD genotype was eliminated by insertion-specific primer and the inclusion of 5% dimethylsulfoxide. No significant differences were found in carotid wall thickness between the three genotypes; the overall mean IMT were 1.18 +/- 0.30, 1.22 +/- 0.24, and 1.08 +/- 0.40 mm in genotypes of II, ID, and DD, respectively. Similarly, the ACE genotypes and allele frequencies did not differ significantly between the subjects with and those without carotid atherosclerosis. There was no association in the subgroups among only nonsmoking subjects or subjects without chronic medication. The present data indicate that the I/D polymorphism of the ACE gene is not related to carotid IMT and is unlikely to play a major role in carotid atherosclerosis.

Oxidized low-density lipoprotein is chemotactic for arterial smooth muscle cells in culture.

The effects of human native and Cu2(+)-oxidized low-density lipoprotein (LDL) were tested on the migration of cultured bovine aortic smooth muscle cells (SMCs) in blind-well chambers. LDL oxidation was controlled by measuring the formation of conjugated dienes and lipid hydroperoxides, and by agarose gel electrophoresis. Oxidized LDL stimulated SMC migration, and the effect was dose-dependent up to 200 microgram/ml. The stimulation was chemotactic in nature. Native LDL was without significant activity. The results suggest that oxidized LDL may contribute to the migration of medial SMCs into the intima during atherogenesis.

Nitric oxide donor GEA 3162 inhibits endothelial cell-mediated oxidation of low density lipoprotein.

The effect of a nitric oxide (NO) donor GEA 3162 on the endothelial cell (EC)-mediated oxidation of low density lipoprotein (LDL) was studied. In comparison to LDL incubated with EC without GEA 3162, the presence of GEA 3162 inhibited LDL oxidation by EC, as indicated by the following findings. (a) The degradation rate of LDL in macrophages was reduced to control levels. (b) The electrophoretic mobility of LDL decreased in a dose-dependent manner. (c) The concentrations of thiobarbituric acid-reactive substances and hydroperoxide-derived hydroxy fatty acids were lower. (d) The breakdown of apolipoprotein B was reduced. The results indicate that GEA 3162 prevents EC-mediated oxidation of LDL.

High expression of human 15-lipoxygenase induces NF-kappaB-mediated expression of vascular cell adhesion molecule 1, intercellular adhesion molecule 1, and T-cell adhesion on human endothelial cells.

Expression of 15-lipoxygenase (15-LO) is induced over 100-fold in early fatty streak lesions. 15-LO activity leads to the production of specific lipid hydroperoxides, which can have major effects on the expression of proinflammatory genes involved in atherogenesis. We have used retrovirus-mediated gene transfer to achieve stable high expression of 15-LO in human endothelial ECV304 cells. These cells were used to study the effects of 15-LO on the expression of vascular cell adhesion molecule 1 (VCAM-1) and intercellular adhesion molecule 1 (ICAM-1), activation of nuclear factor kappa B (NF-kappaB), and T-cell adhesion on endothelial cells. NF-kappaB activation was greatly potentiated by increased 15-LO activity in the stably transduced cells, and both VCAM-1 and ICAM-1 were significantly induced in these cells in response to tumor necrosis factor-alpha (TNF-alpha) and phorbol 12-myristate 13-acetate (PMA) stimulation, as studied by flow cytometry. The induction of ICAM-1 was sensitive to antioxidants in a dose-dependent manner. The adherence of Jurkat T cells on the 15-LO-expressing endothelial cells was markedly induced after PMA stimulation. These results indicate that 15-LO activity may be involved in the early pathogenesis of atherosclerosis by inducing VCAM-1 and ICAM-1 expression and by increasing T-cell adhesion on the endothelium.

Serum fatty acids in 8-year-old Finnish boys: correlations with qualitative dietary data and other serum lipids.

To survey risk factors in coronary heart disease in Finnish children, fasting serum specimens from 244 healthy 8-yr-old boys were analyzed for the fatty acid composition of cholesterol esters (CE), triglycerides (TG), free fatty acids (FFA), and phospholipids (PL). A qualitative dietary survey was made by asking parents to answer a questionnaire including, among others, a question on the kind of fat usually used on bread by the child. The mean percentages of linoleate (18:2) in serum cholesterol esters, triglycerides, free fatty acids, and phospholipids were 53.1, 13.5, 11.5, and 22.7%, respectively, which represent an international average. The quality of dietary fat had a clear influence on serum fatty acids, eg, the content of 18:2 in CE was 56.8 +/- 3.6% in boys using soft vegetable margarine and 50.5 +/- 3.6% in those using butter. The former had also a marginally lower serum total cholesterol (4.87 +/- 0.86 mmol/l) than the latter (5.08 +/- 0.80 mmol/l). Serum total cholesterol showed significant negative correlations with the percentage of 18:2 in all four lipid fractions, the highest r values being with PL-18:2 (-0.41) and CE-18:2 (-0.24). Accordingly, serum cholesterol was lower in the highest CE-18:2 quartile (4.67 +/- 0.76 mmol/l) compared with the lowest (5.30 +/- 0.70 mmol/l; p less than 0.001). The results indicate that when serum fatty acids are used as indicators of the quality of dietary fat, a negative association between polyunsaturated fat and serum cholesterol may be demonstrable even within a free-living population.

Fatty acid composition of serum cholesteryl esters in 3- to 18-year-old Finnish children and its relation to diet.

The composition of fatty acids in serum cholesteryl esters (CE) was analyzed gas chromatographically in 1348 boys and girls aged from 3 to 18 yr. A dietary survey was carried out simultaneously using the 48-h recall method. The dietary P/S ratio had highly significant correlations with CE fatty acids: positive with linoleate (0.567) and total omega 6 fatty acids and negative with saturated, monounsaturated, and omega 3 polyunsaturated fatty acids. The highest mean percentage of CE-linoleate was found in 15-yr-old girls (52.7, SD 4.68%) and lowest in 3-yr-old girls (48.1, SD 5.00%). Age, sex, and the degree of puberty had no independent effect on CE-linoleate after it had been adjusted for the effect of dietary P/S ratio by analysis of covariance. These results indicate that the fatty acid composition of serum CE depends on the quality of dietary fat and that CE-linoleate is a useful reflector of the dietary P/S ratio. The negative correlation between CE omega 3 fatty acids and dietary P/S ratio may be due to displacement of the omega 3 acids in serum CE by the much higher proportion of dietary linoleate.

Vitamin E and cancer prevention.

Some animal experiments and human studies suggest that vitamin E may protect against cancer. Serum alpha-tocopherol concentration was studied for its prediction of cancer in a cohort of 36,265 adults in Finland. During a mean follow-up of 8 y, cancer was diagnosed in 766 persons. The levels of serum alpha-tocopherol were determined from stored serum samples (at -20 degrees C) taken from these cancer patients and from 1419 matched control subjects. Individuals with a low level of alpha-tocopherol had about a 1.5-fold risk of cancer compared with those with a higher level. The strength of the association between serum alpha-tocopherol level and cancer risk varied for different cancer sites and was strongest for some gastrointestinal cancers and for the combined group of cancers unrelated to smoking. The association was strongest among nonsmoking men and among women with low levels of serum selenium. The findings agree with the hypothesis that dietary vitamin E in some circumstances protects against cancer.

The effect of short-term fasting, apolipoprotein E gene polymorphism, and sex on plasma lipids.

The effect of 1 wk of supervised fasting on plasma lipid concentrations in subjects with different apolipoprotein E (apo E) phenotypes was studied in 58 healthy free-living volunteers. The participants consumed an 870-kJ(208 kcal)/d liquid diet containing fruit and berry juices, tea, and water. The decline in plasma total cholesterol during 1 wk of fasting was 0.46 mmol/L in women and 0.35 mmol/L in men. The decreases were significant in both women and men. The response patterns of plasma total cholesterol were not significantly different between the sexes. In men, the changes in plasma low-density-lipoprotein cholesterol during the fast differed significantly (P = 0.0181) between the apo E phenotypes, whereas in women there were no differences due to phenotype (P = 0.695). The magnitude of the change in plasma triacylglycerol during the fast was different between the sexes (P = 0.0099). The changes in plasma triacylglycerols differed significantly between apo E phenotype groups in men (P = 0.0295) but not in women (P = 0.0661). Statistical comparison between different apo E phenotypes was performed with and without the small apo E3,2+E2,2 group, with essentially similar results. During fasting, plasma high-density-lipoprotein cholesterol concentrations decreased slightly but not significantly. The study shows significant differences in the associations of apo E alleles and sex on plasma lipid responses during fasting and illustrates the importance of gene-diet interactions in the regulation of lipid metabolism in humans.

Enhanced growth of smooth muscle cells from atherosclerotic rabbit aortas in culture.

It has been suggested that cells in atherosclerotic lesions differ from normal arterial smooth muscle cells. The present study was aimed at elucidating this possibility. The morphology and growth characteristics of cells cultured from atherosclerotic rabbit aortas were compared with those obtained from control rabbits of the same age. Primary cultures were started from aortic intima-medial explants and they were subcultured after trypsinising the outgrowths. There were no differences in morphology between cells cultured from atherosclerotic and control animals as judged with phase contrast and electron microscopy. However, both the primary and subsequent growth rates of the cells cultured from atherosclerotic aortas were higher than those of control cells. Also the uptake of [3H]thymidine by cells derived from the atherosclerotic aortas was enhanced both in the exponential and stationary phases of cell growth. The results suggest that although the cells from atherosclerotic aortas are morphologically similar to normal arterial smooth muscle cells, atherogenesis is associated with the development of metabolic changes in the arterial cells that are carried over to daughter cells and persist even in culture.

Enhanced synthesis of collagen and total protein by smooth muscle cells from atherosclerotic rabbit aortas in culture.

Protein synthesis in smooth muscle cells from normal and atherosclerotic rabbit aortas was studied. Cultures of cells from the third passage were incubated with [3H]proline and the synthesis rates of collagen and total protein measured from the radioactivities incorporated into protein-bound hydroxyproline and proline, respectively. The synthesis ratio of collagen types I and III was studied by separating the radioactive procollagens of the culture media by DEAE column chromatography. The synthesis rates of both collagen and total protein were higher in cells cultured from atherosclerotic rabbit aortas. There was no difference in the synthesis ratios of types I and III collagen between cells from the two origins. In addition to procollagens, a third protein eluted before procollagen I in the DEAE chromatography of medium proteins. Compared with control cells, cells from atherosclerotic aortas incorporated relatively less radioactivity into this protein. No differences were detected in the amino acid compositions of cellular proteins between cells from both origins. It was concluded that cells grown from atherosclerotic rabbit aortas synthesize more collagen and total protein than those from normal aortas, the synthesis of collagens I and III being equally enhanced. The results also suggest that there are proteins which are not involved in the general enhancement of protein synthesis.

Effect of post-mortem time on the biochemical composition of coronary arteries.

The effects of post-mortem autolysis on the biochemical composition of coronary intima-medias were studied by keeping the coronary arteries of 25 slaughterhouse pigs at +4 degrees C for periods up to 7 days. The samples were analysed for DNA, total protein, collagen, glycosaminoglycans (GAGs), free and esterified cholesterol, triglycerides, phospholipid fractions, and fatty acid composition in various lipid classes. The content of sulphated GAGs decreased steadily, whereas that of hyaluronic acid remained unchanged. These alterations led to a significant decrease in the content of total GAGs and to an increase in the percentage of hyaluronic acid in total GAGs. The results indicate a continuous post-mortem degradation of arterial proteoglycans. After 7 days storage, there was a significant increase in the content of free fatty acids, and the composition of this fraction was changed by increases in the relative amounts of oleic and linoleic acids. These changes were accompanied by an increase in lysolecithin and a decrease in lecithin, which suggests a post-mortem action of arterial phospholipases.

Lipoprotein uptake in primary cell cultures of rabbit atherosclerotic lesions. A fluorescence microscopic and flow cytometric study.

To characterize the lipoprotein metabolism of lipid-filled cells of atherosclerotic lesions, uptake of 3,3'-dioctadecylindocarbocyanine (DiI)-labelled low density lipoprotein (LDL), acetylated LDL (Ac-LDL) and beta-very low density lipoprotein (beta-VLDL) was studied by fluorescence microscopy and flow cytometry in primary cultures of enzymatically dispersed aortic cells from cholesterol-fed rabbits. Most of the foam cells were identified as macrophages on the basis of Fc-receptors and high activities of nonspecific esterase and acid lipase, although cholesteryl ester (CE) inclusions were found by filipin staining also in smooth muscle cells (SMCs). During the culture only SMCs proliferated and were confluent in about 1 week. After incubation with DiI-Ac-LDL most macrophage foam cells were brightly fluorescent, but also many SMCs accumulated fluorescence. In SMCs, an excess of LDL inhibited the uptake of DiI-beta-VLDL and DiI-LDL, indicating that these lipoproteins were taken up by the apoB,E receptor; the activity of this receptor was low 2 days after cell isolation but increased considerably during SMC proliferation. DiI-beta-VLDL was not taken up by the macrophage foam cells until after 7 days' culture, when their CE content had decreased, reflecting a feed-back regulation of these receptors as well. Our results indicate that, in primary cultures of enzyme-dispersed cells from rabbit atherosclerotic lesions, most of the foam cells have lipoprotein receptors resembling those described in macrophages and that also many SMCs accumulate Ac-LDL.

Serum fatty acids in Finnish men.

The fatty acid compositions of 4 serum lipid fractions were analysed from 244 randomly selected 30-59-year-old Finnish men from 4 areas involved in a population survey ('Mini-Finland') in 1979-80. Men in eastern Finland had significantly lower mean percentages of linoleate (18:2) in CE, TG, FFA and PL (45.1, 10.3, 9.3 and 18.8%, respectively) than men in the western part of the country (48.4, 12.5, 10.6 and 20.2%, respectively). Very low values of 18:2 were encountered in the North Karelian community of Ilomantsi, especially in men aged 50-59 (40.9, 8.0, 7.5 and 16.8%, respectively). The percentage of alpha-linolenate tended also to be lower and those of saturated and monounsaturated fatty acids higher in the east, but there were no or only inconsistent differences in the contents of the prostaglandin precursors dihomo-gamma-linolenate, arachidonate and eicosapentaenoate. Eighteen men were studied in November and the following April. Only minor changes in the mean composition of serum fatty acids took place during this period and the correlation coefficients between the percentages of 18:2 recorded at the two time points ranged from 0.70 to 0.81. The low concentrations of 18:2 in serum lipids in Finnish men obviously reflect a low dietary P/S ratio and may contribute to the high prevalence of IHD in Finland and to its regional differences.

Ultrastructural, immunochemical and electrophoretic study of smooth muscle cells in internal mammary arteries of patients undergoing coronary bypass surgery.

The internal mammary artery (IMA) is used widely in bypass grafting for coronary artery disease because of its resistance to atherosclerotic obstruction. Since there are no data on the ultrastructure of IMA or the phenotype of its smooth muscle cells (SMC), we studied the distal parts of left IMA obtained at the time of surgery from 14 coronary bypass patients, aged 43-67 years. Eight IMA were examined by transmission electron microscopy. The distribution of the cytoskeletal proteins actin, vimentin, and desmin in the intima-media of 6 IMA was studied by immunofluorescence microscopy, polyacrylamide gel electrophoresis, and two-dimensional gel electrophoresis. The intimas were very thin, from 3 to 32 microns. The thinnest regions contained no cells. Most intimal cells had the ultrastructural features of SMC; no foam cells were found. The majority of both intimal and medial SMC had a myofilament-rich phenotype. Cells reacting to antibodies of vimentin, desmin and alpha-actin were found in both intima and media. alpha-Actin formed 67% of all actin isoforms in the intima-medial extracts. Our study confirms ultrastructurally the reported scarcity of atherosclerosis in the human IMA and shows that the majority of SMC in the IMA of even severely atherosclerotic coronary bypass patients are both ultrastructurally and biochemically in a differentiated state, which agrees with their resistance to atherosclerosis.