Mark L. Armstrong

Atherosclerosis impairs endothelium-dependent vascular relaxation to acetylcholine and thrombin in primates.

To test the hypothesis that atherosclerosis impairs endothelium-dependent vascular relaxation, we examined the effect of the endothelium-dependent vasodilators acetylcholine and thrombin and the endothelium-independent vasodilator nitroglycerin on iliac arteries from normal cynomolgus monkeys and cynomolgus monkeys with diet-induced atherosclerosis. Rings of iliac artery were suspended in organ chambers at their optimal length for generating tension. After preconstriction with prostaglandin F2, cumulative concentration-response curves to acetylcholine, thrombin, and nitroglycerin were examined The presence of endothehum was confirmed in each vessel by scanning electron microscopy. Atherosclerotic vessels showed morpholigic evidence of moderate to severe atherosclerosis. Acetylcholine produced a maximal relaxation of 65 ± 10% in the normal group and 27 ± 10% in atherosclerotic vessels (P < 0 05). Thrombin (10 0 U/ml) produced relaxation of 39 ± 9% in the normal group and 13 ± 7% in atherosclerotic iliac arteries (P < 0.05). Nitroglycerin relaxed both normal and atherosclerotic blood vessels to an equal extentmaximal relaxation was 92 ± 4% in normal vessels and 98 ± 2% in atherosclerotic vessels To determine if hypercholesterolemia alone produces an abnormality in endothelium-dependent relaxation, we performed two additional studies First, because veins are exposed to hypercholesterolemia, but do not develop atherosclerosis, we studied relaxation responses to acetylcholine and thrombin in veins from normal monkeys and monkeys with diet-induced atherosclerosis. Veins from normal and atherosclerotic monkeys relaxed to a similar extent upon exposure to the endothelium-dependent vasodilators acetylcholine and thrombin Second, we studied relaxation responses to acetylcholine, thrombin, and nitroglycerin in left circumflex coronary arteries from normal dogs and dogs fed a hypercholesterolemic diet for 4–5 weeks when serum cholesterol levels were elevated (serum cholesterol 442 ± 14 mg/dl), but before the onset of atherosclerosis. The endothelium-dependent vasodilators acetylcholine and thrombin produced equivalent degrees of relaxation in arteries removed from normal and hypercholesterolemic dogs These studies demonstrate that atherosclerosis impairs endothelium-dependent relaxation in primate iliac arteries, and that this impairment is not due to a generalized defect in the endothelium caused by hypercholesterolemia, but requires the presence of atherosclerosis

Augmented responses to vasoconstrictor stimuli in hypercholesterolemic and atherosclerotic monkeys.

We examined effects of hypercholesterolemia and atherosclerosis on vasoconstrictor responses to norepinephrine and serotonin. Responses were compared in normal, atherosclerotic, and hypercholesterolemic but non-atherosclerotic cynomolgus monkeys. The hindlimb was per fused at constant flow so that changes in perfusion pressure indicated changes in vascular resistance. We measured the pressure gradient from the iliac to the dorsal pedal artery so that responses of the large artery segment could be determined. Serotonin decreased total hindlimb resistance in normal and hypercholesterolemic monkeys, but increased total resistance in athero sclerotic monkeys. There was a greater than 10-fold increase in constrictor responses of large arteries to serotonin in atherosclerotic monkeys, compared with normal and hypercholesterolemic monkeys. In contrast, we found that vasoconstrictor responses tc norepinephrine are normal in atherosclerotic monkeys and increased in hypercholesterolemic monkeys prior to development of atherosclerosis. Hypercholesterolemia augmented responses of small vessels to norepinephrine. We conclude that, during early stages of hypercholesterolemia in cynomolgus monkeys, vasocon strictor responses to norepinephrine are increased in small vessels. At a later stage, as atheroscle rosis develops, responses to norepinephrine return to normal, but vasoconstrictor effects of large arteries to serotonin are greatly potentiated.

Restoration of endothelium-dependent relaxation by dietary treatment of atherosclerosis.

Atherosclerosis results in impaired relaxation to acetylcholine, thrombin, and the calcium ionophore A23187, all agents that require the presence of endothelium. We now report that dietary treatment of atherosclerosis in monkeys not only produces morphological improvement of the atherosclerotic lesion but restores endothelium-dependent vascular relaxation to normal. Because the intima remains thickened after regression of atherosclerosis, these studies suggest that intimal thickening which is present in both atherosclerotic vessels and after regression of atherosclerosis does not prevent the endothelium-derived relaxing factor from reaching the underlying vascular smooth muscle.

Regression of Coronary Atheromatosis in Rhesus Monkeys

Rhesus monkeys subjected to the atherogenic stimulus of a high-fat, high-cholesterol diet showed significant coronary atheromatosis at the end of 17 months. Smaller fibrotic lesions with scant stainable lipid were found in animals that were subsequently fed either of two cholesterol-free diets for 40 months. The average cross-sectional area of the lumen was more than 80% greater in regression animals than in monkeys with baseline atherosclerosis. The data support the hypothesis that uncomplicated coronary atheromas may regress in primates in appropriate dietary settings.

Cholesterol balance and fecal neutral steroid and bile acid excretion in normal men fed dietary fats of different fatty acid composition

Six normal men were fed formula diets containing either highly saturated fat (cocoa butter, iodine value 32) or polyunsaturated fat (corn oil, iodine value 125). The sterol balance technique was used to compare the changes in serum cholesterol concentration with the excretion of fecal steroids. The method used for the analysis of fecal steroids was chemical, with a final identification and quantification by gas-liquid chromatography. It was confirmed that the chemical method for fecal steroid analysis was accurate and reproducible. The three dietary periods were each 3 wk in length. In sequence, cocoa butter (period I), corn oil, and cocoa butter (period III) were fed at 40% of the total calories. All diets were cholesterol free, contained similar amounts of plant sterols, and were identical in other nutrients. Corn oil had a hypocholesterolemic effect. Mean serum cholesterol concentrations were 222 mg/100 ml (cocoa butter, period I), 177 during corn oil, and 225 after the return to cocoa butter. Individual fecal steroids were determined from stools pooled for 7 days. Both neutral steroids and bile acids were altered significantly by dietary polyunsaturated fat. The change in bile acid excretion was considerably greater than the change in neutral steroids. Corn oil caused a greater fecal excretion of both deoxycholic and lithocholic acids. The total mean excretion (milligrams per day) of fecal steroids was 709 for cocoa butter (period I), 915 for corn oil, and 629 for the second cocoa butter period. The enhanced total fecal steroid excretion by the polyunsaturated fat of corn oil created a negative cholesterol balance vis-à-vis the saturated fat of cocoa butter. The hypocholesterolemic effect of polyunsaturated fat was associated with total fecal sterol excretion twice greater than the amount of cholesterol calculated to leave the plasma. This finding suggested possible loss of cholesterol from the tissues as well.

Endothelium-dependent vascular relaxation is abnormal in the coronary microcirculation of atherosclerotic primates.

Atherosclerosis impairs endothelium-dependent relaxation of large conduit arteries. Because coronary resistance vessels are spared from the development of overt atherosclerosis, endothelium-dependent responses were examined in these vascular segments. Malaysian cynomolgus monkeys (n = 6) were made atherosclerotic by being fed a 0.7% cholesterol diet for 18 months. Control monkeys (n = 6) were fed a standard diet. Coronary microvessels (122-220 microns) were studied in a pressurized (20 mm Hg), no-flow state using a video-imaging apparatus. Relaxations of microvessels, preconstricted with the thromboxane analogue U46619, were determined in response to acetylcholine, bradykinin, the calcium ionophore A23187, adenosine, and sodium nitroprusside. Microvascular relaxations to bradykinin and A23187 were reduced in atherosclerotic monkeys compared with controls, whereas acetylcholine produced additional contraction in atherosclerotic monkeys. Responses of preconstricted microvessels to adenosine and sodium nitroprusside were identical in atherosclerotic and control animals. Indomethacin did not alter responses in control or atherosclerotic animals. Histologic examination revealed neither intimal thickening nor plaque formation in microvessels of this size class despite marked changes in conduit arteries. Electron microscopy showed minor alterations of endothelial cell morphology in microvessels of atherosclerotic animals. In conclusion, long-term hypercholesterolemia markedly impairs endothelium-dependent vascular relaxation in the coronary microcirculation where overt atherosclerosis does not develop. These changes in endothelial cell function may significantly alter regulation of myocardial perfusion by neurohumoral stimuli.

Coronary arterial remodeling studied by high-frequency epicardial echocardiography: An early compensatory mechanism in patients with obstructive coronary atherosclerosis☆☆☆

Coronary arterial remodeling is a compensatory mechanism that may limit the adverse effects of coronary obstructive lesions by expansion of the entire vascular segment. To determine if this compensatory anatomic change occurs in patients, high-frequency epicardial echocardiography using a 12 MHz transducer was performed during open heart surgery in 33 patients (10 with normal coronary arteries undergoing valvular surgery and 23 with coronary atherosclerosis). From stop-frame videotape high-frequency epicardial echocardiographic images, cross-sectional measurements of luminal area and total arterial area (lumen, intima, media and dense adventitia) were made in the patients with atherosclerosis at the site of arterial lesions and from the most proximal portion of the same artery. Remodeling was defined as enlargement of the total arterial area. In normal arteries measurements were made from proximal and midarterial locations. In the patients with normal coronary arteries, total arterial area, as determined by high-frequency echocardiography, decreased from the proximal site to the midportion of the artery (from 10.4 +/- 0.9 to 8.4 +/- 1.0 mm2, p less than 0.05); luminal area also decreased (from 6.0 +/- 0.6 to 4.5 +/- 0.7 mm2, p less than 0.05). In patients with coronary arterial lesions, luminal area also decreased from the proximal site to the arterial lesion site (from 5.3 +/- 0.6 to 2.3 +/- 0.3 mm2, p less than 0.05), but total arterial area increased (from 11.6 +/- 1.0 to 13.0 +/- 1.0 mm2, p less than 0.05). Of the 25 coronary arteries evaluated, only 4 had angiographic evidence of coronary collateral formation. These data indicate that coronary arterial remodeling is an important compensatory mechanism in obstructive coronary disease.(ABSTRACT TRUNCATED AT 250 WORDS)

Lipid Depletion in Atheromatous Coronary Arteries in Rhesus Monkeys after Regression Diets

Lipids were measured in the coronary arteries of monkeys on an atherogenic diet and in the arteries of matched monkeys on the atherogenic diet followed by regression diets. Cholesterol content was 51 mg/g dry weight in the arteries of monkeys with atheromatosis; after 40 months on the regression diets it was 18 mg/g. Cholesteryl ester was 69% lower and free cholesterol 53% lower after the regression diets. Decreases in triglycerides and phospholipids were not significant. The cholesterol content of the arteries of two monkeys autopsied after 20 months on the regression diets was close to the mean value after 40 months. The data show that cholesterol in both its free form and its ester form is depleted from experimentally induced coronary atheromatosis by dietary regression regimens. The data also suggest that most of the cholesterol depletion occurs during the first half of regression; the susceptibility of the residual excess arterial cholesterol to mobilization from the vessel wall by dietary means is questionable.

Structural and hemodynamic response of peripheral arteries of macaque monkeys to atherogenic diet.

The arteries of monkeys given atherogenic diets develop marked intimal thickening and medial thinning, but luminal size apparently changes minimally. The hemodynamic significance of the atherosclerotic changes is therefore uncertain. To evaluate vascular function in atherosclerotic arteries, we studied the hind-limb vessels of adult male rhesus and cynomolgus monkeys to assess the structural and hemodynamic responses to an atherogenic diet given for about 1.5 years or for much longer periods (6.5 years for rhesus and 4.3 years for cynomolgus monkeys). The intimal crosssectional area greatly increased after the atherogenic diet, but there was no significant luminal narrowing after either the 1.5-year diet or the longer diet periods. The media of atherosclerotic arteries showed focal atrophy and focal thinning after pressure fixation, but the total medial mass was not decreased even after the long diet periods. Hemodynamic studies indicated mild functional impairment in the atherosclerotic vessels; resting resistance increased and vasodilator responses decreased, but adrenergic responses were preserved. Thus, the marked changes that occur in the arterial wall in experimental primate atherosclerosis include adaptations to lesion formation that permit a long prestenotic phase of atherosclerosis in which vascular dysfunction is minimal.

Vasa vasorum in atherosclerotic coronary arteries: responses to vasoactive stimuli and regression of atherosclerosis.

The goals of this study were to determine whether vasa vasorum in atherosclerotic coronary arteries respond to vasoactive stimuli and to examine effects of regression of atherosclerosis on blood flow through vasa vasorum in coronary arteries. We studied three groups of monkeys: normal, atherosclerotic, and regression. Blood flow to vasa vasorum was measured with microspheres. Blood flow to intima-media (ml/min ± 100 g) was 5 ± 1 (mean ± SEM) in normal and 47 ± 7 in atherosclerotic monkeys (p<0.05). Infusion of phenylephrine or serotonin did not alter flow through vasa in normal monkeys. In atherosclerotic monkeys, phenylephrine decreased flow through vasa vasorum in intima-media of coronary arteries to 24 ± 4 (p< 0.05), and serotonin decreased flow to 27 ± 5 (p< 0.05). In regression monkeys, blood flow to intima-media was sixfold less (7 ± 2 ml/min ± 100 g) than in atherosclerotic monkeys (p<0.05). During infusion of adenosine, blood flow to vasa was fourfold greater in atherosclerotic monkeys than after regression of atherosclerosis. This finding suggests that loss of vessels, not constriction of existing vessels, accounts for the decrease in flow through vasa in intima-media after regression of atherosclerosis. We conclude that vasa vasorum in atherosclerotic coronary arteries respond to vasoconstrictor stimuli and that there is loss of vasa vasorum and a large decrease in blood flow through vasa to intima-media of coronary arteries after regression of atherosclerosis.