Hugh B. Lofland

Influence of Duration of Cholesterol Feeding on Esterification of Fatty Acids by Cell-Free Preparation of Pigeon Aorta

Influence of duration of cholesterol feeding on esterification of fatty acids and hydrolysis of cholesteryl esters was studied in cell-free preparations of aorta from White Carneau pigeons. Esterification of fatty acids required ATP and CoA; greater than 80% of the esterifying activity was located in the particulate fraction obtained by centrifugation at 105,000×g (after a preliminary centrifugation at 1000×g). Fatty acids were incorporated most efficiently into phospholipid, primarily (82%) lecithin. Greater than 87% of the fatty acid was esterified at the 2-position. During 8 months of cholesterol feeding, incorporation of oleic acid into phospholipids and triglycerides increased relatively little (less than double that of controls); no changes were seen before 1 month. Esterification of oleic acid to cholesterol was increased after 2 weeks of cholesterol feeding (before gross lesions were seen), eventually reaching a maximum increase of 30- to 50-fold. Cholesterol was esterified by transfer of fatty acyl-CoA to cholesterol, a mechanism similar to that described for liver and adrenal cortex. Little if any cholesterol esterification occurred when lecithin labeled at the 2-position with oleic acid-l-14C was used as substrate. The relationship between duration of cholesterol feeding and hydrolysis of cholesteryl oleate could not be evaluated since results depend directly on an unknown extent of equilibration of substrate with pre-existing cholesteryl ester pools.

A semiautomated procedure for the determination of triglycerides in serum

Abstract A procedure is described for the determination of triglycerides in serum, using an AutoAnalyzer and the same isopropanol extract of serum that is used in the automated determination of cholesterol. The method gives results identical to those obtained with a manual procedure, and enables one to carry out 100 or more triglyceride determinations per day.

Carbon monoxide-aggravated atherosclerosis in the squirrel monkey

Abstract Twenty-two squirrel monkeys, fed a cholesterol-containing diet, were placed in two groups: 12 were exposed to CO and 10 were not. In this 7-month study the atherosclerosis induced in the coronary arteries was aggravated by carbon monoxide; aortic atherosclerosis was not. The greater severity in the carbon monoxide group was shown by suggestive electrocardiographic changes, relative increases in heart weights, and a greater degree of coronary arterial stenosis as a result of intimal lipid. Marked deviations from normal values were not observed in several clinical characteristics often associated with the formation of atherosclerotic lesions of the arteries. We propose several mechanisms by which carbon monoxide could have aggravated the cholesterol-induced coronary artery lesions. In the CO-treated monkeys there was more intimal lipid in each atherosclerotic coronary artery rather than more coronary arteries with intimal lipid, as compared with the control group. Few such animal studies have been reported, so it is difficult to compare these results for the squirrel monkey with those for other species.

Comparative primate atherosclerosis: I. Tissue Cholesterol Concentration and Pathologic Anatomy

Abstract Stump-tailed macaques ( Macaca arctoides ), African green monkeys ( Cercopithecus aethiops ), squirrel monkeys ( Saimiri sciureus ), and woolly monkeys ( Lagothrix lagothricha ) were fed control, solid atherogenic (1 mg cholesterol/cal) or liquid diets containing 0, 0.5, or 1 mg cholesterol/cal. Stump-tailed macaques fed the solid atherogenic diet had the highest tissue and serum cholesterol concentration (about 700 mg/dl) and the most extensive atherosclerosis. These monkeys appeared to respond differently to diets containing 1 mg cholesterol/cal. Those animals fed the liquid diet had higher liver cholesterol concentration but lower serum cholesterol concentration than animals fed the solid diet. African green monkeys fed the solid atherogenic diet had serum cholesterol concentrations of about 450 mg/dl. A greater percentage of the abdominal aorta was covered by plaque than the thoracic aorta. Coronary artery atherosclerosis was focal with the largest plaques being found in the left main coronary artery. The microscopic appearance of these plaques was similar to that of plaques from people. Squirrel monkeys fed the atherogenic diet were the most variable group. The average serum cholesterol concentration averaged about 450 mg/dl (range: 291 to 716). The percentage of aorta covered by plaque ranged from 0 to 55% with more thoracic than abdominal aortic atherosclerosis. There were findings consistent with hemorrhage in plaques from two animals. These monkeys, like stump-tailed macaques but unlike African green monkeys had relatively high liver cholesterol concentrations. Woolly monkeys appeared to develop atherosclerosis when fed 1 mg cholesterol/cal but did not have greatly elevated serum cholesterol concentrations.

Influence of atherosclerosis on the composition, synthesis, and esterification of lipids in aortas of squirrel monkeys (Saimiri sciureus)

Summary The composition and synthesis of lipids from aortas of squirrel monkeys were studied following perfusion with [1- 14 C]acetate or [2- 14 C]mevalonate. Twenty-nine cholesterol-fed and 6 control monkeys were used. After 1 year on diet, the cholesterol-fed group had only slightly more aortic atherosclerosis than controls. In both groups fatty streaks and raised plaques were seen. Fatty acid composition of phospholipids and triglycerides was similar in both groups. Substantial differences were noted in the cholesteryl esters; the principal change was an increase in cholesteryl oleate in aortas from cholesterol-fed monkeys. In control aortas perfused with [l- 14 C]acetate phospholipid and triglyceride comprised approximately 90 % of lipid radioactivity. Among cholesterol-fed monkeys there was a slight increase in incorporation of newly synthesized fatty acids into aortic phospholipids with a slight decrease into triglycerides. Cholesteryl ester synthesis was increased approximately 6 times; oleic acid showed the greatest relative increase in synthesis. [2- 14 C]Mevalonate incorporation into aortic lipids was similar for both groups. More than 95 % of the total lipid radioactivity was recovered in two fractions, one has been identified as squalene. Perhaps most significant in these studies is the demonstration of metabolic changes (primarily an increase in cholesteryl ester synthesis) occurring in arteries only slightly more diseased than controls.

Interactions Among Dietary Fat, Protein, and Cholesterol in Atherosclerosis-Susceptible Pigeons

Atherosclerosis-susceptible pigeons fed cholesterol developed coronary atherosclerosis much more frequently than pigeons maintained on a cholesterol-free diet. Fewer of the cholesterol-fed pigeons fed a low protein diet developed coronary atherosclerosis than did cholesterol-fed pigeons on a high protein diet. Corn oil decreased the frequency of coronary atherosclerosis in cholesterol-fed pigeons. Limited evidence suggests that the coronary arteries and aortae in noncholesterol-fed birds are independent of each other in their susceptibility to atherosclerosis.

Effects of Regression of Atherosclerotic Lesions on the Content and Esterification of Cholesterol by Cell-Free Preparations of Pigeon Aorta

This study was designed to determine the effect of regression of atherosclerotic lesions on the incorporation of 1-14C-oleic acid into phospholipids, triglycerides, and cholesteryl esters and to compare these metabolic alterations with changes in the extent of atherosclerosis and the content of cholesterol and cholesteryl esters in the lesions. Aortic atherosclerosis was produced in White Carneau pigeons by feeding them an atherogenic diet for 1−8 months. The birds were then switched to a cholesterol-free diet for 6 months for the regression phase of the experiment. Following the regression phase, no changes were noted in the atherosclerotic index or the free-cholesterol content of aortas from pigeons that had received the atherogenic diet for 1−5 months; however, a reduction in both of these parameters was seen in aortas from pigeons fed the atherogenic diet for 8 months. There was a marked reduction in the content of cholesteryl esters in the aortas following the regression phase paralleled by a decrease in the rate of cholesterol esterification. No change was seen in the rate of incorporation of fatty acid into phospholipids or triglycerides. These studies suggest that local cholesterol esterification might be of considerable importance in maintaining the large amount of cholesteryl esters found in the atherosclerotic lesion and that changes in the rate of cholesterol esterification are associated with the early events in both progression and regression of atherosclerosis.

Atherosclerosis in Cebus albifrons monkeys: III. Clinical and Pathologic Studies☆

Abstract “Young and adult Cebus albifrons monkeys were fed either a control or an atherogenic diet for periods of up to 2 years. Some of the monkeys were necropsied at the end of 1 year. Coronary artery atherosclerosis was evaluated by determining the percentage of diseased arteries and the percentage apparent luminal stenosis in five-step cryostat sections from each of three blocks of ventricular myocardium. Arbitrary scores from 0 to 4 were made on gross stained carotid and renal arteries. Sudan IV and hematoxylin-stained cryostat sections were prepared from portions of basilar, middle cerebral, and posterior cerebral arteries. At 1 year serum cholesterol and age but not sex were related to the extent and prevalence of coronary artery disease. Atherosclerosis was slight in young test monkeys but relatively severe in adult test monkeys. Observations on the adult monkeys after 2 years were not much different from those made at 1 year. Young test female monkeys had five times as many coronary artery lesions as young test males after the 2-year period which was not true at 1 year. Single coronary artery lesions were found in one control adult male and one young control female. Carotid artery atherosclerosis was more prevalent among males than females. One adult control male had a small carotid artery lesion. One adult male test monkey had cutaneous and tendinous xanthomas. No lesion was found in the cerebral arteries of any animal. Sections of the tongue, uterus, kidney, and liver of the second-year test animals were examined for arterial lesions. Lesions, very similar to those of the small coronary arteries, were found in the intramuscular arteries of the tongues. Intrarenal arterial lesions were present in some of the cholesterol-fed monkeys of both age and sex groups. The group with the most pronounced coronary artery atherosclesosis, the adult test group, had the greatest frequency and magnitude of S-T segment deviation in the terminal electrocardiogram. One monkey of this group had discordant T waves in all leads.

The effect of insulin deficiency, hypothyroidism, and hypertension on atherosclerosis in the squirrel monkey☆

Abstract Squirrel monkeys with induced insulin deficiency, hypothroidism, and hypertension, as well as controls were fed a diet containing 1 mg of cholesterol per calorie for over 3 years. The hypothyroid and insulin-deficient monkeys had significantly greater concentrations of serum cholesterol and β-lipoprotein than did the controls, while the controls and hypertensive monkeys did not differ in these regards. The insulin-deficient, hypothyroid, and hypertensive groups all had more extensive coronary arterial and aortic atherosclerosis than did the controls. Atherosclerosis was especially severe in the insulin-deficient monkeys. The level of systolic blood pressure, the concentration of serum cholesterol, and the concentration of serum β-lipoprotein were significantly and positively correlated with coronary arterial and aortic atherosclerosis. The rate of disappearance of intravenously administered glucose was inversely related to serum cholesterol concentration and the indices of atherosclerosis. The squirrel monkey may be a good animal in which to study the mechanisms by which these disorders affect atherosclerosis, since these syndromes appear to affect atherosclerosis in this animal similarly as in man.

Pathologic characteristics and composition of diet-aggravated atherosclerotic plaques during “regression”

Abstract Atherosclerosis of White Carneau pigeons was exacerbated by feeding an atherogenic diet for 1 yr. A subgroup of the birds was examined to determine the baseline extent and severity of atherosclerosis, the frequency of myocardial infarction, and the biochemical composition of the atherosclerotic lesions. The remaining birds were divided into groups to be examined at 2, 4, 8, 12, and 16 mo after changing the diet to one low in fat and free of cholesterol. The extensiveness of aortic atherosclerosis decreased for 8 mo and then remained unchanged through 16 mo. The free and esterified cholesterol concentration of plaques changed markedly during regression. Both decreased markedly for four months and then remained essentially unchanged for the remainder of the year. As the cholesterol content decreased the collagen content increased. The degree of stenosis from coronary artery atherosclerosis declined during the first 8 mo of regression and then remained essentially unchanged. The regression in coronary artery atherosclerosis was associated with a marked decrease in the frequency of myocardial infarction.