Allan J. Day

Uptake and Metabolism of 14C-Labeled Oleic Acid by Atherosclerotic Lesions in Rabbit Aorta

The uptake of 14C-labeled oleic acid and its incorporation into combined lipids by aortic intimas from normal and cholesterol-fed rabbits has been investigated in vitro. More than five times as much oleic acid was taken up by the atherosclerotic intima as by the normal intima. About twice as much oleic acid was incorporated into phospholipid, and twenty times as much into cholesterol ester by the atherosclerotic intima as by the normal. Lecithin was the major phospholipid synthesized from oleic acid in both normal and atherosclerotic intimas. Radioautographs of the atherosclerotic vessels show that the 14C-labeled oleic acid and its metabolic derivatives, principally phospholipid and cholesterol ester, were localized in sudanophilic cells in the intima in both early and advanced lesions. It is concluded that intimal foam cells are primarily responsible for the lipid synthesis that occurs in the atherosclerotic lesion.

Incorporation of Oleic Acid into Lipid by Foam Cells in Human Atherosclerotic Lesions

The incorporation of 14C-labeled oleic acid into phospholipid, cholesterol ester, and triglyceride in human arterial intima (normal and atherosclerotic) obtained from kidney transplant donors has been investigated in vitro. Most of the oleic acid was incorporated into phospholipid, in both the normal intima and the atherosclerotic lesion, but a greater proportion of label was diverted to cholesterol ester in the lesion. Representative sections of the vessels used for metabolic studies were taken for radioautography, and these demonstrated localization of 14C to foam cells, but very little localization in the region of spindle-shaped cells. It was concluded that lipid synthesis in the human atherosclerotic lesion takes place predominantly in the intimal foam cells.

Hypertension-accelerated atherogenesis in cholesterol-fed rabbits

Entry of 125I-labelled low density lipoprotein ([125I]LDL) into the aortic intima was studied over 6 hours in normotenisve and hypertensive rabbits fed a 1% cholesterol diet for 9 and 4 weeks respectively. Studies were also made in hypertensive and normotensive cholesterol-fed rabbits in which blood pressure was reduced acutely with parenteral hydralazine. In all groups the entry of E1125I]LDL was greatest in the aortic arch and significantly less in both the descending thoracic and abdominal regions. Lipoprotein entry into the aorta of cholesterol-fed rabbits was increased some 10-fold over the corresponding value previously found in rabbits fed a normal diet [1]. This increase was due to increased vascular permeability as well as to increased plasma LDL concentration. The hypertensive cholesterol-fed rabbits did not show significantly greater entry of [125I]LDL than the normotensive cholesterol-fed rabbits. Comparison of the rate of LDL entry over 6 house and the quanitity of cholesterol accumulated in the aortic segments over the period of cholesterol feeding indicated that lipoprotein fractions other than LDL must contribute singificant amounts of cholesterol to the developing lesion. The finding that LDL entry paralledled accumulation during cholesterol feeding, together with the finding that acute reversal of hypertension did not reduce the entry of [125I] LDL suggest that mechanisms other than increased filtration of plasma low density lipoprotein contribute significantly to the accelerated development of atherosclerosis in hypertension.

Differential uptake of cholesterol and of different cholesterol esters by atherosclerotic intima in vivo and in vitro

Abstract Cholesterol-fed rabbits were intubated with a single dose of [3H]cholesterol and the entry of the resultant labelled free cholesterol, cholesterol ester and of the individual cholesterol esters in the serum into the aortic intima and media was determined 4 days after ingestion of the cholesterol. The entry of cholesterol into the aortic intima exceeded that of cholesterol ester; the entry of monounsaturated cholesterol ester exceeded that of polyunsaturated cholesterol ester. The greater entry of cholesterol relative to cholesterol ester was confirmed in experiments in vitro in which atherosclerotic aortas were incubated with hypercholesterolaemic serum labelled with [3H]cholesterol ester and 3 H 14 C-labelled free cholesterol in the lipoprotein. It was not possible, however, to demonstrate any significant increase in entry of monounsaturated cholesterol ester over polyunsaturated cholesterol ester in these experiments in vitro. By using hypercholesterolaemic serum containing [3H]cholesterol ester and 3 H 14 C-labelled free cholesterol in the lipoprotein, as incubation medium, it was possible to demonstrate in the experiments in vitro that hydrolysis of the labelled cholesterol ester entering the intima could not account for the greater relative influx of free over ester cholesterol. The factors associated with the entry and accumulation of cholesterol and of various cholesterol esters in the atherosclerotic lesion are discussed.

The effect of hypertension on the accumulation of lipids and the uptake of [3H]cholesterol by the aorta of normal-fed and cholesterol-fed rabbits

Abstract The relationship between blood pressure and arterial wall lipids was studied in normal-fed and cholesterol-fed rabbits with and without renal hypertension. 1.(a) Cholesterol feeding influenced neither the diastolic blood pressure of the normotensive rabbits nor the degree of diastolic hypertension developed by rabbits made hypertensive by cellophane perinephritis. 2.(b) Hypertension produced a significant increase in the cholesterol content of the intima of both normal-fed and cholesterol-fed rabbits, but did not affect the serum cholesterol levels of rabbits on either diet. The increase in intimal cholesterol with hypertension in rabbits on a normal diet was associated with a parallel rise in the intimal dry defatted weight. In cholesterol-fed rabbits, however, hypertension did not produce this effect on intimal weight. Hypertension also had no effect on the percentage cholesterol ester or cholesterol-phospholipid ratio in the intima or media of cholesterol-fed rabbits.In normal-fed animals hypertension had little effect on intimal phospholipid fatty acid composition, but was associated with a shift towards unsaturation in the fatty acid composition of intimal cholesterol ester and triglyceride similar to that produced by cholesterol feeding alone. In addition, cholesterol feeding alone produced a significant increase in the linoleic (18:2) and decrease in the arachidonic (20:4) acid components of the intimal phospholipids. In cholesterol-fed rabbits hypertension did not have any further effect on cholesterol ester and triglyceride fatty acids, but was associated with a significant increase in the palmitic (16: 0) and decrease in the arachidonic (20:4) acid components of the intimal phospholipids. 3.(c) Hypertension was associated with a significant increase in the influx of [ 3 H]cholesterol into the aortic intima in vivo in normal-fed rabbits but not in cholesterol-fed rabbits. However, in both normal-fed and cholesterol-fed rabbits there was a significant correlation between influx and the greater intimal cholesterol of hypertensive animals. 4.(d) The findings indicate that both increased entry and altered metabolism of intimal lipid occur during the acceleration of atherosclerosis by hypertension.

Cholesterol ester and phospholipid composition of normal aortas and of atherosclerotic lesions in children

Abstract Gas chromatographic techniques have been used to study the lipid composition of childrens arteries over the first decade of life. Childrens aortas were separated into intima and media and, where present, into intimal fatty streak lesions and media underlying such lesions, and the level of cholesterol ester and of phospholipid and the fatty acid composition of these fractions in these various portions of the aortas was determined. The ester cholesterol concentration of the normal intima, but not of the media, increased with age. The ester cholesterol concentration of the fatty streak lesions exceeded that of the normal intima and also increased with age. No changes with age in phospholipid content for normal intima or media or for the intimal lesion were observed. The cholesterol ester fatty acid composition of the normal intima and of the media was similar to that of the lesions studied, and all tissues manifested a change in composition with age; cholesterol linoleate increasing and cholesterol palmitate decreasing during the first decade. These age changes for the aortic fractions were shown to resemble changes with age in the serum cholesterol ester fatty acids. The phospholipid fatty acid pattern of normal intima and media resembled those of the lesion but no change with age in phospholipid fatty acid pattern was found. No correspondence of the aortic phospholipid pattern with the serum pattern was demonstrated. These data are consistent with the view that arterial cholesterol ester, but not phospholipid, in both the normal intima and in the early fatty streak lesion in children arises initially from serum.

Cellophane Perinephritis Hypertension and Its Reversal in Rabbits: Effect on Plasma Renin, Renin Substrate, and Renal Mass

Plasma renin activity and renin substrate levels were measured during the development of cellophane perinephritis hypertension induced by either a bilateral renal wrap procedure or a wrap-nephrectomy procedure in 28 rabbits. Unoperated and unilaterally nephrectomized controls were also studied. Plasma renin concentration was derived from plasma renin activity and plasma renin substrate levels using a measured Km of 2.3 × 10−6M. All surgical operations were followed by a transient increase in plasma renin substrate levels and a fall in plasma renin concentration. Unilateral nephrectomy in normotensive controls suppressed plasma renin activity and concentration for a 20-day period. Similar suppression occurred during the onset of hypertension in rabbits subjected to the wrap-nephrectomy procedure. Decapsulation accompanied by a fall in blood pressure again suppressed plasma renin activity and concentration. With bilateral wrapping, hypertension developed gradually without a change in renin levels. Decreased renin secretion during the evolution of perinephritis hypertension probably reflects a decreased renal excretory capacity for salt and water that overrides the renin-releasing effect of renal compression.

Effect of hypertension on the entry of 125I-labelled low density lipoprotein into the aortic intima in normal-fed rabbits

The entry of [125I]-labelled low density lipoprotein (LDL) into different regions of the aortic intima has been studied over a 6 hour period in both normotensive and renal hypertensive rabbits fed a normal diet. Studies have also been carried out in previously hypertensive rabbits in which the blood pressure was normalized with parenteral hydralazine during the six hour period, in which entry was studied. In the normotensive rabbits entry into the aortic intima was less than 1 mug of LDL protein/100 mg dry defatted weight over the 6 hour period with greatest entry into aortic arch intima and significantly less into both the thoracic and abdominal aortic intimae. Hypertension increased the entry into the arch and into aortic arch intimae. Hypertension increased the entry into the arch and into the thoracic and abdominal segments but this was only statistically significant for the aortic arch. The entry of [125I] LDL into the intima in those rabbits in which the hypertension had been normalized was similar to that for the hypertensive rabbits. The results suggest that hypertension in the normal fed rabbit increases lipoprotein entry into the arterial wall by an effect on vessel wall permeability rather than by a direct effect of filtration pressure.

Phospholipase A in the venom of the Australian bulldog ant Myrmecia pyriformis

Abstract The venom of M. pyriformis was examined for the presence of phospholipase A activity. Incubation of the venom with highly purified samples of plant lecithin and of liver lecithin demonstrated that the venom possesses lecithin-splitting activity. The fatty acid distribution in the hydrolysis products after incubation of the ant venom with liver lecithin with different fatty acid composition at the 1- and 2-positions was the same as after incubation with a purified snake venom phospholipase A preparation. Hence the venom hydrolyses the ester bond in the 2-position on the lecithin molecule, and possesses phospholipase A activity.

Incorporation of [14C]oleic acid into lipid by foam cells and by other fractions separated from rabbit atherosclerotic lesions☆

Abstract [ 14 C]oleic acid was incubated in vitro with atherosclerotic aortas obtained from cholesterol-fed rabbits; following incubation, foam cells and other fractions of the aortic wall were separated and studied. Most of the [ 14 C]oleic acid was incorporated into cholesterol ester in the intima whereas most was incorporated into triglyceride and phospholipid in the media. About 10 % of the oleic acid taken up and incorporated into phospholipid, triglyceride or cholesterol ester in the intima was found in the intact foam cells separated. The specific activity of the oleic acid incorporated into both phospholipid and cholesterol ester in the foam cells was considerably higher than that in the other portions of the arterial wall. The role of the foam cell in the synthesis of lipid in the atherosclerotic arterial wall and the possible pathways involved in such synthesis by foam cells are discussed.