Long-term induction and regression of diet-induced atherosclerotic lesions in rhesus monkeys. II. Morphometric evaluation of lesions by light microscopy in coronary and carotid arteries.

Atherosclerotic lesions were induced in rhesus monkeys (Macaca mulatta) by feeding them a high-saturated fatty acid and high-cholesterol diet. After 5.4 years the extent of lesions in three major coronary arteries and the right carotid artery was evaluated morphometrically by light microscopy in one group of animals (group P). The remaining animals were switched to a low-cholesterol diet that remained high in saturated fatty acids and provided the same percentage of total calories as did the atherogenic diet. Lesion regression was then evaluated in one group of monkeys 1.9 years (group R4) and in another group of monkeys 3.7 years (group R5) after withdrawal of cholesterol alone from the diet. In group P, the mean intimal thickness varied between 26 and 47 microns, maximum intimal thickness between 70 and 92 microns, and luminal reduction between 9% and 12% in the three major coronary arteries. Luminal reduction varied between 1% and 11% in right carotid artery segments. After 1.9 years of consuming the basal diet, group R4 animals were no different from group P animals with respect to morphometric measures. Total intimal and medial areas of the left anterior descending (LAD) coronary artery in groups P and R4 were also similar. In contrast, after 3.7 years of consuming the basal diet, group R5 animals showed consistently although not statistically significantly lower values than those in group P for the morphometric measures in coronary arteries and total intimal area in the LAD. Similar results were obtained for the common carotid and external carotid arteries. Thus, our study shows that long-term diet-induced lesions in coronary arteries and in common and external segments of the right carotid artery regressed only when the animals were fed the basal diet for 3.7 years. We conclude that atherosclerotic lesions induced in coronary and carotid arteries can regress toward normal to a certain extent, but they require a longer time for regression than do other arterial segments. These findings support the results of clinical trials in human subjects.

Long-term induction and regression of diet-induced atherosclerotic lesions in rhesus monkeys. I. Morphological and chemical evidence for regression of lesions in the aorta and carotid and peripheral arteries.

Atherosclerotic lesions were induced in rhesus monkeys by feeding a high-saturated fatty acid and high-cholesterol diet. After 5.4 years the extent of fatty streaks and raised lesions was evaluated in one group of animals (group P) by visual estimation in 10 arterial segments and chemically in four arterial segments. The remaining animals were switched to a basal regression diet low in cholesterol but high in saturated fatty acids for up to 3.7 years. Regression of lesions was evaluated in one group for 1.9 years (group R4) and in another for 3.7 years (group R5) after deleting cholesterol from the diet. The atherogenic diet increased serum cholesterol levels in all animals from a mean of 150 mg/dL to a mean of about 430 mg/dL. The atherogenic diet produced lesions in group P in all arterial segments (involving up to 50% of the arterial intimal surface) and increased cholesterol content in four arterial segments (varying between 443 and 506 micrograms/cm2). Switching to the basal regression diet decreased serum cholesterol levels to normal after 12 to 18 weeks. The switch to the basal diet significantly decreased the extent of fatty streaks in most arterial segments in both groups. Although differences in the mean extent of raised lesions among groups were not statistically significant, 7 of 10 arterial segments in group R4 and 9 of 10 segments in group R5 showed a lesser extent of raised lesions than in group P. Cholesterol content was lower (P < .05) in all four arterial segments in group R5 than in group P.(ABSTRACT TRUNCATED AT 250 WORDS)

Use of serum lipid and apolipoprotein concentrations to predict extent of diet-induced atherosclerotic lesions in aortas and coronary arteries and to demonstrate regression of lesions in individual rhesus monkeys.

In most studies, the assessment of lesion "regression" is based on comparisons of group means between the "progression" and regression groups. This comparison depends on the assumption that the extent and distribution of lesions produced by the end of the lesion-induction period in the regression animals are equal to those observed in the progression group. To determine whether significant regression of lesions occurs in an individual regression animal, it is necessary to obtain a measure of the lesions produced in these animals at the end of the lesion-induction period. We achieved this goal by developing models using multiple stepwise regression analysis that related steady-state serum cholesterol and apolipoprotein B and A-I concentrations measured during a lesion-induction period in 27 rhesus monkeys fed an atherogenic high-saturated-fat/high-cholesterol diet for 2 years. The models were developed to estimate the percent of intimal surface with lesions, the esterified cholesterol content (micrograms/cm2) for the artery segments, and three histomorphometric measures (mean intimal thickness, mean maximal intimal thickness, and mean percent stenosis) for the coronary arteries. In these models, multiple R2 ranged from 0.42 to 0.74 for the aortas and peripheral arteries, indicating that approximately one half to three fourths of the variance in lesions was accounted for. For the three histomorphometric measures in coronary arteries, however, the multiple R2 was 0.27 or 0.28, indicating that only approximately one fourth of the variance in lesions was accounted for.(ABSTRACT TRUNCATED AT 250 WORDS)

Changes in plasma lipoprotein concentrations and compositions upon feeding cholesterol in high- and low-responding rhesus monkeys.

When fed cholesterol, the high-responding rhesus monkeys develop severe hypercholesterolemia, whereas low-responding rhesus monkeys show only slight increases in plasma cholesterol levels. We report changes in plasma lipoprotein concentrations and compositions along with changes in plasma lipid concentrations in high- and low-responding rhesus monkeys fed a high-cholesterol diet. On low-cholesterol diet, the concentrations and compositions of plasma lipoprotein fractions were similar in the two groups. Upon feeding cholesterol, plasma very-low-density (VLDL), intermediate-density (IDL) and low-density (LDL)-lipoprotein concentrations increased in both groups, but the increases were significantly (p less than 0.01) higher in high-responders than in low-responders. Plasma HDL concentration decreased significantly (p less than 0.01) in high responders but not in low responders. In high responders, percent cholesterol increased in both VLDL and IDL fractions but in low responders, it decreased in VLDL and increased in IDL. Percent triglycerides decreased in VLDL, IDL and LDL fractions in high responders, while in low responders it tended to increase in VLDL, remained unchanged in IDL and decreased in LDL. The composition of HDL did not change in the two groups upon feeding cholesterol. Thus, when fed cholesterol, the high- and the low-responding monkeys respond distinctly differently in plasma lipoprotein concentrations and compositions. The responses occurred simultaneously, suggesting metabolic interrelationships between various lipoproteins.

Reduction of preterm birth in an HMO.

In 1984 a preterm birth prevention project was introduced into a large, well-established HMO. Five years of resultant experience and data are compared to metropolitan area and state data. The method described by Dr. Robert Creasy was followed. Some recommendations for change of the screening tool are made. The impact on the preterm birth rate in the HMO setting was substantial, with the rate of decline exceeding that of the metropolitan area and the state.

Effect of cholesterol feeding on lipolytic activities in high- and low-responding rhesus monkeys.

We compared the activities of lipoprotein (LPL) and hepatic lipase (HL) in postheparin plasma in groups of high- and low-responding rhesus monkeys fed a low cholesterol diet followed by a high cholesterol diet. Cholesterol feeding resulted in a two-phase response in lipolytic activities: an initial phase lasting about 21 days that was similar in the two groups, followed by the second phase in which major differences became apparent between the groups. In the initial phase, LPL and HL activities increased along with plasma cholesterol and high density lipoprotein (HDL) cholesterol, but there was no change in plasma triglycerides or apolipoprotein (apo) A-I levels. These changes suggest that both high and low responders respond in a similar manner to metabolize an increased lipid load and that both groups continue to remove triglyceride efficiently while cholesterol accumulation begins. The second phase (between 21 and 42 days and thereafter) in high responders was characterized by a decrease in LPL and HL activities along with plasma HDL cholesterol and apo A-I levels, continued increase in plasma cholesterol, and a slow increase in plasma triglycerides. In low responders, LPL activity was maintained at a high level, HL activity decreased to the basal level, and plasma lipids were unchanged. All changes occurred simultaneously, suggesting metabolic relationships between plasma lipolytic activities, plasma lipids, and apolipoprotein.

Differences in fecal excretion of cholesterol and bacterial degradation products in high- and low-responding rhesus monkeys: implications in colon cancer.

This paper reports the differences in the fecal excretion of cholesterol and its degradation products in high- and low-responding rhesus monkeys fed diets with and without extra cholesterol. The high-responding monkeys had a great increase in plasma cholesterol concentration when fed a high-cholesterol diet, whereas the low-responders had a small increase when fed the same diet. The results show that low-responding monkeys, when fed high-cholesterol diets, excrete nearly two to three times the amount of cholesterol and its bacterial degradation products in the feces than the high-responding monkeys. We suggest that these two select groups of monkeys may be useful models for the study of the role cholesterol and its bacterial degradation products play in colon cancer.

Studies on the mechanism of high intestinal absorption of cholesterol and campesterol in high-responding rhesus monkeys.

To understand better the mechanism of higher absorption of cholesterol and campesterol in high-responding than in low-responding rhesus monkeys, we measured the concentrations of the two sterols in the micellar fraction isolated from small intestinal content, and also determined their rates of esterification by cholesterol esterase prepared from the small intestinal mucosa. The results show that the concentrations of both cholesterol and campesterol in the micellar fraction were significantly higher in the high- than in low- and intermediate-responding rhesus monkeys. Also the rates of esterification of both sterols are higher in the proximal segment of the small intestine in high-responders than the other two groups. We conclude that the two necessary steps in the process of sterol absorption, namely, the amounts of sterols solubilized in micelles and their esterification within mucosal cells which are higher in high- than in low-responders are responsible for the higher absorption of the sterols in the high-responding rhesus monkeys.

Relationships between dietary cholesterol, cholesterol absorption, cholesterol synthesis, and plasma cholesterol in rhesus monkeys.

The relationships between the cholesterol content of the diet, plasma cholesterol concentration, cholesterol absorption, and cholesterol synthesis (measured indirectly by desmosterol suppression technique) are explored in groups of high- and low-responding rhesus monkeys fed diets containing 0.02, 0.15 and 0.75 mg cholesterol/kcal. The cholesterol content of the diet (expressed as mg/kcal) is positively correlated with plasma cholesterol concentration and is negatively correlated with percent cholesterol absorption in both groups. Also the cholesterol content of the diet is negatively correlated (r = -0.66) with cholesterol synthesis only in low-responders but not in high-responders. Similarly, the correlation between percent cholesterol absorption and cholesterol synthesis is significant in the low-responders (r = 0.82, P less than 0.01) but not in the high-responders (r = 0.12, P greater than 0.1). Further, a negative correlation (r = -0.61) is observed between cholesterol synthesis and plasma cholesterol concentration in the low-responders but in the high-responders, there is no relationship between the parameters. From these correlations we conclude that the higher cholesterol absorption in the high-responders than in the low-responders is one important mechanism responsible for the development of severe hypercholesterolemia in the high-responding monkeys fed cholesterol. The results also suggest that there might be a lower sensitivity in the feedback inhibition mechanism of cholesterol biosynthesis in the high-responding than in the low-responding rhesus monkeys.

Regression of experimental atherosclerotic lesions in rhesus monkeys consuming a high saturated fat diet.

Atherosclerotic lesions were induced in rhesus monkeys by feeding them a high-fat, high-cholesterol diet for 2 years. Arteries were examined after autopsy of a subgroup of animals (group P) and cholesterol was removed from the diet of the remaining animals. Lesions were examined in other subgroups after 30 weeks (group R1) and after 52 weeks (group R2). A control group (group C) was fed the diet without cholesterol throughout the study. The mean total serum cholesterol concentration before, during, and after lesion induction was 151, 390, and 157 mg/dl, respectively. The mean percent of surface area with fatty streak or fibrous plaque and the free and esterified cholesterol content of the artery increased in all six arterial segments examined in group P. The means for percent of surface with fatty streak and for arterial cholesterol content or concentration (but not for extent of surface with fibrous plaque) were consistently less in groups R1 and R2 than in group P, although they remained greater in groups R1 and R2 than in group C. The mean intimal thickness for coronary arteries was 10-fold greater in group P than in group C and 60 percent less in groups R1 and R2 than in group P; there was, however, much variability among animals and these differences among groups were not statistically significant. By using several measures in several arterial systems, we have shown that there was regression of diet-induced atherosclerotic lesions in rhesus monkeys while they were fed a diet high in saturated fat but without cholesterol for 30 or 52 weeks.

Effect of dietary cholesterol level on plasma campesterol concentration in rhesus monkeys.

High- and low-responding rhesus monkeys were fed diets containing 0.02, 0.15 and 0.75 mg/kcal cholesterol without or with 2% of a mixture of plant sterols for 6 weeks. No plant sterols were detected in the plasma of either high- or low-responding animals fed the cholesterol-containing diets but with low levels of plant sterols. After addition of 2% plant sterols to the diets, only campesterol was found in the plasma of both groups of animals. The plasma campesterol concentrations in either the high- or the low-responding groups were highest on the diet with 0.75 mg/kcal cholesterol and lowest with 0.02 mg/kcal cholesterol. Also, on all three diets, the plasma campesterol concentration was higher in the high responders than in the low responders. We conclude, based on the plasma campesterol levels, that the level of cholesterol in the diet affected the plasma level by affecting the intestinal absorption of the sterol. The higher the dietary cholesterol level, the higher the absorption of campesterol, and thus the higher the plasma level. Also, the high responders absorb higher amounts of campesterol than the low responders, a finding similar to cholesterol absorption in the high-responding rhesus monkeys.

The rate of biliary cholesterol secretion in high- and low-responding rhesus monkeys.

The rates of secretion of cholesterol in bile measured by an isotope ratio method were found similar in cholesterol-fed high- and low-responding rhesus monkeys. The results indicate that the failure on the part of the high-responders to increase proportionately the fecal excretion of neutral steroids to compensate for the higher absorption of cholesterol than the low-responders, as suggested earlier, is not due to a difference in the rate of biliary cholesterol secretion but must lie in some other aspect of cholesterol metabolism.

Effects of feeding cholesterol and mixed plant sterols on the fecal excretion of acidic steroids in rhesus monkeys.

The effects of feeding diets with high or low amounts of cholesterol and with low or high levels of mixed plant sterols (sitosterol: campesterol: stigmasterol, 60:35:5) on the daily fecal excretion of acidic steroids were studied in rhesus monkeys. During periods of low dietary plant sterol, total fecal acidic steroid excretion was 43% lower (P less than 0.01) during low dietary cholesterol than during high dietary cholesterol. During periods of high dietary plant sterols the fecal acidic steroid excretion was 113% higher (P less than 0.01) with low dietary cholesterol than with high dietary cholesterol. Addition of mixed plant sterols to the low-cholesterol diet produced nearly a 2-fold increase (P less than 0.005) whereas, such an addition to the high cholesterol diet produced a significant decrease by about 53% (P less than 0.025) in the total fecal acidic steroid excretion. The results suggest that the effect of cholesterol feeding on fecal acidic steroid excretion depends on the level of plant sterols in the diet. This interaction of the effects of cholesterol and plant sterols on the fecal acidic steroid excretion is probably related to the inhibitory effect of plant sterols on cholesterol absorption.

Effect of triparanol on cholesterol absorption in rhesus monkeys.

The drug, triparanol, a known inhibitor of cholesterol biosynthesis, was found to interfere with absorption of cholesterol in rhesus monkeys. The percent luminal cholesterol absorbed decreased by 13 and 21%, respectively, in the high- and low-responding monkeys when the drug was fed along with the low-cholesterol, low-plant sterol diet. When the diet contained large amounts of plant sterols, feeding of the drug also reduced the percent cholesterol absorption by the same order of magnitude. The results showed that triparanol decreased the absorption of endogenous cholesterol in rhesus monkeys. The effect of the drug on cholesterol absorption was independent of that of plant sterols. The mechanism(s) of the observed inhibitory effect of triparanol on cholesterol absorption is not clearly understood.

Lipoprotein profiles in rhesus monkeys with divergent responses to dietary cholesterol.

From a group of 53 rhesus monkeys, we selected 12 animals, the six with the highest and the six with the lowest response to a high cholesterol diet, and we made detailed analyses of their cholesterol and apolipoprotein profile. The high responders differed from the low responders in several ways. During the high cholesterol diet period, the high responders had much higher plasma apolipoprotein B and E concentrations and much lower plasma apolipoprotein A-I concentrations than did the low responders. Nearly all the increase in plasma cholesterol and apolipoproteins B and E concentrations in the high responders occurred in the lower density fractions (d = 1.006-1.030 g/ml), while the decrease in plasma apolipoprotein A-I concentrations in the high responders was confined to the lower density fraction of the high density lipoproteins (HDL), i.e., HDL2 (d = 1.063-1.125 g/ml). In the low responders, on the other hand, the slight increase in cholesterol concentrations was evenly distributed between the lower density fractions and HDL, and the increase in apolipoprotein A-I concentration of the lower density fractions is related to the decrease in the concentration in the HDL2 in the high responders.

Mechanism of the variability in plasma cholesterol response to cholesterol feeding in rhesus monkeys.

The response of plasma cholesterol to cholesterol feeding in rhesus monkeys varies widely among individual animals. Some develop severe hypercholesterolemia (the high-responders) while some show only a mild increase in plasma cholesterol concentration (the low-responders) with similar cholesterol intake. We have reviewed our studies on cholesterol absorption and metabolism in the high- and low-responding rhesus monkeys and conclude that the consistent observation that high-responders absorb significantly higher percentage of intestinal luminal cholesterol than the low-responders is one primary mechanism responsible for the differential response of plasma cholesterol to dietary cholesterol. We have discussed the significance of these findings in rhesus monkeys, an animal species used extensively as a model for atherosclerosis research, as they may relate to cholesterol metabolism in humans.

Survey of serum cholesterol and triglyceride concentration and lipoprotein electrophoretic pattern in rhesus monkeys (Macaca mulatta).

Serum lipids and lipoproteins were surveyed during quarantine of a group of 811 wild-caught rhesus monkeys. No abnormalities indicative of dyslipoproteinemia analogous to the familial hyperlipoproteinemias in man were observed. Males had significantly lower mean cholesterol (134 mg/dl) and higher mean triglyceride (41.9 mg/dl) concentrations than females (155 and 37.5 mg/dl, respectively). Electrophoretic distribution of lipoproteins differed with sex. Cholesterol concentration and electrophoretic pattern had not attained steady state by ten weeks either in animals recently imported or in those conditioned in another laboratory for six months or more after import.