Endothelial damage and thrombocyte adhesion in pigeon atherosclerosis.

Scanning electron microscopy studies of spontaneously occurring atherosclerosis in pigeons reveal dramatic alterations in endothelial integrity. An irregular endothelium at the intimal cushion region of 5-week-old birds gives rise to extensive areas of pitted endothelium and subendothelial exposure. Thrombocytes, thrombocyte aggregates, and leukocytes are associated with the developing lesion.

Effect of neonatal modulation of cholesterol homeostasis on subsequent response to cholesterol challenge in adult guinea pig.

Experiments were designed to study whether or not the mechanism of handling dietary cholesterol in adulthood can be modulated by the manipulation of cholesterol homeostasis during neonatal period. The effects of enhancing cholesterol degradation (cholestyramine feeding), high dietary cholesterol intake, and early weaning during neonatal period of guinea pigs on their subsequent plasma cholesterol levels and the response to dietary cholesterol challenged in adulthood were investigated. Pretreatment of neonatal guinea pigs with cholestyramine resulted in (a) a lower plasma cholesterol level, (b) an increased excretion rate of fecal bile acids and total steroids, (c) an expanded bile acid pool, (d) an increased activity of cholesterol 7 alpha-hydroxylase, and (e) no change in the hepatic 3-hydroxy-3-methylglutaryl coenzyme A (CoA) reductase activity when challenged with cholesterol in adulthood. Cholesterol pretreatment during neonatal period resulted in (a) no alteration in the plasma cholesterol level, (b) no alteration in the fecal excretion of steroids, or (c) no alteration in the cholesterol 7 alpha-hydroxylase activity when they were challenged with a high cholesterol diet. Early weaning did not influence the fecal excretion of steroids or cholesterol 7 alpha-hydroxylase activity but resulted in a slight decrease in the hepatic 3-hydroxy-3-methylglutaryl-CoA reductase activity when they were challenged with a high cholesterol diet. These results suggest that stimulation of cholesterol catabolism rather than cholesterol feeding or early weaning during neonatal period can influence the response to dietary cholesterol challenge in adulthood.

Effect of coconut oil on plasma apo A-I levels in WHHL and NZW rabbits.

Age-matched Watanabe (WHHL) and New Zealand White (NZW) rabbits were fed a coconut oil-enriched diet (14%, w/w) for 2 weeks. Lipid and apolipoprotein (apo) A-I levels in plasma and lipoprotein fractions were monitored. Within 3 days after the start of the coconut oil diet, plasma apo A-I and high-density lipoprotein (HDL)-apo A-I levels increased 3-fold in the WHHL rabbits. A smaller but significant increase (63%) in apo A-I and HDL-apo A-I levels was also observed in the NZW rabbits. HDL cholesterol levels also increased from 16 +/- 3 mg/dl during a regular diet to 46 +/- 16 mg/dl (288%) during the coconut oil diet in the WHHL rabbits and from 37 +/- 7 mg/dl to 69 +/- 19 mg/dl (186%), respectively, in the NZW rabbits. Apo A-I and HDL cholesterol levels fell sharply to the original levels soon after switching back to a regular diet (within 3 days for WHHL rabbits and within 5 days for NZW rabbits). The fractional catabolic rate calculated from 125I-HDL kinetic studies indicated that the turnover rate for HDL was significantly slower in WHHL rabbits fed the coconut oil diet than the control diet (0.018 +/- 0.004 h-1 vs. 0.027 +/- 0.007 h-1, P less than 0.01). No changes were found in the NZW rabbits fed either diet. Trilaurin, the main component of the coconut oil (46.9%) supplemented diet (6.5%, w/w), was also used in this study. The effect of trilaurin on plasma apo A-I and HDL-cholesterol levels is discussed.

Tween-20 increases the immunoreactivity of apolipoprotein A-I in plasma.

Tween-20 (polyoxyethylene 20), a trademark for a sorbitan polyoxyalkalene derivative is used as an emulsifier and detergent. In present studies, we report that Tween-20 is able to reduce the nonspecific binding of radiolabeled apolipoprotein A-I to test tubes. It also enhances the specific binding of 125I-labeled apolipoprotein A-I to anti-apolipoprotein A-I antibodies. Maximal enhancement was achieved at a Tween-20 concentration greater than 0.08%. The results were confirmed by using two different antisera raised in rabbits and goats. Since the immunoreactivity of apolipoprotein A-I in plasma cannot be fully detected by a conventional radioimmunoassay procedure, we have, therefore, studied the effect of Tween-20 on the immunoreactivity of apolipoprotein A-I in plasma. As determined by inhibition radioimmunoassay, we found that Tween-20 drastically increased the immunoreactivity of apolipoprotein A-I using both rabbit and goat antisera. The maximal reactivity was achieved at a Tween-20 concentration of 0.32%. However, Tween-20 did not enhance the immunoreactivity of delipidated plasma. Thus, the results indicate that the plasma lipids may hamper apolipoprotein A-I immunoreactivity and suggest that the nonionic detergent, Tween-20, somehow interacted with HDL lipids or the apolipoproteins and then facilitated the reaction of antigenic reactive site(s) of apolipoprotein A-I with their antibodies.

Rapid regulation of apolipoprotein A-I secretion in HepG2 cells by a factor associated with bovine high-density lipoproteins.

The effect of fetal bovine serum (FBS) on the secretion of apolipoprotein A-I (apo A-I) by HepG2 cells was studied. The cells incubated with FBS always secreted more apo A-I than the cells incubated with serum-free medium. The changes in the rate of apo A-I secretion were observed within 1 h after addition or depletion of serum. The high-density lipoproteins (HDL) or the lipoprotein-deficient serum (LPDS) obtained from FBS also stimulated apo A-I secretion rapidly to the same level as obtained with FBS. Addition of low-density lipoproteins did not have any effect. The rate of general protein synthesis was not affected by short-term incubations with or without serum or HDL. The rate of apolipoprotein E secretion by these cells did not change significantly, parallel to the changes in apo A-I secretion in the presence or absence of FBS. It is concluded that serum may have a factor that plays a specific role in the regulation of apo A-I secretion by the liver cells and this factor is associated with the HDL fraction.

Studies on the effect of storage of normolipidemic plasma on the metabolism of very-low-density lipoprotein cholesterol in human skin fibroblasts.

Very-low-density lipoproteins (VLDL) isolated from fresh normolipidemic plasma were able to compete with labeled low-density lipoproteins (LDL) for receptor binding as efficiently as LDL in human skin fibroblasts. However, VLDL from fresh plasma failed to stimulate acyl-CoA: cholesterol acyltransferase (ACAT) (EC 2.3.1.26) activity significantly. When plasma was stored at 4 degrees C for 24-48 h before ultracentrifugation, the VLDL were able to stimulate ACAT activity significantly. Proteolytic and lipolytic enzymes do not appear to play a role in this phenomenon. The increased capacity of VLDL from stored plasma to stimulate ACAT activity appears to be due partly to the increase in the internalization and degradation processes. Both the larger and the smaller VLDL subclasses were able to stimulate ACAT activity in the cells. The actual changes that take place in the VLDL molecule during the storage of plasma are not known.

Metabolism of glycosylated very-low-density lipoproteins in human skin fibroblasts.

Very-low-density lipoproteins (VLDL) were glycosylated using glucose and sodium cyanoborohydride. The binding and rate of degradation of glycosylated VLDL in human skin fibroblasts were reduced significantly compared to native VLDL. These glycosylated VLDL were not able to activate acyl-CoA:cholesterol acyltransferase or inhibit cholesterol synthesis in fibroblasts. Glycosylated VLDL showed higher mobility on agarose electrophoresis, and apolipoprotein E in these lipoproteins showed higher molecular weight on SDS-polyacrylamide gel electrophoresis. The physiological significance of glycosylation of VLDL in hyperglycemic subjects is speculated.

Physical, chemical, and immunochemical studies of apolipoprotein A-I from pigeon plasma high density lipoproteins.

Pigeon plasma high density lipoproteins (HDL) were isolated by ultracentrifugation between the densities of 1.063 and 1.21 g/ml. Gel filtration of delipidated HDL in 5 M guanidine-HCl on Sephadex G-150 yielded a major fraction which eluted at the same position as human apolipoprotein A-I isolated from HDL. In SDS-gel electrophoresis, the isolated apolipoprotein co-migrated with human apolipoprotein A-I with a molecular weight of approx. 28 000. The amino acid composition was similar to the apolipoprotein A-I isolated from human and hen plasma. The isolated apolipoprotein from pigeon plasma had therefore been designated as apolipoprotein A-I. As judged by circular dichroism (CD), the apolipoprotein A-I displayed a maximum mean residue ellipticity of approx. -3 000 at 222 nm while at concentrations greater than 0.2 mg/ml. Calculations of alpha-helicial content gave values of 85%. Lowering the concentration of apolipoprotein A-I was found to concomitantly decrease the ellipticity (absolute value) suggesting that there was some conformational change when the apolipoprotein A-I concentration varied. The isolated pigeon apolipoprotein A-I was found bound to the phospholipid (dimyristoyl phosphatidylcholine) and there was no significant conformational change upon lipid binding as judged by CD. Under the same experimental conditions, human apolipoprotein A-I exhibited a drastic conformational change by increasing its helicity in the presence of phospholipid. The helical content of human apolipoprotein A-I was increased from 48 to 85%. This finding suggests that the apolipoprotein may not necessarily increase its helical content during lipid binding. Moreover, immunochemical studies showed that rabbit antiserum prepared against pigeon apolipoprotein A-I could partially react with human apolipoprotein A-I determined by quantitative radioimmunoassay.

Immunochemistry of human plasma apolipoprotein C-II as studied by radioimmunoassay.

The immunoreactivity of human plasma apolipoprotein C-II was investigated using a specific radioimmunoassay. In whole plasma, the mean value quantitated was 2.21 +/- 0.415 mg/dl, while in delipidated plasma, a mean value of 3.84 +/- 1.186 mg/dl was obtained, suggesting that the antigenic sites of the apolipoprotein were not fully detected in unmodified plasma by our antibody preparation. Two detergents, Tween-20 and Triton X-100, were studied to determine if they could enhance the immunoreactivity of apolipoprotein C-II in whole plasma. At concentrations of 0.012-0.06%, Tween-20 markedly increased the immunoreactivity of whole plasma, but not of delipidated plasma, indicating that antigenic sites of plasma apolipoprotein C-II has been exposed by Tween-20. In contrast, Triton X-100 had no effect on the immunoreactivity of whole plasma apolipoprotein C-II. A radioimmunoassay conducted in the presence of 0.06% Tween-20, resulted in a mean value in whole plasma (3.39 +/- 1.11 mg/dl) that was not significantly different from that obtained when the assay was done on delipidated samples. The immunoreactivity of VLDL apolipoprotein C-II was also drastically enhanced following lipolysis by bovine milk lipoprotein lipase, supporting the hypothesis that antigenic sites are masked by the lipids. Finally, the mechanism responsible for the effect of Tween-20 on apolipoprotein C-II immunoreactivity was investigated. The results obtained from circular dichroism and ultracentrifugation suggest that the detergent may dissociate the apolipoprotein from lipoprotein particles, thus fully exposing the antigenic sites for reaction with antibodies.

Cholestyramine treatment in early life of low-density lipoprotein receptor deficient Watanabe rabbits: decreased aortic cholesteryl ester accumulation and atherosclerosis in adult life.

Effect of cholestyramine treatment in early life of Watanabe heritable hyperlipidemic rabbits (an animal model lacking low-density lipoprotein receptor activity) on subsequent (6 months recovery) occurrence of natural atherosclerotic lesion and arterial cholesterol metabolism was investigated. Initial cholestyramine treatment decreased both plasma total cholesterol and HDL-cholesterol levels which normalized within 4 weeks after treatment was discontinued. At 9 months of age (age of occurrence of spontaneous atherosclerotic lesions), the extent of aortic atherosclerosis in cholestyramine pre-treated animals was modestly lower (P less than 0.05), as compared to controls, with a significant (P less than 0.05) decrease in aortic cholesteryl ester content. Furthermore, at the end of the recovery period aortic activity of acyl-CoA: cholesterol acyltransferase and neutral cholesterol esterase activity was significantly (P less than 0.05) lower in cholestyramine-pretreated animals. These studies show that early cholestyramine pre-treatment in a low-density lipoprotein receptor-deficient animal model causes persistent changes which might influence cholesteryl ester accumulation and atherogenesis in adult life, even after cholestyramine treatment is discontinued.

Primary inferior vena cava thrombosis: report of nine cases.

All cases of inferior vena cava obstruction diagnosed at the Mayo Clinic between 1950 and 1973 were reviewed. Diagnosis was confirmed by surgery, phlebography, or postmortem examinations in 64 cases; the cause in 55 cases is described. Carcinoma of the kidney was the most common cause (31% pf cases). In nine cases, extensive laboratory investigation failed to reveal the cause of the process. These cases were considered to be primary inferior vena cava thrombosis and are reviewed in detail.

Insulinemia in children at low and high risk of NIDDM.

OBJECTIVE: Fasting hyperinsulinemia in the presence of normoglycemia usually indicates insulin resistance and is characteristic of populations at high risk for developing NIDDM. Hyperinsulinemia predicts the development of impaired glucose tolerance and NIDDM in Pima Indians, a population with a high incidence of NIDDM. Insulin concentrations in population-based samples of children who have different risks of developing NIDDM later in life have not been reported previously. RESEARCH DESIGN AND METHODS: We compared fasting insulin concentrations in two populations of nondiabetic children, 6-19 yr of age: Pima Indians from southern Arizona and Caucasians from Minnesota. RESULTS: Insulin concentration varied with age, sex, glucose concentration, and relative weight. Mean fasting insulin concentration was 140.3 pM in Pima Indian males, 94.4 pM in Caucasian males, 171.5 pM in Pima Indian females, and 107.1 pM in Caucasian females. For each sex, the mean fasting insulin concentration, controlled for age, glucose, and relative weight, was significantly higher in the Pima Indians than in the Caucasians (P < 0.001). CONCLUSIONS: From a young age, Pima Indian children have higher fasting insulin concentrations than Caucasian children. As hyperinsulinemia predicts subsequent NIDDM, these data suggest that the susceptibility to NIDDM is manifest at a young age as fasting hyperinsulinemia.

Lipoprotein compositional changes in the fasting and postprandial state on a high-carbohydrate low-fat and a high-fat diet in subjects with noninsulin-dependent diabetes mellitus.

Our aims were 1) to examine the effects of a high-carbohydrate low-fat diet on fasting and postprandial plasma lipids, apolipoproteins (apo), and lipoprotein composition in noninsulin-dependent diabetes mellitus, and 2) to determine whether postprandial shift of apo between triglyceride-rich lipoproteins (TRLP) and high density lipoproteins (HDL) is affected by diet. A cross-over study, of 4 weeks duration, of a high-carbohydrate (60% carbohydrate, 20% fat) and a high-fat (40% carbohydrate, 40% fat) diet was performed in seven subjects with noninsulin-dependent diabetes mellitus. TRLP, low density lipoproteins (LDL), and HDL were separated by fast protein liquid chromatography. The high-carbohydrate diet resulted in a decrease of fasting total, HDL, and LDL cholesterol and a trend toward an increase in plasma triglycerides. The apo composition of fasting TRLP and HDL was similar on both diets. TRLP apo CII, CIII, and E increased whereas HDL apo CII, CIII, and E decreased postprandially on both diets. In contrast, TRLP apo CI increased, and HDL apo CI decreased only after the high-carbohydrate diet. We conclude that 1) a high carbohydrate diet results in a decrease in total, LDL, and HDL cholesterol and a trend toward an increase in plasma triglycerides; 2) fasting TRLP and HDL apo composition was similar on a high-carbohydrate or a high-fat diet; and 3) on both diets, apo CII, CIII, and E transfer from HDL to TRLP postprandially. However, only the high-carbohydrate diet induced postprandial transfer of apo CI from HDL to TRLP. This may explain in part the changes in lipoproteins observed with this diet.

Bile acid metabolism in ascorbic acid-deficient guinea pigs.

Sterol balance techniques have been used to determine the effect of short-term ascorbic acid (AA) deprivation on bile acid excretion in the guinea pig. The effects of a brief (2-week) AA deficiency on bile acid pool sizes and the activity of the rate controlling enzyme in bile acid biosynthesis have been determined. It was found that, while food intake and body weight were not affected by the short-term AA deficiency, liver AA levels had fallen to 25% of control levels. At the same time, the rate of excretion of bile acids and the size of the bile acid pool were both reduced by about 50% in guinea pigs deficient in AA. These results were supported by a decrease in the activity of cholesterol 7 alpha-hydroxylase in the deficient animals. It is concluded that an AA deficiency will significantly impair bile acid metabolism independent of any side effects of clinical scurvy.

Postprandial exchange of apolipoprotein C-III between plasma lipoproteins.

Three healthy male subjects were given a high fat meal after fasting for 12 h. Blood samples were drawn at hourly intervals over 6 h. The plasma triglyceride levels reached peak values within 3 to 6 h postprandially. Plasma cholesterol concentration, however, remained constant in the three subjects as well as in a fasting normal subject. Apolipoprotein C-III (apoC-III), a major apoprotein in triglyceride-rich lipoprotein particles and known to be exchangeable between plasma high density lipoproteins (HDL) and triglyceride-rich particles, was studied in regard to its net transfer between lipoproteins during meal absorption and postabsorptive lipolysis. ApoC-III levels in total plasma as quantitated by radioimmunoassay were stable regardless of the increase of plasma triglycerides. When triglyceride levels increased after a meal, apoC-III in the d. less than 1.063 lipoprotein fraction increased concomitantly, while apoC-III in HDL decreased. The converse was observed during lipolysis as plasma triglycerides fell. In any case, apoC-III levels in d. less than 1.063 lipoproteins were positively correlated with plasma triglycerides (r = 0.82, p less than 0.01) after the meal. The finding suggests that the apoC-III concentrations in HDL are very dynamic in vivo. ApoC-III can transfer from HDL to triglyceride-rich particles during meal absorption and can transfer from triglyceride-rich particles to HDL during postabsorptive lipolysis.

Differential health benefits of weight loss in upper-body and lower-body obese women.

Upper-body obesity (UB Ob) is more strongly associated with adverse health consequences; however, few obesity-treatment studies have examined outcome according to body-fat distribution. To examine whether diet and formal- or informal-exercise instruction causes differential changes in health and lipid profiles, ten LB Ob and nine UB Ob premenopausal women received dietary intervention (2.1 MJ-deficit/d for 16 wk) and were randomly assigned to either formal- or informal-exercise instruction. Weight loss was similar between groups (approximately 8 kg), and no change occurred in lean body mass or basal metabolic rate. Baseline cholesterol and triglycerides were greater (P < 0.01) in UB Ob than LB Ob women and decreased more (P < 0.01) in response to treatment in UB Ob women. Formal exercise instruction increased high-density-lipoprotein cholesterol (P < 0.05) especially in UB Ob women. Future studies on treatment of obesity should include consideration of regional fat distribution.

Sucrose sensitivity of patients with coronary artery disease.

Various loads of simple carbohydrates were fed to 148 patients with known coronary-artery disease (CAD) for 4 days in the Clinical Research Center. The 148 patients were grouped according to the diet regimen tested-sucrose (low and high), glucose, and fructose. A high-sucrose diet was fed to 29 control subjects. Diets containing 2 g of simple carbohydrate (predominantly either sucrose or glucose) per kilogram of body weight per day had no significant effect on fasting plasma glucose, serum triglycerides, or serum free fatty acids. However, diets containing 4 g of simple carbohydrate (predominantly sucrose) or 2 g fructose per kilogram of body weight per day produced a significant rise in serum triglycerides with decreases in fasting plasma glucose and free fatty acids. Serum cholesterol diminished in all the diet groups, probably because of the decrease fat and cholesterol intakes. The increase of serum triglycerides in CAD patients receiving simple carbohydrate at the 4-g/kg rate was significantly greater than in the normal control subjects fed the same diet, suggesting a sensitivity of CAD patients to this stimulus. No significant correlation could be demonstrated between changes in serum triglycerides and the extent of CAD (one, two, or three vessels) as determined from coronary angiograms.

Plasma lipids and apolipoprotein A-I and A-II levels in alcoholic patients.

This prospective study compared total plasma lipids, high-density lipoprotein cholesterol (HDL-C), and apolipoproteins A-I (apo A-I) and A-II (apo A-II) in 72 alcoholic patients and in 285 nonalcoholic controls. The HDL-C in the alcoholic group was not significantly different from that in the nonalcoholic controls. Alcoholic men had significantly higher levels of cholesterol and triglycerides and lower levels of apo A-I when compared with nonalcoholic controls. Alcoholic women had significantly higher levels of cholesterol and apo A-II when compared with nonalcoholic controls. Serial measurements in 25 alcoholic patients showed a significant decline in HDL-C, apo A-I, and apo A-II levels during the 4-wk hospital stay. HDL-C demonstrated its expected inverse relationship with plasma triglyceride level and its direct relationship with apo A-I, apo A-II, and the hepatic enzyme aspartate aminotransferase.

Sterol excretion and cholesterol absorption in diabetics and nondiabetics with and without hyperlipidemia.

Fecal neutral and acidic sterols and cholesterol absorption were measured in 12 normal control subjects, 40 diabetic subjects with and without hyperlipidemia, and 27 subjects with hyperlipidemia but without diabetes mellitus. All subjects were on a low-cholesterol diet (less than 300 mg cholesterol/day). Fecal excretion of neutral and acidic sterols was increased in patients with hypertriglyceridemia and was more marked in diabetic patients with hypertriglyceridemia. Cholesterol absorption was decreased in diabetic patients with hypertriglyceridemia. Otherwise, there were no significant differences in sterol excretion or cholesterol absorption in diabetic and nondiabetic subjects compared with control groups with similar lipid levels. The best predictors of fecal neutral- and acidic-sterol excretion and of estimated cholesterol synthesis were very low [corrected]-density lipoprotein triglycerides and high-density lipoprotein cholesterol. Correction of hyperlipidemia may be beneficial in decreasing cholesterol synthesis and, thereby, in decreasing the risk of atherogenesis.

Cholesterol excretion studies in familial hypercholesterolemic children and their normolipidemic siblings.

Twenty children ages 3 to 17 yr, eight with normal lipids and 12 with familial hypercholesterolemia were studied on a metabolic unit for 14 days to evaluate fecal bile acid and fecal neutral sterol excretion. The diet contained a moderately low cholesterol content, 180 to 200 mg/day. Stools were collected in three separate, 3-day pools. Fecal bile acids and fecal neutral sterols were measured using two stool markers and thin-layer, and gas-liquid chromatography techniques. Fecal neutral sterol and fecal bile acid excretion were the same for normal and familial hypercholesterolemic children on a mg/kg basis. Fecal neutral sterols in familial hypercholesterolemic children decreased with age, p less than 0.001; fecal bile acid excretion also appeared to decrease with age, but less significantly, p less than 0.07. Although the familial hypercholesterolemic children have significantly increased plasma and potentially elevated tissue or total body cholesterol, the excretion of fecal bile acids and fecal neutral sterols did not differ between familial hypercholesterolemic and normal children.