Lipid droplets in atherosclerotic fatty streaks of human aorta.

Preparations of lipid droplets and droplet-free tissue residue (cytoplasm + membranes + nuclei) were obtained by homogenization and centrifugal separation from intimal fatty streak lesions of aortic atherosclerosis of 21 adults who had died suddenly. Neutral lipids and phospholipids were analyzed by quantitative thin-layer chromatography and cholesteryl ester fatty acids by gas-liquid chromatography. Optical properties of droplets were evaluated by differential counting and sizing procedures with the polarizing microscope. The droplets occurred in mixtures of two forms distinguished by their optical properties, anisotropic (i.e. liquid crystals) and isotropic (true liquids). Both forms had average diameters of about 1.8 mu, with a range of 0.5-5mu. The proportions of the two forms varied with temperature as individual droplets changed their form; anisotropic forms averaged 83.7% at degrees C and 37.8% at 37 degrees C, with isotropic forms being 16.3 and 62.2% respectively. The proportions of anisotropic forms at 22 degrees C decreased with age. These forms were not separated for chemical analysis. The droplets contained about half the lipid in the lesions. The composition of the lipids of the droplet mixture was remarkably uniform and strikingly different from that of the droplet-free residue, respectively: cholesteryl esters 94.9% vs. 38.7%, free cholesterol 1.7% vs. 18.6%, total phospholipids 1.0% vs. 38.6%, and triglycerides 2.4% vs. 4.0%. The proportions of individual phospholipids, with the exception of lysolecithin, were also different between the preparations. In the droplets only the proportions of lecithin correlated positively with the proportion of anisotropic forms (at 22 degrees C). Droplet cholesteryl esters were particularly rich in oleic acid and when compared to residue esters had more palmitoleic (+0.7%), oleic (+12.3%), and eicosatrienoic (+2.4%) and less palmitic (-2.2%), linoleic (-12.4%), and arachidonic (-1.6%) acids. The proportions of most individual fatty acids of droplets and residue correlated positively. The lipids of the residue closely resemble those reported for the normal intima.The observations that these droplets are prominent in the morphology of the fatty streak lesions, and that their high content of oleate-rich cholesteryl esters is similar to that reported for analysis of the whole lesions, suggest that the droplets may be involved in the pathogenesis of the fatty streak lesions of artherosclerosis in man.

Studies on the purification of antihemophilic factor (factor VIII). II. Separation of partially purified antihemophilic factor by gel filtration of plasma.

A high degree of purification of antihemophilic factor was achieved by filtration of chylomicronpoor human plasma through columns of agarose. The final product contained, on the average, 67 units of antihemophilic activity per mg of protein, and was 3360-fold purified compared with the filtered plasma. The molecular weight of antihemophilic factor appeared to be at least two million. Preparations separated by gel filtration were contaminated with appreciable amounts of plasma thromboplastin antecedent (PTA), and traces of Christmas factor and Hageman factor, but no detectable fibrinogen was present. Similar fractions of plasma prepared from the blood of patients with classic hemophilia, von Willebrand's disease, or a circulating anticoagulant directed against antihemophilic factor contained, on the average, somewhat less protein than normal plasma; whether this difference was significant is not yet known. The purified fractions were partially stabilized by the addition of 1% gelatin. Adaptation of the technique of gel filtration to purification of antihemophilic factor for clinical use remains to be explored.

Studies of arteriosclerosis in Japanese and American men. I. Comparison of fatty acid composition of adipose tissue.

The proportions of fatty acids in lipids of subcutaneous adipose tissue was compared in closely age-matched, urban men from two populations with a great difference in mortality from arteriosclerosis, namely in 50 Americans and 56 Japanese aged 15-65 yr who had died suddenly and unexpectedly. Specimens from both groups were analyzed side by side for fatty acids by gas-liquid chromatography. Compared with Japanese, Americans had significantly (P<0.01) higher proportions of lauric (+ 0.2%), myristic (+ 0.4%), palmitic (+ 1.4%), stearic (+ 2.2%), and oleic (+ 5.3%) and lesser of palmitoleic (- 1.8%), linoleic (-6.3%), and linolenic (-0.4%) acids. Japanese had higher proportions of longer chain polyunsaturated fatty acids. The distributions of fatty acids for the groups at ages 35-44 yr had significant differences. With age, Americans showed significant increases of palmitic and oleic acids and decreases of lauric, myristic, stearic, and linoleic acids; Japanese showed no correlations of proportions of fatty acids with age. The significant correlations between per cent standard body weight and fatty acids in Americans were positive for palmitic and negative for lauric and stearic acids, and in Japanese, negative for myristic acid. The patterns of interacid correlations were dissimilar for the groups. These patterns may be stable characteristics of these groups providing further insight into their fatty acid metabolism. The relationships with the fatty acid compositions of the American and Japanese diets are discussed.

Dose-response study of bezafibrate on serum lipoprotein concentrations in hyperlipoproteinanemia.

The effect of bezafibrate in the dosages 450, 900 and 1350 mg daily on serum lipoprotein concentrations were studied in 20 subjects with primary hyperlipoproteinaemia (16 of type IV, 2 of type II B, 1 of type III). Except for LDL cholesterol maximum effects were obtained with 450 mg daily. Total serum cholesterol and triglycerides (TG) fell significantly. Mean very low density lipoprotein (VLDL) TG fell by 54%. The maximum effect on low density lipoprotein was obtained with 900 mg daily. The effect was highly dependent on initial concentrations, decreases being observed above 4 mmoles/l and increases below that concentration. High density lipoprotein cholesterol increased by 31% (P less than 0.01) independently of the VLDL decrease. Three subjects suffered gastrointestinal side-effects on 1350 mg daily. Only benign reversible changes were noted on non-lipid measurements. Bezafibrate is a well-tolerated drug with a good VLDL TG lowering effect. It is particularly effective in increasing HDL cholesterol concentrations.

One-year study of the effect of bezafibrate on serum lipoprotein concentrations in hyperlipoproteinaemia.

The effect of bezafibrate (2-(4-[2-(4-chlorobenzamido)-ethyl]-phenoxy)-2-methylporopionic acid) (450 mg daily) on serum lipoprotein concentrations was studied for 1 year in 14 subjects with hyperlipoproteinaemia (3 of type II A, 2 of type II B, 2 of type III and 7 of type IV). After 2 months serum cholesterol decreased from 7.12--6.08 mmoles/l (15% P less than 0.01), triglycerides from 2.52--1.96 mmoles/l (22%, P less than 0.01), very low density lipoprotein triglycerides from 1.48--0.86 mmoles/l (42%, P less than 0.01) and low density lipoprotein cholesterol from 5.00--3.81 mmoles/l (24%, P less than 0.01). While triglyceride concentrations of serum and lipoproteins remained constant for 1 year, serum and LDL cholesterol concentrations increased slightly after 6 and 12 months. The former rise was partly due also to a rise of the possibly beneficial high density lipoprotein cholesterol from 1.27--1.68 mmoles/l (32%, P less than 0.01) after 6 months, making the effect on total cholesterol less pronounced. Subjective side-effects were nausea in one patient. No severe biochemical side-effects were noted.

Effect of bezafibrate during 4.5 years of treatment of hyperlipoproteinaemia.

Bezafibrate in a dosage of 200 mg 3 times daily was given to 24 patients with type II A (n = 8), II B (n = 1) and IV (n = 15) hyperlipoproteinaemia for 4.5 years. In type II A the content of total cholesterol and that of low density lipoprotein in serum decreased by 10-23 and 11-34 percent over the years compared to pretreatment. In type IV the content of total triglycerides and that of very low density lipoprotein decreased by 28-39 and 38-52 percent, respectively, over the years. High density lipoprotein cholesterol increased in both types. The "atherogenic index" was reduced by the drug. The effect remained through the 4.5 years of treatment. Compliance to the drug was good. Changes in safety laboratory parameters were minor and reversible. No subjective side-effects occurred and no cases of gallbladder disease or cancer were noted. It is concluded that bezafibrate is a safe, convenient and effective serum lipid-lowering drug suitable for the use in primary and secondary prevention of atherosclerotic disease.

Fatty acid composition of adipose tissue in patients with coronary heart disease.

In order to study the relationship between the fatty acid composition of adipose tissue and coronary heart disease (CHD), 34 consecutive male patients with acute myocardial infarction and 33 hospitalized men free of CHD were compared. Patients with diabetes mellitus, endocrine disorders, liver and kidney diseases, recent changes in body weight and deviations from the "normal", customary diet were exlcuded. A statistically significant difference between the two groups was observed only in stearic acid, its proportion being lower in CHD patients (3.25% vs. 4.13%). Using multivariate discriminant analysis, age discriminated best between the groups, followed by stearic acid. The signs observed were positive for the former and negative for the latter. All other acids, relative body weight, and skinfold measurements did not significantly contribute to the discrimination. Age did not correlate with the proportion of stearic acid. Blood lipids from samples taken within 24 h of admission did not significantly differ between the groups. Three months later they had risen considerably in the infarct patients. The metabolic basis of the relationship between CHD and stearic acid is not clear at present. Additional studies are necessary to substantiate the importance of this acid as an indicator of CHD.

Effect of BM 15.075 on lipoprotein concentrations in different types of hyperlipoproteinaemia.

The four-week lipoprotein lowering effect of 0.2 g t.i.d. of BM 15.075 and of 0.5 g t.i.d. of clofibrate was studied in 29 subjects with different types of hyperlipoproteinaemia in a single blind crossover fashion. BM 15.075 decreased very low density lipoproteins (VLDL), triglycerides (TG) and cholesterol concentration in all types of hyperlipoproteinaemia the effect being dependent on initial lipoprotein concentrations. BM 15.075 decreased VLDL triglyceride concentrations on average 20% more than did clofibrate. BM 15.075 decreased low density lipoproteins (LDL) cholesterol concentrations in Type IIA and IIB but did not significantly affect this lipoprotein lipid in type IV hyperlipoproteinaemia. Regression analysis showed that the drug tended to decrease LDL cholesterol if initial concentrations were above 157 mg/100 ml and to increase initially lower levels. No significant differences between BM 15.075 and clofibrate was found in the effect of LDL cholesterol. High density lipoproteins (HDL) cholesterol concentrations were not influenced by BM 15.075. No subjective side effects were noted on BM 15.075. S-ASAT increased and alcaline phosphatases decreased on both treatments.

The effect of bezafibrate on the fibrinolytic enzyme system and the drug interaction with racemic phenprocoumon.

The influence of a new hypolipidaemic agent, bezafibrate, on anticoagulant requirements and fibrinolysis was studied in 15 patients with hyperlipidaemia on long-term treatment with racemic phenprocoumon. Our results suggest a dose-dependent augmentation of the anticoagulant response to the coumarin drug. Treatment with bezafibrate at 450 and 600 mg daily required a reduction of the phenprocoumon dose by 18.5 and 33.5%, respectively. Correspondingly, the serum level of phenprocoumon decreased by 11.6 and 35.3%. No evidence for an altered drug elimination of racemic phenprocoumon could be found during treatment with bezafibrate. The results support the hypothesis that bezafibrate and analogous hypolipidaemic drugs enhance the response to oral anticoagulant drugs by increasing the affinity of the receptor site for coumarins or the rate of degradation of the vitamin-K-dependent clotting factors. The investigation of the fibrinolytic enzyme system demonstrated an increase of the fibrinolytic activity by enhancing the activity of the plasminogen activator. The lysis time for euglobulin clot was reduced significantly, plasma fibrinogen only moderately. The antiplasmin activity could not be altered substantially by a decrease of alpha1-antitrypsin and a slight increase of alpha2-macroglobulin. In contrast with the inhibition of platelet function the effect of bezafibrate on the fibrinolytic enzyme system showed no dose dependence.

Reduced LDL- and increased HDL-apoproteins in patients with hypercholesterolaemia under treatment with bezafibrate.

The effect of bezafibrate on serum lipids, lipoproteins and the apoproteins A-I, A-II and B was studied in 18 patients with primary hypercholesterolaemia. Total cholesterol was lowered by 20% (P less than 0.05), LDL-cholesterol by 24% (P less than 0.05), and apo B by 14% (P less than 0.05), which is comparable to the effect obtained with anion exchange resins but with far fewer side-effects. HDL increased significantly during bezafibrate treatment both by measurement of HDL-cholesterol (+54%, P less than 0.05) and by the determination of HDL-apoproteins A-I (+ 19%, P less than 0.05) and A-II (+ 23%, P less than 0.05). This increase of HDL and the decrease of triglycerides was maintained for 6 weeks of placebo treatment after cessation of bezafibrate, while serum total and LDL cholesterol as well as apo B returned to their baseline levels.

[Effect of methylpyrazole-carboxylic acid on carbohydrate and lipid metabolisms in patients with diabetes mellitus]. / Die Wirkung der Methylpyrazolcarbonsäure auf den Kohlenhydrat- und Fettstoffwechsel von Patienten mit Diabetes mellitus

The effect of 3-methylpyrazole-5-carboxylic acid (MPC) on carbohydrate and lipid metabolisms was studied in 18 patients with diabetes mellitus. In addition to diet 17 patients had basic treatment with sulfonylureas with or without biguanides, one patient was treated with insulin. In all patients carbohydrate metabolism was not well controlled, 14 patients had elevated triglycerides. Following a control period of 2 weeks the patients received increasing doses of MPC in addition to basic treatment (25825825 mg; 50,25,25 mg; 50, 50, 25 mg). Blood samples were taken in the fasting state before the first dose of MPC. Free fatty acids almost doubled under the influence of MPC. This was due to a rebound effect at night following suppression of lipolysis during the day. Blood glucose levels showed a tendency to fall, urinary glucose excretion, separately examined for day and night ,did not change consistently. Triglycerides fell markedly by 25%, but this reduction was not statistically significant. Cholesterol decreased by 5%. 40% of the patients showed an increase in urinary ketone bodies. Body weight did not change. Side effects due to MPC included flushing, gastrointestinal distress and cardiovascular complaints and were observed in 75% of the patients. Due to the high frequency of side effects it does not seem to be worthwhile to further investigate the therapeutic effect of MPC in a larger number of patients with different dosage regimens.

[Dose-effects studies with bezafibrate in patients with hypercholesterolemia and hypertriglyceridemia (author's transl)]. / Dosis- Wirkungsuntersuchung mit Bezafibrat an Patienten mit Hypercholesterinämie und Hypertriglyzeridämie.

The dose-effect relationship of bezafibrate, a new clofibrate derivative, was investigated in 24 outpatients with a primary hyperlipidemia and a Type lla lipoprotein pattern. The patients were divided into two groups at random and in each case received 3 X 50, 3 X 100, 3 X 150 and 3 X 200 mg bezafibrate daily in increasing or falling dosage for four weeks. For rising doses a statistically significant dose-effect relationship could be demonstrated for cholesterol and triglycerides. This was only true for cholesterol with falling doses while triglycerides remained at a low level. The optimal dose was found to be 600 mg bezafibrate daily. The substance was well tolerated, and the usual laboratory parameters remained within the normal range in all patients.

[Lipid-lowering effect of bezafibrate in patients with diabetes mellitus and hyperlipidaemia (author's transl)]. / Der lipidsenkende Effekt von Bezafibrat bei Patienten mit Diabetes mellitus und Hyperlipidämie.

600 mg of bezafibrate daily were administered to 13 well controlled diabetics with hyperlipidaemia for 12 weeks. Placebo was given before and after the treatment period. Compared with pretreatment placebo, bezafibrate reduced triglycerides (between 37 and 47%) and cholesterol (between 12 and 19%) significantly. Blood glucose levels during treatment were significantly lower at 8 and 12 weeks compared with post-treatment placebo values. Urinary glucose excretion did not change. Hypoglycaemia was not observed. No change in antidiabetic medication was necessary. Bezafibrate was well tolerated. It lowered blood lipids effectively in diabetics with hyperlipidaemia. No additional precautions have to be taken to control carbohydrate metabolism during bezafibrate treatment.

Bezafibrate: lack of effect on creatinine excretion and muscular proteins.

The effect of bezafibrate (200 mg t.i.d.) on renal and, to some degree, on muscular function was studied in nine patients with primary hyperlipoproteinemia. No signs of a reduction in creatinine excretion or an increase in muscular proteins could be found serving as a possible explanation for the slight increase in serum creatinine observed in a bezafibrate long-term study in more than 1,000 patients. Further studies are necessary for clarification.

Diurnal lipid and lipoprotein profiles in hypertriglyceridemic patients with bezafibrate and clofibrate.

Thirty hypertriglyceridemic patients were treated under metabolic ward conditions for 10 days with either placebo, clofibrate (500 mg t.i.d.) or bezafibrate (200 mg t.i.d.) in a randomized, double-blind study. In addition, patients received an isocaloric prudent diet. On day 10 a diurnal lipid and lipoprotein profile was carried out. Compliance to medication was good. Each treatment led to significant reductions of fasting triglycerides and cholesterol. Lowering of fasting and integrated diurnal triglycerides was greatest with bezafibrate. HDL-cholesterol profiles were highest with this drug. A strong correlation between fasting and diurnal triglycerides was observed. Triglyceride-lowering therapy must therefore aim at fasting triglyceride values as low as possible.