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The lipid contents of the plasma and liver and enzyme activities in the aorta were determined in rats given the following diets: Group I, normal diet (ND); Group II, high cholesterol diet (HCD, ND plus 1% cholesterol and 0.5% cholic acid); Group III, HCD plus 0.5% gamma-oryzanol; Group IV, HCD plus 2% gamma-oryzanol in the last days of the 4th, 8th and 12th week of the feeding period. Total cholesterol, low-density lipoprotein-cholesterol and very low-density lipoprotein-cholesterol in the plasma were significantly higher in rats in groups III and IV than in those in group II. High-density lipoprotein-cholesterol was increased in groups III and IV. There were no differences in the triglyceride or phospholipids concentrations in the plasma of rats in the four groups. The liver weight was higher in the HCD groups (groups II, III and IV) than in the ND group (group I). The cholesterol ester and triglyceride contents of the liver were higher in the HCD groups than in the ND group, and the increase was the least in group IV of the HCD groups. Changes in acid cholesterol esterase activity and acyl CoA: cholesterol acyltransferase activity in rat aorta homogenates to compensate for lipid deposition were greater in groups III and IV than in group II.

Captopril (37.5 mg daily) was administered to 64 hypertensive patients for 16 weeks. During treatment, systolic and diastolic blood pressures decreased significantly (from means of 164/98 mmHg before treatment to 150/90 mmHg at four weeks and 142/86 mmHg at eight weeks; P less than 0.001), but serum levels of total cholesterol, triglycerides, lipoprotein cholesterol, lipoprotein triglyceride, and apolipoproteins showed no significant changes. Scores on the atherogenic index did not change. Patients with high initial total cholesterol levels and low high-density lipoprotein cholesterol levels tended to improve their lipid levels. It is concluded that captopril does not adversely affect serum lipoprotein metabolism.

The characteristics and properties of lipid metabolism in the aorta and the brain microvessels of rabbits were investigated to clarify the role of lipid metabolism in formation of atherosclerosis. In rabbit aorta, cholesterol esterase and lipase each had an acidic and a neutral optimum pH, whereas acyl-CoA synthetase, acyl-CoA: cholesterol acyltransferase, triglyceride synthesizing activity and choline phosphotransferase each had one neutral optimum pH. These pH optima were similar to those in rats. High cholesterol diet induced atheromatous lesions in the aorta, but not in the brain microvessels. In atheromatous aorta, the acid lipase and acid phosphatase activities were higher than in controls, but not the acid cholesterol esterase activity. Moreover the activities of neutral lipase, acyl-CoA synthetase, acyl-CoA:cholesterol acyltransferase, triglyceride synthesis and choline phosphotransferase were increased, but neutral cholesterol esterase activity was normal. These data suggest that lipis metabolism in the atheromatous aorta is changed in a manner favoring accumulation of lipids, especially cholesterol esters. In controls, most of the above enzyme activities in the brain microvessels were higher than those in the aorta. However, these enzyme activities in the brain microvessels were not changed by cholesterol feeding. Thus it is suggested that the properties of lipid metabolism in the aorta and brain microvessels, including permeability of lipoproteins into the vessel walls, are important in formation of atherosclerosis in addition to the serum factors.

Lipid metabolism in the arterial wall of spontaneously hypertensive rats (SHR) fed on tocopherol-deficient diet, high-cholesterol diet or both was studied. Serum tocopherol was greatly decreased in tocopherol-deficient SHR. Lipoperoxide, determined as thiobarbituric acid-reactive substances, was higher in tocopherol-deficient SHR than in normal SHR. Tocopherol-deficient SHR showed a decrease in acid cholesterol esterase activity, but no change in neutral cholesterol esterase, acid and neutral lipase, acyl CoA synthetase, cytidine-diphosphate choline-1-2-diacyl glycerol choline phosphotransferase (CPT) or triglyceride synthesizing activity.

Pirozadil (pyridine 2,6-dimethanol-bis(3,4,5-trimethoxybenzoate), Pemix) was administered for 16 weeks at a dose of 1.5 or 2.0 g/d to 15 adults with Type IIa hyperlipidemia. All patients had achieved a stable weight and were given a diet before treatment. Plasma lipids, lipoproteins and apolipoproteins were measured at 4-week intervals. Pirozadil reduced the mean total cholesterol (TC) level from 291.0 mg/dl to 260.8 mg/dl in 16 weeks (p less than 0.05). The mean percentage decrease was 8.5% in week 16. Low density lipoprotein (LDL)-cholesterol and apolipoprotein B (apoB) decreased 9.7% and 8.5%, respectively, in week 16. A decrease of more than 10% in TC, representing a decrease of more than 30 mg/dl, was observed in 12 of the 15 patients. These 12 patients, who were regarded as responders, showed mean decreases in the levels of TC, LDL-cholesterol and apoB of 13.7%, 10.4% and 11.7%, respectively, but no significant change in the level of high density lipoprotein (HDL)-cholesterol or serum triglycerides. Eight of 11 patients in the lower dose group (1.5 g/d) and all 4 patients who had obtained the higher dose (2.0 g/d) were responders. Suspected side effects of pirozadil were edema, numbness of the extremities and palpitation. The results obtained suggest the beneficial effect of pirozadil on serum lipids in patients with Type IIa hyperlipoproteinemia.

The dithioester analog of dihexanoylphosphatidylcholine, rac-1,2-S,S-dihexanoyl-3-phosphocholine-1,2-dimercapto-3-propanol was synthesized and compared to the corresponding acyl-oxyester lipid as a substrate for bovine milk lipoprotein lipase. The apparent maximal reaction velocity (Vmax) for dihexanoyldithiophosphatidylcholine was considerably lower than that for dihexanoylphosphatidylcholine (0.12 vs. 5.0 mumol product released/min per mg lipoprotein lipase, respectively). The apparent Km values were 1.9 and 4.0 mM, respectively. 3-Butyrylthio-1,2-dibutyryloxypropane was also compared to tributyrylglycerol as a substrate for lipoprotein lipase; hydrolysis of the acyl-thioester bond was insignificant when compared to the corresponding oxyester derivative. Apolipoprotein C-II, the activator protein of lipoprotein lipase for long-chain fatty acyl substrates, had no effect on the hydrolysis of either the thio- or oxyester short-chain substrates. The low lipoprotein lipase activity for the thioester substrates is discussed in relation to the structure of the lipid and the active site of the enzyme.

Several enzyme activities involved in lipid hydrolysis (acid and neutral cholesterol esterase and lipase) or synthesis (acyl-CoA synthetase, acyl-CoA: cholesterol acyltransferase and cholinephosphotransferase) were assayed in the aortas of rats with nephrosis induced by daunomycin and of rats fed on high cholesterol diet. In nephrotic rats activities of some enzymes involved in lipid hydrolysis, but not in synthesis, were increased. On the contrary, in rats fed on high cholesterol diet, the activities of all enzymes involved in lipid synthesis were significantly increased, with some increase in those involved in lipid hydrolysis. In nephrotic rats fed on high cholesterol diet all enzyme activities were markedly increased. From the view point of accumulation of cholesterol ester (CE), the ratio of hydrolysis of CE in lysosomes to CE incorporated from the blood and the ratio of hydrolysis of CE to reesterification of free cholesterol in microsomes were considered to be important. From this point of view, nephrotic hyperlipidemia was not so atherogenic as hyperlipidemia induced by the diet. The role of serum high density lipoproteins in lipid metabolism in the aorta was discussed.

Acid and neutral cholesterol esterase activities in rat arterial wall were released from the lysosomal fraction and microsomal fraction respectively into the 105,000 X g supernatant fraction by treatment with Triton X-100, heparin and dextran sulfate. The percentage releases of acid cholesterol esterase by Triton X-100 (0.1%), heparin (50 micrograms/ml) and dextran sulfate (1 mg/ml) were 21%, 18% and 4%, respectively, while those of neutral cholesterol esterase were 66%, 56% and 39%, respectively. The cholesterol esterase released by dextran sulfate, especially that from the microsomal fraction, hydrolyzed cholesterol ester in artificial lipid mixtures with similar lipid compositions to those of the deposits in fatty streaks and fibrous plaques of atheromatous lesions.

Four local irritation studies of cefodizime sodium (THR-221), a new developed cephem-type antibiotics, were carried out with NZW rabbits. 1. In eye irritation test, 25% THR-221 water solution had no irritancy on eye mucosa in rabbits. 2. In single injective intramuscular irritation study, regardless of solvents (water or 0.5% lidocaine), 25% THR-221 solution had irritancy equal to 0.75% acetic acid. But recovery process of the muscle injured with THR-221 was faster and better than with 0.75% acetic acid. 3. In five days injective muscular irritation study, the irritancy of 25% THR-221 water solution on the muscle was milder than that of CTT or CET. Histopathological damage with THR-221, necrosis/degeneration of muscle fibers and edema/hemorrhage in interstitium etc., were well recovered. 4. In vessel irritation study, 10% or more THR-221 water solution had irritancy on ear vessel. THR-221, as same as CTT, caused organized thrombi and inflammation at the surrounding area. The degree of irritation of 20% THR-221 solution was slightly stronger than that of 20% CET, but weaker than that of 20% CTT. 5. In a clinical phase, it is to be desired that THR-221 like as CTT or CET shall be avoided repeated intramuscular or intravenous injections at the same site.

Peri- and postnatal study of cefodizime sodium (THR-221), a new developed cephem-type antibiotics, was carried out with ICR mice. THR-221 at dose levels of 0, 300, 1000 and 3000 mg/kg/day were administered intravenously to pregnant and delivered dams from day 15 of gestation through day 21 of lactation. No changes in body weights of dams in all treated groups but slight decrease in food consumptions of 3000 mg/kg/day group were observed. Treated sites, tails of a few dams in this group, were affected with inflammatory lesions because of repeated dosing. Neonates from dams treated with 3000 mg/kg/day were slightly decreased in body weight at birth. At term sacrifice of F 1 of 10 weeks age, absolute and relative spleen weights were decreased in male 3000 mg/kg/day group and in female 1000 and 3000 mg/kg/day group. No effects on other physical, behavioral or reproductive ability examinations of F 1 offspring were showed. It is suggested that no effect dose level of THR-221 is 1000 mg/kg/day and that of F 1 offspring is 300 mg/kg/day.

Cholesterol ester is present in lipid deposits in atherosclerotic lesions, such as fatty streaks, fibrous plaques and complicated lesions. The possibility of hydrolysis of cholesterol ester in lipid deposits and its mechanism were examined by studying the effects of the various components of lipid deposits on cholesterol ester hydrolysis. Studies were carried out using artificial lipid samples prepared by sonication of mixtures of the components of lipid deposits. Results suggested that phospholipids, especially phosphatidylcholine, play an important role in the hydrolysis and that alteration of lipid components, other than cholesterol ester, influences cholesterol ester hydrolysis in lipid deposits.

Apolipoprotein C-II, the activator protein of lipoprotein lipase, contains 78 amino acids with a single residue of arginine at position 49. Chemical modification of apolipoprotein C-II with 1,2-cyclohexanedione or 2,3-butanedione results in a loss of both the arginine residue and the ability of the protein to enhance the activity of bovine milk lipoprotein lipase toward a trioleoylglycerol substrate; removal of the modifying group restores arginine and more than 70% of the activating property of the apolipoprotein. Arginine modification of apolipoprotein C-II does not effect its lipid-binding properties as assessed by its association to sonicated vesicles of dimyristoylphosphatidylcholine. Furthermore, secondary structure associated with complex formation with dimyristoylphosphatidylcholine are nearly identical for the unmodified, 1,2-cyclohexanedione-modified or modified-reversed proteins. These results suggest that arginine-49 of apolipoprotein C-II is situated at or near an amino acid sequence domain involved in the activation of lipoprotein lipase. However, a guanidinium group is not required for lipid binding.

Highly purified cis-5,8,11,14,17-20:5 (n-3) or cis-4,7,10,13,16,19-22:6 (n-3) was administered to rats for 2 weeks, and serum lipoprotein lipid levels, platelet aggregation, and lipid metabolism in the aorta in vivo were investigated. 20:5 (n-3) decreased the level of low density lipoprotein phospholipid. 22:6 (n-3) increased the level of high density lipoprotein cholesterol and decreased those of low density lipoprotein triglyceride and phospholipid. Both 20:5 (n-3) and 22:6 (n-3) markedly inhibited platelet aggregation. 20:5 (n-3) decreased the acid and neutral cholesterol esterase activities, but did not affect either acyl-CoA synthetase or acyl-CoA: cholesterol acyltransferase activity. 22:6 (n-3) had no effect on any of these enzyme activities. From these results, the roles of 20:5 (n-3) and 22:6 (n-3) in the formation of atheromatous lesions and the mechanism of the decrease in cholesterol esterase activity by 20:5 (n-3) were discussed.

The effect of apolipoproteins on the hepatic lipase-catalyzed hydrolysis of high density lipoprotein (HDL) triacylglycerols was studied in an in vitro system consisting of purified human post-heparin hepatic lipase, HDL2 and albumin. The apparent values of the Michaelis constant (Km) and maximal velocity (Vmax) for the hepatic lipase-catalyzed hydrolysis of HDL2-triacylglycerols were 0.18 mM and 86 nmol free fatty acids released/mg hepatic lipase per min, respectively. The addition of purified human plasma apolipoprotein A-I, A-II, E, C-I or C-III2 (containing 2 mol of sialic acid) to HDL2 caused inhibition of hepatic lipase activity. At a 1:1 weight ratio of added apolipoprotein to HDL2-protein, inhibition was 50% for apolipoprotein E and over 75% for the other apolipoproteins tested. Inhibition of enzyme activity occurred with both the unfractionated HDL2 and the HDL which were reisolated by ultracentrifugation. The major alteration in the composition of the reisolated HDL was an increase in the protein to phospholipid ratio. Based on these results, we speculate on the possible role of the apolipoproteins in the metabolism of HDL2 by hepatic lipase.

A patient with an extremely high level of high density lipoprotein (HDL)-cholesterol and HDLc-like particles in the serum is discussed. The patient was a 46-year-old female with a serum total cholesterol concentration of 382 mg/dl and HDL-cholesterol level of 214 mg/dl. The HDL-cholesterol levels of her mother, brother, sister and 2 of her daughters were 82 mg/dl, 82 mg/dl, 74 mg/dl, 82 mg/dl and 82 mg/dl, respectively (mean HDL-cholesterol levels of control subjects: 52 +/- 6 mg/dl in males and 55 +/- 8 mg/dl in females). Her serum apolipoprotein A-I and E levels were elevated. Zonal ultracentrifugal analysis of her serum lipoproteins showed that the increased level of HDL-cholesterol was mainly due to HDL2; HDLc-like particles were also recognized between the LDL and HDL fractions. The incorporation of the patient's HDL and HDLc-like particles into cultured HepG2 cells was almost the same as that of HDL (1.063 less than d less than 1.21) from normal control serum. The incorporation of normal control HDL into the patient's peripheral blood lymphocytes was markedly less than that into lymphocytes from normal controls. These findings are discussed in terms of the reason for hyperalphalipoproteinemia in this patient.

Arteriosclerotic lesions were formed in rat aorta by the administration of vitamin D2, a high-fat diet and a thyroid suppressing agent. This treatment increased the serum total cholesterol level to 12 times the control level. In the arteriosclerotic lesions that were induced the activities of lysosomal enzymes, such as acid phosphatase and acid lipase, were higher than in controls, that of acid cholesterol esterase was decreased, those of microsomal lipid-synthesizing enzymes--such as acyl-CoA synthetase and cholesterol ester synthesizing activity--were increased and that of neutral cholesterol esterase was decreased. These data suggest that lipid metabolism in arteriosclerotic lesions was changed, resulting in the accumulation of cholesterol esters in the aorta. Administration of high-fat diet and thyroid suppressing agent also increased the serum cholesterol levels to 12-fold the control level, but did not induce arteriosclerotic lesions. After this treatment the activities of hydrolyzing enzymes, such as acid and neutral cholesterol esterase and lipase, in the aorta increased, but the activities of lipid synthesizing enzymes also increased. These data suggest that lipid metabolism in the aorta in this condition changed to compensate for the large influx of serum lipids and to prevent arteriosclerosis. The roles of the serum lipid level, cell injury and lipid metabolism in the aorta in forming arteriosclerotic lesions are discussed on the basis of these results.

Teratogenicity of 1,1,3-trimethyl-5-phenylbiuret (ST-281), a new anti-rheumatic agent, was evaluated in rabbits. ST-281 at doses of 0, 50, 100, 200 and 400 mg/kg/day were administered orally to pregnant NZW rabbits from day 6 to day 18 of pregnancy. Body weight and food consumption at the administration and the subsequent periods were significantly decreased in 400 mg/kg/day group, and 5 dams (41.7%) affected severely were dead. No remarkable changes were investigated in findings at near-term caesarean section in any dosed group including 400 mg/kg/day. In visceral and skeletal examinations, no significant increase in incidence of abnormal fetuses were observed. This report suggests that ST-281 has no embryotoxicity or teratogenicity in rabbits.

The effects of hypertension on various enzyme activities involved in lipid metabolism and beta-oxidation in rat brain microvessels and aorta were studied. The purity of the brain microvessel preparation was confirmed immunologically and microscopically. Activities involved in lipid synthesis, such as triacylglycerol synthesizing activity, acyl-CoA synthesizing activity, acyl-CoA: cholesterol acyltransferase and cytidine diphosphate choline:1,2-diacylglycerol cholinephosphotransferase, were significantly higher in brain microvessels than in aorta in both normotensive and hypertensive rats; lipid hydrolyzing activities, such as lipases and cholesterol esterases, were similar in the two preparations. beta-oxidation in brain microvessels was more active than in aorta in both groups. Hypertension did not alter these enzyme activities in either aorta or brain microvessels, or change beta-oxidation in the aorta. However beta-oxidation in brain microvessels was significantly lower in hypertensive rats than in normotensive rats. These results suggest that brain microvessels are metabolically more active than aorta, and that their beta-oxidation activity is more susceptible to effects of hypertension. Reduced beta-oxidation in brain microvessels might lead to angionecrosis by derangement of energy production, which in turn may cause cerebral bleeding.