The effect of substrate condition on cholinephosphotransferase (CPT) was investigated in rat aorta microsome in terms of kinetics using various species of diacylglycerols (DG) prepared differently as substrate.
In use of phosphatidylcholine (PC)-emulsified diolein (18:1) as substrate, CPT activity was higher with unsaturated PC's such as 1-linoleoyl-2-linolenoyl PC (LLPC), dioleoyl PC (DOPC), and 1-palmitoyl-2-arachidonyl PC (PAPC) than with saturated PC's such as dipalmitoyl PC (DPPC), distearoyl PC (DSPC), and dimyristoyl PC (DMPC). This higher CPT activity with unsaturated PC-emulsified diolein was due to higher affinity of the substrate to the enzyme according to the kinetic study.
PC liposomes themselves act on the enzyme differently: Saturated PC liposomes inhibited the CPT activity by reducing the affinity of diolein to the enzyme, while the saturated PC's did not inhibited the activity when used for emulsifying diolein. With respect to DG species, when DG's were emulsified with Tween 20, diolein showed much higher CPT activity as compared with distearin (18:0) and dipalmitin (16:0), while diolein and dipalmitin gave the comparable CPT activity when emulsified with DOPC. This effect of DOPC seems to be brought by increasing V_max, that is, transfer activity between the two substrates, CDP-choline and dipalmitin: K_m was increase with DOPC. It implies that phospholipid might regulate the interaction between CDP-choline and DG during the product synthesis.
In this paper it was shown that PC synthesis was greatly affected by the condition of DG.

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The purpose of this study was to investigate the significance of platelet volume in cerebrovascular disease. We already reported the platelet production was increased and its survival time was shortened in stroke prone SHR (SHRSP) with aging after 10 weeks of age. The platelet volume of SHRSP was larger than that of stroke resistant SHR (SHRSR). On the other hand the platelet volume in patients of apoplexy was larger than in patients of control and essential hypertension. Hence it was suspected platelet consumption was increased at the damaged endothelial cells by adherence and aggregation of platelets in patients of apoplexy. The increased platelet volume could be suppressed by treating with plateletinhibiting drugs. Therefore it was suggested that the plateletinhibiting drugs were effective in the patients of apoplexy to prevent the second stroke.

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1. Culture medium conditioned by endothelial cells contained anticoagulant activity. The concentrated serum-free conditioned medium, not only directly inhibited Factor X_a activity but also enhanced the inactivation of X_a activity by antithrombin III. Acidic glycosaminoglycans (GAGs) isolated from conditioned culture medium showed X_a inhibition only in the presence of antithrombin III.
2. On the cultured endothelial cell monolayer, thrombin activity was inhibited by antithrombin III. This inhibitory effect was absent on the endothelial cells pre-treated with F. heparinum enzyme.
These results suggest that GAGs (probably heparan sulfate) on the endothelial cells exert heparin-like activity both in the liquid phase after secretion and in the solid phase on the cell surface.

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With the endothelium directly and the smooth muscle cell indirectly exposed to intravascular pressure and flow, blood pressure is an important measurement in the investigation of vascular disease. The culture of endothelial cells and smooth muscle cells was studied in constant pressure chamber. The following results were obtained: The endothelial cell growth was maximal at 80mmHg, minimal at 0mmHg (atmosphere pressure) on the second day of incubation. Cell degeneration began to occur at 120mmHg. Marked degeneration was noted at 160mmHg. On the contrary, production of prostacyclin was maximal at 0mmHg and minimal at 80mmHg. Smooth muscle cell growth was not affected by the pressure chamber environment.

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Effects of cholesterol feeding on plasma lipoproteins and on the complexes of lipoproteins with connective tissue macromolecules in the aorta were studied in rabbits fed a diet containing 0.5 cholesterol for six weeks. Plasma lipoprotein distribution was monitored by rate-zonal ultracentrifugation and electrophoresis.
Changes in plasma lipids and lipoproteins in cholesterol-fed rabbits were as follows:
1) Plasma levels of cholesterol, phospholipids and free fatty acids were increased significantly, whereas triglyceride level remained unchanged.
2) Plasma very low density lipoproteins and materials emerged in low density lipoprotein density range increased significantly, and they were cholesterol-rich and triglyceride-poor.
3) High density lipoproteins decreased below level of detection.
Changes in aortic composition and distribution of accumulated cholesterol among connective tissue components in the aortas from cholesterolfed rabbits were as follows:
1) Cholesterol was increased significantly in atherosclerotic aortas, while triglyceride and phospholipid levels remained unchanged.
2) Concentrations of glycosaminoglycans in the atherosclerotic aortas were significantly increased, whereas collagen was slightly decreased. No change in elastin content in the aortas were observed.
3) Contents of cholesterol bound to glycosaminoglycans and to elastin were significantly increased 1.5- to 2-fold and 3- to 4-fold, respectively, whereas that to collagen remained unchanged.

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A study was made of the relationship between aortic pulse wave velocity (PWV) in terms of the degree of arteriosclerosis as observed non-invasively from vascular properties and hyperlipidemia a risk factor in arteriosclerosis. Subjects were a total of 317 cases (male 158 and female 159) with hyperlipidemia, hypertension, cerebral vascular disease, ischemic heart disease, diabetes melitus and others with an average age of 64.1 years. Serum lipids determined were 8 kinds of total cholesterol (TC), β-lipoprotein (β-LP), triglyceride (TG), free fatty acid (FFA), cholesterol ester (CHO-E), lipid peroxide (LPO), HDL-cholesterol (HDL-C) and phospho lipid (PL). PWV was 7.5, 8.4, 9.1, 10.2 and 11.5m/sec for the age level of 40, 50, 60, 70 and 80 years, respectively, with increasing means and deviations with ageing. TC, β-LP, TG, CHO-E and PL showed tendencies of high contents for the age-levels of 50, 60 and 70 years and low for those of 40 and 80 years, while HDL-C, FFA and LPO showed no constant tendencies for each of these age-levels. Correlation by age-level “r” between PWV values and serum lipids was 0.18 for CHO-E at the age level of 60 years, -0.21 for HDL-C at that of 50 years, 0.21 for TC at that of 60 years, 0.24 for PL at that of 60 years, 0.25 for LPO at that of 60 years, 0.25 for FFA at that of 60 years, 0.25 for β-LP at that of 50 years, 0.41 for FFA at that of 50 years and 0.47 for TG at that of 50 years, thus being observed between age-level of 50 years and that of 60 years. No correlation was observed at the age-levels of 40, 70 and 80 years between PWV values and serum lipid values. This seemed to be due to organic arteriosclerotic lesions caused at the age-levels of 50 and 60 years by hyperlipidemia as a result of lipid dysmetabolism observed at the age-level of 40 years.

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With the purpose of establishing the degree of cerebral atherosclerosis non-invasively, we have developed an ultrasonic quantitative blood flow measurement system (QFM). The QFM system is designed to measure the absolute blood flow volume and wave form of the common carotid artery. Parameter Rp (the degree of cerebral circulation resistance) was measured non-invasively on the carotid arteries by the QFM system and simulation technique. “Atherosclerosis” is a pathological concept. It is important to justify the correlation between clinical parameter Rp and the degree of atherosclerosis in histological sections obtained by autopsy. Degree of Atherosclerosis in Autopsy Subjects: The circle of Willis was taken for study at autopsy of patients who had been measured by the QFM system while alive. The grade (0, +1, +2, +3, +4) of stenosis, based on Baker's method, was determined according to the ratio of the area enclosed by the intima to the area circumscribed by the internal elastic lamina in each histological section. The degree of atherosclerosis was determined as the average grade of 5 segments (2 from anterior, 2 from middle cerebral, and 1 from internal carotid arteries). Results: Materials consisted of 37 vessels obtained by autopsy of 19 subjects. Rp correlated well with the degree revealed by autopsy (r=0.79). Histopathological findings confirmed the QFM system to be of great use in defining the cerebral atherosclerosis noninvasively.

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The fatty acid composition of plasma, plasma phospholipids, erythrocytes and erythrocyte phospholipids were measured in 9 survivors of cerebral infarction (CI) in the distribution of a cortical artery (7 males, 2 females) and in 9 healthy subjects (4 males and 5 females).
The plasma lipids were extracted with chloroform-methanol. The lipids of erythrocytes were extracted with iso-propanol and chloroform. The lipid fractions were separated on silica gel chromatoplates using heptane: ethyl ether: acetic acid (80:20:2) as a solvent. The fatty acid methyl esters were analysed with a gas chromatograph (Shimadzu GC-7A).
The CI patients showed a significantly higher proportion of palmitic acid in their plasma phospholipid fatty acids and a lower proportion of linoleic acid in their erythrocyte fatty acids. Analysis of the composition of erythrocyte phospholipids showed a significantly lower proportion of linoleic acid and a higher proportion of palmitic acid in the survivors.
The relative amounts of the polyunsaturated fatty acids, particularly arachidonic acid and eicosapentaenoic acid showed no significant differences between the two groups.
These findings suggest that the fatty acid pattern might be an important factor in the development of cerebral infarction.

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Serum complements are principal components in immunological reactions. Recently, serum components have been suggested to have influences on atherosclerosis, and aggregations of platelets and leucocytes. The aim of this study was to investigate the correlations between serum complements and atherogenic factors in healthy subjects. Subjects were 230 examinees (172 males and 58 females) at the Center of AMHTS (automated multiphasic health testing and services) in Tokai University Hospital.
The average age was 45.3 years old (male 44.0, female 46.7). Serum C₃C, C₄ and C₃A (B_b) levels were measured by single radial immunodiffusion (SRID) methods. Correlation coefficients were calculated between serum complements levels and following factors as systolic blood pressure, obesity index, erythrocytes sedimentation rate (ESR), WBC count, fasting blood suger (FBS), uric and, total cholesterol (C), triglycerides (TG), HDL-C, LDL-C, and atherogenic index (AI=TC-HDL-C/HDL-C). Plasma HDL-C levels were estimated by the precipitation method with dextran-Mg⁺⁺. Plasma LDL-C levels were obtained by the calculation with Friedewalds' equation.
Serum C₃C levels had significant positive correlations with systolic blood pressure, obesity index, ESR, FBS, TC, TG, LDL-C and AT, and had a significant negative correlation with HDL-C. Serum C₄ levels had significant positive correlations with uric acid, TG and AI. Serum C₃A had significant positive correlations with systolic blood pressure, obesity, index, WBC count, uric acid and TG.
Therefore, serum C₃ levels had the closer relations with atherosclerosis advancing factors than C₃A and C₄.

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Serum VLDL and triglyceride of male SHR were higher than those of male WKR (Wistar Kyoto rat) which is a mother strain of SHR, and HDL and cholesterol were lower in SHR than in WKR. Apoprotein E/C ratios in VLDL and HDL of SHR were low whem compared with those of WKR.
Castration of SHR induced the decrease of VLDL and triglyceride together with the increase of HDL, cholesterol and VLDL apo E protein. The effects of castration on those of WKR were less than those on SHR.
Testosterone administration to the castrated SHR restored these changes of serum lipids and lipoproteins by the castration to the levels of control SHR.
It is concluded that testosterone influences markedly lipoprotein metabolism and SHR is a animal species sensitive to tetsosterone.

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It has been reported that Glycyrrhizin decreases plasma total cholesterol level and reduces the formation of atheroma.
To clarify the mechanisms of the antiatherosclerotic action of Glycyrrhizin, the effects of Glycyrrhizin on lysosomal and microsomal functions which were involved in the lipid metabolism of rat arterial wall, were investigated.
Glycyrrhizin inhibited phospholipase A₂ activity of rat arterial wall.
Glycyrrhizin increased the membrane-bound activity of acid cholesterol esterase in peritoneal macrophages obtained from cholesterol fed rats, when Glycyrrhizin was intraperitoneally administered.
These results, as observed in arterial wall experiments, confirmed that Glycyrrhizin maintains lysosome function by inhibiting phospholipase A₂ activity.
In smooth muscle cells, the incorporation of [¹⁴C] acetate or [¹⁴C] linoleic acid into cholesterol ester was inhibited by adding Glycyrrhizin to the culture medium, i. e. Glycyrrhizin inhibited the esterification of cholesterol in microsome.
From these results, it was suggested that Glycyrrhizin may prevent the accumulation of lipids originated form lysosome and microsome.

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In chronic male schizophrenic patients treated with chlorpromazine, the high density lipoprotein cholesterol (HDL-C) levels were significantly low (p<0.001) compared to normal control men. HDL subfractionation revealed what significantly low (p<0.005) was HDL₃-C, but not HDL₂-C. Both serum apo A-I and apo A-II levels were also low in schizophrenics treated with chlorpromazine, however, no significant differences in apo A-I and apo A-II were found between HDL₂ and HDL₃ fractions. No differences in total cholesterol, triglycerides and HDL-C were found between users and nonusers of benzodiazepines in schizophrenics treated with chlorpromazine.

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We studied the effect of Ca-antagonist nicardipine hydrochloride on experimental atherosclerosis in cholesterol-fed rabbits.
White Japanese male rabbits (n=18) weighing 2-3kg were fed on a 1% cholesterol diet for 2 weeks, and then divided into two groups so as not to make a statistical difference in the serum total cholesterol levels between the two groups. One group (n=9) was continued on a 1% cholesterol diet as a control. The other group (n=9) was fed on a 1% cholesterol diet plus 0.06% nicardipine hydrochloride (60mg/day). At the end of the 14-week period, all animals were killed and autopsied.
There was no difference in serum lipids and body weight between the two groups throughout the study period. Atheromatous aortic lesions covered 13.9±4.4 (mean±S.E.) % of the intimal surface in the control rabbits, and 18.6±2.5% in the nicardipine-treated rabbits.
Although we observed that LaCl₃ suppressed experimental atherosclerosis in rabbits without reducing hypercholesteremia, nicardipine was not effective. An experiment similar to these experiments is now being performed using 0.21% diltiazem.

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Because of its high specificity for cerebral arteries, nicardipine HCl is considered a suitable hypotensive for the elderly patients. We clinically used 60mg/day nicardipine HCl over at least 12 weeks for hypertensive patients, with the following results:
1) Nicardipine HCl lowered total-cholesterol (T-C), triglyceride (TG) and atherogenic index (A. I.), while elevating high density lipoproteincholesterol (HDL-C).
2) The above effects of nicardipine HCl were particularly marked in patients with high blood T-C, low blood HDL-C and high A. I.
3) Blood pressure was significantly lowered in hypertensive patients, but hardly changed in normotensive patients.
The above results suggest that nicradipine HCl is useful not only in the hypotensive treatment but in improving the risk for arteriosclerosis too. It seems likely that nicardipine HCl improves lipometabolism by stimulating the activities of such enzymes as LPL and LCAT and playing a part in the metabolisms of HDL apoenzymes and prostanoids.

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An acid stable phosphoprotein was formed as a reaction intermediate of Ca²⁺-dependent ATPase of bovine aortic microsomes. The Ca²⁺-dependence of phosphoprotein formation was not affected by the addition of diltiazem (10μg/ml), nifedipine (100ng/ml), vanadate (10μM), nicotinic acid (10μg/ml) or nicotinamide (10μg/ml). Nccomol increased the phosphoprotein level at 1μM Ca²⁺. The rate of phosphoprotein formation was not changed, but the rate of its decomposition was decreased by 65% in the presence of 10μg/ml of nicomol. A high activity (3.89unit/mg) of adenylate kinase in the mitochondrial fraction isolated from bovine aortic smooth muscle was observed. The adenylate kinase activity was not affected by nicomol (10μg/ml) and nicotinamide (10μg/ml), but it was stimulated by 158% by 10μg/ml of nicotinic acid.

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We studied the effect of Ca²⁺-antagonists LaCl₃ and diltiazem on aortic smooth muscle cells in culture. Smooth muscle cells in subculture were grown in the Waymouth medium containing 10 fetal bovine serum for 14 days. The smooth muscle cells cultured with 10^-3M LaCl₃ or 10^-4-10^-5M diltiazem showed oval or polygonal endothelial cell-like forms, keeping a paving-stone-like monolayer until the end of the 14-day culture. By fluorescent staining with the FITC-labelled antifactor VIII serum, the factor VIII-antigen, which is regarded as characteristic of endothelial cells in general, was proved. When the medium was changed into the Waymouth medium without Ca²⁺-antagonist, the usual state of smooth muscle cells showing spindle-shaped multi-layer proliferation was recovered. 10^-3M theophylline showed a similar ‘differentiation’ effect on the smooth muscle cells in culture. Embryologically, both smooth muscle cells and endothelial cells are derived from the mesenchymal cells and this phenomenon is of interest in consideration of mutual differences.

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Recently special interest has been focussed on the anti-atherosclerotic effect of Ca-antagonist in various species. However the mechanism of such an effect is obscure.
To study the effect of Ca-antagonist on the release of PGI₂ of rat aortic smooth muscle cells (SMCs) in culture, we planned this experiment.
By the method of Russel Ross, aortic smooth muscle cells were isolated from medial explants of thoracic aortas of male Wistar rats of 5-8 weeks old and cultured in the Eagle's minimum essential medium containing 10% fetal calf serum under 95% air and 5% CO₂. Cells between 5-7 subcultures were used in this experiment.
Ca-antagonists (Diltiazem, Nicardipine, Nifedipine, 10^-9-10^-5M respectively) were added and the effects on the growth curve and the release of PGI₂ into the medium were checked. To determine the PGI₂ content in the medium, we used the New England Nuclear 6-keto PG F_1α³H-kits.
As conclusion,
1) Diltiazem, Nicardipine and Nifedipine (except 10^-7M Diltiazem) stimulated the growth of SMCs in culture at 2 days of culture and Diltiazem significantly inhibited their growth at 9 days of culture.
2) Release of PGI into the medium was significantly increased from the SMCs treated with higher concentration (10^-5M) of each Caantagonist.

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Phospholipids that accumulate in atheromatous lesion are considered to be de novo synthesized. But intracellular metabolism of LDL-phospholipids has not been clarified.
We prepared LDL-DLPC by associating DLPC (1-linoleoyl-2-[¹⁴C] linoleoyl-phosphatidylcholine) with LDL, and studied the metabolism of LDL-DLPC incorporated into smooth muscle cells. Smooth muscle cells (SMC) were prepared from rabbit thoratic aorta according to the method of Wissler et al.
(1) Phospholipase A₁ in SMC showed pH optima at 4.0 and 7.0, and phospholipase A₂ was optimum at pH 4.5, 6.5 and 8.5. It has been reported that LDL-cholesterol ester (LDL-CE) is hydrolyzed by cholesterol esterase only in the acid range, suggesting that LDL-CE is hydrolyzed only in lysosomes. By contrast, LDL-DLPC was hydrolyzed not only in acid range, but also in neutral or alkaline range, suggesting that LDL-DLPC might be hydrolyzed not only in lysosomes, but also in the other subcellular organellas.
(2) After 48hrs of incubation of 3rd passage smooth muscle cells with 10% lipoprotein deficient serum, LDL-DLPC was added to the medium and the metabolism of LDL-DLPC incorporated into smooth muscle cells was studied. The metabolites of LDL-DLPC were found as Phospholipids, triglycerides or cholesterol esters etc. To study the role of lysosomes in LDL-DLPC metabolism, chloroquine was added to the medium to inhibit lysosomal functions. By addition of chloroquine, [¹⁴C] phospholipids in lysosomal fraction were not increased.
The metabolites of LDL-[¹⁴C]CE (prepared by the method of Krieger) incorporated into SMC were studied, and found as cholesterol esters, free fatty acids or triglycerides etc. By addition of chloroquine, [¹⁴C]CE in lysosomal fraction was extremely increased. This suggested that chloroquine inhibited the hydrolysis of LDL-[¹⁴C]CE in lysosomes. In contrast to this, LDL-DLPC was hydrolyzed under the condition when lysosomal function was inhibited. These results suggest that the metabolic pathway of LDL-DLPC might differ from that of LDL-CE.
Two experimental results described above suggest that LDL-phospholipids might be hydrolyzed not only in lysosomes but also in the other subcellular organellas.

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The effect of Probucol on serum lipoprotein and cholesterol metabolism in the WHHL-Rabbit was studied. After the observation period of 2 weeks, 500mg/kg of Probucol in olive oil was given orally for 3 weeks during which the serum was serially taken and analyzed. The results obtained were as follows.
1) The cholesterol levels in serum were markedly lowered (-47%), Phospholipid was mildly lowered but Triglyceride was not changed. The VLDL cholesterol, LDL cholesterol and HDL cholesterol were also reduced.
2) Apo A₁ and Apo B were decreased but Apo E was not changed by the drug.
3) Fecal excretion of ³H labelled cholesterol after the administration of the drug showed no increase, compared with those before the drug administration. Fractional Catabolic Rate of ³H labelled cholesterol did not show any changes after the administration of the drug.
4) In vitro experiments, Probucol inhibited the cholesterol biosynthesis in the liver at the concentration of 50 and 100μM of the drug.
From these results it is postulated that Probucol reduces the serum cholesterol level in WHHL-Rabbit, which has been proposed as an animal model for human familial hypercholesterolemia, through the inhibition of cholesterol biosynthesis in the liver.

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In order to evaluate influences of probucol on serum and lipoprotein concentrations of cholesterol, triglyceride and phospholipid and the composition of apoproteins in very low density lipoprotein, we studied 10 patients with heterozygous familial hypercholesterolemia. Probucol was given twice daily for 8 weeks at the dose of 1g per day. Lipoprotein fractions were separated by serial ultracentrifugation into VLDL, IDL, LDL and HDL. The apoprotein compositions of VLDL fractions were determined with isoelectric focusing.
After 8 weeks treatment with probucol, mean serum cholesterol level decreased significantly by 14%, from 313±15mg/dl to 270±19mg/dl. IDL, LDL and HDL cholesterol levels decreased significantly by 27%, 9% and 29%, respectively. Serum, VLDL and IDL triglyceride concentrations showed no significant changes. HDL triglyceride level decreased significantly from 17mg/dl to 13mg/dl, but LDL triglyceride level increased significantly from 34mg/dl to 43mg/dl. Phospholipid concentration in serum, LDL and HDL fractions decreased significantly.
Cholesterol/triglyceride ratio in LDL decreased significantly from 6.57 to 4.45. However, these ratios in VLDL, IDL and HDL did not change significantly. LDL cholesterol/HDL cholesterol ratio increased from 3.90±0.43 to 4.78±0.58.
With probucol treatment apo E-2/E-3 ratio did not change significantly, but apo C-II/C-III₁ ratio increased from 0.61 to 0.88 and apo C-II/C-III ratio increased significantly from 0.22 to 0.34. These changes may relate to the reported decrement of lipoprotein lipase activity by probucol. Apo E/C ratio decreased significantly from 0.56 to 0.30. This change suggests that probucol retards VLDL catabolism.
In conclusion, probucol not only lowers cholesterol levels in serum, IDL, LDL and HDL fractions, but also changes the apoprotein composition of VLDL fractions.

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The effect of probucol (750-1, 000mg/day) on serum lipids and clinical symptoms was studied in 35 patients with familial hypercholesterolemia (FH) including 5 cases of homozygote. Although the study was designed to investigate the effect during a 24-month period, as most patients have already been medicated for more than 12 months, we report the effect of Probucol in this paper as an interim report.
In 4 of the 5 patients of homozygote, serum total cholesterol, ranging from 477 to 754mg/dl, was markedly decreased over the first 6 months. In these 4 patients, the mean value of the decrement was 128mg/dl and 97mg/dl after two and 6 months, respectively. However, little change was noted in serum lipid levels throughout 12 months in another patient.
In 30 cases of heterozygote, serum total cholesterol (the pretreatment value was 359±55mg/dl, mean±SD) was significantly (p<0.001) decreased and the mean level of all samples measured over a 12-month period was 294mg/dl. The mean per cent decrease throughout this period was 18.1%. The number of patients being medicated more than 15 months were less than 20, but their mean total cholesterol levels showed constantly low values around 300mg/dl. Serum HDL-cholesterol was also decreased significantly, but the decrease was most remarkable during the first 6 months and tended to increase slightly thereafter. As the decrement of LDL-cholesterol was much larger than that of HDL-cholesterol, the difference between the two was significantly diminished during the treatment. Serum triglyceride was slightly but significantly (p<0.05) decreased during the 4-12th month of the treatment.
Tuberous xanthomas were diminished in 17 of the 26 patients, and completely disappeared in some cases. The thickening of Achilles tendon (some were observed by Xeroradiography before and during the treatment) was also improved in most cases.
These observations, though they were incomplete at present, suggest that probucol may be a useful drug for the treatment of FH including homozygotes, which have been considered to be resistant to any existent drugs.

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It has been reported that the simplified fat emulsion tolerance test (FETT) has been developed for the analysis of metabolism of triglyceride rich lipoprotein and pathogenesis in hyperlipoproteinaemia. In the present study, reliability and reproducebility of the removal rate (K₂) in FETT were investigated.
One hundred and one subjects consited of healty subjects and patients with primary and secondary hyperlipoproteinaemia except type I and type V. The subjects aged between 21 and 72 years of both sex were fasted for 14 hours. An indwelling catherter was inserted into the antebrachial vein, and then 0.25cc/kg body weight of 10% Intralipid was injected intravenously in 90 seconds. Timing was started at the midpoint of the injection. The blood was sampled before the injection and at 3, 5, 7, 9, 11, 14, 17, and 20 minutes after the injection. Light scattering index (LSI) of the serum diluted 1:100 with physiological saline was determined by nephelometer (Kyoto Daiichi Kagaku Co., Ltd., DN-2110). The samples were read in duplication, and LSI was calculated by subtraction of serum zero-time value from the estimated mean value after the injection.
For calculation of the removal rate (K₂) of fat emulsion, LSI were plotted against time in a semilogarithmic plot. Slope of the regression line was determined by the method of least squares and equation log Y=bx+a was calculated. The removal rate, K₂, of fat emulsion was expressed as-b±Sb where Sb is standard error of b (Fig.1). Student's t-value calculated as b/Sb ratio was distributed between 5 to 80 in 101 subjects. This high t-value sustained high liniarity of the semilogarithmic disappearance curves of fat emulsion. In other words, high t-value indicated high reliability of K₂ (Fig. 2).
Twenty one subjects were received FETT repeatedly with an interval from one day to 7 days. The correlation coefficient between K₂ in the first and second FETT was 0.85 (p<0.001).
K₂ was highly reproducible (Fig. 3). These findings suggests that simplified FETT could be a good tool for the study of hyperlipoproteinaemia and lipoprotein metabolism.

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We have investigated a double precipitation procedure for the determination of serum HDL₂-and HDL₃-cholesterol by Gidez et al (1982), and its clinical availability was evaluated. The procedure is composed of a initial precipitation of apo B containing lipoprotein with heparin-MnCl₂ and a further precipitation of HDL₂ by adding dextran sulfate (MW 15, 000, Sochibo, Boulogne, France) to the initial supernatant.
1. The concentrations of serum total HDL-cholesterol by the precipitation method with heparin-MnCl₂ (final concentration of MnCl₂: 0.091M) were compared with those of the preparative ultracentrifuge. The correlation coefficient of the values between two methods was r=0.940, and the difference of the means was 1.1mg/dl.
2. With regard to the second precipitation, we have found that a optimal final concentration of dextran sulfate required to precipitate HDL₂ was 0.30g/dl or above, which differed from the concentration of 0.13g/dl originally recommended by Gidez. By the precipitation with dextran sulfate concentration of 0.30g/dl, the values of HDL₂-and HDL₃-cholesterol gave a best correlation compared with those of the ultracentrifuge method, and their correlation coefficients were r=0.933, 0.927 respectively. But the precipitation method showed slightly higher results for HDL₂-cholesterol and slightly lower values for HDL₃-cholesterol.
3. By using this method, we have confirmed that mean HDL₂-cholesterol values of the patients with coronary heart disease (18.1±7.3) and hypertriglycemia (16.1±6.0) were significantly lower than that of control (29.2±13.3mg/dl), and mean HDL₃-cholesterol value of the patients with cirrhosis of the liver (7.8±3.0) was characteristically lower than that of control (23.3±4.9mg/dl).
We have concluded that this precipitation procedure for determination of HDL subclasses was very useful in clinical studies, because not only the results by this method correlated well with those obtained by the preparative ultracentrifuge, but also the procedure was simple and rapid.

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We already reported the variability of serum phospholipid levels both in normal subjects and in patients suffering from vascular diseases such as diabetes mellitus, cerebral infarction, cerebral hemorrhage, and myocardial infarction. However, the method used until now was so complex and time-consuming that we used HPLC for measurement of serum choline-containing phospholipid (PL). Elution pattern of HPLC was detected separately according to their densities, i. e. chylomicron, VLDL, LDL, HDL₂, HDL₃, and albumin-PL. In 52 healthy males, we measured the levels of serum PL, in which total PL 193mg/dl, LDL-PL 109mg/dl, HDL₂-PL 39mg/dl, HDL₄-PL 32mg/dl and albumin-PL 14mg/dl were detected respectively. Single correlation were as follows; TG vs. LDL-PL, Tch vs. LDL-PL, HDL-ch vs. HDL₂-PL, HDL₂-PL vs. HDL₃-PL were positive whereas TG vs. HDL-ch, HDL-ch vs. A. I., LDL-PL vs. HDL₂-PL were negative significantly. From these results, it was supposed that HPLC was easy to handle and to detect choline-containing phospholipid separately and clearly.

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Recent trends in biological science indicate that there is close correlation between biological function and shape of macromolecules. We consider this also as relevant in the study of serum lipoproteins. However, the morphological aspects of lipoprotein metabolism have not been widely studied, because of the technical difficulties and instrumental limitations.
We approached this field by employing scanning electron microscopy (SEM), and explored in this study the methodology for SEM of serum lipoproteins. Using Chylomicron and VLDL as lipoprotein model, we checked the following five points: i) effect of isolation procedures of lipoproteins, ii) suitable fixation, iii) influence of carbon and gold coating, iv) comparison for configurations between transmission electron microscopy (TEM) and SEM, and v) variance due to different serum lipid levels.
Both lipoproteins isolated by ultracentrifugation at 9, 500g for 45 minutes for chylomicron and at 82, 000g for 16 hours for VLDL and those isolated by gel-filtration with Bio-Gel A-150m had no deterious effects, while lipoproteins gained by precipitation with heparin-managenese showed the surface fused with several particles. There was no difference in size and shape of SEM between the single fixation with 1% OsO₄ for 16 hours and the double fixation with 3% glutaraldehyde for 30 minutes plus 1% OsO₄, 16 hours. However, there was a great difference in shape of lipoproteins when we compared TEM of the lipoproteins negatively stained with 3% phosphotungstic acid for 60 seconds and SEM of the lipoproteins fixed with 1% OsO₄ for 16 hours. The former had occasionally collapsed, polygonal particles with the surface shrinked and uneven, while the latter revealed the spheres with the surface plump and smooth.
From these results, we conclude that SEM gives a better undistorted, three dimensional forms of lipoproteins than TEM of negatively stained lipoproteins. The methods for the SEM of serum lipoproteins in the final form are, thus, as follows; chylomicron (d<0.9) and VLDL (d<1.006) were isolated by ultracentrifugation, and washed with physiological saline solution three times, then fixed with 1% OsO₄ for 16 hours. The fixed lipoproteins were washed with distilled water more than 10 times, and mounted on a specimen holder to coat with carbon and gold, then viewed under SEM at 10, 000-30, 000x magnifications.
A preliminary application of this method revealed that VLDL from normolipidemic subjects had an average diameter of 561±197 Å, while those from hypertriglyceridemic patients had a greater diameter ranging from 1995±1355Å to 3040±922Å. SEM of serum lipoprotein will be used as a useful tool for the morphological studies on serum lipoproteins.

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In order to investigate the underlying mechanism to maintain low plasma and LDL cholesterol levels in Pima Indians who have high prevalence of diabetes and obesity, simultaneous studies of VLDL and LDL apolipoprotein B (apo B) turnover study has been performed, which examine the kinetics of VLDL, IDL and LDL and the extent of interconversion of VLDL to LDL.
Using autologus ^131I-VLDL and ¹²⁵I-LDL, specific activities of VLDL, IDL and LDL were followed for 14 days. Kinetic data were analyzed by multicompartmental model.
Ten Pima males who had mean (±SEM) age 23±2yrs, plasma cholesterol 153±9mg/dl and triglycerides 157±22mg/dl were investigated. Production of VLDL apo B was 17.2±1.3mg/kg/day, but production of LDL apo B was only 9.2±0.6mg/kg/day. 8.5±1.0mg/kg/day (48.9±3.9%) of VLDL apo B was removed without conversion to LDL apo B. Thus, in Pima Indians, incomplete conversion of VLDL to LDL may represent a mechanism for the maintenance of low LDL and plasma cholesterol.

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Diurnal changes in plasma apoprotein levels were measured in 6 healthy volunteers. The same meal was given on two consecutive days and 30gr of fat (P/S ratio: 1.2) was administered with each meal on the second day. Blood samples were taken at fasting, at 1 and 3 hours after breakfast, at 2 and 4 hours after lunch and supper. Plasma total cholesterol, triglyceride, HDL-cholesterol and apoprotein A_I A_II, B, C_II, E levels were measured. Plasma apoprotein levels were measured with the method of single radial immunodiffusion (SRID).
Plasma triglyceride levels reached the peak at 2 hours after lunch and declined slightly afterwards. Fat administration increased the postprandial triglyceride levels but had no effects on the curve of the diurnal change. Plasma total cholesterol and HDL-cholesterol levels did not show any diurnal changes and were not affected by oral fat administration. The levels of plasma apoprotein A_I, A_II, B, C_II, E did not show any diurnal changes and were not influenced by oral fat administration. Therefore, the postprandial levels of total cholesterol, HDL-cholesterol and apoproteins can be considered as the same values as the fasting values.

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Apolipoprotein C-II is generally accepted as the major activator of lipoprotein lipase which is considered to be a key enzyme in the hydrolysis of triglyceride from chylomicron and VLDL. Recent biochemical advances have made it possible to determine easily the concentration of serum apolipoprotein C-II in clinical practice. The recently developed analytical method will promote further clinical studies on participation of apolipoproteins in lipoprotein metabolism relating to atherogenesis. This study was intended to clarify clinically the agedependent difference of lipoprotein metabolism in association with apolipoprotein C-II.
A hundred and thirty-nine healthy adults with normal physical findings, blood pressures, urinalysis, blood sedimentation rates, chest films, ECGs and liver function tests were selected for this study in the annual screening for circulatory diseases. Fasting blood specimens were drawn early in the morning. Serum apolipoprotein C-II level was determined by single radial immunodiffusion method using agar plate containing specific antiserum. Mean values and standard deviations of serum apolipoprotein C-II in healthy subjects aged 20 to 39 were 2.9±0.77mg/dl in 14 males and 3.1±0.86mg/dl in 29 females. Neither of age groups indicated significant sex difference. The highest levels in males and females were exhibited in subjects aged 40 to 59 and in those aged over 60 years respectively.
The significant correlation between apolipoprotein C-II and triglyceride was displayed in subjects aged 20 to 39 and 40 to 59 years. In subjects aged above 60 years, apolipoprotein C-II was directly proportional to total cholesterol, phospholipid, apolipoprotein A-I and A-II, showing no significant correlation between this apolipoprotein and triglyceride, and phospholipid correlated positively to total cholesterol, apolipoprotein A-I and A-II.
In present results, direct relationship between apolipoprotein C-II and triglyceride in young age groups confirms the generally recognized property of apolipoprotein C-II which plays an important role in the hydrolysis of triglyceride by lipoprotein lipase. Apolipoprotein C-II is an obligatory factor of phaspholipase A₁ activity on lipoprotein lipase, close associations of apolipoprotein C-II with total cholesterol, phospholipid, apo A-I and A-II in aged subjects suggest strongly the participation of apolipoprotein C-II in action of phospholipase A₁ as apolipoprotein C-II does not correlate to triglyceride. These results reveal clearly the agedependent diversity of apolipoprotein C-II in lipoprotein metabolism. Therefore, in conclusion, clinical observation of lipoprotein metabolism should be performed taking the age difference into account.

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Correlations between concentrations of apolipoproteins (apo A-I, A-II, B, C-II and E) and lipids in milky plasma found in donated blood were investigated. The concentrations of apo C-II and E closely correlated with those of cholesterol, triglyceride, and phospholipid in donated plasma, while the concentrations of apo A-I, A-II and B did not correlate with those of lipids. Average concentrations of apo A-I, A-II and B in donated blood containing 15% of acid-citrate-dextrose were 111±21, 28±6, and 97±33mg/dl, respectively. The slopes of lines showing the linear-relationship between concentrations of lipids and apo C-II and between those of lipids and apo E resembled each other. The result suggests that the property of binding of apo E to lipoprotein surface was similar to that of binding of apo C-II. Excess surface area caused by an increase in lipid content of plasma was occupied by apo C-II, E, and possibly by apo C-III. The milky plasma found in donated blood seemed to be due to a catabolic disorder of triglyceride-rich lipoproteins, chylomicrons and very low density lipoproteins, because the content of apo B was not significantly affected by the increase in lipid content of the plasma. The exceptional subjects, in which the contents of apo C-II and/or E markedly deviated from the values predicted from the values of lipids by using the linear-relationships between apolipoproteins and lipids, might have some disorders other than the abnormalities in the catabolism of triglyceride-rich lipoproteins.

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Hyperlipidemia can be often seen in diabetics. In order to evaluate the effect of clinofibrate on serum lipid levels, twenty-four hyperlipidemic diabetics were investigated for twenty-four weeks. We also compare the potency of clinofibrate on lowering serum lipids of the well controlled diabetics with that of the uncontrolled diabetics. Clinofibate (six-hundred mg per day) induced no significant decrease in serum total cholesterol in all cases for twenty-four weeks, although, in many cases, which had exhibited normal levels from the start.
HDL-cholesterol was increased significantly, resulting in significant reduction of atherogenic indices.
Serum triglyceride and phospholipid were decreased significantly.
The effect of clinofibrate on lowering serum lipid levels were more potent in well controlled diabetics than in uncontrolled diabetics.
Nearly all laboratory parameters analyzed were unchanged.
Therefore it can be concluded that clinofibrate would be useful to lower serum lipid levels in the hyperlipidemic diabetics, especially in the well controlled diabetics.

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Changes in serum lipids concentration induced by either trichlormethiazide alone or combination therapy with propranolol were observed for 2 years in 91 patients with established essential hypertension. The effects of clinofibrate on such changes was also studied in 28 patients who had already received trichlormethiazide at least for 6 months.
Patients were divided into 3 groups according to dosis of trichlormethiazide (TM) used and with or without propranolol.
Group A; 23 patients treated with 2mg of TM
Group B; 41 patients treated with 4mg of TM
Group C; 27 patients treated with TM and propranolol
Total cholesterol, HDL cholesterol, triglyceride and beta lipoprotein as well as fasting blood sugar, serum electrolytes and uric acid were meassured in 6 months interval up to 2 years. Patients whose body weight changed by ±2kg during observation were all excluded on this study.
Significant increases in total cholesterol, triglyceride and beta lipoprotein were observed at the 6th month after diuretic therapy and kept significantly high values for another 12 to 18 months. Concomitant increases in FBS and uric acid were also noted. Dose response relations were clearly demonstrated on triglyceride and beta lipoprotein between group A and B, showing earlier elevation and higher levels in Group B than in Group A. Additive use of propranolol accelerated above tendencies. Characteristic change with combination therapy (Group C) was significant and continuous decrease in HDL cholesterol which was not found by TM alone, with resultant decrease in atherogenic index. Concomitant use of clinifibrate, 600mg daily, with TM lowered total cholesterol and triglyceride significantly to their control values or below but had little effects on HDL cholesterol and beta lipoprotein.
Increase in inactive insulin secretion to stimulate triglyceride synthesis in the liver as suggested by Olefsky or hemodynamic effect to lower blood pressure with use of diuretics might be responsible for metabolic consequence after diuretic therapy. Clinofibrate effectively reversed the increased serum lipid concentration to the control level even continuing TM therapy. Above observation indicated that clinofibrate might have some protective effects against arteriosclerotic vascular disease in patients with diuretic therapy.

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In the present study, effect of probucol (500mg×2, daily) on serum lipids and lipoproteins in S. F. family who showed typical familial hyper-cholesterolemia (FH) were investigated. The results were as follows:
1) The values of serum cholesterol (TC) before and after treatment of probucol were 348±14mg/dl and 220±12mg/dl, respectively. These data shows significant reduction of serum cholesterol after treatment of probucol. The percent of reduction of serum cholesterol was 36.4%. This value was significantly higher than the value (17%) in the other FH.
2) Though levels of LDL- and HDL-C were decreased after treatment of probucol, LDL-C level was markedly decrease in S. F family as compared with other FH. Reduction of HDL-C after treatment of probucol was not significant difference between in S. F family and other FH.
3) Total biliary lipids after treatment of probucol increased from 68.8μmol/ml to 141.9μmol/ml in a patient of S. F family. Level of bile acid increased after treatment of probucol, especially, in this case. On the other hand, the change of biliary lipids after treatment of probucol was not seen in other FH.
From these data, though mechanism of marked reduction of LDL-C in S. F family after treatment of probucol was still unclear, metabolism of cholesterol to synthesis of bile acid might have especially involved to marked reduction of serum LDL-C in S. F family.

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It has been accepted by some investigators that the mechanism of cholesterol-lowering action is associated wth action of Probucol converting cholesterol to bile acid. Because Probucol changes biliary lipid composition and increases percent bile acid in bile.
Therefore, the effects of this drug on biliary lipids were further investigated in this study.
1. When 750mg a day Probucol was given to 21 hyperlipoproteinemic patients for eight to twelve weeks. The molar percent cholesterol (NS) and phospholipids (p<0.1) of bile slightly decreased and therefore percent total bile acid increased slightly (p<0.1). The lithogenic index tended to be lowered but not statistically significant. Individual bile acid composition was not altered.
2. When four familiar hyperlipoproteinemic patients (two type IIa, IIb and IV) were treated with same dose of Probucol for two weeks fecal output of neutral and acidic sterols was not increased in spite of dramatic decrease of serum cholesterol.
Therefore, it is possible to conclude that although Probucol induces slight change in biliary lipid composition, there is no evidence that its mechanism of action is converting cholesterol to bile acid resulting in increasing fecal output of sterols.

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The effects of probucol on serum lipids, lipoprotein lipids, serum α-tocopherol and Co-Q₁₀ were studied in 28 patients with type II hyperlipo-proteinemia.
The patients received 2×250mg of probucol per day for 16 weeks.
Total cholesterol, triglyceride and phospholipid were significantly reduced with probucol (20%, 17%, 18% respectively). LDL and HDL cholesterol were also reduced significantly (14%, 10% respectively), but atherogenic index (VLDL+LDL-Ch/HDL-Ch) was not significantly changed. The extent of the fall in total cholesterol was directly proportional to the pre-treatment total cholesterol level, the higher this level the greater the decrement. HDL-cholesterol fell by 46% in patients whose pre-treatment HDL-cholesterol levels were over 50mg/dl, but did not fall in patients whose pretreatment levels were under 40mg/dl.
There was no significant change in serum α-tocopherol-concentration.
Serum Coenzyme Q₁₀ concentration were significantly reduced from 1, 765±308μg/ml to 1, 109±153μg/ml. Generally the drug was well accepted and free of significant toxic effects.
These results suggested that probucol in a dose of 500mg/day appeared to be useful for the treatment in patients with type II hyperlipoproteinemia.

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Long-term therapy with probucol (750mg/day) involving 65 cases of hyperlipidemic patients resulted in the following observations:
1) Serum total cholesterol was reduced significantly after 1 month of probucol treatment and the reductive effect lasted for the entire 24 month administration period thereafter. The mean reduction rate was from 15.2% to 20.1%. The effect was remarkable in the group whose total cholesterol level at baseline was high.
2) No significant reduction was observed in triglyceride, however, the group whose baseline level was over 150mg/dl showed a significant reduction at the 15 month administration.
3) HDL-cholesterol was also decreased after treatment of probucol and the group whose baseline level was less than 40mg/dl showed no significant reduction.
4) LDL-cholesterol was decreased significantly by probucol therapy at 1 month period and thereafter. The reduced amount varied from 31mg/dl to 43mg/dl in mean and mean reduction rate was 15.0% to 22.1%.
5) No significant change was observed in atherogenic index during 24 month administration period.
6) In a few cases, increases of serum GOT and GPT levels manifested, but most of them were minor of nature.
7) The side effect was diarrhea which was noted in 1 case and the symptom was disapeared by ceasing the therapy.
In conclusion, probucol is considered to be a safe and highly beneficial drug in hyperlipidemic patients.

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Effects of blood pressure, heart rate, body weight biochemical values and especially histo-pathological changes were examined under the treatment of pantethine (PTN) on stroke prone spontaneously hypertensive rat (SHRSP).
The male SHRSP were administered PTN at daily dose of about 250-300mg/kg per orally as drinking water for 13 weeks. At the end of experiment, biochemical studies of plasma and histopathological examinations of organs were performed and the results were summerized as followings:
1. PTN reduced the blood pressure 10 to 20mmHg during the development of hypertension, but did not affect the heart rate. The increasement of body weight was mildly suppressed by PTN.
2. PTN inhibited the increase of blood urea nitrogen and decreased the value of neutral fat in plasma.
3. Histopathological studies revealed the degree and incidence of arteritis in mesentery and arteritis and myocardial fibrosis in heart were mild in PTN treated group compared with control group. Kidney of PTN treated group showed almost normal appearance in contrast to that of control group which presented a number of vascular and glomerular lesions.
4. The measurements of aorta disclosed, though not be significant, the inhibition of medial hypertrophy of aorta in PTN treated group.
These results indicated PTN might inhibit the occurrence of hypertensive vascular lesions in SHR-SP.

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Effect of clinofibrate, a serum lipid reducing agent, upon prolonged administration was studied referring to rise and fall of aortic pulse wave velocity (PWV) that is one of the nonivasive evaluations of arteriosclerosis.
A total of 49 patients including 28 males and 21 females with arteriosclerotic diseases without administration of antiarteriosclerotic agent, their age ranging from 47 to 83 years, 68.5 years on an average, were used as the sclerotic group (S group), while a total of 41 patients including 24 males and females under administration of clinofibrate (Lipoclin^®) at 600-1, 200mg/day, their age ranging from 48 to 83 years, 62.9 years on an average, were used as the administration group (A group).
PWV was applied to all of the patients every 3 months, blood pressure was measured once a month, and blood chemical assay of serum lipid, etc., was carried every 4 months. The observation period ranged from 25 to 129 months (72.2 months on an average) for the S group and from 2 to 25 months (12.5 months on an average) for the A group. PWV was measured by simultaneous tracing of carotid pulse, femoral pulse and heart sound as well as distance in a straight line from the right sternal border connected with the second costa to the position whereat femoral pulse was traced, using the following formula:
PWV=[D×1.3/t+t_c]_p
wherein:
D: Distance in a straight line from the right sternal border connected with the second costa,
1.3: Anatomical smoothing value to the true length of aorta,
t: Starting time difference between carotid and femoral pulse tracings,
t_c: Starting time difference between aortic second sound and carotic notch, and
p: Minimum brachial pressure upon measurement.
As PWV depends on the minimum blood pressure, correct it to a PWV at minimum blood pressure of 80mmHg using a pressure calibration to have individuals comparable.
When changes of PWV (ΔPWV) on an average of both groups are compared every 5 months, it was 0.17m/sec by 5 months, 0.31m/sec by 10 months, 0.56m/sec by 15 months, 0.82m/sec by 20 months and 0.90m/sec by 25 months respectively in the S group, while in the A group it was 0.06m/sec by 5 months, 0.05m/sec by 10 months, 0.19m/sec by 15 months, 0.13m/sec by 20 months and 0.1m/sec by 25 months respectively. PWV varied within the low level, and after 20 and 25 months the A group showed the lower value at 5% and 1% levels of significance respectively.
It was suggested that upon prolonged administration of clinofibrate, organic arteriosclerotic lesion is inhibited.

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To elucidate the mechanism by which KCD-232 inhibits cholesterol absorption from intestinal lumen, we studied the influence of KCD-232 on cholesterol or fatty acid absorption, esterification of absorbed cholesterol and protein synthesis of lymph chylomicron in thoracic-duct fistura rats.
The results obtained are summaried as follows.
1) KCD-232 significantly depressed [¹⁴C] cholesterol absorption into lymph by 43.6% during the 8-hr period and by 49.5% during the 24-hr period, but did not [³H] oleic acid absorption at all.
2) KCD-232 had no effect on the distribution of absorbed [³H] oleic acid among the lipid fractions of lymph, however showed a significant decrease percentage of [¹⁴C] cholesterol associated with esterified form.
3) KCD-232 had no effect on the protein level of lymph chylomicron and [¹⁴C] leucine incorporation into chylomicron protein, and did not alter the apoprotein patterns of chylomicron as judged from polyacrylamide gel electrophorogram.
These results suggest that the inhibition of cholesterol absorption by KCD-232 may be attributed to a decrease of cholesterol esterification within mucosal cells perior to formation of chylomicron, rather than to a suppression of protein synthesis of chylomicron.

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The effects of Dilazep Dihydrochloride on platelet and arterial arachidonate metabolism in vitro and in vivo were studied in patients with cerebral atherosclerosis and age-matched controls. Platelet phospholipase activity was measured as described elsewhere (Thrombos. Res., 24, 131, 1981). Thromboxane B₂ and 6-keto prostaglandin F_1α concentrations were measured by ¹²⁵I-radioimmuno assay. Platelet phospholipase activity was significantly higher (p<0.005) in patients with cerebral atherosclerosis than in the controls (11.93±4.84, 8.72±3.88n mol/mg prot/30 sec, respectively). The values indicate mean±S. D. Nearly same extent of concentration-dependent suppressions were observed in platelet phospholipase activity and thromboxane B₂ synthesis in those patients by in vitro addition of Dilazep. The former was suppressed to 60.0±32.7% of control (p<0.005), and the latter to 57.6±3.3 (p<0.005) by the addition of Dilazep (100μg/ml). Plasma thromboxane B₂ was decreased to 84.9±23.0% of control by the administration of Dilazep (150-300mg/day) for 2 to 4 weeks, however, 6-keto prostaglandin F_1α was not. These results suggest that the effects of Dilazep previously reported by other investigators, such as platelet membrane stabilization, suppression of thromboxane B₂ synthesis and platelet aggregation, may be due to the inhibition of platelet phospholipase.

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Mace-83 extract obtained from seed covering of nutmeg is expected to show an antiarteriosclerotic effect due to its potentiation with unsaturated fatty acid, various vitamins, etc. In this study an effect of Mace-83 was examined from serum lipid and six constituents of thoracic aorta.
A total of 16 male rabbits of 38 months, including 8 with the Mcce-83 (M group), 4 with cholesterol (C group) and 4 with Mace-83 plus cholesterol (MC group) were used. to produce arterosclerosis cholesterol was orally given at 1g to the C and MC group. To the M and MC groups Mace-83 was given at 500mg/day. Sixteen weeks later referring to all the rabbits, total cholesterol (T. CHO), triglyceride (TRIG), β-lipoprotein (β-LIPO), HDL-cholesterol (HDL), 4 in total of serum lipids, were measured. Furthermore, all the rabbits were killed and constituents media of thoracic aorta were stained and observed histochemically, i. e., SMC with Azocarmine G stain, EL with Weigert's stain, CL with Van Gieson's stain, AMPS with Alcian blue stain and GP with PAS stain. T. CHO was 90.5mg/dl in M group, 369.8mg/dl in C group and 81.3mg/dl in MC group respectively. The MC group showed no difference with M group but the significantly lower value than that of C group (p<0.01). TRIG was 108.5mg/dl in M group, 83.3mg/dl in C group and 94.4mg/dl in MC group respectively. No significant difference was observed among these 3 groups. Beta-LIPO was 184.3mg/dl in M group, 995.3mg/dl in C group and 151.5mg/dl in MC group respectively. Compared with C group, M and MC groups showed significantly low value (p<0.001-p<0.01). HDL was 18.9mg/dl in M group, 22.3mg/dl in C group and 26.5mg/dl in MC group respectively. There was no significant difference among those 3 group, although it was highest in MC group. When histochemically observed, C group was strongly disturbed, while MC group showed slight swelling of SMC and cell nucleus that was not remarkable though, and damage of EL and CL was minimal. AMPS and GP remained unchanged. From changes of serum lipid and observation of arterial tissue, it was suggested that Mace-83 has an antisclerotic effect.

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