To assess the atherogenic significance of the particle size abnormalities of HDL, the particle size of HDL was measured by high performance liquid chromatography in 69 male diabetics, 60 male patients with coronary heart disease (CHD), and 151 male normal subjects. The particle size of HDL was represented by the elution volume at the peak of HDL.
In normal subjects the particle size of the cholesterol monitored HDL (C. HDL) was larger than that of the triglyceride monitored HDL (G·HDL). This suggests that the smaller G rich HDL becomes the larger C rich HDL taking up cholesterol. In diabetics the particle size of the G·HDL was larger than that in normal subjects, and there was no difference between the particle size of the G·HDL and the C·HDL. These results suggest that the function of HDL taking up cholesterol is disturbed in diabetics. In patients with CHD the particle size of the G·HDL was larger than that in normal subjects, which was similar to the result in diabetics. This suggests that the larger G·HDL is related to atherogenesis.

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Diabetic patients and diabetic animals are known to exhibit an increased tendency to develop vascular disease. Blood vessels and platelets obtained from diabetic patients and diabetic animals also exhibit altered production of eicosanoids.
The leukotrienes are a recently discovered group of eicosanoids which are produced by vascular tissue as well as neutrophil, eosinophil and mast cell.
Leukotriene B4 and C₄ (LTB₄ and LTC₄), members of leukotrienes, have shown to constrict vessel, increase the vasopermiability, stimulate the production of superoxides and so on. Therefore, the closed relationship between leukotrienes and vascular damage could be exist in diabetes mellitus. Then, we measure the levels of whole blood leukotrienes in NIDDM patients and STZ-diabetic rats.
The first results are that the value of whole blood LTB₄ and LTC₄ are higher in NIDDM patients than in control group and that the mean level of whole blood LTC₄ is higher in patients with ST-T change on ECG or with increased pulse wave velocity (PWV) than in patients without ST-T change or with normal range of PWV.
The Second results are that the levels of whole blood LTB₄ and LTC₄ are higher in STZ-diabetic rats than in normal rats, but the effect of duration of diabetes on the levels of LTB₄ and LTC₄ are not observed in STZ-diabetic rats.
These results made us strongly speculate a meaningful relationship between leukotrienes level, diabetic control and vascular damage in NIDDM patients and STZ-diabetic rats.

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Lipids accumulate in atheromatous lesions not only intracellularly but also extracellularly. Major parts of extracellular lipids are bound to extracellular matrix (ECM). In this paper we established a model system to investigate lipoprotein bindings to ECM prepared from cultured aortic smooth muscle cells and studied the effects of pancreatic elastase on such bindings.
Low density lipoprotein (LDL) was incubated with ECM in a CO₂ incubator at a concentration of 25 to 100μg/ml (total cholesterol) for 1 to 24h.
The binding of LDL increased dose-dependently up to 8h and reached plateau. The modified form of LDL which was generated by incubating with human umbilical vein endothelial cells bound to ECM 4.38 times as much as native LDL. Elastase reduced the binding of LDL to ECM when it was added to the culture either at the same time as or before LDL addition. Moreover, elastase increased the release of LDL from ECM which was already bound to ECM.
The above results indicated that the ECM prepared in vitro could be used as a model of in vivo extracellular matrix which could bind lipoproteins. The effects of elastase observed in the model system suggested a mechanism of antiatherogenecity of the drug other than hypolipidemic effect.

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We have been using atherosclerotic risk factor index as (TC-HDL-CHO)/TC. This index was founded to be related with serum triglyceride, T-cholesterol, uric acid and obesity, and negative relation was noted with HDL-cholesterol. In this report, we have selected 128 persons who had risk factor index over 0.8, more than twice, among the total number of 7, 211 persons who had checked during 1979 to 1984.
Control group was chosed 126 persons who had risk factor index below 0.8. The persons with high risk also showed hypertension in 34, diabetes 3, hyperlipemia 10, liver diseases 3, while hypertension 10, diabetes 1, liver diseases 2, and hyperlipemia 0 in the low risk group.
The frequency of food intake were observed much more frequently of cheese and milk, vegetable, fruits, fish and rice in low risk group. Tabaco is inhaled high frequently by high risk group.

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The relationship between ChE activity and coronary artery stenosis was investigated. Serum ChE levels were positively correlated with T. Chol, Apo B, C-II, C-III, VLDL-C and LDL-C, but not with serum albumin level, suggesting that ChE in serum might be secreted from the liver accompanied with VLDL synthesis and secretion. To evaluate the role of ChE on coronary stenosis, we investigated the relationship between ChE levels and coronary stenosis score in 32 patients (Men 22, Women 10) who were carried out coronary angiography under the suspect of ischemic heart disease. In comparison between High ChE group and Low ChE group, there were no difference between both groups in the levels of T. Chol, VLDL-C+LDL-C and Apo B, however, coronary stenosis score in High ChE group exhibited a statistically significant low levels, as compared with that in Low ChE group. These results suggest that the serum ChE may play an important role on lipoprotein metabolism and work as an antiatherogenic factor.

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The relationship of plasma lipids, lipoprotein and apolipoprotein (apo) concentrations to angiographically defined coronary artery disease (CAD) was investigated in 69 male patients above the age of 50. Selective coronary angiography (CAG) was performed by the technique of Sones and Judkins and CAD was defined as 75% or greater luminal diameter narrowing of one or more of the major coronary arteries. According to the results of CAG, the patients were divided into three groups: patients with multivessel CAD (n=18), single vessel CAD (n=29) and normal coronary artery (NCA: n=22). All subjects were normolipidemic (total cholesterol<230mg/dl, triglyceride<150mg/dl) and the coronary risk factors of age, blood pressure, smoking, body mass index and glucose tolerance were matched among three groups. Patients with multivessel CAD had significantly higher levels of plasma triglyceride and lower levels of HDL-cholesterol. No significant differences in plasma cholesterol and LDL-cholesterol were observed among three groups. CAD groups had significantly lower levels of plasma apo A-I and higher levels of plasma apo B and only apo A-I/apo B ratio was related significantly to the severity of CAD. Hyperlipidemia increases the risk for CAD, but about half of the Japanese patients with CAD have normal plasma lipid levels. Our data indicated that apolipoproteins, which are the protein parts of lipoproteins, are more important than the lipid component in discriminatinb between CAD and NCA in normolipidemic males.

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Fatty acids of plasma total lipids, abdominal subcutaneous adipose tissue and omental adipose tissue were analyzed in 27 subjects including IGT and mild NIDDM patients who received surgical treatment of cholecystolithiasis. Their dietary habits were estimated by a combined recall and interview method. The subjects were divided into meat-eating group, fish-eating group and intermediate group. Meat group was shown to have consumed an increased amount of oleic acid (C 18:1) when compared with fish group. The latter group was rather characterized by the increased intake of long-chain polyunsaturated fatty acids such as C 20:5 and C 22:6. These long-chain polyunsaturated fatty acids were found to be significantly increased in fish group as compared with meat group. The fish intake accelerated the distribution of C 20:5 and C 22:6 more strongly in plasma than in adipose tissues.
Interestingly, C 18:1 showed rather constant body distributions among the 3 groups despite the marked difference in the rate of its intake, and C 18:1 was rather poor in plasma and rich in depot fat tissues in all groups.

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We examined the effect of splenectomy on experimental atherosclerosis in cholesterol-fed rabbits. Following cholesterol feeding the splenectomized rabbits showed significantly higher levels of serum cholesterol, triglyceride, and phospholipid in contrast to lower levels of highdensity lipoprotein cholesterol, as compared with sham-operation rabbits. The percentage of aortic plaque area in the splenectomized group tended to be higher than that in the sham-operation group. Our results suggest a possible role of the spleen in lipid metabolism, in particular an exsitence of splenic factor which can suppress diet-induced hyperlipidemia.

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It is noted that there are two types of WHHL rabbits; the one with high incidence of coronary atherosclerosis (type 1), and the other with low incidence of coronary atherosclerosis (type 2). In order to analyze the difference between type 1 and type 2 WHHL rabbits, in this study, the atherogenesity of some classes of lipoproteins were estimated by incubation with mouse peritoneal macrophages.
Among the several classes of lipoproteins, VLDL from type 1 WHHL rabbit could stimulate cholesteryl [14C]-oleate synthesis 10.5 fold than did VLDL from type 2 WHHL rabbit. VLDL from normal rabbit showed no significant stimulatory effect. LDL did not stimulate esterification of cholesterol in macrophages, unless it became modified form.
Mass ratios (cholesterol/protein) of these lipoproteins were calculated for their characterization. The calculated ratios of β-VLDL, VLDL from type 1 WHHL rabbit, from type 2 WHHL rabbit and from normal rabbit were 13.6, 5.69, 2.05 and 0.82, respectively.
These data suggest that the high incidence of coronary atherosclerosis in type 1 WHHL rabbit is partially due to the cholesteryl ester rich VLDL, and that the atherosclerosis observed in WHHL rabbit is caused not only by high concentration of plasma LDL, but also by cholesteryl ester rich VLDL.

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The purpose of the present study was to analyze the hypotensive mechanisms of ketanserin (a serotonergic receptor antagonist) in hypertension. The perfused mesenteric vasculatures were prepared in spontaneously hypertensive rats (SHR, Okamoto and Aoki, 20-22 weeks old) and age-matched Wistar Kyoto rats (WKY), and the effects of ketanserin on norepinephrine (NE) overflow from the sympathetic nerve endings as well as on vascular responsiveness were investigated.
(1) Pressor responses to electrrcal nerve stimulation or exogenous NE were inhibited by ketanserin in a dose-dependent manner.
(2) NE overflow evoked by electrical nerve stimulation was also decreased by ketanserin in both SHR and WKY. The suppressive degrees were greater in SHR than in WKY.
These results demonstrate that ketanserin could affect presynaptic neurotransmitter release from the sympathetic nerve endings in resistance vessels. These sympatho-depressive actions might contribute, at least partially, to the hypotensive mechanisms of ketanserin.

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1) We studied the influences of a stable TXA₂ analogue, STA₂, on cultured bovine aortic smooth muscle cells.
2) STA₂ (10^-7-10^-5M) inhibited the PDGF-induced migration of SMC dose-dependently.
3) STA₂ (10^-8-10^-5M) inhibited the proliferation of SMC dose-dependently.
4) STA₂ injured SMC only in a high concentration (10^-5M) when the damage was estimated by LDH release.
5) STA₂ did not affect the viability of SMC examined by the trypan blue exclusion test.
6) STA₂ stimulated PGI₂ release of SMC; however, the effect of STA₂ was not suppressed by a TXA₂ receptor antagonist, ONO-3708.

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To investigate the effect of an HMG-CoA reductase inhibitor, CS-514, on the serum levels of lipid, lipoprotein including Lp (a) and apoprotein, CS-514, in a dosage of 10mg a day, was administered for 12 weeks to 15 patients with hypercholesterolemia of which 5 patients had heterozygous familiar hypercholesterolemia. Before and after 4, 8 and 12 weeks of treatment, blood was collected after 12 hours of fasting. Serum total cholesterol and triglyceride levels were determined by enzymatic methods. HDL cholesterol level was determined by the heparin-Ca method. Apoprotein A-I, A-II, B, C-II, C-III, E and Lp (a) lipoprotein levels were measured by single radial immunodiffusion (SRID) methods. The total cholesterol level significantly decreased from 311±61 to 267±57mg/dl, whereas the HDL cholesterol level significantly increased from 52.1±16.1 to 59.2±20.3mg/dl. Both triglyceride and Lp (a) levels remained virtually unchanged (from 164±131 to 150±115mg/dl, from 23.0±17.2 to 23.6±18.3mg/dl, respectively). The serum levels of apo A-I and A-II significantly increased whereas that of apo B significantly decreased. No significant changes were found in the serum levels of either apo C-II, C-III, or E. No significant untoward reactions were observed.

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Studies in animal models suggest that monocyte accumulation in the arterial intima is an important early event in the initiation of atherosclerosis. It has been reported that chemotactic factors for monocytes can be extracted from vessels of cholesterol fed animals. The present study indicated that extracts from intima and media of normal portion of human thoracic aorta induced monocytes chemotaxis. Checker board analysis showed that the activity was due to chemotaxis and not to chemokinesis. We investigated the chemotactic activity of aortic extracts for monocytes from normal subjects and familial hypercholesterolemia (FH). The data indicated that chemotactic index (C. I.) in FH was 2-3 times greater than that in normal subjects.
These findings indicated that there were monocyte-chemotactic factors in the intima and media of normal aorta and that control of monocyte recruitment into artery would involve not only alteration of the arterial wall but also functional changes in the circulating monocytes.

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We made the intimal thickening of jugular vein (JV) and common carotid artery (CCA) in rabbits by placing a polyethylene cuff. And the intimal thickening was studied by light and electron microscopy.
Even in JV, the thickening was formed probably by the migration of medial smooth muscle cells (SMC) into the intima with subsequent proliferation.
The thickening of CCA consisted of tightly packed SMC and a few elastic fibers. While that of JV was filled with abundant collagen fibers, some layers of SMC and few elastic fibers.
Some arterioles were often observed in the area of intimal thickening of JV.

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Myocardial infarction can experimentally be produced in cholesterol-fed rabbits by flushing of thromboxane A₂ (TXA₂) into the aortic root. Using this model, we studied the preventive effects of the anti-platelet agents such as aspirin, phthalazinol-a cyclic AMP phosphodiesterase inhibitor and ticlopidine-adenylate cyclase activator.
Eighteen male albino rabbits were fed with one percent cholesterol containing pellets for 20 weeks and then with a commercial diet for 5 weeks. Fourteen rabbits were handled as the treated groups. Four rabbits were given 10mg/kg of aspirin orally every morning for seven days before the experiment. In the same way, six rabbits were treated with 400mg/kg of phthalazinol and four rabbits with 200mg/kg of ticlopidine. Other four rabbits were treated for the same period with placebo capsules of potato starch as placebo controls. Twenty-five μg of prostaglandin H₂ and 2.5mg protein of microsome obtained from cow platelets were mixed and flushed into the coronary circulation through an indwelling balloon catheter, which was inserted through carotid artery into the aorta to the levels of the orifice of the aortic valve. ECG, BP, plasma levels of thromboxane B₂ (TXB₂), 6-keto prostaglandin F_1α (6-keto PGF_1α), cyclic AMP (CAMP), cyclic GMP (cGMP), GOT, GPT, LDH and CPK were monitored for 48 hours.
In the placebo control rabbits, all rabbits exhibited typical ST elevation in II, III, aVF and/or precordial leads immediately after injection of TXA₂ and Q waves followed. Plasma levels of TXB₂ increased prominently immediately after flushing of TXA₂. Parallel to these changes, 6-keto PGF_1α levels were also high immediately after flushing but former levels were reverted to 5 minutes after injection. Histological and histochemical studies, 48 hours after flushing, revealed an ischemic areas of various size, scattered irregularly from the endocardial to epicardial lesions of the ventricles. Microthrombi, what we call “shower thrombi”, were observed in many intramuscular small coronary vessels.
On the other hand in the treated groups, Q waves were recorded in only one of four in the aspirin, only two of six in the phthalazinol and none of four in the ticlopidine treated rabbits. Plasma levels of TXB₂ of these three treatedgroups increased immediately after flushing, but their maximal values were lower than those in control group during the experimental period. Histological studies demonstrated that ischemic lesions were smaller and less frequently encountered in the three treated-group rabbits.
Plasma levels of cAMP and cGMP arose at the maximum interestingly five minutes after flushing in the three groups except the aspirin, although the levels of prostanoid did the maximum just immediately after flushing.
These data demonstrate that thromboxaneinduced myocardial infarction could be prevented by the treatment with the anti-platelet agents such as aspirin, phthalazinol, ticlopidine-treated rabbits.

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We investigated the effects of elastase on cyclic AMP accumulation in cultured smooth muscle cells from pig coronary artery. The cells were stained with HHF 35, a muscle-actin-specific monoclonal antibody.
Although elastase alone did not have significant effects on cyclic AMP accumulation in the cells either in the presence or in the absence of isobutylmethylxanthine, a phosphodiesterase inhibitor, it enhanced cyclic AMP accumulation in response to forskolin or isoproterenol. Elastase failed to suppress cholesterol accumulation by the cells during the incubation period of 5 days with hyperlipidemic serum. However, it prevented the reduction of cyclic AMP levels in the cells exposed to hyperlipidemic serum.
These results suggest that elastase may alter cyclic AMP accumulation in the coronary smooth muscle cells and such may partly explain the antiatherosclerotic properties of elastase.

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It is our purpose to investigate serum lipids, especially serum apoproteins in order to clarify the precipitating factors in atherogenesis in healthy elderly.
In this investigation 38 healthy Male elderly subjects aged 64.8±5.4 years (M±SD) and 12 healthy male young subjects aged 23.1±3.3 years were used, who all showed normal blood pressure levels, normal findings in ECG and normal renal or hepatic function tests. Fasting blood sample was obtained by venipuncture for measuring serum lipids and so on.
There were found the significantly increased values of serum cholesterol (Chol), triglyceride (TG), (VLDL+LDL)-Chol and beta-lipoprotein in healthy male elderlys as compared to healthy male youngs. Moreover, apoprotein B, C-II, C-III and B/A-I ratio were higher significantly in healthy male elderlys than in healthy male youngs. Apoprotein B and B/A-I ratio were also higher significantly in 28 healthy male elderlys than in 12 healthy male youngs, who all showed less than 240mg/dl of serum cholesterol and less than 200mg/dl of serum triglyceride.
In the group of male 50 cases consisting of 38 healthy male elderlys and 12 healthy male youngs, there were found significant positive correlations between apoprotein A-I and HDL-Chol, between apoprotein B and Chol, TG or body mass index (BMI), between apoprotein C-II and C-III, between Chol and BMI, and between TG and apoprotein C-II or C-III. Even in the elderly group of 38 healthy male elderlys there were also observed significant positive correlations between apoprotein A-I and HDL-Chol, between apoprotein B and Chol, TG or BMI, between apoprotein C-II and C-III, and between TG and apoprotein C-II or C-III.
It was concluded that healthy male elderlys showed the relatively increased values of serum apoprotein B, C-II, C-III and B/A-I ratio in addition to the increments of serum cholesterol and triglyceride as compared to healthy male youngs.

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We have previously reported the important role of a vascular elastase-like enzyme for pathogenesis of arterial diseases. In this experiment, in order to clarify the pathogenesis of arterial changes in atherosclerosis and hypertensive arterial diseases, the effects of sodium and a high fat diet in addition to hypertension on aortic elastase-like activity and elastin content were examined using stroke-prone rats (SHRSP) which are a suitable model for vascular diseases. Furthermore, the effects of exogenous elastase on endogenous elastase-like activity and elastin content were also examined.
Long-term (6 weeks) loading of salt and a high fat diet resulted in an increase in elastase-like activity and in a decrease in elastin content in aorta of SHRSP. These results indicate that the vascular changes could be stimulated by the loading of such risk factors for vascular diseases due to the increase in elastase-like activity. On the other hand, vascular elastase-like activity was lower in SHRSP administered exogenous elastase in comparison with untreated control, whereas the elastin content was higher in the former than in the latter. This indicates that the administration of elastase may prevent the degradation of elastin due to the lowering of elastase-like activity.
Although the mechanisms of increasing or decreasing in elastase-like activity are still obscure, the results confirmed the important role of vascular elastase-like enzyme or elastase for arterial changes.

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This study was performed to analyze coronary risk factors by using multi-factorial analysis (Hayashi-III) and to find a method to predict the occurrence of ischemic heart disease in apparently healthy persons. Subjects included 4, 949 apparently healthy industry employees and 9 patients with myocardial infarction selected for comparison with healthy persons. Items of measurements used in this study included serum lipids, resting blood pressure, resting ECG, and exercise ECG.
Results were as follows:
(1) From the results of a quantification method for multi-dimensional qualitative data, the major factor of the first component was blood pressure.
(2) The major factor of the second component were serum lipids.
(3) In scatterd diagram, persons having scores under the discriminant line are at risk for the occurrence of ischemic heart disease and prophylactic measures should be started.

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A low level of high-density-lipoprotein (HDL) has been reported to be a predictive indicator of coronary heart disease (CHD). Apolipoprotein A-I is a principal protein constituent of HDL and decreased apolipoprotein A-I level is also regarded as a risk factor of CHD. We studied the difference in a Restriction Fragment Length Polymorphism (RFLP) of apolipoprotein A-I gene with endonuclease Pst I digestion between 30 CHD-patients and 24 normolipidemic control groups. Using 2.2kb fragment of human apolipoprotein A-I gene as a probe, 2.2kb and 3.3kb bands were detected. There was no significant difference in the frequency of 3.3kb band between patients with CHD and controls, 0.100 and 0.063, respectively. In this study the polymorphism of apolipoprotein A-I gene is different from those reported by Ordovas. Such difference would be expected to show racial and regional influences. In Japanese population there was no significant association between this RFLP and CHD. The 24 patients with CHD were divided into two groups according to the presence or absence 3.3kb band and patients with 3.3kb band have been proved to have higher triglycerides and lower HDL cholesterol levels.

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In the present study, we evaluated the effects of lecithin (LC) and eicosapentaenoic acid (EPA) administration on the changes of the plasma apoproteins and lipids in subjects with normo- or hyperlipidemic non-insulin-dependent diabetes mellitus (NIDDM).
EPA significantly reduced the level of plasma LDL-cholesterol in cases with normolipidemic NIDDM.
When EPA was administrated to persons with hyperlipidemia, total cholesterol and apoprotein-B slightly but not significantly decreased (281mg/dl to 240mg/dl, 132mg/dl to 118mg/dl, respectively).
LC tended to increase plasma level of apoprotein A-I in normo- and hyperlipidemic NIDDM.

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We have described a new family of apolipoprotein C-II deficiency, and examined restriction enzyme fragment length polymorphism (RFLP) of apo C-II gene. Patient was 32-year-old woman, and suffered from acute pancreatitis during pregnancy. Her triglyceride level was 8, 670mg/dl. Apo B and C-III levels were increased, but apo C-II level was negligible by single radial immunodiffusion analysis. Isoelectrofocussing gel electrophoresis showed no band corresponding to apo C-II. RFLP of apo C-II gene with EcoR I, Tq I, Bgl I, and MSP I showed almost same bands with that of control subject, suggesting that there was no major deletion in apo C-II gene of this apo C-II deficient patient.

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With a view to elucidating the actual aspect of serum lipids in healthy normal Japanese living in cities, on those managers of commerce and industry and their family members living in principal cities of Japan, aged from the twenties to the seventies including 18, 873 males and 5, 471 females, 24, 344 in total (excluding those cases with past history such as hypertension, heart disease, hepatic failure, etc.), the recent levels of serum lipids for healthy normal Japanese were examined from changes with ageing in distribution characteristics by sex and age of total cholesterol, triglyceride, β-lipoprotein, and HDL-cholesterol and in their mean values. The results revealed more remarkable individual characteristics than ageing characteristics and practically no shift to higher values with ageing on frequency distribution. In terms of the correlation by sex of the above levels of serum lipids with age, males showed higher values than females, and in terms of changes with age and by sex in their mean values, each of these serum lipids showed a remarkable difference by sex in age-related tendency, with those distribution characteristics which can not be observed for other nutrients such as blood sugar, serum total protein, etc., but can be observed only for serum lipids. HDL tended to decrease with ageing for females compared with males showing a gradually increasing tendency.

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On healthy normal Japanese living in principal cities of Japan, examination was made of the relationship between levels of 4 serum lipids, i.e. total cholesterol (TC), triglyceride (TG), β-lipoprotein (β-LP) and HDL-cholesterol (HDL), and ischemic ECG changes or aortic pulse wave velocity values (PWV) in recent (1984) epidemiological cases of 18, 873 males and 5, 471 females, 24, 344 in total, aged from the twenties to the seventies.
1) Incidence of ischemic ECG changes in each serum lipid value showed better and weaker positive correlations for TC from the thirties to the fifties of males and females, respectively. For TG, no definite tendency was seen except for the weak positive correlation observed for males from the thirties to the forties.
2) In terms of the relationship between each serum lipid value and PWV value indicative of organic changes in the aorta, no definite tendency was seen except for positive correlation for TC at each age-group of both sexes. Correlation coefficiency showed low values, and each serum lipid tended to show higher values for females than for males. However, mean PWV value in each serum lipid value was only a little higher for males than for females.
3) Change in PWV valve showed a significantly high value for each age-group of both sexes in the ischemic ECG change-positive group, with the difference by sex being only a little greater for males. Incidence of ischemic ECG changes in each PWV value showed an exponential function increase, suggesting a progress of PWV value preceded by some ischemic ECG changes.

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