The role of macrophages in cholesterol ester accumulation in atheromatous aorta have been discussed. Lipoprotein infiltrate across the arterial endothelium, and degraded in macrophages and/or modified smooth muscle cells in subendothelial space of the arterial wall. The cells probably transform to foam cells during the progressive accumulation of cholesterol ester. If the successive endocytosis occurs, foam cells may become overloaded with cholesterol ester and ruptured. Lipophilic materials spread out in extracellular space may not be regulated by living matters. Extracellular lipids are possibly endocytized by scavenger cells such as macrophages and modified smooth muscle cells to maintain hydrophiric environment and produce further foam cells. Thus, foam cell formation seems to be one of the protective mechanisms against excess lipids in extracellular space in atherosclerotic plaques.

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The appearance of the human aortic wall and coronary arterial wall was examined by Scanning (SEM), Transmission electron microscopy (TEM) and chemical analysis in order to clarify the mechanism of removal of lipids deposited in the aorta.
The findings of SEM and TEM showed that circulating liucocytes penetrated into the arterial wall through endotherial marginal folds in the early atherosclerotic lesion. In some specimens many spherical bodies with a diameter of 10-30μ were observed in the subendotherial space and intimal media. After the leucocytes had phagotized a large quantity of the lipids in the arterial wall, they were transformed into lipid-including cells, which we named “lipid-containers.” Furthermore, we found that some lipid-containers returned from the subendotherial space into the blood stream. Possibly, many lipids in the atherosclerotic lesion can be removed by this process. From those new findings, it is considered that the leucocytes (macrophage) play an important role in regression and prevention of atherosclerosis of human arteries at the cellular level. The human arterial wall was divided into three sections such as intima, intimal media and media by cryostat.
The small lipids particles in the macrophages were isolated by ultracentrifugation after the three sections were digested with proteolytic enzymes. The lipid particles isolated from intima were small in diameter and very low density (1.006). But those from media were big and high density (1.210). The latter could not be so easily removed from arterial wall.
From above findings, it suggested that mobility of macrophages played an important role of lipids deposition in the arterial wall.

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ATP-dependent Ca²⁺ uptake by microsomes, isolated from bovine aortic and mesenteric smooth muscle, was inhibited by Ca²⁺ ionophore: A23187, and Ca²⁺ release from the microsomes was induced immediately after adding A23187. However, Ca²⁺, Mg²⁺-ATPase activity of those microsomes was not affected. Rate of A23187-induced Ca²⁺ release was not slowered by divalent cations: Ca²⁺, Mg²⁺, and Sr²⁺ at high concentrations (1mM). LDL also inhibited ATP-dependent and ATP-independent Ca²⁺ uptake by the microsomes, but did not induce Ca²⁺-release from the microsomes. Thus, site of the LDL action of the microsomes was suggested to be outer membrane binding sites of Ca²⁺.

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It is well known that there are various oxidized derivatives of cholesterol. Recentry, some types of the oxidized cholesterol have been shown to be mutagenic to cultured cells. However, the mechanism of formation and metabolism of the oxidized cholesterol in vivo have not been yet clarified.
In the present study, the contents of cholesterol and oxidized cholesterol in human atherosclerotic lesion were determined in order to examine the relationship between the oxidized cholesterol and the progression of atherosclerosis.
Human aorta obtained at autopsy was sectioned into intima, intimal media and media. The lipids in each section were extracted with chloroform-methanol mixture and analyzed using thin-layer chromatography and gas-liquid chromatography.
In intact human aorta, the contents of cholesterol and cholesterol-5α, 6α-epoxide decreased in proportion to the distance from luminal surface. In atherosclerotic lesion, the contents of cholesterol and cholesterol-5α, 6α-epoxide in intimal media were larger than in intima and in media. The content of cholesterol-5α, 6α-epoxide in intimal media and media were about 2.9 times and 2.4 times larger than in intact aorta, respectively.
Additionaly, larger amount of cholesterol-5α, 6α-epoxide was detected in the serum of elderly subjects however it was not detected in the serum of cord blood.
The present study revealed that oxidized derivatives of cholesterol were related to the progression of atherosclerosis and aging.

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Proteoglycans (PG) were extracted from bovine aorta (intima-media) with 4M guanidium chloride in the presence of protease inhibitors following through cetylphridinium complexes. The crude PG, thus obtained, were fractionated by CsCl density gradietnts centrifugation into eight fractions, and the bottom fraction 1 and 2-5 were pooled as PG-A and PG-B, respectively. The ratio of glycosamino-glycans (GAG) to proteins of PG-A was higher than that of PG-B. Chondroitin sulfate is the main GAG in both PG-A and PG-B and that of PG-A and PG-B are 68.6% and 65.1% of the total GAG, respectively. On the other hand, heparan sulfate of PG-A and PG-B are 1.4% and 9.3% of the total GAG, respectively, and the heparan sulfate in PG-A is less than one-seventh of heparan sulfate of PG-B.
The effects of PG-A, PG-B and standard GAGs on platelet aggregation were studied. The PG-A did not affect platelet aggregation induced by ADP, collagen, but inhibited that induced by thrombin, whereas PG-B did not affect platelet aggregation induced by ADP, collagen, epinephrine and thrombin. Of the standard GAGs investigated, hyaluronic acid and chondroitin sulfate had only little inhibitory action on thrombin-induced platelet aggregation, while the inhibitory action of dermatan sulfate and heparan sulfate was markedly potent in this respect. It is suggested from the results of this study that the differences of action on platelet aggregation between PG-A and PG-B would not be due to thier GAG compositions.
In order to clarify the affector side of PG on platelet aggregation, we will further study on the details of characteristic differences of PGs such as sulfur content, the ratio of glucosamine to galactosamine and molecular weight.

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We measured by immunoassay the accumulation of apo B in seven separate segments of coronary arteries from swine to determine the relationship between apo B accumulation in coronary arteries and coronary atherosclerosis. Twenty-two swine were used as experimental animals. Twelve of them were fed 1.5% cholesterol diet for 10 to 15 weeks. Ten age-matched swine were fed control diet. Apo B accumulation was greater in coronary artery segments from cholesterol-fed group than from control group. Especially proximal segments of coronary arteries demonstrated increases in apo B accumulation and appearance of atherosclerotic lesions. Even in distal portions of coronary arteries, apo B accumulation and foam cell lesions were observed in branch point in cholesterol fed group. Apo B accumulated in edematous zones just below endotherium that were also positive for alcian blue, presumed to be sulfated glycosaminoglycans (S-GAG). These findings suggest that LDL deposition in arteries plays a important role to develop atherosclerotic lesions.

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It has been widely recognized that arteriosclerosis is a silent disease for a long period of time during its progressive stage. Therefore, it seems reasonable to think very important for early stage of it to be investigated in children and young adults.
This study was aimed to make the differences clear between coronary arteries and aortae for the purpose of elucidation of local factor (s) of progression of sclerosis.
Arteriosclerotic index was caliculated by the method of point counting and Gore's score given to severity of arteriosclerosis, was significantly higher in the aortae (398 cases) than in the coronary arteries (314 cases). In contrast, microscopically proven intimal thickening had significant tendencies that coronary arteries (126 cases) was stronger than aortae (233 cases). Coronary arteries has characteristics of fibrocellular proliferation and weaker lipid deposition in thickened intima, while aortae was characterized by significant lipid deposition in thinner intima. Vasa vasorum became more prominent microscopically in coronary arteries, as intimal thickening progressed, but they did not so in aortae. Circulatory disturbances within the wall might be one of local factors as to coronary arteries.

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Using spontaneously hypertensive rats with different degrees of hypertension, the influence of the duration of hypertension on platelets was examined through changes in platelet serotonin contents. Their blood pressure was in the order of m-SHRSP>SHRSP>SHR>WKY. Serotonin content in normotensive WKY platelets was maintained in the range of 0.715±0.048 (17) n mole/10⁸ cells through ages (5-50 weeks) in both sexes. In contrast, a significant decrease has begun to be observed in male m-SHRSP at 18 weeks of age, in female m-SHRSP and male SHRSP at 22 weeks, and in female SHRSP at 32 weeks respectively, compared with age-and sex-matched WKY. The content in SHR of both sexes was unaltered upto 36 weeks of age. Time of the appearance of these exhausted platelets coincided with the reported time of scan electron microscopic observation of vascular injuries in each strain of rats.
Then, stroke was induced in SHRSP by 1% NaCl loading starting at 10 weeks of age. About 2/3 of them showed stroke signs with concomitant decrease in body weight, and increase in urinary volume and protein within 2-3 weeks. They were killed in 3 weeks and cerebral lesion was confirmed by autopsy. Platelet counts in stroke group was reduced to 36%, and thrombin-induced aggregation to 66% of those of healthy group. ADP and serotonin contents in platelets diminished by 45 and by 65% respectively due to stroke while plasma TBARS in stroke group was two fold of healthy group. There was a singificant inverse correlation between platelet serotonin content and plasma TBARS.
It is concluded that long duration of hypertension causes platelets degranulated and exhausted due to in vivo activation of platelets at injured arteries, thus the changes of platelet contents could be prediction of vascular injuries.

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We observed platelet aggregation induced by several kinds of aggregating agents being both added to PRP. Furthermore, we compared platelet aggregabilities induced by a mixture of epinephrine and collagen between healthy persons and patients with effort angina.
ADP or collagen-induced platelet aggregations were potentiated by epinephrine. Weak potentiation by ADP and collagen was also found in platelet aggregation. Moreover, very strong potentiation of epinephrine, ADP and collagen was detected.
A lower concentration of the mixture of epinephrine and collagen caused more obvious platelet aggregation in patients with effort angina than in healthy persons.
The results in present experiments indicate that very low concentrations of aggregating agents can easily induce platelet aggregation by potentiation especially in patients with effort angina, because many aggregating agents are present in vessel bed and circulating blood.

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The percentages of apo C-II and C-III proteins in very low density lipoprotein (VLDL) were high in the patients with hypertriglyceridemia (type IIb and IV) when compared with normal control subjects and those of apo A-I, A-II and B proteins were low. Positive correlation was shown between serum VLDL concentration and the percentage of apo C proteins, and the inverse relationship was between VLDL concentration and the percent apo B protein. Consequently, C/B and C/E ratios were higher in these patients than in healthy control.
By contrast, there were no significant difference in high density lipoprotein apoprotein composition between normal subjects and the patients with high triglyceride concentration.

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To examine the relationship of lipoprotein particle size distribution to atherosclerosis, we subfractionated serum lipoproteins by using “high-performance” liquid chromatography (HPLC). The HPLC columns were TSK-G 5000 PW+TSK-G 5000 PW. We calibrated columns by using as standards globular proteins and serum lipoproteins separated by ultracentrifugation. The particle diameter of lipoproteins were determined by electron microscopy with negative staining. The elution volume was determined by monitoring at Also. This column system was suitable for analysis of the particles with diameters from 20 to 200nm. In normal and type ha hyperlipoproteinemic subjects, elution pattern of serum lipoprotein (d<1.063 fraction) showed one peak with narrow range (20-40nm). In type IIb and IV hyperlipoproteinemic subjects, however, lipoproteins distributed wide range from 20 to 150nm. As the percentage composition of cholesterol and protein was increased, lipoprotein particle size became smaller. The percentage composition of triglyceride was decreased with lipoprotein paticle diameter. The heterogeneity of triglyceride-rich lipoprotein was more prevalent and remarkable in hypertriglyceridemic subjects than in normotriglyceridemic subjects. The HPLC method was useful for the subfractionation of serum lipoproteins, especially in hypertriglyceridemia.

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Heparin-Sepharose affinity chromatography was employed to separate serum very low density lipoprotein (VLDL) into their subclasses both in hemodialysis patients and in age or sex-mathed controls. The VLDL obtained by sequential ultracentrifugation below density 1.006g/ml and dialysed against 2mM sodium phosphate buffer containing 0.05M NaCl, pH 7.4 was applied to Heparin-Sepharose column (1.5×20cm) equibrated with 2mM sodium phosphate buffer containing 0.05M NaCl, pH 7.4, and eluted stepwise at 0.05, 0.15 and 1M NaCl. The procedure of apolipoprotein (apo) VLDL delipidated was identical exept that the buffer contained 6M urea and the elution was performed at 0.05 and 0.5M NaCl.
The VLDL sufractions were used for chemical determinations, 10% polyacrylamide-gel electrophoresis in 0.1% sodium dodecyl sulfate (SDS-PAGE) and Biogel A-15m column chromatography.
The unbound fraction at 0.05M NaCl in both VLDL and apo VLDL of hemodialysis patients was much larger than those of controls. On the other hand, the bound fraction at 0.15M NaCl was similar between two groups. These lipoprotein fractions differed in their chemical compositions, the bound fraction at 1M NaCl in the patients containing more protein and less triglyceride than that in controls. The heterogeneity of the VLDL in hemodialysis patients were comfirmed by the fact that on gelfiltration the bound fraction at 0.15 and 1M NaCl of the patients were eluted slower than those of controls although the unbound fraction at 0.05M NaCl in both groups showed similar elution patterns. In SDS-PAGE, the bound fractions contained apprecicable amounts of apo E in both groups.
These results indicate that VLDL from hemodialysis patients shows the low affinity to heparin, probably depending upon low concentrations of apo E.

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In this study, we assayed VLDL receptors on fresh human peripheral lymphocytes and monocytes and examined whether the abilities of these cells to bind (4°C) and take up (37°C) ¹²⁵I-VLDL might be affected by the divalent cations (Ca²⁺ and Mg²⁺), pH and DNase in incubation media.
1. The amount of ¹²⁵I-VLDL taken up by both cells was dramatically suppressed by calcium in a concentration of more than 260μM, but the binding to both cells was not affected by the presence of calcium.
2. Although the uptake of ¹²⁵I-VLDL by lymphocytes was dependent on magnesium with a fixed dose of calcium (109μM), the increment by magnesium of ordinary dose contained in tissue medium was within 30%. The binding of ¹²⁵I VLDL to lymphocytes was not affected by magnesium.
3. When pH exceeded 6.0, the amount of ¹²⁵I-VLDL uptake reduced markedly, and the binding to lymphocytes instead of monocytes showed a tendency to decrease.
4. DNase contained in the media (<0.5μg/ml) did not affect both the binding and the uptake.

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Low density lipoprotein (LDL) receptor has been partially purified from rat liver membrane as following. The hepatic LDL receptor was activated by five-day injection of estradiol (5mg/kg body weight). LDL receptor containing proteins were solubilized with use of detargant, n-octylglucopyranoside, from the membrane fraction, which was prepared ultracentrifugally as 8, 000g-100, 000g fraction. The solubilized membrane proteins were loaded on DE52 and eluted with an NaCl gradient containing n-octylglucopyranoside. The LDL receptor activity, which was expressed as ¹²⁵I-LDL binding activity precipitated with phosphatidylcholine-liposomes, appeared at about 100 to 150mM NaCl.
The ligand blotting of LDL receptor revealed that the molecular weight of LDL receptor of rat liver membrane is about 130, 000 and this coincident closely with that of bovine adrenal cortex.

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The exact determination of LDL cholesterol by ultracentrifugation is time-consuming and expensive procedure, whereas the Friedewald formula which can be determined LDL cholesterol using the concentration of serum triglyceride gives an imprecise value for LDL cholesterol in case of hypertriglyceridemia.
Precipitation method by heparin-citrate solution for the determination of LDL cholesterol was tested and compared to the data of ultracentrifugation and Friedewald's methods in the present study.
1) Precipitation of LDL except other serum lipoproteins was determined by the methods of immunodiffusion, electrophoresis and additional recovery examination of VLDL and LDL.
2) For the determination of LDL cholesterol, there was a good correlation between ultracentrifugation and the present methods (r=0.9695, y=1.0349x+5.4411) as well as that between ultracentrifugation and Friedewald's methods (r=0.9707, y=1.0316x-2.5561) in normolipidemia.
3) In hypertriglyceridemia exceeding 400mg/dl, a slight deteriolation was observed between ultracentrifugation and the present methods (r=0.7971, y=0.8926x-4.4083, r=0.7582, y=0.9911x-25.4388, respectively).
4) In hypertriglyceridemia above 600mg/dl below 1, 000mg/dl, the present method was superior to that of Friedewald, although there was no significant difference between two methods in hypertriglyceridemia above 400mg/dl below 600mg/dl.
5) Determination of LDL cholesterol in supernatant after addition of heparin-citrate solution was superior to that of LDL cholesterol in pellet.
From these results we conclude that the present method may be suitable for the determination of LDL cholesterol.

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Four patients on maintenance hemodialysis who have appreciable amounts of apolipoprotein (apo) B-48 in very low density lipoprotein (VLDL) and intermediate density lipoprotein (IDL) were chosed to evaluate the effect of hemodialysis itself on apo B-48 metabolism.
After 12-14hrs overnight starvation, venous blood of 15ml was drawn at predialysis and at 2hrs later during hemodialysis and separated into serum which contained 1mM EDTA and 0.01% Tobramycin.
The serum was used for chemical analyses and sequential ultracentrifugation to isolate VLDL and IDL. The detection and densitometric scanning of apolipoprotein in those lipoproteins were performed by the staining with Coomassie brilliant blue after 3.5% polyacrylamide-gel electrophoresis in 0.1% sodium dodecyl sulfate.
After hemodialysis, serum triglyceride (TG) levels dramatically decreased while serum cholesterol and phospholipid levels remained unchanged. The reduction of serum TG levels resulted in a decrease in VLDL with more protein, cholesterol and less TG on chemical composition, but not in IDL. The apo B-48 of VLDL in all cases and IDL in some cases disappeared after 2hrs hemodialysis.
These results suggest that hemodialysis itself has an effect on a removal of apo B-48 in TG-rich lipoproteins in patients on maintenance hemodialysis.

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We previously revealed that hyperinsulinism (insulin hypersecretion after glucose) develops in the young obese people during the maturation years of 10-20, and the insulin levels were shown to be normal in the obese group aged 10 years or younger. In this study, 24-hour urine insulin and C-peptide were estimated in order to judge hyperinsulinism in 28 obese young people aged 7-16 years and in 16 non-obese group aged 7-18 years. There was significant positive correlation between age and urine insulin (r=0.687, p<0.001) or between age and urine C-peptide (r=0.488, p<0.02) in the 28 obese people.
Both plasma triglyceride and total cholesterol (TC) increased slightly but significantly in the obese group aged 10 years or more as compared with the obese group aged 9 years or younger and the nonobese people. Plasma HDL-cholesterol (HDL-C) and apoproteins B and A-I showed a considerable but insignificant change in the young obese people aged more than 10 years as compared with other groups. Thus, the atherogenic cholesterol index ((TC—HDL-C)/HDL-C) and atherogenic apoprotein index (apoproteins B/A-I) were shown to be increased with age. There was a significant positive correlation between urine insulin and the cholesterol index (r=0.406, p<0.01) or between urine insulin and the apoprotein index (r=0.314, p<0.05).
Therefore, during the sex-maturation, atherogenic dyslipoproteinemia seems to develop in parallel with insulin levels.

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Macrophages may play an important role in the atherogenic process because they appear to be precursors of some arterial wall foam cells. It has been reported that human peripheral monocytes in culture synthesize and secrete lipoprotein lipase (LPL).
1) The present studies show that human monocytes and guinea pig alveolar macrophages in culture synthesize and secrete LPL. The rate of secretion from human monocytes was low initially but increased with time in culture. During 14 days in culture, maximal activity was attained after 12 days. On the other hand, Guinea pig alveolar macrophages increased from the beginning. During 8 days in culture, maximal activity was attamed after 4 days.
2) We investigated the erect lipoprotein on LPL activity of guinea pig alveolar macrophages. Alvolar macrophages were cultured for 2 days with either RPMI-1640 containing 10% lipoprotein deficient serum (LPDS) or that medium with the addition of VLDL, LDL and HDL respectively in the concentrations of 300μg protein/ml. When VLDL was added to LPDS after one day, LPL activity was significantly reduced to 48.3% within 2 days of exposure when compared with LPDS controls (p<0.02).

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Blood stasis occurred as the result of sclerotic changes in the arteries accelerates the formation of ischemic lesions with respect to decrease in blood supply to the tissues. The rheological properties of blood and its components play an important role in the physiology of blood flow, and the augmentation of blood fluidity elicited by improvement in the properties of plasma and corpuscular components prevents tissues from thrombotic diseases even in the aged with atherosclerotic changes. The alterations in the structure of the erythrocyte membrane which is mainly composed of lipids and proteins give influence on erythrocyte deformability. Mature erythrocyte is not capable of the de novo synthesis of cholesterol. One of the notable features of blood cholesterol is ability to move between erythrocyte membrane and plasma under regulation by LCAT. This study was intended to clarify the participation of LCAT in blood fluidity disclosed by the measurement of deformability index in connection with the observations of the interaction of lipids between plasma and erythrocyte membrane.
Fasting blood specimens were drawn from 24 healthy persons with the age of 20 to 23 years. Deformability index was measured by the method of Reid et al. Erythrocyte membrane was separated by the method of Dodge et al. for the analysis of free cholesterol, phospholipids and total protein. Free cholesterol (FC) and phospholipids (PL) were determined by enzymatic methods, total protein (TP) by Lowry's method. The fractionation of phospholipids was carried out by thin-layer chromatography following the extraction of lipid by Folch's solution. Sum of phosphatidylcholine (PC) and sphingomyeline (SM) in the outer layer, and the value of phosphatidylethanolamine (PE) and phosphatidylserine (PS) in the inner layer of the membrane were calculated. Then, the ratio of PC+SM to PE+PS (PC+SM/PE+PS) was investigated in order to clarify the interactions of lipids in the membrane. LCAT activity was assessed by Nagasaki and Akanuma's method.
The properties of blood in subjects were tabulated in Table 1. The ratio of FC to PL (FC/PL) in plasma was directly proportional to LCAT activity, FC/PC in erythrocyte membrane was directly to the ratio in plasma. FC in the membrane was inversely proportional to PC+SM/PE+PS and FC inversely to TP. Deformability index was directly proportional to membrane TP, inversely to TP/FC+PL, and LCAT activity.
Results obtained in this study by observations of the influence on deformability index indicated close relations of plasma cholesterol, phospholipids and LCAT to hemorheological behavior. The intimate association of LCAT with blood fluidity was confirmed by direct connection of this enzyme with deformability index in addition to the indirect participation through the effect on FC/PL in plasma which affects the ratio in membrane. Although the important role of LCAT in lipid metabolism has been established, there have been conflicting data dealing with participation of this enzyme in the myocardial infarction which is greatly affected by abnormal lipid metabolism. Some authors have refered to severe atherosclerotic changes in not all familiar LCAT deficiency cases and have illustrated a little association of LCAT with occurrence of atherosclerotis in abnormal lipid metabolism. No articles have been found evidence that contradict the significance of high viscosity and low filtrability observed in blood of patients with myocardial infarction. Therefore results obtained in this study emphasize the necessity of versatile observations from the view point of hemorheology in addition to lipid metabolism for investigation of role of LCAT in atherosclerotic diseases.

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The effect of phosphatidylcholine liposomes on the activity of lysosomal acid lipase purified from rabbit liver was studied.
The activity using 4-methylumbelliferyl oleate as substrate was enhanced in the presence of phosphatidylcholine liposomes. Both acyl chain length and degree of unsaturation of phosphatidylcholine were involved in the activation. In a series of cholesterol-saturated phosphatidylcholine liposomes, relative enzyme activity was decreased in parallel with the ratio of cholesterol to phosphatidylcholine. The relative activity, however, increased in a series of cholesterol-unsaturated phosphatidylcholine liposomes.
If one thinks about the abnormal lipid composition of atheromatous lysosomal membranes (for example, content of saturated fatty acid in phospholipid and free cholesterol was increased in this fraction), it may be possible that lysosomal acid lipase in atheromatous lesion cannot sufficiently hydrolyze the substrate, such as cholesterol ester.

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The present studies was to investigated the effects of the elastase like enzyme on the plasma LCAT activity. Rat plasma LCAT activity was inhibited by the addition of the elastase like enzyme (100units) in the reaction mixture, and increasing of the free cholesterol was obserbed. The inhibitor of elastase like enzyme repressed the increasing of free cholesterol in the reaction mixture. Inhibition of LCAT activity was due to digestion of albumin, apoprotein A, C or D which stabilizing factor of LCAT by elastase like enzyme. Also, we recongized the existance of cholesterol esterase activity in rat plasma. Cholesterol esterase activity in the reaction mixture was not inhibited by elastase like enzyme. Therefore, free cholesterol increased in the reaction mixture.

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We have studied the effects of elastase like enzyme on the lipid metabolism in rat. 660units/kg/day of elastase like enzyme were injected for 13 weeks. The lipids of plasma, lipoprotein and liver lipids were decreased by elastase like enzyme.
In apo-lipoprotein, apo-E of VLDL decreased, but on the other hand the increasing of apo-E of HDL was obserbed. Lecithin: Cholesterol acyltransferase activity was decreased about 20%. Although LCAT activity decreased, but also decreasing of plasma free cholesterol was suggested that the disappearance of plasma free cholesterol was increased. Significant decrease were obserbed in lipids of liver as same as plasma lipids. Liver cholesterol ester was decreased about 65%. Acid cholesterol esterase and AcylcoA: Cholesterol acyltransferase activity were decreased by elastase like enzyme treatment. However, neutral cholesterol esterase increased about 2 times. The ratio of LDL-c to acid cholesterol esterase activity was about 1/2, and the ratio of ACAT activity to Neutral cholesterol esterase activity was about 1/10 by elastase like enzyme treatment. Cholesterol 7α-hydroxylase activity raised 4 times. These findings suggest that the metabolism of lipoprotein might be accelerated in plasma, that accumulation of cholesterol ester might be repressed in liver, and free cholesterol of liver was carried away through the lipoprotein of bile acid by elastase like enzyme.

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For the purpose of estimating physiological role of elastase, elastase activity in serum in various diseases was determined using purified elastin as the natural substrate with fluorophotometric method. NaCl was used for the discriminating assay method of the pancreatic and leukocytic elastase activities, and then the elastase activities of different origins in variuos physiological conditions were estimated.
Separation and purification of activities of serum elastase, leukocyte elastase and pancreatic elastase on CM-Sephadex column chromatography and by chromatofocusing technique revealed that there are two types of elastase activities: that is, leukocyte type and pancreatic type elastase in human serum.
It was found in this study that the leukocyte elastase activity is activated by 0.5M NaCl while pancreatic elastase activity is suppressed by 0.5M NaCl.
Effect of NaCl on the elastase activities of different origins was applied to know the ratio of leukocyte type and pancreatic type elastase activities in serum. It was found as the result that the total elastase activity in serum in patients with hypothyroidism was so low as compared with that in patients with hyperthyroidism. More leukocyte type elastase activity was involved in patients with hypothyroidism. In the senile group, total serum elastase was low as compared with that in the young group, and less leukocyte type elastase activity was involved.
It may be said from these experiments that the serum elastase is essential for the normal metabolism of the connective tissues. Elastase activity in serum is so variable related to age, disease and other factors that it may be important to estimated the serum elastase activity and the ratio of leukocyte type to pancreas type elastase activities in order to evaluate the condition of the connective tissues in connection with diseases.

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Effect of soybean crude fiber on the serum concentrations of lipids and apolipoproteins in normolipemic and hyperlipemic subjects were examined, the concentrations of serum triglycerides, VLDL-triglycerides, VLDL-cholesterol and apo C-II were decreased singificantly following administration of soybean crude fiber for 2 months. No significant changes were found in total cholesterol, LDL-cholesterol, HDL-cholesterol, apo A-I, A-II and B levels. Also, no significant changes were observed in weight before and after administration of soybean crude fiber.

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Effects of mild exercise therapy on serum lipids (TC, TG, PL and HDL-C) and apoproteins (Apo A-I, A-II, B, C-II, C-III and E) were examined in 25male patients with hyperlipidemia (n=16), coronary artery disease (n=11) and hypertension (n=11). Mild exercise program consisted of the exercise intensity of WBPLA₁, 60minutes/time and 3times/week for 10 weeks. Serum levels of HDL-C and total cholesterol increased following exercise therapy (p<0.1) and the changes were significantly correlated (r=0.52, p<0.02). Serum levels of phospholipid and Apo E were significantly elevated following exercise therapy (p<0.001, p<0.05). There were no changes in TG and apoproteins (A-I, A-II, B, C-II and C-III). Those who had higher levels of TG and lower levels of HDL-C before exercise therapy showed a little increase in HDL-C levels following exercise therapy compared to those of lower TG and higher HDL-C levels. Patients with hypertension showed a little increase in HDL-C levels following exercise therapy compared to those without. There were significant correlation between changes of HDL-C and Apo A-I following exercise therapy (r=0.42, p<0.04).

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A method determining the lipolytic reactivity of plasma very low density lipoprotein (VLDL) with use of purified milk lipoprotein lipase is presented for studying the triglyceride clearing from the vascular compartment and the mechanisms of hypertriglyceridemias.
A significant inverse relationship between plasma triglyceride levels and VLDL lipolytic reactivities was observed in subjects with hypertriglyceridemia.
There were an apparent increase in percentage of triglyceride and a decrease in percentage of protein in VLDL particles with increasing triglyceridemia.
The relation between VLDL lipolytic reactivities and percentage compositions of apolipoprotein C-II, a cofactor for lipoprotein lipase, showed no significant correlation, whereas a significant negative correlation was found between VLDL lipolytic reactivities and percentage compositions of apolipoprotein C-III, an inhibitor for apolipoprotein C-II-activated lipoprotein lipase, in the cases with higher concentrations exceed 1.2% of apolipoprotein C-III in VLDL.
These results suggest that low lipolytic reactivity of VLDL may be one of the causes of hypertriglyceridemias.

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We studied the effects of single oral aspirin ingestion (500mg/day, 1 day) on ADP-, collageninduced platelet aggregation and plasma β-thromboglobulin (β-TG) for 14 days in healthy and diabetic subjects. The difference of mean serum salicylic acid, before and 6hrs, 24hrs after ingestion, between these two groups were not significant. Maximal inhibitory effect in ADP-induced platelet aggregation was observed at 6hrs in both groups to a same extent. Mean plasma β-TG in diabetic subjects was significantly (61.2±13.1ng/ml, Mean±SEM) higher than in healthy subjects (17.6±2.1ng/ml). After ingestion, mean plasma β-TG in diabetic subjects elevated to 179.8±42.9ng/ml at 6hrs, and varied from 44.2ng/ml to 78.2ng/ml, so it indicated 105.6±23.0ng/ml at 14 days. Mean platelet count were not different between two groups. These results might suggest that single oral aspirin ingestion every 3 or 4 days is useful in its antiaggregatory effect for antiplatelet therapy.

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Antisclerotic effect of clinofibrate (lipoclin®) was studied referring to rise and fall of aortic pulse wave velocity (PWV) and observation of aortic wall constituents of rabbits with experimental arteriosclerosis.
Twenty-seven male rabbits of 38 months were divided into the healthy group (H group) of 13 rabbits, the sclerotic group (S group) of 9, and the clinofibrate (200mg/day) administration group (A group) of 5 respectively. Arteriosclerosis was produced by contaminant use of three method, namely, hypoxic method (N₂ gas inhalation), tissue injury method (intramuscular injection of norepinephrine) and intimal lesion formation and vaso vasorum obstruction (administration of 0.5g cholesterol) for S and A group. PWV and blood pressure measured every two weeks to see their rise and fall. After 16 weeks all the rabbits were killed and 6 components in media of thoracic aorta, namely, smooth muscle cell (SMC), elastin (EL), collagen (CL), acid mucopolysaccharides (AMPS), glycoprotein (GP) and calcium phosphate (Ca) were observed and assayed histochemically by microspectrophotometric procedure. Furthermore, a principal component analysis was conducted to generally evaluate differences among those 3 groups as to changes of those 5 components.
When changes of PWV (ΔPWV) on an average of 4 weeks were studied, no change was observed in the H group, but in the S group, it increased about 1.0m/sec after the 10th week in S group and it decreased about 1.0m/sec after 4 weeks in the A group. Upon comparison between the S and A groups, after 4 weeks with the level of significance at 1%, a significant difference was noted, the effect of which lasted until the 16th week.
Upon histochemical observation, atrophy, disappearance and decrease of SMC, fragmentation and disappearance of EL and CL, uneven distribution and increase of AMPS and GP, and increase of calcium phosphate were noted in the S group, while in the A group either component received only mild disturbances. Amount of the components with the H group, S group and A group respectively were as follows: SMC: 40.6±6.6, 32.6±4.4, 35.7±4.7%E, EL: 52.8±4.3, 42.3±2.6, 46.3±4.3%E, Cl: 38.3±4.0, 33.3±3.7, 39.0±3.3%E, AMPS: 28.4±6.6, 35.7±5.4, 34.2±3.4%E, GP: 33.2±3.7, 39.7±3.7, 36.9±3.5%E. Upon principal component analysis and a study of their distribution on the quadratic coordinates in comparison of those 3 groups, it was suggested that distribution of the A group being near the region of the H group. It was suggested that clinofibrate inhibits a organic arteriosclerotic lesion.

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To elucidate the clinical significance of hyper-HDL-cholesterolemia, the subjects with HDL-cholesterol above 100mg/dl were studied.
Among 22, 876 persons (male 17, 657, female 5, 219) who visited Keio Health Councelling Center for their health check from January 1981 to October, 1984, we found 177 subjects whose HDL-cholesterol was above 100mg/dl. The incidence was 0.8% for the total (0.5% for male, 1.6% for female). Their percent body weight deviation averaged as -5%, while their blood pressure, liver function tests, fasting blood sugar, and uric acid were within the normal range. Their total cholesterol was 226mg/dl, triglycerides 70mg/dl, HDL-cholesterol 109mg/dl. Their average alcohol intake was 18g/day for the total (28g/day for male, 7g/day for female). The most common diseases in their past history were appendicitis and gastroduodenal ulcers.
These indicated that the subjects with hyper-HDL-cholesterolemia were slim in physical shape with occasional gastro-intestinal disorders and hypo- or normolipidemias. It remains to be studied whether this type of subjects has a close relation to some particular ilness in their adulthood or in their senescense.

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Nine-hundred-ninety-eight obese children aged 6 to 15 years participated in the present study. Fasting serum lipids and lipoproteins cholesterol values were measured and the relationships between obesity and abnormalities of serum lipids and lipoproteins were assessed.
The results were as follows.
1. Obese children were much more likely than non-obese children to have elevated serum cholesterol, triglyceride and LDL-C (low density lipoprotein cholesterol) levels and reduced HDL-C (high density lipoprotein cholesterol) levels.
2. The more the relative body weight index increased, the worse the abnormalities of serum lipids and lipoproteins became. This tendency was marked in junior high school boys.
3. Hypercholesterolemia in obese children was mostly accounted for by LDL-C only and elevated HDL-C which accounted for their hypercholesterolemia could not be detected.

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