In summary, it was shown that platelets enhance macrophage cholesteryl ester accumulation. We have demonstrated that activation of the platelet was required, that the macrophage response was specific for platelets and that it may be mediated or enhanced by specific platelet/macrophage interactions. The macrophage response to platelets involved the release of cholesterol-rich microvesicles from platelets that have a molecular weight>300, 000. Finally we have shown that platelets do not stimulate macrophage endogenous cholesterol synthesis and that macrophage lipoprotein receptors were not involved. Therefore, platelets may play a contributing role to the development of advanced fibrous plaques from fatty streak lesions and may be important with respect to the early stages of atherosclerosis.
I would like to acknowledge the efforts of my contributors: Dr. Cheryl A. Dyer, Dr. Carole L. Banka, Dr. Edward F. Plow, Ms. Audrey Black and Dr. Yasushi Takagi.

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The effects of physical exercise on the concentrations of serum lipoproteins and apolipoprotein, particularly on the composition of high-density lipoprotein, were investigated in this study. Our subjects consisted of 9 weightlifters (mean age 21 years), 12 endurance runners (mean age 21 years), and 10 age-matched untrained students as a control group.
Apolipoprotein levels were measured by a single radial immunodiffusion method. Endurance runners showed lower LDL cholesterol values (80mg/dl) and higher level of HDL cholesterol (73mg/dl) than did the control group (107, 46mg/dl respectively). We found no differences in serum HDL cholesterol levels between weightlifters (45mg/dl) and the control group. Serum apoprotein A-I concentrations and the Apo A-I/A-II ratio were higher in top athletes than in untrained men.
Our results show that vigorous physical exercise may influence serum lipid and apoprotein levels independently.

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Kanazawa et al. and Muraoka et al, reported that cholesterol concentrations of ultra-water-soluble LDLs (UWS-LDL) separated by dialysis from tap-water were higher in cerebral infarction or ischemic heart disease patients than in healthy persons.
In this experiment, internalization into macrophages and cell injury caused by UWS-LDLs obtained from patients with arteriosclerotic disease were investigated.
1) The internalization rates of UWS-LDLs such as proteins, cholesterol esters (CE), triglycerides (TG), free fatty acids (FFA) and free cholesterol (FC) into macrophages were greater than those of native LDL.
2) The internalization rate of UWS-LDLs such as CE and FC into macrophages was enhanced by adding heparin to the medium, although no heparin-related differences were found for protein, TG and FFA.
Thus, we concluded that the metabolic pathway as well as the phagocytic pathway might be the mechanism of internalization of UWS-LDLs into macrophages.
3) The scanning electron microscope findings of macrophages did not change after incubation with native LDLs and macrophages at 37°C. However, after 2 hours of incubating UWS-LDLs and macrophages, cell swelling and rough change of the cell centrum due to internalization of UWS-LDLs were observed. After 6 hours, cell injury was more advanced and the central part of the cells were destroyed. Finally, floating cells were observed in the medium.
Conclusion:
The concentration of UWS-LDL-cholesterol was closely related to the incidence of arteriosclerotic disease. We also confirmed that UWS-LDLs were easily internalized into macrophages and that they injured the cells.
Thus, we feel that this study on UWS-LDLs was very important to clarify the causes of arteriosclerosis.

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Bifemelane hydrochloride is a new substance which exhibits antianoxic and memory retrieval effects in cases of ischemic cerebral damage. Recent clinical trials have also revealed favorable effects on vascular dementia.
This study was designed to comfirm neuropathologically the effects of this substance on ischemic changes in the brains of Mongolian gerbils caused by ligated left carotid arteries. Two-hundred 8-week-old Mongolian gerbils were divided into 4 groups of 50 animals each. Bifemelane hydrochloride was administered intraperitoneally to the animals in 1.0% physiological saline at doses of 25mg/kg body weight. Bifemelane hydrochloride was given to group II 30 minutes before, and twice to group III 30 minutes before and after the ligation procedure. Group I served as control. This group was administered intraperitoneally the same volume of physiological saline as bifemelane hydrochloride solution used for animals with the same body weight. Group IV was prepared for an observation of vascular permeability after administering bifemelane hydrochloride by intravenous injection of Evans blue. All the animals were sacrificed 3 hours after the ligation procedure by perfusion fixation and were observed with a microscope.
The survival rates and the neurological deficits did not vary significantly between groups. The incidence of prominent neurological changes in the brains of group I, II and III were 62, 54 and 54% respectively. Group I displayed a generalized reduction of stainability and widely dispersed severe ischemic lesions with a spongy state and edema, especially, in white matter and the caudate nucleus. These morphological changes expanded out from periventricular areas, and were demarcated from deeply stained normal tissues.
Hippocampus samples from group I subjects showed decreased neuronal cells, vacuolar degeneration of these cells and irregular dendritic trees. Electron microscopic observations of group I specimens revealed degeneration of endothelial cells, intracellular edema, and swelling of neuronal mitochondria. Histopathological changes of group II and III subjects administered bifemelane hydrochloride were generally not serious, and boundary areas of the ischemic lesions of these subjects appeared to be intermingled with normal histological figures. Electron microscopic examination of group II and III subjects revealed that a considerable amount of organelles, such as mitochondria, microtubules, synaptic vesicles, and membranes of neuronal cells, kept their ordinal structures in the intermingling state. Evans blue transdated slightly to ischemic lesions of the animals administered bifemelane hydrochloride. Neuropathological changes observed in group I were somewhat similar to those in the brains of patients with vascular dementia; therefore, slight changes manifested in groups II and III should be regarded as evidence suggesting the utility of bifemelane hydrochloride in treating vascular dementia.

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We compared the long-term effects of pravastatin (CS-514) (CS), probucol (PB) and cholestyramine (CH) on serum lipids and regression of Achilles tendon xanthomas in 66 heterozygous patients with familial hypercholesterolemia (FH).
During the single drug regimen, treatment with CS proved to be the most effective in reducing LDL-cholesterol (LDL-C) (CS;-27±9%, PB; -19±8%, CH; -22±10%). Combined use of two of these drugs produced an additive effect. This effect was especially strong when combining CS and PB (CS+PB; -41±9%, CS+CH; -38±9%, PB+CH; -28±14%). By using the three drugs together, LDL-C was reduced much more than with the other regimen (-48±2%) and, consequently, serum lipid levels were normalized.
Single drug treatment caused substantial xanthoma regression which was approximately equal among the three drugs (CS; -5.8%, PB; -4.0% and CH; -5.2%). Furthermore, the regression was greater with combined treatment (CS+PB; -9.6%, CS+CH; -7.1% and PB+CH; -8.0%). The extent of xanthoma regression was most closely correlated with reduction of LDL-C and duration of treatment (r=0.45, p<0.02). Moreover, in patients treated with PB, there was a significant correlation between maximum reduction of HDL-C and the extent of xanthoma regression (r=0.59, p<0.05).
We concluded that intensive drug treatment using a combination of those three drugs, which have different lipid-lowering mechanisms, is useful in heterozygous FH.

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A large part of serum lipid peroxides (LPO) are believed to be carried in lipoproteins. It has been reported that serum LPO levels increase at the time of vascular accidents. Little is known about the distribution of LPOs in lipoproteins, however, or the relationship between LPOs and thrombus formation.
We studied LPOs in oxidatively modified lipoproteins and their effect on platelet aggregation. Lipoproteins were isolated from 20 patients with arteriosclerotic diseases. The lipoproteins were oxidized by dialysis against tap water and against water containing Cu⁺⁺. LPO was measured using a thiobarbituric acid reacting substance (TBARS) assay. Platelet aggregation was measured by the Born method, and lipid constituents of the lipoproteins were analyzed using thin layer chromatography (TLC).
1) TBARS levels of very low density lipoproteins (VLDL), low density lipoproteins (LDL) and high density lipoproteins (HDL) increased after dialysis. After tap water dialysis, the profile of lipid peroxide levels of LDL was almost the same as that of VLDL, and much higher than that of HDL. After dialysis against Cu++, lipid peroxide in the LDLs was higher than that in the VLDLs and much higher than that in the HDLs.
2) Tap-water-dialyzed lipoproteins which were preincubated with platelet-rich plasma potentiated adenosine diphosphate (ADP)-induced platelet aggregation. Among tap water dialyzed lipoproteins with the same LPO levels, LDLs had the strongest potentiation affect.
3) Using TLC, new spots were detected between triglycerides (TG) and free fatty acids (FFA) and between FFA and free cholesterol (FC) in tap-water-dialyzed lipoproteins. These spots were not detected when ethylenediamine tetraacetate or butylated hydroxytoluene was added to the dialysate. The spot between TG and FFA was detected only with oxidized LDLs and HDLs. Spots of the same rate of flow as these were identified in oxidized products of cholesterol esters in native LDLs, but they were not detected in oxidized products of other lipid constituents.These spots were made of oxidized products of cholesteryl linoleate.
Our data suggest the following: 1) oxidized lipoproteins have different biological activities; 2) the oxidized cholesteryl linoleate product plays an important role in the potentiation of platelet aggregation by oxidatively modified LDLs.

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The purpose of this study is to investigate whether MgCl₂ supplement can influence blood pressure and prevent hypertensive vascular lesions. Twelve female stroke-prone SHR (SHRSP), 6 female stroke-resistant SHR (SHRSR) and 5 female Wistar Kyoto rats (WKY), all aged 2 months were used. In these rats 6 female SHRSP were given 2% solution of MgCl₂ for 17 months. As the control, 6 SHRSP, 6 SHRSR and 5 WKY were given tap water for 17 months. After anesthesia with intraperitoneal infusion of 10% urethan solution, blood samples were taken, and then rat mesenteric arteries were removed and microscopically observed with H-E staining and also for electron microscopic observation.
Rat body weights on the average were heaviest in WKY and lightest in SHRSP, but did not differ significantly in SHRSP between 2% MgCl₂ solution and tap water groups. Systolic blood pressure levels were highest in SHRSP and lowest in WKY, but did not differ in SHRSP between 2% MgCl₂ solution and tap water groups. Heart weight per 100g of body weight and kidney weight per 100g of body weight had the same tendencies as the systolic blood pressure levels.
Macroscopic findings of the mesenteric arteries were bead-like lesions in 5 of 6 SHRSP fed on tap water, while these changes were not shown in SHRSP fed on MgCl₂ solution or in SHRSR and WKY fed on tap water. Microscopic findings of the mesenteric arteries were periarteritis nodosa in 4 of 5 SHRSP fed on tap water., while no such lesion was found in the other rats. Electron microscopic findings of the mesenteric arteries showed slight proliferations of endothelial cells, subendothelial accumulation of fibrin and destroyed internal elastic membranes.
Serum Mg increased significantly in SHRSP fed on 2% MgCl₂ solution as compared to the other groups. Serum cholesterol, fructosamine and creatinine values did not show significant differences in SHRSP between 2 % MgCl₂ solution and tap water groups. From these findings we concluded that lesions of mesenteric arteries were not related to systolic blood pressure, serum cholesterol level, disturbed glucose metabolism or impaired renal function. Some other mechanism of MgCl₂ against hypertensive mesenteric arterial lesions is suggested.

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We examined age-related changes of serum lipids and their distribution in each lipoprotein fraction in normolipidemic healthy adults (182 males, 191 females) including 59 octogenarians and 23 nonagenarians. Lipid distribution was analyzed using the agarose gel electrophoresis-enzymatic staining method. The relationship between the lipoproteins, postheparin lipolytic activity and sex hormones was examined in 76 randomly chosen subjects.
The total cholesterol (TC) levels peaked in the 7th decade in males (190mg/dl) and in the 6th decade in females (203mg/dl). It then decreased gradually in both groups. The triglyceride (TG) levels in males showed no changes except for a slight decline in the 10th decade. The TG levels in females increased with age and exceeded those of males after age 70.
The TG distribution showed characteristic changes after age 80. In males, VLDL-TG (%) remained at about 60% for ages 20-59, then decreased to 36.8% at the 9th decade and 25.6 at the 10th decade. LDL-TG (%) was nearly 30% for ages 20-59, and then increased to 50.2% at the 9th decade and 63.5% at the 10th decade. In females, the changes of VLDL-TG (%) and LDL-TG (%) were similar to those in males, but the degree of change was smaller.
Lipoprotein lipase activities of postheparin plasma did not change remarkably in either sex. Hepatic triglyceride lipase (H-TGL) activites in males were constant at about 8μmols FFA/ml/hr for ages 20-59. They then decreased remarkably to 3.3μmoles after age 80. In females, the activities also decreased after age 60. H-TGL activities showed a positive correlation to VLDL-TG (%) (male: r=0.679, p<0.001, female: r=0.323, p<0.05) and a negative correlation to LDL-TG (%) (male :r=-0.574, p<0.001, female: r=-0.347, p<0.05).
The levels of DHEA-S decreased steadily with age in both sexes, and were very low after age 80. DHEA-S showed a significant correlation to VLDL-TG (%) (r=0.516, p<0.01), LDL-TG (%) (r=-0.501, p<0.01) and H-TGL activities (r=0.596, p<0.001) in males.
In conclusion, the increase in LDL-TG (%) and the decrease in VLDL-TG (%) detected after age 80 were characteristic age-related changes in the distribution of serum lipids. We suspect that these changes resulted from the stagnation of the remnant derived from TG-rich lipoproteins, and that the stagnation was affected by the decline of H-TGL activities and DHEA-S levels.

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Abnormal lipid metabolism has not yet been established as a risk factor of cerebral infarction. Serum lipid levels (Total cholesterol, TC; Triglyceride, TG; HDL cholesterol, HDLC; LDL cholesterol, LDLC) and apolipoproteins (apo A-I, B, E) were measured in 127 normal healthy subjects (N), 31 patients with normolipidemic hypertension (HT), 20 patients with normolipidemic diabetes mellitus (DM), and 71 patients with hyperlipidemia (HL). Ages ranged from 20 to 78 years (46.0±12.3).
Our results were as follows.
1. In N subjects, apo B and the B/A-I ratio were elevated, and the LDLC/apo B ratio was reduced with age.
2. TG and apo B were significantly elevated, and the LDLC/apo B ratio was significantly lower in the group with a family history of cerebrovascular disease (CVD).
3. In HT patients, apo B was significantly higher than in N subjects, and the LDLC/apo B ratio was significantly lower.
4. HDLC was significantly lower in DM patients than in N subjects.
5. LDLC, the Atherogenic Index (TC-HDLC/HDLC), apo B, apo E and the apo B/A-I ratio were all significantly elevated, and HDLC was significantly decreased in HL patients.
This data suggests that apo B and the LDLC/apo B ratio are useful indicators for the development of cerebral infarction.

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