This study investigated the effect of atrial natriuretic peptide (ANP) on endothelin (ET) secretion from cultured human endothelial cells. Confluent umbilical venous endothelial cells were incubated with experimental agents in multi-well plates, and the level of immuno-reactive ET in the medium was measured by radioimmunoassay. There was no significant effect of ANP (10^-8, 10^-7 and 10^-6 M) on ET secretion after a 3-or 6-hour incubation. However, with 24-hour incubation, ANP significantly inhibited ET secretion from cultured human endothelial cells (control, 139.0± 7.2 fmol/well ; 10^-8 M, 89.4 4.7 fmol/well ; 10^-7 M, 79.4 8.2 fmol/well ; 10^-6 M, 71.0 ±10.1 fmol/well, P< 0.01). Furthermore, the addition of 8-bromo-cyclic GMP to the medium inhibited ET secretion (control, 147.2 ± 2.9 fmol/ well ; 10^-5 M, 140.9 ± 2.3 fmol/well ; 10^-4 M, 143.0±1.0 fmol/well ; 10^-3 M, 96.6± 6.3 fmol/ well, P< 0.01). These findings demonstrate that ANP inhibits accelerated ET secretion from cultured human endothelial cells, probably due to augmentation of intracellular cyclic GMP levels by ANP-activated guanylate cyclase. J Atheroscler Thromb, 1994 ; 1 : 76-79.

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We report evidence for the presence of 7α- and 7β-hydroperoxycholest-5-en-3β-ols (cholesterol 7-hydroperoxides, Ch 7α-OOH and Ch 7β-OOH, respectively) in human plasma lipoproteins in vivo, which had been reported to be markers of aging in rat skin. A comparative study was carried out focusing on the detection of Ch 7-OOHs in plasma lipoproteins from diabetic patients whose plasma has been suggested to be under high oxidative stress. Blood samples were collected from healthy volunteers (control) and diabetics with and without hypercholesterolemia. Ch 7-OOHs were isolated from low and high density lipoproteins (LDL and HDL, respectively) in the plasma of these subjects, identified, and determined by high-performance liquid chromatography with a chemiluminescence detector. The percent detection of Ch 7-OOHs in LDL from diabetics without hypercholesterolemia was similar to that in the control group. However, it was significantly higher in diabetics with hypercholesterolemia than in those without hypercholesterolemia.The percent detection of Ch 7-OOHs in HDL from diabetics without hypercholesterolemia was higher than both that in LDL from the same group and that in HDL from the control group. J Atheroscler Thromb, 1994 ; 1 : 80-86.

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Oxidation of low density lipoproteins (LDL) has been shown to lead to enhanced uptake by macrophages mediated by the scavenger receptor. In the present study, changes in LDL induced by copper-catalyzed oxidation were investigated using gel permeation chromatography (GPC), and the results were compared with several parameters of oxidized LDL (ox-LDL). When LDL at 200 μg/ml was oxidized with 10 μ M Cu²⁺ at 37°C for up to 24 hours, increases in thiobarbituric acid-reactive substances and electrophoretic mobility were first observed within 3 hours. An increase in fluorescence and a decrease in intact apolipoprotein B (apoB) were then observed in parallel with an increase in ¹²⁵I-LDL degradation by macrophages after 6 hours. Finally, LDL aggregation separated by liquid chromatography was observed after 24 hours. The aggregated and monomeric fractions of ox-LDL were analyzed and the results compared with the monomeric fraction of native LDL. Both fractions of ox-LDL contained hardly any intact apoB and showed an intense fluorescence. The electrophoretic mobility increment of aggregated ox-LDL was almost half that of monomeric ox-LDL, yet the lysine residues of aggregated ox-LDL were more extensively decreased than those of monomeric ox-LDL. Degradation of aggregated ox-LDL by macrophages showed a slightly greater increase than that of monomeric ox-LDL. GPC analysis is a useful method to estimate the LDL aggregation, and these results provide a basis to investigate the formation of aggregated LDL. J Atheroscler Thromb, 1994 ; 1 : 87-97.

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To investigate whether the remnant like particles (RLP), separated from serum by an immunoaffinity gel mixture of anti-apo B-100 and apo A-I monoclonal antibodies, are relevant to the initiation or progression of atherosclerosis, the incorporation of RLP into mouse macrophages was studied using histochemical and biochemical techniques. Remnant lipoproteins such as RLP are reported to contain a large quantity of chyloniron and very low density lipoprotein (VLDL) remnants, especially in diabetic patients. The RLP separated from the sera of 32 diabetic patients were found to be predominantly taken up into macrophages harvested from mouse abdominal cavities by the staining method applying oil red 0. Furthermore, using ¹⁴C-oleate to prove the uptake of lipoproteins by macrophages, the uptake of RLP-VLDL, a VLDL fraction of RLP by ultracentrifugation, was the next highest to that of the oxidized LDL, which suggests that RLP-VLDL is also aggressively taken up by macrophages. The degree of uptake of RLP-VLDL by macrophages was positively correlated with HbA_1c of these diabetic patients (r = 0.556, p < 0.01), irrespective of the ways of the treatment of diabetes. In conclusion, RLP can contribute to the foaming of macrophages, which in turn may explain the acceleration of atherosclerosis in diabetic patients. J Atheroscler Thromb, 1994 ; 1 : 98-102.

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The short-term effects of low-density lipoprotein (LDL) apheresis using a dextran sulfate cellulose (DSC) column equipped with a plasma separator using a polysulfone (PS) membrane filter on the serum total cholesterol, lipoprotein (a) (Lp (a)), C4b binding protein (C4bp), protein C and protein S and complement components levels were examined in a patient with familial hypercholesterolemia (heterozygote, type lla). PS/DSC-LDL apheresis markedly lowered the total cholesterol by 69.9 ± 2.9%, serum Lp (a) level by 80.2±3.4%, and C4bp by 81.1± 2.5% after 8 apheresis sessions. All the above parameters gradually returned to the baseline levels within 10 days. The free protein S level was not significantly changed, while the C4bp/protein S complex level was markedly decreased relative to the decrease in C4bp. The protein C level was moderately reduced by 29%. Almost all serum complements and complement co-factors as well as C4bp were moderately to markedly decreased after LDL apheresis, but returned to the initial levels within a few days. PS/DSC-LDL apheresis effectively eliminated both serum LDL and Lp (a), and did not have an adverse effect on fibrinolysis or complement cascade in the blood. J Atheroscler Thromb, 1994 ; 1 : 103-107.

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To evaluate platelet activity in patients with non-insulin-dependent diabetes mellitus (NIDDM), we measured the mean platelet volume (MPV) and 24-hour urinary excretion of 11-dehydro-thromboxane B₂ (11-dTXB₂) and 6-keto-prostaglandin F_1α (6-kPGF_1α), stable metabolites of thromboxane A₂ and prostacyclin, respectively. The MPV of the 103 subjects in the NIDDM group were 10.72 ±0.82 fl for males and 10.52 ±1.01 fl for females (mean ± SD), significantly higher than those of normal controls (9.95 ± 0.75 fl for males and 9.84 ± 0.72 fI for females). The MPV of patients with NIDDM showed positive correlations with fasting plasma glucose level and HbA1c (r = 0.234, P < 0.05 ; r = 0.267, P < 0.01, respective-ly). The urinary excretion of 11-dTXB2 was greater in the NIDDM group (7.58±4.42 μg/day for males and 5.65 ± 2.38 μg/day for females) than in the normal controls (4.61± 2.31 and 3.83±1.60, respectively), suggesting that the synthesis of thromboxane A₂ by platelets may be accelerated in vivo in patients with NIDDM. The urinary 6-kPGF_1α, was not different between the NIDDM group and normal controls among the males. but was greater in the NIDDM group among the females. As MPV showed a positive correlation (r = 0.364, P < 0.05) with urinary excretion of 11-dTXB₂, MPV may be related to platelet activity. These findings suggest that the platelets of patients with NIDDM may be in a hyperactive state. J Atheroscier Thromb, 1994 ; 1 : 108-112.

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To clarify the significance of increased blood sialic acid in atherosclerotic cardiovascular disease, we investigated the relationship between serum sialic acid level and atherosclerotic risk factors such as serum uric acid, fasting blood glucose, systolic and diastolic blood pressure, atherogenic index, and white blood cell count. By simple regression analysis, the serum sialic acid level was found to correlate significantly with these parameters. The mean sialic acid level was significantly higher in the highest quartile for serum uric acid than in its lowest quartile and also for fasting blood glucose concentration in comparison with its other three quartiles. A tendency was noted for the mean serum sialic acid level of each quartile to become higher with an increase in the quartiles of systolic or diastolic blood pressure, atherogenic index, and white blood cell count. Multiple regression analysis showed the correlations to be significant for the relationship of serum sialic acid to atherogenic index, mean arterial pressure and white blood cell count. From these results and previous findings of higher serum sialic acid levels in smokers, patients with diabetic angiopathies, and patients with hyperlipidemia, it is suggested that serum sialic acid reflects the degree of atherosclerotic progress involving inflammation processes. J Atheroscler Thromb, 1994 ; 1 : 113-117.

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Rats with alloxan-induced diabetes developed severe atherosclerotic lesions when they were maintained on a 0.25% cholesterol diet for one year. The atheromatous changes developed at the aortic arch, appeared as early as 3 months after the start of the experiment, and increased thereafter. The diabetic rats also developed atherosclerosis when they were fed standard rat chow, but the area of the atheromatous lesion was about one tenth of that in rats fed the high-cholesterol diet. Normal rats did not develop atherosclerosis even when fed the high-cholesterol diet for one year. The alloxan diabetic rats showed no increase in body weight, but developed serum glucose levels as high as 600-800 mg/dl as well as high serum cholesterol levels and lower serum HDL-cholesterol levels. The development of atherosclerosis in these rats was significantly related to an increase in the serum cholesterol/phospholipid ratio, the atherogenic index (TC-HDLC/HDLC), and the serum total cholesterol level, but was not related to the serum glucose, HDL-cholesterol, triglyceride, or lipid peroxide levels, These relationships were found as early as 8-16 weeks after the start of the experiment. These data suggest that the serum cholesterol/phospholipid ratio, the atherogenic index, and the total cholesterol level are important risk factors for the development of atherosclerosis in rats with alloxan diabetes. J Atheroscler Thromb, 1994 ; 1 : 118-128.

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We report a 46-year-old man with familial hypercholesterolemia who simultaneously developed angina pectoris and left hemiplegia. Angiography revealed complete tapering occlusion of the right internal carotid artery and a 75% stenosis of the right coronary artery. In addition to hypercholesterolemia, his serum Lp (a) levels were very high, with a mean (± SE) of 62±2 mg/dl. J Atheroscler Thromb, 1994 ; 1 129-131.

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