Tsugumichi Uemura

Evaluation of Oxidized Low-Density Lipoprotein and Large Molecular Size Low-Density Lipoproteins in Atherosclerosis

To study the roles of modified low-density lipoprotein (LDL) and the molecular size of LDL in atherogenesis, the following studies were carried out. Eight white male rabbits fed a standard oriental diet with 1% cholesterol were used to isolate LDL and to observe changes in the molecular size of LDL due to cholesterol feeding. The tissue LDLs in the aorta were analyzed to confirm the existence of modified LDL (namely, LDL with peroxidized cholesteryl ester) by thin-layer chromatography. In addition, plasma LDLs were isolated from 18 patients with myocardial infarction and 11 patients with angina pectoris to confirm the existence of LDL with peroxidized cholesteryl ester. Each LDL separated consisted of 3 fractions; namely, IDL (1.006-1.018), LDL1 (1.019-1.052) and LDL2 (1.053-1.063) by sequential ultracentrifugation. The molecular sizes of LDL were measured by a planimeter from electron microscopic photographs, with negative staining. The estimation of peroxidized cholesteryl linoleate in LDL was performed using our method. The modified LDLs with peroxidized cholesteryl ester were poorly estimated in the LDL separated from the plasma of cholesterol-fed rabbits and from the aorta extraction after 16 weeks of feeding. The peroxidized cholesteryl ester was clearly identified in the plasma LDLs of the patients with myocardial infarction and angina pectoris, and in whole extracts from human aortic atheroma, although it was not clearly identified in the tissue LDL fraction. The molecular sizes of LDL1 enlarged week by week with cholesterol feeding, but two fractions of IDL and LDL2 did not change in size. The infusion of cholesterol-rich LDL of large molecular size or LDL with peroxidized cholesteryl ester into the vessels led to fixation, on the surface of the arteries of many platelets, red cells, and white cells, and to marked irregularities in the endothelial folds. The evidence suggests that atheromas, formed in a short period in rabbits with cholesterol feeding, are caused mainly by the increase in LDL1 of large molecular size, and that foam cells, formed in human atheromas, are caused mainly by the production of modified LDL with peroxidized cholesteryl ester.

Acceleration of platelet aggregability due to modulation of native LDL.

The aim of this experiment was to clarify whether low density lipoprotein (LDL) causes an acceleration of platelet aggregability. Native LDL was separated into two fractions by filtered tap-water dialysis, namely, water-soluble LDL (WS-LDL) and non-water-soluble LDL. Although native LDL did not enhance the platelet aggregability, WS-LDL made it markedly increased. WS-LDL consisted of the lipid constituents that were not found in native LDL. Namely, in thin-layer chromatography of WS-LDL, an unknown spot between triolein and free fatty acid was clearly stained. This unknown spot in the WS-LDL was produced by the peroxidation of cholesteryl ester in native LDL. It was confirmed that the spot has the same Rf value as the peroxidate of cholesteryl linolate in thin-layer chromatography. If native LDL is modulated by divalent metal ions and oxygen in the fluids, LDL with biological activity such as an increase of platelet aggregability is produced.

A new approach to prevention and treatment of atherosclerosis by dyslipoproteinemia.

A number of papers’.’ have been published on the treatment and prevention of atherosclerosis. The main points of those papers were to reduce the concentrations of low density lipoprotein cholesterol (LDL-CH), apoprotein B (apo B), and lipid peroxide (LPO) or to increase that of high density lipoprotein cholesterol (HDL-CH). Furthermore, in several human and nonhuman primate experiment^^.^ it was shown that atherosclerotic lesions regressed by reducing the concentration of LDL-CH for an extended period of time. Recently, Steinberg’ and Kitah reported that the foramtion of atheroma was suppressed in WHHL rabbits by probucol treatment. They pointed out that the suppression of LPO was more important for the treatment and prevention of atherosclerosis than the reduction of LDL-CH. In this paper, new aspects for the treatment and prevention of atherosclerosis will be discussed from the viewpoint of the composition and physical character of LDL.

Chemical Characterization of Peroxidized Low-Density Lipoprotein in Plasma and Aortic Atheroma

Hydroperoxidized cholesteryl linoleate (HPO-CL, spot X1 ) was produced by peroxidation of normal LDL isolated from plasma of healthy persons. Spot X1 stained on

Transformation of Cultured Human Monocytes by Peroxidized Low-Density Lipoprotein

To study the roles of peroxidized low-density lipoprotein (pox-LDL) during the formation of foam cells from human monocytes, monocytes isolated from normal human blood were incubated with RPMI alone, normal low-density lipoprotein (n-LDL) and copper pox-LDL. The concentrations of total cholesterol (TCH) and lipid peroxide in the human monocyte medium did not change significantly after incubation with RPMI medium alone or n-LDL, but those of TCH decreased slightly after incubation with pox-LDL for 24 h. Triglyceride (TG) concentrations in the culture medium of human monocytes decreased after incubation with all above substances. Ultrastructural studies showed that the monocytes changed to macrophages after incubation with RPMI alone or n-LDL and to foam cells after incubation with pox-LDL for 48 h. We conclude that TGs may be metabolized for ATP production by human monocytes. The energy may play a role in the transformation of monocytes to macrophages, and pox-LDL can induce transformation of monocyte-derived macrophages to foam cells.