Correlation between cholesterol content in circulating immune complexes and atherogenic properties of CHD patients' serum manifested in cell culture

Blood serum of most patients with coronary heart disease (CHD) caused a 2-5-fold increase in the total cholesterol content of smooth muscle cells cultured from unaffected human aortic intima, i.e. possessed an atherogenic potential manifested in culture. Removal of immunoglobulins G and M from an atherogenic serum brought about a fall in its atherogenic potential. The serum deficient in immunoglobulins A retained its ability to induce the cholesterol accumulation in cells. Treatment of the CHD patients' serum with 2.5% polyethylene glycol 6000 removed the circulating immune complexes. The serum subjected to this treatment lost its atherogenicity, i.e. failed to increase the cholesterol content in cultured cells. Incubation of smooth muscle cells derived from human aortic intima with circulating immune complexes isolated from an atherogenic patients' serum caused a 1.5-3-fold rise in the intracellular cholesterol. Blood sera of most (89%) CHD patients was characterized by a high cholesterol content in circulating immune complexes. More than 75% of healthy subjects and patients without stenosis of coronary arteries had low level of cholesterol in immune complexes. Blood sera atherogenicity manifested in culture directly correlated with the cholesterol level of circulating immune complexes (r = 0.90). These findings suggest that the atherogenicity of CHD patients blood serum is due to cholesterol-containing immune complexes.     

Effects of lovastatin therapy on the level of low density lipoproteins and atherogenic potential of serum in patients with ischemic heart disease and hypercholesterolemia]

The serum atherogenic potential in patients with coronary heart disease (CHD) concurrent with hypercholesterolemia (LDL cholesterol more than 200 mg/dl), which is able to cause accumulation of intracellular cholesterol in cultured cells has been recently shown to be directly related to the level of total cholesterol and LDL cholesterol. The study was undertaken to examine how a lovastatin-induced decrease in LDL levels affects serum atherogenicity in patients with CHD and hypercholesterolemia. It was shown that the therapy of 22 patients with CHD and hypercholesterolemia led to a reduction in total and LDL cholesterol levels on an average by 24% and 32%, respectively. There were 3- and 1.5-2-fold decreases in circulatory immune complexes and the atherogenic potential, respectively. The findings suggest that the significant reduction in serum LDL cholesterol levels in patients with CHD concurrent with hypercholesterolemia who take hypolipidemic therapy is followed by a decrease in the atherogenic potential.     

Atherogenic properties of sera and low-density lipoproteins in patients with diabetes mellitus]

The effects of sera and low density lipoproteins (LDL) on cholesterol levels were evaluated in cultured aortic intimal cells from patients with Types 1 and 2 diabetes mellitus. About 80% Type 1 diabetic patients' sera and 90% of Type 2 diabetic patients' sera caused a 1.5-4-fold increase in intracellular cholesterol levels. LDL isolated from atherogenic sera induced intracellular cholesterol accumulation. There was a close correlation between the atherogenic effect of the sera and LDL. The atherogenic properties of the sera and LDL from patients with diabetes mellitus were unassociated with the examinees' age. There was a relationship between the atherogenic serum effect and the compensation of carbohydrate metabolism in male patients with Type 1 diabetes mellitus. The atherogenic effect of the sera and LDL failed to increase in patients with Types 1 and 2 diabetes mellitus concurrent with coronary heart disease. The property of the sera to accumulate intracellular cholesterol is likely to be typical of diabetes mellitus and mainly due to disturbed LDL metabolism.     

Triggerlike stimulation of cholesterol accumulation and DNA and extracellular matrix synthesis induced by atherogenic serum or low density lipoprotein in cultured cells

A 72-hour incubation of cultured cells with blood sera or plasma of patients suffering from coronary heart disease (CHD) with angiographically assessed coronary atherosclerosis caused a threefold to fourfold elevation of intracellular cholesterol. An elevated cholesterol level in the cells precultured with patients' sera was retained several days after the removal of the examined serum from culture. The accumulation of intracellular cholesterol was accompanied by enhanced synthesis of DNA, total protein, collagen, sulfated glycosaminoglycans, and hyaluronic acid. Enhanced DNA and total protein synthesis was retained for at least 9 days after the serum had been removed from culture. The obtained results suggest that the sera of CHD patients possess an atherogenic potential that manifests itself at the arterial cell level in the stable stimulation of atherosclerotic cellular processes: proliferation, lipidosis, and fibrosis. The examined sera of healthy donors were devoid of such an atherogenic potential. The low density lipoprotein (LDL) fraction (density, 1.030-1.050 g/cm3) obtained from an atherogenic serum had the same atherogenic potential as a whole serum. Atherosclerotic alterations in cultured intimal cells caused by atherogenic LDL were retained for at least 3 days after the removal of the lipoprotein from culture. Preincubation of intimal cells with LDL obtained from healthy donors had no effect on the intracellular cholesterol level or the synthesis of DNA and extracellular matrix. One may assume that the atherogenic potential of CHD patients' sera is related to the presence of LDLs that are qualitatively different from the LDL of healthy subjects.     

Blood serum atherogenicity associated with coronary atherosclerosis. Evidence for nonlipid factor providing atherogenicity of low-density lipoproteins and an approach to its elimination

To reveal the presence of atherogenic potential in the blood serum obtained from patients with angiographically assessed coronary atherosclerosis we used primary cultures of subendothelial cells isolated by collagenase from unaffected human aortic intima. Earlier, we have demonstrated that such cultures are made up mostly of typical and modified smooth muscle cells. Within 24 hours of cultivation with a 40% sera of patients suffering from coronary atherosclerosis, the total intracellular cholesterol level increased twofold to fivefold. Cultivation with the sera of healthy subjects had no effect on the intracellular cholesterol level. The sera of patients were separated by ultracentrifugation into two fractions: total lipoprotein fraction containing the main classes of lipoproteins and a lipoprotein-deficient fraction. The former, but not the lipoprotein-deficient fraction, was characterized by atherogenicity (i.e., the ability to induce the accumulation of intracellular cholesterol). Lipoproteins of the patients' serum were separated into main classes: low density lipoproteins (LDL), very low density lipoproteins (VLDL), and high density lipoproteins (HDL2 and HDL3). An atherogenic component of the serum capable of stimulating the deposition of intracellular cholesterol was represented by LDL and, in one case, by VLDL, but not by other classes of lipoproteins. LDL and other lipoproteins isolated from the blood serum of healthy subjects failed to raise the cholesterol content in cultured cells; that is, they were nonatherogenic.(ABSTRACT TRUNCATED AT 250 WORDS)     

Serum from type IIA hyperlipoproteinemic patients does not stimulate proliferation of and collagen synthesis in human fetal aortic smooth muscle cells in culture

The effect of serum from type IIA hyperlipoproteinemic patients on the proliferation and synthesis of collagen and other proteins of human fetal aortic smooth muscle cells (SMC) was studied. The activity of lysyl oxidase secreted into the culture medium was also measured. Type IIA hyperlipoproteinemic sera did not affect the proliferation of human aortic SMC as compared to normolipidemic sera, regardless of incubation time. The findings were similar for 3 different human fetal aortic SMC lines and one fibroblast line from adult human skin. The synthesis of collagen and other proteins was inhibited rather than stimulated in the presence of type IIA hyperlipoproteinemic sera. The activity of lysyl oxidase was not affected by type IIA hyperlipoproteinemic sera. The atherogenicity of hypercholesterolemia cannot be explained either by its direct effect on the proliferation of arterial SMC, as has been suggested by animal cell studies, or by its direct effect on the fibrogenicity of these cells.     

Atherogenic properties of sera from patients with ischemic heart disease demonstrated in vitro

The sera of coronary patients with angiographically-documented stenosis of 1 to 3 coronary arteries caused a two-to-fivefold accumulation of intracellular cholesterol in intact human aortic intimal cell culture. This property, qualified as "atherogenic", was exhibited by the sera of 90% of the coronary patients examined (97 males and females, aged 29 to 55 years). Only 20% of normal subjects revealed atherogenic properties in cell cultures. The capacity of the sera to accumulate intracellular cholesterol was unrelated to the levels of total cholesterol, high density lipoprotein cholesterol, apo-B or apo-AI, taken separately, but showed a weak correlation to the apo-B/apo-AI ratio. The sera of coronary patients caused lipid accumulation in human aortic medial smooth-muscle cells and blood mononuclear cells, in human and mouse peritoneal macrophages, but never in human aortic or umbilical vein endothelial cells, nor human embryo fibroblasts.     

Surgical correction of hyperlipoproteinemia in patients with ischemic heart disease

The experimental results obtained in 102 animals showing a beneficial effect of partial ileac bypass on hyperlipoproteinemia and experimentally-induced atherosclerosis prompted the authors to perform this surgical intervention in 34 patients with coronary heart disease (CHD) and attendant hyperlipoproteinemia resistant to therapeutic agents. The follow up period was up to five years. The patients exhibited a persistent and substantial decrease in blood lipids and atherogenic lipoproteins, a reduced incidence of angina attacks and an improved CHD course. Furthermore, the surgery resulted in a considerable and stable elevation of the cholesterol levels of antiatherogenic high density lipoproteins (by an average of 85,3%) and in a 45.8% decrease of the cholesterol coefficient of atherogenicity which contributes to atherosclerosis regression.     

Use of cultured atherosclerotic cells for investigation of antiatherosclerotic effects of anipamil and other calcium antagonists.

Calcium antagonists have been shown to exhibit an antiatherosclerotic action in primary cultures of human aortic atherosclerotic cells by causing a reduction in intracellular lipid content, proliferative activity and synthesis of the extracellular matrix. Verapamil and nifedipine exhibited the highest efficacy in this respect. The new calcium antagonist, anipamil (racemate and enantiomers), has been tested in cell cultures. At 10(-6) M and higher concentrations, anipamil and its enantiomers produced considerable decrease in intracellular content of cholesteryl esters, triglycerides and free cholesterol, suppressed cell proliferation and inhibited synthesis of the extracellular matrix. The efficacy of anipamil (enantiomers and racemate) was similar to that of verapamil and greater than that of nifedipine. Plasma obtained from patients after administration of 80 mg verapamil or 20 mg nifedipine significantly lowered the cholesterol content of cultured cells. Blood plasma of most atherosclerotic patients possesses atherogenic properties, i.e., it is able to increase the cholesterol content in cultured cells. Plasma atherogenicity seen in cultures decreased considerably or even disappeared after both nifedipine and verapamil administration. After 28 days of nifedipine therapy, plasma atherogenicity was lower compared with the initial value at the beginning of the treatment. These observations suggest that control of plasma atherogenicity after drug administration may provide an additional tool for optimisation of direct antiatherogenic and antiatherosclerotic therapy.     

Transgenic overexpression of human lecithin: cholesterol acyltransferase (LCAT) in mice does not increase aortic cholesterol deposition

Results from several atherosclerosis studies using morphometric procedures have proven controversial with regard to whether over-expression of human LCAT in transgenic (Tg) mice is atherogenic. The purpose of the present study was to determine the effect of 10-fold over-expression of human LCAT on aortic free and esterified cholesterol (EC) deposition as well as plasma lipoprotein cholesteryl ester (CE) fatty acid composition in mice fed an atherogenic diet containing cholic acid. C57Bl/6 (control) and human LCAT-Tg mice were fed chow or an atherogenic diet (15% of calories from palm oil, 1.0% cholesterol and 0.5% cholic acid) for 24 weeks before measurement of aortic cholesterol content. Compared with the chow diet, control and LCAT-Tg mice fed the atherogenic diet had a 2-fold increase in plasma total, free and EC, a 7-fold increase in plasma apoB lipoprotein cholesterol, and a 40-50-fold increase in hepatic cholesterol content. The aortic EC content was increased in control (0.7 vs. 1.2 mg/g protein) and LCAT-Tg (0.3 vs. 1.5 mg/g protein) mice fed the atherogenic diet compared with those consuming the chow diet; however, there was no difference in aortic free (14.4+/-6.8 vs. 18.5+/-7.7 mg/g protein) or esterified (1.2+/-1.0 vs. 1.5+/-1.2 mg/g protein) cholesterol content between atherogenic diet-fed control and LCAT-Tg mice, respectively. LCAT-Tg mice fed the atherogenic diet had a 2-fold increase in the ratio of saturated+monounsaturated to polyunsaturated CE species in plasma apoB lipoproteins compared with control mice (9.4+/-2.4 vs. 4.9+/-0.7). We conclude that over-expression of human LCAT in Tg mice fed an atherogenic diet containing cholic acid does not result in increased aortic cholesterol deposition compared with control mice, even though the CE fatty acid saturation index of plasma apoB lipoproteins was doubled.     

Atherogenic properties of beta adrenergic blockaders evident on vascular wall cells

In cultured human cells, beta-adrenoblockers such as propranolol, alprenolol, metoprolol, atenolol, pindolol, and thymolol, as well as sera from patients with coronary heart disease were examined for atherogenic activity following a single administration of propranolol (80 mg) and pindolol (10 mg). Addition of the beta-adrenoblockers to the cell culture was demonstrated to enhance cellular total cholesterol levels and to stimulation their proliferation. Propranolol and pindolol given in a single dose was found to result in the appearance of atherogenic properties of the patients' sera.     

Beta-blockers: propranolol, metoprolol, atenolol, pindolol, alprenolol and timolol, manifest atherogenicity on in vitro, ex vivo and in vivo models. Elimination of propranolol atherogenic effects by papaverine.

The addition of the beta-blockers propranolol, metoprolol, atenolol, pindolol, alprenolol and timolol to a culture of peritoneal macrophages or smooth muscle cells induced an increase in the intracellular cholesterol content. Blood serum obtained from a rabbit after a peroral administration of beta-blockers also induced cholesterol accumulation. This property of drug or blood serum obtained after peroral administration is conventionally referred to as atherogenic potential or atherogenicity. Regular administration of propranolol during a 21-day period evoked stable atherogenicity of rabbit blood serum. This was accompanied by stimulation of manifestations of atherosclerosis in the aorta deendothelialized with a balloon catheter. Propranolol increased neointimal thickening, lipid accumulation, an increase in cell number and in the collagen content. In vitro, the combination of propranolol with papaverine eliminated the atherogenic effect of propranolol which manifested itself as stimulation of cholesterol accumulation in cultured cells. Simultaneous peroral administration of propranolol and papaverine prevented the appearance of serum atherogenicity. Papaverine eliminated neointimal thickening, an increase in cell number and in the lipid and collagen contents evoked by propranolol. Papaverine itself had no effect on these parameters. Thus, the atherogenicity of propranolol as well as capacity of papaverine to eliminate beta-blocker atherogenicity revealed in cell culture was confirmed in vivo. We hope that these results may be useful in the development of new drugs and optimization of antiatherosclerotic drug therapy.     

Effects of beta adrenergic blocking agents and calcium antagonists on atherogenic properties of blood serum of patients with ischemic heart disease

The beta-adrenoblockers anapriline and visken and the calcium antagonist corinfar were studied for effects on the atherogenic properties of the sera from patients with coronary heart disease who took the drugs. The study was performed by using cultured atherosclerotically altered human aortic smooth muscle cells. A single dose of anapriline, 80 mg, and that of visken, 20 mg, were found to give rise to or to enhance atherogenic properties of the patients' sera, while that of corinfar, 20 mg, yielded their antiatherosclerotic properties.     

Remnant lipoproteins and atherogenesis

Recent clinical studies have revealed that increased serum triglyceride (TG) levels are closely related to atherosclerosis, independently of serum levels of high-density lipoproteins (HDL) and low-density lipoproteins (LDL). Among triglyceride-rich lipoproteins (TRLs), remnant lipoproteins (RLPs) are considered to be atherogenic and an independent coronary risk factor. We previously reported that monocytes cultured in the presence of RLPs increased their adhesion to vascular endothelial cells. The underlying mechanism involved activation of RhoA, a member of small GTP binding proteins, resulting in activation of focal adhesion kinase (FAK) and s1-integrin. It is also known that RLPs enter vessel walls. In another study, we reported that RLPs induced smooth muscle cell (SMC) proliferation, independently of oxidative stress. Recently, we identified the molecular mechanisms, in which RLPs from hypertriglyceridemic patients stimulated SMC proliferation via epidermal growth factor (EGF) receptor transactivation and heparin-binding EGF-like growth factor (HB-EGF) shedding. More recently, we reported that apoB48 receptor was involved in RLP-induced foam cell formation in macrophages. The current review focused on the molecular mechanisms for the atherogenicity of RLPs.     

年龄及地区因素对年轻人动脉壁的影响及其和动脉粥样硬化早期病变发生发展的关系

本研究在动脉粥样硬化（ＡＳ）和冠心病（ＣＨＤ）高发区（北京）、低发区（南宁），于１９８６年～１９８９年间，共收集了意外死亡年轻人（１５～３９岁）新鲜尸检标本３２４例。用形态测量和生化分析的方法研究了两地同龄年轻人主动脉内膜脂质浸润、内膜厚度、平滑肌细胞（ＳＭＣ）和基质中蛋白聚糖（ＰＧｓ）的变化以及地区差异，以探索其与ＡＳ发生发展的关系，为ＡＳ的防治提供依据。主要结果：主动脉内膜ＳＭＣ核的密度尤其面积密度均是北高于南（Ｐ＜０．０１）。ＳＭＣ核密度与同组切片阿新蓝染色阳性物质量呈负相关；同组标本的生化分析ＰＧｓ总量和有抑制ＳＭＣ增殖的ＨＳ－ＰＧ含量均北低于南（Ｐ＜０．０１）；主动脉内膜脂质浸润的嗜苏丹病变（ＳＬ），除男性腹主动脉ＳＬ面积百分比北高于南（Ｐ＜０．０１）外，两地接近。上述结果提示有抑制ＳＭＣ增殖的ＨＳ－ＰＧ含量降低可能为北京主动脉内膜ＳＭＣ增殖的原因之一，还提示随南宁地区近年来人群血情胆固醇水平上升，在主动脉内膜ＳＬ已有所反映，这一变化趋势从预防ＡＳ或ＣＨＤ角度应引起注意。     

HDL cholesterol lipoprotein levels and the atherogenicity coefficient in males with diabetes mellitus and ischemic heart disease

A hundred males aged 40-69 years receiving treatment at the Institute of Experimental Endocrinology and Hormone Chemistry for type I and II diabetes mellitus were examined. Thirty-eight patients had coronary heart disease (CHD). The diagnosis of CHD was based on the data of the WHO Cardiological Questionnaire and the presence of ECG changes corresponding to the categories of the Minnesota Code 1-1,2,7; 4-1,2 and 5-1,2 (without 3-1,3). The blood levels of total cholesterol (CS), triglycerides (TG), high density lipoprotein cholesterol (HDLC) and the atherogenicity coefficient (AC) were determined in all patients. The results of the study showed that males with diabetes mellitus irrespective of its type or the presence of CHD had significantly higher mean values of total CS, TG and AC than in control. The mean levels of HDLC in males with type I diabetes mellitus without CHD did not differ from those in normal subjects whereas the level of HDLC in patients with type II diabetes mellitus was lowered. The patients with CHD showed a significant decrease in HDLC in both type I and II diabetes. The highest mean values of TG, AC and the lowest levels of HDLC were characteristic of the males with type II diabetes mellitus and CHD.     

Value of hemosorption and essential phospholipids in the complex treatment of patients with ischemic heart disease

Eighty-three patients were treated with hemosorption and essential phospholipids for coronary heart disease (CHD) and followed up for 3-5 years. The new approach to the treatment of CHD involves simultaneous removal of cholesterol-rich atherogenic lipoproteins by hemosorption and administration of essential phospholipids to increase cholesterol withdrawal from the cells. This combined treatment of coronary patients is shown to normalize cholesterol metabolism, improve microcirculation, increase stress tolerance and bring about an obvious clinical remission. The good clinical effect is corroborated by the results of bicycle ergometry and data on lipid spectrum, cholesterol metabolism, microcirculation and radio-isotope studies.     

Stable analogues of prostacyclin and thromboxane A2 display contradictory influences on atherosclerotic properties of cells cultured from human aorta. The effect of calcium antagonists

We examined the influence of stable prostacyclin analogues (carbacyclin) and thromboxane A2 (U46619) on atherosclerotic properties of cells: [3H]thymidine incorporation and intracellular cholesterol content. A primary culture of human aortic subendothelial cells derived from atherosclerotic plaques was used. Carbacyclin exerted a direct anti-atherosclerotic effect, significantly reducing atherosclerotic manifestations of cells, while agent U46619 stimulated proliferation and cholesterol accumulation, i.e. demonstrated atherogenic potency in culture. Calcium antagonists (verapamil and diltiazem) markedly enhanced anti-atherosclerotic properties of carbacyclin and restricted the atherogenicity of U46619.     

Circulating immune complexes of Hodgkin's disease contain an antigen that is present in Hodgkin and reed-sternberg cells

Summary Circulating immune complexes (CIC), isolated from the serum of a patient with Hodgkin's disease (HD) and from control serum (CS) of healthy adults, were used to generate heterologous antisera in rabbits. The antiserum directed against CIC from HD (AS-HD) and the antiserum directed against CIC from CS (AS-CS) were used to identify immunoglobulins, complement factors and alpha₂-macroglobulin as immune complex components. After adsorbing both antisera with normal human sera, we found that the adsorbed AS-HD was immunoreactive with radiolabelled CIC from HD serum but not with radiolabelled CIC from CS. Sera of patients with different diseases and sera of healthy adults were assessed for the occurrence of this Hodgkin immune complex-associated antigen (HIC-Ag). The HIC-Ag was present in 37% (12/33) of sera from patients with HD, 8% (8/101) of sera from patients with nonmalignant diseases, and 0% (0/6) of sera from healthy adults. This antigen was equally distributed among HD patients with and without symptoms, but its occurrence correlated with an advanced clinical stage of the disease. Using the adsorbed AS-HD in the immunoperoxidase technique, we identified the HIC-Ag as a cytoplasmic antigen in Hodgkin and Reed-Sternberg cells; whereas, the adsorbed AS-CS did not reveal any staining. These data indicate the presence of an HIC-Ag in the sera of patients with HD and suggest that the adsorbed AS-HD might be useful for isolation and characterization of this antigen for future use as a tumour marker.     

Inheritability of atherosclerosis and the role of lipoproteins as risk factors in the development of atherosclerosis in WHHL rabbits: risk factors related to coronary atherosclerosis are different from those related to aortic atherosclerosis.

Inheritability of atherosclerosis and the influences of serum lipids on atherosclerosis were examined by following its progression in selectively bred WHHL rabbits. Our studies indicate (1) coronary atherosclerosis is clearly inherited from parents by offspring whereas inheritability of aortic atherosclerosis is uncertain; (2) coronary stenosis is positively correlated to serum cholesterol level, although the correlation coefficient is markedly low: in contrast, no relationship between serum lipid levels and aortic atherosclerosis was observed; (3) cholesterol-rich VLDL showed atherogenicity in aorta, but not in coronary arteries; (4) an unknown lipoprotein detected by 3.6% polyacrylamide gel electrophoresis was related to coronary atherosclerosis, although no relationship between the unknown lipoprotein and aortic atherosclerosis was observed. These findings suggest that there are two types of genetic factors involved in atherosclerosis, one of which is unique to coronary atherosclerosis whereas the other is related to only aortic atherosclerosis.