Soybean protein diet increases low density lipoprotein receptor activity in mononuclear cells from hypercholesterolemic patients.

The effect of two diets containing different protein sources (animal vs. soybean) on the low density lipoprotein (LDL) receptor activity was tested in freshly isolated mononuclear cells from 12 individuals with severe type II hyperlipoproteinemia. The two diets, both taken for 4 wk in a crossover design were of otherwise identical composition. During the soybean protein diet period, total cholesterol was reduced by 15.9% and LDL-cholesterol by 16.4%. The diet containing animal proteins exerted no significant change in plasma lipid levels vs. the baseline findings. The soybean diet regimen dramatically affected the degradation of LDL by mononuclear cells. Degradation was increased 16-fold vs. the basal activity and 8-fold compared with the standard low lipid diet with animal proteins. There was, however, no clear relationship between the reduction of total and LDL-cholesterolemia and the increased LDL degradation. These findings confirm similar data previously obtained in cholesterol-fed rats and suggest that some factor/s, most likely of a protein nature, may regulate the expression of lipoprotein receptors in peripheral cells, particularly when receptor activity is suppressed by experimental diets and/or spontaneous hypercholesterolemia.

Reduced platelet aggregability and increased vascular prostacyclin formation in a variant rat strain (IVA-SIV) with endogenous hypertriglyceridemia.

The Ivanovas-Sieve (IVA-SIV) rat represents the only available animal model of endogenous hypertriglyceridemia, in the absence of obesity and/or overt diabetes. Since plasma lipids/lipoproteins can modulate platelet reactivity and eicosanoid metabolism, these were examined in two groups of Charles River (CR) and IVA-SIV rats of identical age. The IVA-SIV rats had 2-fold higher plasma triglycerides and a 55% higher number of circulating platelets; the number of platelets was significantly correlated with triglyceridemia. Platelet reactivity to ADP and to collagen was significantly reduced in these animals, whereas the formation of thromboxane B2 did not differ from that of the CR. After perfusion of platelet-rich plasma (PRP) through the aortas of animals of the two strains, platelet aggregability, already lower in the IVA-SIV, was reduced to a higher extent compared to the CR. Increased levels of the prostacyclin metabolite 6-keto-PGF1 alpha were identified in the perfusate from the aortas of IVA-SIV rats. Platelets from these animals also showed an increased sensitivity to Iloprost, a stable prostacyclin analogue, with an IC50 1.7-fold lower compared to CR rats. Spontaneous hypertriglyceridemia in the IVA-SIV model is not associated with platelet hyperresponsiveness, but rather with a reduced sensitivity to major aggregants.

Pantethine reduces plasma cholesterol and the severity of arterial lesions in experimental hypercholesterolemic rabbits.

Pantethine (P), a coenzyme A precursor, was administered to cholesterol-fed rabbits (0.5% cholesterol diet + 1% pantethine) for 90 days. At the end of treatment, plasma total cholesterol levels were reduced 64.7% and the HDL/total cholesterol ratio increased in P-treated animals; a significant rise of the apo A-I/A-II ratio was detected in HDL. VLDL lipid and protein levels were, on the other hand, reduced by P. The cholesterol-ester content of both liver and aortic tissues was not significantly affected by P. Although the total aortic area with evident plaques was reduced only 18.2%, the microscopical examination of sections from the major vessels of P-treated animals, showed a reduction in the severity of lesions, both in the aorta and in the coronary arteries. These findings suggest that P, in addition to significantly lowering plasma cholesterol levels in rabbits on an experimental diet, may modify lipid deposition in major arteries, possibly by affecting lipoprotein composition and/or exerting an arterial protective effect.

Effect of gemfibrozil treatment in hypercholesterolemia on low density lipoprotein (LDL) subclass distribution and LDL-cell interaction.

Gemfibrozil, a widely used fibric acid derivative, corrects hypercholesterolemia in a non-negligible fraction of patients. To investigate the mechanism of the cholesterol-lowering activity of fibric acids, a study was performed in 12 type IIa hyperlipidemic patients treated with gemfibrozil for 12 weeks. Changes in low density lipoprotein (LDL) structure and composition, agonist capacity of LDL against the LDL-receptor in human skin fibroblasts, LDL-receptor activity in mononuclear cells, lecithin:cholesterol acyltransferase (LCAT) and cholesterol ester transfer protein (CETP) activity, were evaluated. Plasma total and LDL cholesterol levels decreased by 17% and 20% after 12 weeks of treatment, the reduction being directly correlated with the baseline levels (r = 0.75 and 0.78, respectively). The mean LDL diameter increased significantly, from 25.5 to 26.1 nm, while the relative content of small LDL particles (< 25.1 nm) increased from 23.4% to 32.8% of total LDL. Neither the apolipoprotein (apo) B secondary structure nor the affinity of LDL for the LDL-receptor of fibroblasts were affected. The LDL-receptor activity in patients' mononuclear cells increased 3-fold, the rise being unrelated to the plasma cholesterol reduction. LCAT activity did not change, while CETP activity was reduced by 25% (P = 0.13) after treatment. These findings indicate that gemfibrozil causes significant changes in LDL structure that do not, however, affect the LDL interaction with peripheral cells.

Reduced apoprotein B and increased lipoprotein turnover in cholesterol-fed rabbits after partial ileal bypass.

Plasma lipid and lipoprotein levels and composition of the very low-, low-, and high-density lipoproteins (VLDL, LDL, HDL) were studied in white New Zealand rabbits in the control state, 8 weeks after initiation of a 2% cholesterol-enriched diet, and sequentially for 16 weeks following a partial ileal bypass (PIB) procedure. Turnover and aortic uptake of 125I-labeled VLDL, obtained from the rabbits before and 16 weeks after PIB, were also analyzed. A significant reduction of plasma cholesterol from the postdiet levels to well below the prediet mean, despite continuation of the high cholesterol diet, followed PIB. A very early reduction of the apoprotein B percentage content in both VLDL and LDL was observed after the procedure, even when a relative cholesterol ester enrichment of these lipoproteins was still present. HDL cholesterol levels were somewhat lowered by the experimental diet and further reduced after PIB. 125I-labeled VLDL after PIB showed a reduced arterial uptake, as compared to the same lipoproteins from animals before PIB. These findings may provide an interpretation for the decreased atherogenesis after PIB.

Endogenous hypertriglyceridemia in a nonobese rat model: plasma lipoproteins and dietary sensitivity.

Sprague-Dawley male rats from the Ivanovas-Sieve colony (IVA-SIV) show higher plasma triglyceride levels compared to the standard Charles River (CR) rats. The triglyceride enrichment occurs primarily in the very-low-density lipoproteins (VLDL), otherwise of a normal % composition, suggesting that the number of particles is increased, rather than their triglyceride (TG) content. High-density lipoprotein (HDL) particles, corresponding to HDL1, appear to be increased in the IVA-SIV rats, as confirmed by rate-zonal ultracentrifugation. The apoprotein composition of isolated lipoproteins (apo B content and isoelectric focusing pattern), does not differ in the two strains. The IVA-SIV rats are remarkably more TG inducible, compared to the standard CR. This can be shown with fructose loading and with a cholesterol-cholic acid diet. The plasma TG increase, after Triton administration, indicative of the VLDL-TG production, is fourfold higher in the IVA-SIV, compared to the CR rats. These findings provide evidence for some similarities between the IVA-SIV rat and human endogenous hypertriglyceridemia, and suggest an increased TG biosynthesis in this model.

Pathogenetic mechanisms of the endogenous hypertriglyceridemia in a nonobese rat model.

Male Sprague-Dawley IVA-SIV rats were compared to male Sprague-Dawley Charles River rats of the same age, body weight, and daily food intake. The IVA-SIV rats demonstrated hypertriglyceridemia (182 +/- 9.4 v 131 +/- 9.4 mg/dL, P less than 0.001), associated with increased fasting plasma glucose (115 +/- 3 v 84 +/- 2 mg/dL, P less than 0.001), and plasma insulin (35 +/- 5 v 19 +/- 2 microU/mL, P less than 0.001) levels. Furthermore, IVA-SIV rats responded to an oral glucose load with higher plasma glucose and insulin levels. Very-low-density lipoprotein (VLDL)-triglyceride (TG) turnover studies were performed, documenting a higher TG production rate, which correlated with the plasma TG concentrations, (r = 0.58, P less than 0.01) in the IVA-SIV rats. Since lipoprotein lipase activity in both adipose tissue and muscle was not significantly different in the two groups of rats, it appears that the hypertriglyceridemia in IVA-SIV rats is due to increased VLDL-TG secretion, associated with hyperglycemia, hyperinsulinemia, and increased plasma FFA levels. The IVA-SIV rats provide a model of endogenous hypertriglyceridemia, independent of obesity.

Differential effects of beclobrate on lipid/lipoprotein distribution in normal and hypercholesterolemic rats.

Beclobrate, a new fibric acid derivative, displays remarkable lipid lowering activity in rodents. In order to evaluate changes in the distribution and liver handling of lipoproteins, beclobrate was tested in rats fed on a normal or hypercholesterolemic diet. On the normal diet, beclobrate lowered total plasma cholesterol by 22-33.4% (10-50 mg/kg); the cholesterol reduction occurred mainly in high density lipoproteins (HDL) (by 24-45% with the three tested doses). The metabolic clearance of 125I-labelled beta-very low density lipoproteins (beta-VLDL) injected into these animals almost doubled (0.20 1/h vs. 0.13 1/h in controls) after treatment with 20 mg/kg of beclobrate. In addition, beclobrate administration dramatically increased the activity of the high-affinity receptors for beta-VLDL in isolated liver membranes (Bmax: 208 +/- 17.6 vs. 146 +/- 2.6 ng/mg of protein for controls). On the hypercholesterolemic diet, beclobrate treatment (50 mg/kg) was associated with a 25% reduction in total cholesterol accompanied, however, by a 166% rise in HDL cholesterol. In these animals, the composition of VLDL, typically cholesterol-enriched, became close to normal. The increased HDL was characterized by a remarkable enrichment with particles containing apolipoprotein E (apo E), which is compatible with either an improved peripheral cholesterol removal or an enhanced direct secretion of apo E. The two models offer different opportunities for evaluating the mechanism of action of this new lipid lowering agent. Lipoprotein catabolism and receptor-mediated clearance were characteristically improved in normolipidemic rats whereas major effects on HDL metabolism could be demonstrated in hypercholesterolemic rats.

New microporous cholestyramine analog for treatment of hypercholesterolemia.

A new, microporous, uniformly reticulated preparation of cholestyramine is described. The preparation, cholpor, has a higher exchange capacity for chloride than does cholestyramine and swells very little in water. It is 15--20% more potent than chloestyramine in the in vitro binding of sodium cholate; moreover, the binding velocity is considerably higher than that of cholestyramine. Colestipol hydrochloride, also used as a reference anion-exchange resin, is about half as potent as the other two resins; its binding velocity is similar to that of cholpor. Cholpor may be prepared in a suspension form of good palatability. Preliminary clinical findings in short-term trials showed a cholesterol-lowering effect similar to that of cholestyramine with lower doses and fewer side effects.

Dietary animal proteins and cholesterol metabolism in rats.

The effects on cholesterol metabolism in rats of diets containing various animal proteins or soy protein were studied. The animal proteins tested were casein, whey protein, fish protein, hemoglobin, plasma proteins, ovalbumin, egg-yolk protein, beef protein and chicken-meat protein. The semi-purified diets were isonitrogenous and balanced for residual fat and cholesterol in the protein preparations. The nature of the dietary protein had no effect on serum cholesterol concentration. Group mean liver cholesterol concentration was increased and fecal excretion of bile acids was decreased by all animal proteins when compared with soy protein. This study suggests that carefully balancing diets for components other than protein in the protein preparations prevents protein effects on serum cholesterol in rats but not on liver cholesterol and bile acid excretion.

Soy proteins and cardiovascular disease.

The soybean diet is the most potent dietary tool for hypercholesterolemia. The United States Food and Drug Administration recently approved the health claim for its role in reducing the risk of coronary disease. The hypocholesterolemic effect is directly correlated to the patient's cholesterolemia, with minimal or no reductions occurring at cholesterol of 6 mmol/L or less, and the most benefit occurring in patients with cholesterol of greater than 7 mmol/L. Hypotheses on the mechanism of action include soy fiber, isoflavones (phytoestrogens), and the protein itself. Although there is no evidence for the effect of fiber, studies with ethanol-extracted soy (devoid of isoflavones) indicated a loss of effect, but the extract itself (isoflavone rich) has no hypocholesterolemic activity. In humans, soy protein activates the low-density lipoprotein (LDL) receptor pathway. Recent data suggest that soy protein subunits, particularly 7S, directly activiate LDL receptors in the human liver, thus providing a novel mechanism of plasma cholesterol reduction different from currently available diets and hypolipidemic drugs.

Mechanisms of lipid-lowering agents.

Lipid-lowering agents are used with the purpose of ameliorating hyperlipoproteinemias, in order to prevent arterial disease. Lipid-lowering drugs can be classified into absorbable agents and into nonabsorbable compounds, acting within the gastrointestinal lumen. Absorbable drugs (fibric acids, nicotinic acid, probucol, HMG-CoA reductase inhibitors) reduce plasma very-low-density lipoproteins (VLDL) and/or low-density lipoproteins (LDL) by a variety of mechanisms. Fibric acids, in particular, act by stimulating the catabolism of VLDL and also, as a consequence, improving LDL delipidation, thus favoring receptor uptake. Nicotinic acid and acipimox interfere with the biosynthesis of LDL and can also improve the clearance of VLDL/LDL. Probucol acts by a newly described mechanism, i.e. accelerating reverse transport of cholesteryl esters from high-density lipoproteins to lower-density lipoproteins. Finally, HMG-CoA reductase inhibitors, interfering with the biosynthesis of cholesterol, can induce an increased expression of liver high-affinity lipoprotein receptors. Nonabsorbable agents (anion-exchange resins, neomycin, beta-sitosterol) interrupt the recirculation of bile acids and/or reduce the absorption of cholesterol with the gut. They display a selective activity on hypercholesterolemia, again by increasing LDL receptor expression. The choice of one or more lipid-lowering agents will depend upon the patient's phenotype, determining responsiveness to the pharmacological treatment.

Soy and cholesterol reduction: clinical experience.

A role of vegetable proteins in reducing coronary artery disease risk was postulated as long ago as 1909 in Russia by Ignatowski. The protein hypothesis of atherosclerosis was pursued by many investigators, who studied the possible role of animal vs. vegetable protein in modifying concentrations of plasma lipids and thus cardiovascular disease risk. Over the past 20 y, our research group has examined the potential of a diet based on vegetable protein (in most cases, textured vegetable protein, or TVP) to modify plasma lipid concentrations. Textured products allow administration of a large percentage of protein (up to 50-60% in the product) and are available in a variety of food items. We studied > 1000 patients. An extensive review of the literature indicates that similar findings have been reported by others when administering TVP or TVP-like items to subjects with well-characterized hypercholesterolemia (Fredrickson type II). Data are less consistent for treatment of patients with marginal hypercholesterolemia or hypercholesterolemia already corrected by a standard diet before administration of soy products. The TVP diet, is, however, effective when normolipidemic individuals are made hypercholesterolemic by dietary cholesterol administration. These and other findings suggest that, in man, similar to experimental animals, soy protein may in some way up-regulate LDL receptors depressed by hypercholesterolemia or by dietary cholesterol administration.

Effects of sub-chronic exposure to SO2 on lipid and carbohydrate metabolism in rats.

Sulfur dioxide (SO2) is a ubiquitous air pollutant, present in low concentrations in the urban air, and in higher concentrations in the working environment. While toxicological reports on SO2 have extensively dealt with the pulmonary system, essentially no data are available on the effects of chronic exposure to this pollutant on intermediary metabolism, although some biochemical changes in lipid metabolism have been detected. The present investigation was aimed at evaluating the effects of sub-chronic exposure to SO2 on concentrations of serum lipids/lipoproteins and on glucose metabolism, in animal models of hypercholesterolemia and diabetes. A specially designed control-inert atmosphere chamber was used, where male Sprague-Dawley rats fed on either standard or cholesterol enriched (HC) diets, as well as streptozotocin diabetics, were exposed to SO2 at 5 and 10 ppm, 24 h per day for 14 days. In rats, both on a standard diet and on a HC regimen, SO2 exposure determined a significant dose-dependent increase in plasma triglycerides, up to +363% in the 10 ppm HC exposed animals. This same gas concentration significantly reduced HDL cholesterol levels. In contrast, exposure of diabetic animals to 10 ppm SO2 resulted in a fall (-41%) of plasma and liver triglycerides and in a concomitant increase (+62%) of plasma HDL cholesterol. This discrepancy could apparently be related to diverging effects of SO2 exposure on plasma insulin levels in the different animal groups. Kinetic analyses of triglyceride synthesis carried out in rats on a standard diet revealed, in exposed animals, a significant reduction in the secretory rate, in spite of the concomitant hypertriglyceridemia. These findings suggest that SO2 exposure can markedly modify major lipid and glycemic indices, also indicating a differential response in normo/hyperlipidemic versus diabetic animals.

7S globulin from soybean is metabolized in human cell cultures by a specific uptake and degradation system.

We examined the biological fate of 7S globulin from soybean in a hepatoma cell line (Hep G2) and in human skin fibroblasts (HSF) to gain new insights into the 7S globulin cell process, the final effect of which is an enhanced expression of the LDL-receptor. The ability of 7S globulin to bind and to be internalized and degraded by both cell types was investigated under different experimental conditions. In all cases, specific uptake (binding + internalization) and degradation of 125I-7S globulin were curvilinear functions of substrate concentration at 37 degrees C. The two processes were saturated at around 80 mg/L, a concentration at which an up-regulation of LDL-receptor was previously reported. The specific uptake of 125I-7S globulin at 37 degrees C was a curvilinear function of time, and achieved equilibrium after 6 and 12 h in HSF and Hep G2 cells, respectively. Binding experiments, conducted at 4 degrees C in Hep G2 cells, showed a specific and saturable association of 7S globulin to the cell membrane. Linear Scatchard analysis demonstrated a single population of binding sites. The amount of 7S globulin bound at saturation (Bmax) was about 2.73 mg/L, with an apparent Kd of 21 micromol/L, assuming 175 kDa as the 7S globulin molecular weight. SDS-PAGE of Hep G2 membrane proteins incubated with 125I-7S globulin revealed a specific interaction of 7S globulin with a cell protein component with molecular weight between 14 and 21 kDa. Further studies are needed to ascertain whether this interaction is directly or indirectly related to the observed stimulation of the LDL-receptor.

Natural history of TCDD-induced liver lesions in rats as observed by transmission electron microscopy during a 32-week period after a single intraperitoneal injection.

Forty-two male rats were injected with a single intraperitoneal dose of TCDD in acetone and corn oil and sacrificed after 2, 4, 8, 16, 24, and 32 weeks, to study the long-term effects of a single injection. The liver lesions become progressively worse up to the 16th week and appear thereafter to slowly regress.

Dietary animal proteins and cholesterol metabolism in rabbits.

The effect in rabbits of giving isonitrogenous purified diets containing casein, ovalbumin, fish protein, milk-whey protein and soya-bean protein were compared. The diets were balanced for cholesterol and for the amount and type of fat. When incorporated into low-cholesterol diets (0.08 g cholesterol/kg), casein, ovalbumin and soya-bean protein produced similar levels of serum cholesterol. With a high background of dietary cholesterol (1.5 g/kg), serum cholesterol concentrations increased with soya-bean protein, whey protein, casein and fish protein, in that order. Thus, the hypercholesterolaemic effect of casein in carefully balanced diets was only seen against a high-cholesterol background. The development of hypercholesterolaemia produced by giving fish protein was different from that produced by casein. First, less cholesterol accumulated in the very-low-density-lipoprotein fractions and more in the lipoproteins of higher density with fish protein than with casein. Second, fish protein, unlike casein, did not increase liver cholesterol. Third, transfer of rabbits from a diet containing soya-bean protein to one containing casein resulted in an immediate marked depression in neutral steroid and bile acid excretion in faeces. However, when rabbits were fed on the diet with fish protein after the diet with soya-bean protein, there was no significant depression in neutral steroid output and the depression in bile acid output was delayed. The present study suggests that different animal proteins cause hypercholesterolaemia by different mechanisms.