RESUMEN
The binding of human 125I-labeled 'anionic polypeptidic fraction' (APF) to purified rat liver plasma membranes was studied. The dissociation constant for this binding was 3.0 micrograms protein/mg membrane protein. Binding was competitively inhibited by unlabeled human APF, but not by human LDL (low density lipoproteins). When unlabeled HDL3 was added, binding of labeled APF was competitively reduced to a level between that of unlabeled APF and unlabeled LDL. Experiments with cultured rat hepatocytes confirmed those obtained with liver membranes and suggested the presence in rat liver of saturable APF-binding sites which seem to be specific for APF. The physiologic significance of these APF binding sites is discussed in relation to the fate of cholesterol in the liver.
Asunto(s)
Hígado/metabolismo , Péptidos/metabolismo , Animales , Sitios de Unión , Unión Competitiva , Membrana Celular/metabolismo , Células Cultivadas , Colesterol/metabolismo , Humanos , Radioisótopos de Yodo , Cinética , Lipoproteínas HDL/farmacología , Lipoproteínas LDL/farmacología , Ratas , Ratas EndogámicasRESUMEN
Computation using a coherent system of units demonstrates simply that no significant specific biological and pharmacological effects can be expected from very highly diluted solutions.
Asunto(s)
Sistema Métrico , Soluciones/normas , Cómputos MatemáticosRESUMEN
The regulating process of cholesterol in the liver was studied in relation to its exogenous contribution in the rats fed high-fat (28%) high-cholesterol (1.2%) diets rich in saturated (S) fat (lard) or polyunsaturated (PU) fat (corn oil). Accordingly, the fate of 14C free cholesterol originating from high- or low-density lipoproteins (LDL) was examined in the biliary, hepatic and plasmatic lipids, as well as the activity of two key enzymes in the metabolism of lipoproteins: lipoprotein lipase (LPL) and lecithin cholesterol acyl transferase (LCAT). The LPL activity increased in the S diet, in comparison to the PU diet or to a low fat (6%) control (C) diet and the LCAT activity increased but not significantly in the PU diet. In bile the secretion of 14C-cholesterol and 14C-bile salts originating from 14C-cholesterol-HDL increased in the S diet compared to the PU diet and a C diet [previous results]. S and PU diets increased to the same extent the hepatic storage of 14C-esterified cholesterol originating from LDL, compared to the C diet. This cholesterol would contribute to a greater extent to the hepatic synthesis of the lipoproteins destined for the plasma in the case of the S diet than that of PU diet. These results may be explained by the adaptation of hepatic acyl cholesterol acyl transferase and cholesterolesterase to both high-fat-diet enzymes acting simultaneously on the two free and esterified cholesterol compartments. It resulted in an important redistribution of the cholesterol of these two compartments between plasma, bile and liver.
Asunto(s)
Colesterol/metabolismo , Grasas de la Dieta/administración & dosificación , Lipoproteínas HDL/análisis , Lipoproteínas LDL/análisis , Animales , Bilis/metabolismo , Colesterol en la Dieta/metabolismo , HDL-Colesterol/metabolismo , Grasas de la Dieta/farmacología , Lipoproteína Lipasa/metabolismo , Liposomas/metabolismo , Hígado/enzimología , Hígado/metabolismo , Masculino , Fosfatidilcolina-Esterol O-Aciltransferasa/metabolismo , Ratas , Ratas EndogámicasRESUMEN
1. Four groups of adult male Sprague-Dawley rats were fed for 6 weeks on a diet with a low-fat content (50 g/kg) and another four groups were given a diet rich in fat (250 g/kg) and cholesterol (12 g/kg). In both cases, the basal diets were either fibre-depleted or supplemented with cellulose (60 g/kg), wheat bran (100 g/kg) or low-methoxyl pectin (100 g/kg). 2. Low-methoxyl pectin displayed the most hypocholesterolaemic effect and decreased the cholesterol content of the very-low-density lipoproteins (VLDL) and low-density lipoproteins (LDL), when the low-fat diet was given. When rats were fed on the high-fat diet, pectin no longer had a hypocholesterolaemic effect but still decreased the VLDL-cholesterol content. Pectin lowered serum triglyceride and VLDL-triglyceride levels only when the low-fat diet was given. 3. Wheat bran exerted no hypocholesterolaemic effect in rats fed on the low- and high-fat diets, but decreased the cholesterol content of VLDL and lowered serum triglycerides and VLDL-triglycerides when the high-fat diet was given. 4. Purified cellulose had no significant effect on plasma lipids. 5. As shown by multivariance analysis, low-methoxyl pectin and wheat bran both beneficially modified the serum triglyceride and cholesterol variables except VLDL-triglycerides. However, the magnitude of the effect of each individual type of fibre was dependent on the fat and cholesterol content of the diet, suggesting the existence of different mechanisms of action.
Asunto(s)
Celulosa/farmacología , Grasas de la Dieta/administración & dosificación , Fibras de la Dieta/farmacología , Lípidos/sangre , Pectinas/farmacología , Animales , Colesterol/sangre , Colesterol en la Dieta/administración & dosificación , Lipoproteínas LDL/sangre , Lipoproteínas VLDL/sangre , Masculino , Ratas , Ratas Endogámicas , Triglicéridos/sangreRESUMEN
The effects of high fat diet and injection of chlorpromazine on bile lipid secretion were studied in the rats fed a control diet (C), a saturated fat, high cholesterol diet (S) and a polyunsaturated fat, high cholesterol diet (PU). As compared to controls, injection of chlorpromazine in the S and PU diet groups caused no appreciable change in the level of bile salts and bile phospholipids. Chlorpromazine did however enhance bile cholesterol, especially in the PU group, and lower secretion of lysosomal enzyme (beta-glucuronidase) into bile. Impairment of lysosomal enzyme secretion but not of bile lipid secretion suggests that the lysosomal activity is not directly involved in the bile secretion mechanism. These data point up the risks of using chlorpromazine therapy in association with a diet high in fat and cholesterol.
Asunto(s)
Bilis/metabolismo , Clorpromazina/farmacología , Grasas de la Dieta/farmacología , Animales , Ácidos y Sales Biliares/metabolismo , Colesterol/metabolismo , Glucuronidasa/metabolismo , Hígado/metabolismo , Masculino , Ratas , Ratas Endogámicas , Transaminasas/sangre , Triglicéridos/metabolismoRESUMEN
Determination of the physicochemical parameters of a lipoprotein class from weight percentage data in light of a recent quantitative biodynamic concept which strictly adheres to the CGS unit system (cm, g, s) and the mass/volume chemical unit, mol.cm-3.
Asunto(s)
Lipoproteínas , Fenómenos Químicos , Química Física , Matemática , Peso Molecular , Pesos y MedidasRESUMEN
Determination of the physicochemical parameters of unilamellar and multilamellar liposomes from concentration data in light of a recent quantitative biodynamic concept which strictly adheres to the CGS unit system (cm, g, s) and the mass/volume chemical unit, mol.cm-3.
Asunto(s)
Liposomas , Fenómenos Químicos , Química Física , Matemática , Pesos y MedidasRESUMEN
Active biological systems can be divided into five phases: the aqueous polar phase, the monolayer and/or bilayer interfacial phase, the apolar or hydrophobic phase, the solid or insoluble phase and the gaseous phase. The micellar phase is a special dispersed state of an interfacial phase. Molecules are distributed among these five phases according to their physicochemical properties. Herein is proposed a standardization in strict compliance with the CGS (cm, g, s) unit system and uses the mass/volume, mole per cm3 (mol X cm-3) chemical unit. This standardization requires a new set of symbols to clearly distinguish the concentrations in the different phases. The numerous implications of this standardization are discussed with respect to the quantitative classification of lipids based upon interphase partition coefficients, a new definition of micelles, simple models for the study of lipid biodynamic behavior and sites of action of lipid metabolism enzymes as well as determination of the physicochemical parameters of circulating lipoproteins. By compartmentalization in an aqueous polar phase, an interfacial phase comprising phospholipids and free cholesterol and an apolar phase comprising triglycerides and esterified cholesterol, this standardization will greatly simplify quantitative research on the factors regulating and disturbing cholesterol homeostasis. The notion of total cholesterol must be foresaken, since the biodynamic behavior of free cholesterol and esterified cholesterol are fundamentally different. Free cholesterol shares the fate of the interfacial phase of which it is a part, this fate being hinged on enzymatic biotransformations and/or ligand--receptor interactions. The proposed standardization gives rise to a new rationale using simple calculations and its advantage will be 2-fold: first, in the design of experimental protocols; and second, in allowing immediate and unambiguous comparison of experimental data based upon strictly defined parameters.
Asunto(s)
Lípidos/fisiología , Animales , Humanos , Peso MolecularRESUMEN
Isolated perfused rat liver was used to study the effects of constant taurocholate perfusion, with or without the addition of phosphatidylcholine unilamellar vesicles, upon both the bile salt-dependent and bile salt-independent secretion of bile. Taurocholate introduction increased bile flow and normalized the bile lipid secretion by restoring the bile salt-dependent secretion. At a flow rate of 30 ml/min, the liver was perfused by a single-pass method. The perfusion medium contained 17.5 microM taurocholate with or without 5.83 microM phosphatidylcholine. In light of a recent quantitative dynamic concept on the interphase partition of lipids, it was calculated that more than 99% of the taurocholate reaches the liver as monomers and/or dimers. It was also deduced that the lipids were secreted in bile as small discoidal lipoprotein structures rather than unilamellar lipoproteic vesicles. During the course of the experiments (2 hr), the excellent criteria of viability of this model make it highly suitable for the investigation of hepatic metabolism. Furthermore, the addition of phosphatidylcholine unilamellar vesicles to the perfusate constitutes a potential vector for various liposoluble molecular species.
Asunto(s)
Bilis/metabolismo , Metabolismo de los Lípidos , Hígado/metabolismo , Animales , Ácidos y Sales Biliares/metabolismo , Liposomas/metabolismo , Perfusión , Fosfatidilcolinas , Ratas , Ácido Taurocólico/farmacologíaRESUMEN
A system is proposed for a quantitative classification of lipids, based on interphase partition coefficients. This system enables calculation of exchanges of lipid molecules between phases. The mass/volume chemical unit mol X cm-3, strictly derived from the CGS system, is used, thus simplifying mathematical relations. Applied to bile salt-lecithin-cholesterol mixed micelles, this dynamic concept gives new insight into the variations of physico-chemical parameters. Experimental results obtained with the glycodesoxycholate and the taurocholate show a striking difference in partition coefficients between aqueous and mixed bile salt-lecithin interfacial phases. A new model applying triangular co-ordinates to a bile salt-lecithin-cholesterol mixed lipid phase is described.
Asunto(s)
Ácidos y Sales Biliares , Colesterol , Lípidos , Fosfatidilcolinas , Fenómenos Químicos , Química , Cinética , Matemática , Micelas , Conformación Molecular , Peso MolecularRESUMEN
The two main proteic constituents of the human Apo-bile lipoprotein complex (BLC), i.e., the anionic polypeptide fraction (APF) and the IgA fragments, were separated by preparative zonal ultracentrifugation using a sucrose gradient containing 1.5 mM glycodesoxycholate. The purification of the APF was verified by sodium dodecyl sulphate (SDS) polyacrylamide gel electrophoresis and immunology, and its amino acid composition then was determined. This procedure was used to obtain a polyclonal antiserum directed solely against the APF.
Asunto(s)
Apolipoproteínas/aislamiento & purificación , Bilis/análisis , Péptidos/aislamiento & purificación , Aminoácidos/análisis , Centrifugación Zonal/métodos , Humanos , Sueros Inmunes , Inmunodifusión , Peso MolecularRESUMEN
Intralipid was incubated with pancreatic lipase (EC 3.1.1.3) and/or phospholipase A2 (EC 3.1.1.4) at two bile salts/phosphatidylcholine molar ratios and at two different triglyceride hydrolysis rates using various amounts of lipase. Incubations were studied by gel filtration. Results show: During lipase action, three phases of lipids coexist: an emulsified phase, a micellar phase and an intermediate heavy phase sized between the two others. The equilibrium between each phase is dependent upon the bile salts concentration. Under these conditions, pancreatic lipase was at 60% bound to the emulsified phase, whereas pancreatic phospholipase A2 was bound at 94% to the micellar phase.
Asunto(s)
Lipasa/metabolismo , Páncreas/enzimología , Fosfolipasas A/metabolismo , Fosfolipasas/metabolismo , Animales , Radioisótopos de Carbono , Cromatografía en Gel , Emulsiones Grasas Intravenosas , Hidrólisis , Cinética , Fosfolipasas A2 , Porcinos , TrioleínaRESUMEN
Most experiments were conducted in the presence of human gallbladder bile; colipase and pancreatic lipase were purified using porcine pancreas. The adsorption of bile salts, phospholipids and cholesterol from the bile, together with that of pancreatic lipase was measured on wheat bran, cellulose, hemicellulose (xylan), slightly methylated pectin (42%) and cholestyramine. In contrast to cholestyramine which intensively binds biliary lipids (61.7-81.7%) and pancreatic lipase (47.5%), the fibers studied only had a low adsorbent power. The direct influence of these fibers and of cholestyramine at concentrations ranging from 0-5% on lipase activity was measured at constant pH, using two conventional assay systems, long chain triglycerides and tributyrin. In the presence of human bile and colipase, a drastic reduction in triglyceride hydrolysis by lipase was observed with cholestyramine (loss of 66-82%) and wheat bran (loss of 77-94%) at 1% concentration. The other fibers did not have any marked effects on enzyme activity. The use of a radio labeled lipase made it possible to demonstrate that the inhibitory effect of bran on enzyme activity was independent of adsorption phenomena on bran. The fraction of bran that can be solubilized in the aqueous phase, in fact, induced this reduction in activity. The presence of protein inhibitor in bran may be responsible for the reduction in pancreatic lipase activity.
Asunto(s)
Resina de Colestiramina/farmacología , Fibras de la Dieta/farmacología , Lipasa/antagonistas & inhibidores , Páncreas/enzimología , Animales , Bilis/metabolismo , Celulosa/farmacología , Humanos , Técnicas In Vitro , Lipasa/metabolismo , Metabolismo de los Lípidos , Pectinas/farmacología , Polisacáridos/farmacología , Unión Proteica/efectos de los fármacos , Porcinos , Triglicéridos/metabolismoRESUMEN
We assayed the lipid content of bile from rats that had been fed either a standard diet (5% fat) or a high fat diet (25% fat, 1.2% cholesterol) in the presence or in the absence of various dietary fibers (namely, wheat bran, pectin and cellulose). The cholesterol concentration in bile from rats fed the high fat diet plus wheat bran or pectin was lower than that of the rats fed the high fat, high cholesterol diet without fiber. Bile phospholipids did not vary significantly from one group to another. In comparison to the standard diet, the high fat, high cholesterol diet led to a greater ratio of primary to secondary bile salts and a higher level of glycoconjugates. The observed differences may be explained by a variation in the metabolism of bile salts brought about by the difference in diet.
Asunto(s)
Bilis/metabolismo , Celulosa/farmacología , Fibras de la Dieta/farmacología , Metabolismo de los Lípidos , Pectinas/farmacología , Animales , Ácidos y Sales Biliares/metabolismo , Peso Corporal/efectos de los fármacos , Colesterol en la Dieta/farmacología , Grasas de la Dieta/farmacología , Masculino , Ratas , Ratas Endogámicas , TriticumRESUMEN
Twenty-one male Wistar rats, 13 weeks old, were fed ad libitum hyperlipidic diets (28% fats) loaded with cholesterol (1.2%) for 5 weeks. One group of 11 rats was fed saturated fats (diet group "S") and another group of 10 rats was fed polyunsaturated fats (diet group "PU"). On the day they were sacrificed 10 of the rats were injected intravenously with 1 mg of filipin. Contrary to the rats in diet group "PU," the rats in diet group "S" treated with filipin presented certain characteristics that were not found in the nontreated group: They provided evidence of biliary cholestasis accompanied by a decline in the level of secretion of bile salts and phospholipids into bile. The concentrations of both free and esterified cholesterol in plasma fell and the amount of (esterified) hepatic cholesterol rose, although there was no change due to the filipin in the amounts of hepatic phospholipids. Explanatory hypotheses for these phenomena were considered, first, at the site of plasma membranes where filipin binds selectively to the cholesterol in the membrane, causing a disruption which probably disturbs the absorbance of circulating lipoproteins, especially that of hepatocyte cells, particularly in diet group "PU." Second, the effects of filipin on subcellular membranes seem to disturb the secretion of lipids and lipoproteins into bile and plasma, especially in diet group "S." Last, at the intracellular level, filipin appears to have a blocking effect on the organelles involved in biliary lipid secretion. The activity of certain enzymes such as cholesterol esterase may also be blocked, particularly in diet group "S," which would explain the accumulation of esterified cholesterol in liver.
Asunto(s)
Bilis/efectos de los fármacos , Colesterol/metabolismo , Filipina/farmacología , Hígado/efectos de los fármacos , Polienos/farmacología , Alanina Transaminasa/metabolismo , Animales , Bilis/metabolismo , Ácidos y Sales Biliares/metabolismo , Membrana Celular/efectos de los fármacos , Colesterol/sangre , Ésteres del Colesterol/metabolismo , Grasas de la Dieta/administración & dosificación , Grasas Insaturadas/administración & dosificación , Glucuronidasa/metabolismo , Hiperlipidemias/etiología , Hiperlipidemias/metabolismo , Hígado/metabolismo , Masculino , Lípidos de la Membrana/metabolismo , Fosfolípidos/metabolismo , Ratas , Ratas Endogámicas , Fracciones Subcelulares/efectos de los fármacos , Fracciones Subcelulares/metabolismoRESUMEN
Phospholipids and cholesterol combine with a protein fraction (IgA and an acid polypeptide) in bile to form the bile lipoprotein complex. We wished to determine whether lysosomes participated only on IgA secretion or if their secretory role also involved the lipid components of the bile complex. This aspect was studied with a single acute injection of chloroquine, a lysosomotropic drug. The results show that a nonnegligible quantity of IgA travels through the lysosomes. In addition, phospholipid and cholesterol levels undergo a significant (P less than 0.05) decrease 1 hr after injection before increasing to normal levels. In contrast to the total inhibition of protein secretion (beta-glucuronidase, acid phosphatase), a transitory decrease of the secretion of bile lipids takes place that suggest secretory mechanisms involving organelles other than lysosomes.
Asunto(s)
Fosfatasa Ácida/metabolismo , Bilis/metabolismo , Cloroquina/farmacología , Glucuronidasa/metabolismo , Metabolismo de los Lípidos , Lisosomas/enzimología , Animales , Bilis/efectos de los fármacos , Cloroquina/administración & dosificación , Colesterol/sangre , Inyecciones Intravenosas , Cinética , Lisosomas/efectos de los fármacos , Masculino , Ratas , Ratas Endogámicas , Albúmina Sérica/metabolismo , Triglicéridos/sangreRESUMEN
The adsorption of pancreatic phospholipase was studied in vitro in the presence of egg yolk lipoprotein emulsion, Intralipid emulsion, and milk fat globules. When the emulsions are incubated with bile salts, the latter dissociate a considerable fraction of the phospholipids initially associated with the emulsions, leading to the coexistence of an emulsified phase and a phase of mixed micelles. After the addition of pancreatic phospholipase A2, gel filtration shows that the enzyme was more than 90% bound to mixed micelles, regardless of the type of emulsion used. Comparable results were obtained by replacing the bile salts with human gallbladder bile. In parallel, pancreatic zymogen was never found to be bound to any of the lipid structures present (emulsion or mixed micelles). When the catalytic site of pancreatic phospholipase A2 was blocked with 4-bromophenacylbromide, there was no fixation on mixed micelles. Fixation was restored by the presence of lysolecithins and fatty acids in the incubation mixtures. The partial transformation of all emulsified substrates to mixed micelles by bile salts in vivo would thus lead to optimum activity of pancreatic phospholipase A2.
Asunto(s)
Páncreas/enzimología , Fosfolipasas A/metabolismo , Fosfolipasas/metabolismo , Adsorción , Animales , Ácidos y Sales Biliares/metabolismo , Bovinos , Cromatografía en Gel , Yema de Huevo , Emulsiones Grasas Intravenosas/metabolismo , Femenino , Humanos , Lipoproteínas/metabolismo , Micelas , Leche/metabolismo , Fosfolipasas A2 , PorcinosRESUMEN
The human bile lipoprotein complex includes an apoprotein fraction of IgA fragments and an acidic polypeptide tightly bound to bile cholesterol and phosphatidylcholines. The fate of the intestinal bile lipoprotein complex was immunohistochemically studied. Its localization in the human duodenum is consistent with selective capture or synthesis. The result of both might be the presence of bile apoprotein in the bloodstream. These two mechanisms may explain the cross-reaction between the lipoprotein complex apoprotein and the high density lipoprotein.
Asunto(s)
Apolipoproteínas/análisis , Bilis/análisis , Mucosa Intestinal/metabolismo , Duodeno/metabolismo , Histocitoquímica , Humanos , Inmunoglobulina A/análisis , Fragmentos de Inmunoglobulinas/análisisRESUMEN
In bile, phospholipids and cholesterol were found in close association with an apoprotein fraction which cross-reacted to antiserum anti-IgA. In order to determine the origin of the IgA part of the apo bile lipoprotein complex and to establish its relationship with IgA, th localizations of these two components in rat and in dog livers were compared. Two immunohistochemical methods were used. Whereas IgA was found on the sinusoidal surface of the hepatocytes, in cytoplasmic particles and in bile canaliculi, the IgA part of the apo bile lipoprotein complex was only detected in the bile canaliculi, and in particles which presented the same pattern of location as lysosomes. These results suggest that the IgA part of the apo bile lipoprotein complex could originate from the transit of a part of the internalized IgA via the lysosomal compartment of the hepatocytes.