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Biomedical subjects

R Dargel

Publications and source records attributed to R Dargel.

At least 37 records · Page 2Linked to original sources
PubMed1994

Zonal differences in lipoprotein formation in the thioacetamide-induced micronodular-cirrhotic rat liver.

The aim of the studies was to answer the question to what extent thioacetamide-induced structural alterations of hepatic architecture leading to fibrosis and micronodular pseudolobuli affect the formation of very low density lipoproteins and the zonation of lipoprotein metabolism observed in normal and acutely injured livers. Therefore, the number of the VLDL particles/Golgi complex and the relative specific volume of Golgi complexes as well as the number and relative specific volume of VLDL-filled vesicles was determined in lobular and nodular zones of normal and the micronodular-cirrhotic livers, respectively. The perinodular and centrinodular regions were morphometrically analysed in nodules with diameters between 0.3 and 0.5 mm. -Generally, in thioacetamide-induced micronodular liver cirrhosis a zonality was observed with respect to the amount of VLDL particles as well as the number and volume of organelles involved in the formation and secretion of hepatic lipoproteins. However, the number of VLDL particles/Golgi complex was significantly reduced to 52% in the centrinodular and to 71% in the perinodular region of cirrhotic livers when compared with the corresponding periportal and perivenous area, respectively. Furthermore, the relative specific volume of the Golgi complexes markedly increased in the perinodular region, thus abolishing the zonal difference observed in the controls.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1993

Low-density lipoprotein catabolism does not respond to estrogen in the fetal and newborn rat.

The elimination of native and methylated low-density lipoprotein (LDL) from serum and the effect of estradiol on the serum LDL-apoB pool, the uptake of homologous [125I]-LDL into liver and adrenals, and the fractional catabolic rate (FCR) of [125I]-LDL was studied in fetal (22nd day of gestation), newborn (15th day postpartum), and adult rats. In fetal rats the receptor-mediated LDL decay accounted only for 47%, whereas in adults it was calculated to 65%. In the latter, estrogen caused (1) a diminution of the serum LDL-apoB pool by 85%; (2) an enhancement of the LDL uptake into the liver and the adrenals by 5- and 10-fold, respectively, and (3) an acceleration of the [125I]-LDL decay in the serum with a rise of the FCR by 3-fold. In contrast, neither the LDL pool and uptake nor the LDL elimination (FCR) did respond to estrogen in fetal and newborn rats. In summary, the LDL catabolism of the rat is insensitive towards estrogen in the late gestational period and during the first 2 weeks of postnatal life [3].

Adrenal Glands↗
PubMed1993

Stimulation of bile flow and inhibition of biliary secretion of taurocholate and leukotriene C4 in thioacetamide-induced liver fibrosis.

In order to study the relation of hepatic fibrogenesis to biliary elimination, female Uje:WIST rats were treated with thioacetamide (TAA). This treatment results in single cell necrosis, fibrosis, nodular parenchyma und proliferation of bile ducts. In isolated perfused livers from pretreated rats, liver hemodynamics, bile flow, and secretion of taurocholate and leukotriene C4 were determined. After TAA-pretreatment, sinusoidal efflux of glucose and pyruvate was reduced increasing lactate/pyruvate ratio threefold. Biliary secretion of [14C]taurocholate and [3H]leukotriene C4 was lowered by 47% and 35%, respectively, compared to controls. In contrast, basal bile flow was increased after TAA-treatment. The beta-adrenergic agonist isoproterenol stimulated bile flow threefold over controls. The increased stimulation was correlated with increased liver weight after TAA-treatment. We postulate that both increased bile flow and stimulation by isoproterenol are due to proliferation of bile ducts and increased ductular bile secretion partially compensating for disturbed hepatocellular secretion.

Animals↗
PubMed1993

[Isolation and characterization of hepatocytes from chronic cholestatic damage to rat livers after bile duct ligation].

The aim of these investigations was the isolation and characterisation of hepatocytes from chronic injured rat livers after bile duct ligation. In three months old female Uje:WIST rats the distal common bile duct was double ligated. 14 days after the ligation 80% of the rats survived. The content of hepatic OH-prolin was increased in relation to shamoperated control animals. The histological examination showed a marked fibrous and oedematous spreading of the portal tract as well as an extensive bile duct proliferation after 14 days. A correlation between OH-prolin content of the liver and index of fibrosis was found. 14 days after ligation the hepatocytes were isolated according to Seglen (7). The yield of hepatocytes isolated from the liver of cholestatic rats and from control animals was on average 4 x 10(8) cells with a viability of 80%. Ultrastructural the most hepatocytes showed intact organells without injury. We can conclude, that it is possible to isolate hepatocytes from chronic cholestatic rat livers with a good yield and viability as well as with an intact ultrastructure. These cells are suitable for additional metabolic investigations.

Animals↗
PubMed1992

The visceral yolk sac--an important site of synthesis and secretion of apolipoprotein B containing lipoproteins in the feto-placental unit of the rat.

Rat fetuses exhibit a high serum LDL concentration at term. Delivery caused a marked decrease of the LDL apolipoprotein (apo) B concentration independent of whether this occurred on days 21, 22 or 23 of gestation. The interruption of the yolk sac circulation by a ligature in situ for 6 h led to the same alterations of the LDL-apo B concentration as Caesarean section. Immunoelectronmicroscopic studies provided evidence that the epithelial cells of the visceral yolk sac exhibited electron dense LDL-sized and apo B containing particles which were localized over the compartments of the Golgi complexes, endoplasmatic reticulum, secretory vesicles and intercellular spaces, but not over the cell nuclei, mitochondria or lysosomes. ApoB containing LDL-sized particles could be obtained by ultracentrifugation from the disrupted material of the microsomal fraction of yolk sac homogenates. Isolated segments of the yolk sac membranes were capable to secrete apoB containing lipoproteins floating in the d less than 1.020 g/ml as well as in the d = 1.020-1.064 g/ml fraction with a 10-fold higher amount of apoB in the higher density class. Incorporation experiments with [35S] methionine gave evidence that these lipoproteins were at least partially provided with newly synthesized apoB predominantly found in the LDL fraction. The size of the negatively stained particles in the d = 1.020-1.064 g/ml fraction secreted from yolk sac segments corresponded to that of LDL from fetal rat serum. In contrast their acylglycerol content was significantly higher, whereas the percentage contribution of total cholesterol and protein was markedly reduced in comparison with serum LDL of the fetus. In summary, biochemical and ultrastructural studies provide clear cut evidence that the rat yolk sac is able to synthesize and to deliver apo B containing lipoproteins in the density ranges of VLDL, IDL and particular of LDL thus contributing to the supply of serum lipoproteins in the rat fetus. By recalculation of recent tracer kinetic data (Plonné et al. (1990) J. Lipid Res. 31, 747) using a mathematical step function model it was possible to assess the contribution of the rat yolk sac to the LDL influx into the fetal serum.

Animals↗
PubMed1992

Metabolic and hemodynamic responses of bivascularly perfused rat liver to nerve stimulation, noradrenaline, acetylcholine and glucagon in thioacetamide-induced micronodular cirrhosis.

Thioacetamide-induced rat cirrhosis was characterized by single-cell necroses, fibrosis, nodular parenchyma, decrease in parenchymal volume density and an increase in liver weight per body weight so that the total amount of parenchyma was not altered. The glycogen content was normal, and signs of decompensation were not found. Isolated livers were single-pass perfused by way of both the hepatic artery and the portal vein. In the normal livers stimulation of the nerve plexuses around the hepatic artery or portal vein (7.5 Hz; 2 msec) and infusions of noradrenaline (1 mumol/L) by way of either vessel and of acetylcholine (10 mumol/L) by way of the artery only increased glucose output, reduced both portal and arterial flow and increased the intravascular pressures. Glucagon (0.5 nmol/L) augmented glucose release and had no hemodynamic effects. In chronically thioacetamide-injured livers all stimuli caused smaller metabolic alterations per gram of liver weight and decreased portal flow more and arterial flow less with stronger enhancements of intravascular pressures than in the controls. The lowered metabolic responsiveness per gram of cirrhotic liver was largely compensated by the increase in liver weight. Thus despite massive histological alterations and pronounced increases in stimulation-dependent resistances - predominantly in the portal system - cirrhotic rat livers responded in their glucose metabolism to nervous and hormonal stimuli in almost the same manner as normal livers.

Acetylcholine↗
PubMed1992

Synthesis, secretion and immunoelectron microscopic demonstration of apolipoprotein B-containing lipoprotein particles in the visceral rat yolk sac.

Electron microscopic investigations on the involvement of the fetal membranes of the rat (visceral yolk sac) in the lipid metabolism revealed the occurrence of lipoprotein-sized particles located in cisternal Golgi stacks, Golgi vesicles and secretory vesicles of the cells of the visceral yolk sac epithelium as well as in distended areas of the intercellular space between adjacent epithelial cells. Application of the protein A-gold technique with specific anti-apoB antiserum resulted in a specific location of immunogold both over the different compartments of the lipoprotein pathway (RER, Golgi complex, secretory vesicles) as well as over the distended intercellular spaces, thus confirming these particles to be lipoproteins in nature. Isolated visceral epithelial cells prepared by a tryptic digestion method exhibited some ultrastructural alterations, such as a loss of apical brush border, a change from columnar to spherical cell shape, a decrease in phagolysosomes, but an increase in autophagosomal structures after 6 h incubation at a vitality rate of at least 85%. Within this period the epithelial cells secreted measurable amounts of apoB-containing lipoproteins into the medium floating in the density classes d less than 1.006 g/ml, d = 1.006-1.020 g/ml and d = 1.020-1.064 g/ml. The production of the lipoproteins was partly inhibited by cycloheximide indicating the secretion of particles with performed as well as newly synthesized apoB. Negative staining of the particles revealed an average diameter of 34 nm of VLDL, 31 nm of IDL and 24 nm of LDL. In summary, our studies demonstrate that in the feto-placental unit of the rat the fetal membranes are capable of synthesizing and secreting lipoproteins. The cells of the visceral yolk sac epithelium were shown to be the producers of apoB-containing particles.

Animals↗
PubMed1992

Quantitative studies on fatty acid metabolism in isolated parenchymal cells from normal and cirrhotic livers in rats.

Parenchymal cells isolated from normal and thioacetamide-induced micronodular cirrhotic rat livers were used to evaluate changes in hepatocellular fatty acid (FA) metabolism in cirrhosis. Exogenous free FA (FFA) are rapidly taken up by hepatocytes obtained either from normal or cirrhotic livers. They are predominantly esterified to triglycerides and accumulate intracytoplasmatically as lipid droplets with increasing cellular FFA uptake. In the parenchymal cells of cirrhotic livers, however, the following changes were observed when compared with controls: (i) decreased cellular output of esterified FA derived both from exogenous sources and from de novo FA synthesis; (ii) increased total ketone body production, mainly as beta-hydroxybutyrate; (iii) decreased cholesterol synthesis; and (iv) enhanced incorporation of newly synthesized FA into phospholipids in spite of an unaffected rate of FA synthesis. In summary, the data provide evidence for intrinsic alterations in the lipid metabolism in the parenchymal cells of cirrhotic livers which are preserved in the isolated hepatocytes under optimum incubation conditions for oxygen and substrate supply.

3-Hydroxybutyric Acid↗
PubMed1992

Lipid peroxidation--a common pathogenetic mechanism?

Lipid peroxidation is considered at present as one of the basic mechanisms involved in reversible and irreversible cell and tissue damage. The current knowledge about the role of peroxidative breakdown of polyunsaturated fatty acids in the pathogenesis of various diseases has been reviewed. Lipid peroxidation leads to degradation of the lipid membrane, interaction of degradation products with intra- and extracellular targets and to the production of new reactive oxygen species during the course of the chain reaction thus leading to damage of cells and tissues. According to our current view lipid peroxidation is implicated in the pathogenesis of cancer, inflammatory processes, atherosclerosis, toxic injury by xenobiotics and ischemic-reperfusion damage.

Arteriosclerosis↗
PubMed1991

Lipid peroxidation in thioacetamide-induced macronodular rat liver cirrhosis.

Microsomes and isolated hepatocytes from thioacetamide (TAA)-induced macronodularly cirrhotic rat livers were analysed for their susceptibility to unstimulated and stimulated lipid peroxidation measured as malondialdehyde (MDA) formation. In microsomes from TAA-induced macronodularly cirrhotic livers the MDA production stimulated either by ascorbate-iron or by ADP-iron in a NADPH-regenerating system was decreased. Hepatic microsomes from TAA-treated rats exhibited a reduced cytochrome P450 content and lowered activities of ethylmorphine N-demethylase, ethoxycoumarin O-deethylase and epoxide hydrolase. Besides this, the microsomal fatty acid pattern of phosphatidylcholine and phosphatidylethanolamine was significantly changed after 6 months of TAA administration. The 18:2/20:4 ratio of phospholipid fatty acids was markedly increased. In contrast to the microsomes, in isolated hepatocytes from macronodularly cirrhotic livers the iron- and ascorbate-iron-stimulated MDA formation was increased. The hepatocellular GSH content was unaffected by TAA pretreatment, whereas the GSSG content exhibited a significant increase, thus leading to a pronounced reduction of the GSH/GSSG ratio. The calcium channel blocker verapamil (200 microM), known to be able to scavenge OH' radicals produced by the Fenton reaction, revealed an inhibitory effect on ascorbate-iron- and ADP-iron-stimulated lipid peroxidation in hepatocytes from normal as well as TAA-treated livers which is attributed to its antioxidative properties. In summary, lipid peroxidation is altered in TAA-induced macronodularly cirrhotic rat livers. Furthermore, the data clearly show that isolated microsomes and parenchymal cells prepared from cirrhotic livers react differently to prooxidant stimuli.

Animals↗
PubMed1991

[Microsomal phospholipid pattern and liver ultrastructure of aging rats following treatment with essential phospholipids].

The treatment of aging rats with 100 or 300 mg/kg "essential" phospholipids (EPL) for 10 weeks significantly changed the microsomal concentration of phosphatidyl serine plus phosphatidyl inositol. The most marked alterations of the phospholipid fatty acid pattern predominantly observed with unsaturated fatty acids were detectable even 7 days after cessation of EPL application. Ultrastructurally, an enrichment in the hepatocytes with rough endoplasmic reticulum was visible which was more prominent in acinus zone 3. The changes after EPL administration are regarded as the expression of an increased membrane fluidity.

Aging↗
PubMed1990

Biochemical and ultrastructural studies on the effect of verapamil on formation and secretion of lipoproteins in rat hepatocyte suspensions.

The effect of the calcium channel blocker verapamil on structure, formation and secretion of very-low-density lipoproteins (VLDL) from rat hepatocytes in suspension was examined. After 30 min incubation at a verapamil dose of 200 microM neither free fatty acid (FFA) uptake nor triglyceride (TG) and phospholipid (PL) secretion into the incubation medium were significantly changed. After 90 min incubation the TG secretion was inhibited by about 60%, whereas the PL output was only insignificantly lowered, indicating the secretion of abnormally composed lipoprotein particles. Morphologically, after 30 min incubation the hepatocytes had lost their microvillous border and exhibited a 2-3-fold increase in volume density and average size of the lysosomes. In the Golgi-containing regions an accumulation of smooth-surfaced microvesicles was regularly evident. The configuration of the Golgi complexes was normal. After 90 min incubation the lysosomes showed a further significant elevation in volume and size. The Golgi complexes exhibited only minor changes, but their content in VLDL particles was reduced per microns 2 Golgi complex by about 75%. Commonly, the VLDL were larger and more heterogenous in size. The diameter of those VLDL secreted into the incubation medium ranged from 31 to 84 nm, thus surpassing the control values by 2-3 times. The secretion of large-sized VLDL was regularly associated with the intracytoplasmic appearance of dilated smooth-surfaced vesicles filled with size-modified VLDL. These vesicles were concentrated within Golgi-containing areas from where they were widely dispersed towards the cell periphery.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1990

The synthesis of glycosaminoglycans in isolated hepatocytes during experimental liver fibrogenesis.

During human and experimental liver fibrogenesis, the pattern of glycosaminoglycans in fibrotic liver matrix is greatly changed by severalfold increases of hyaluronic acid, chondroitin sulfate, and dermatan sulfate, respectively. The present study aimed to determine whether hepatocytes take part during fibrogenesis in the alteration of the glycosaminoglycan profile in liver matrix. Rats received thioacetamide orally for 2 and 10 weeks, respectively. After 10 weeks a typical micronodular cirrhosis had developed. Hepatocytes isolated at these time points were characterized by light and electron microscopy and incubated for up to 4 h in suspension cultures in [35S]-sulfate and [3H]-glucosamine containing medium to study the synthesis and intra-/extracellular distribution of total and specific types of glycosaminoglycans. A biphasic change of glycosaminoglycan synthesis in hepatocytes was found. After 2 weeks of TAA-treatment parenchymal cells synthesized about 25% more labeled glycosaminoglycans than control liver cells, but at 10 weeks the synthesis was reduced by more than 40%. Thus, between 2 and 10 weeks of TAA-treatment hepatocellular glycosaminoglycan synthesis decreased by more than 50%. The major portion of newly synthesized glycosaminoglycans was nitrous acid labile and, hence, identified as heparan sulfate. Its fractional synthesis decreased from 0.90 in control cells to 0.84 (2 weeks TAA) and 0.76 (10 weeks TAA), respectively. Thus, the absolute synthesis of heparan sulfate was reduced by 50% in hepatocytes from cirrhosis liver. Eighty to 90% of labeled glycosaminoglycans remained cell-associated. Hyaluronic acid was detected neither in normal hepatocytes nor in hepatocytes from injured liver. We conclude from these data that parenchymal liver cells will not contribute actively to the accumulation of galactosaminoglycans (chondroitin sulfate, dermatan sulfate) and hyaluronic acid in the extracellular matrix during fibrogenesis. The diminished rate of synthesis of heparan sulfate in hepatocytes from cirrhotic liver might explain its fractional decrease in cirrhotic liver matrix.

Animals↗
PubMed1990

[Quantitative determination of apolipoprotein AI in serum by the use of monoclonal antibodies in a noncompetitive ELISA].

A methodical base for estimation of apolipoprotein AI (apo AI) in the serum using a monoclonal antibody in a non-competitive ELISA is described. The technical approach is adjusted to commercial materials of the DDR industry. The assay allows to determine apo AI within the range of 5-100 micrograms apo AI/I. The Vk was 4% (within), and 8% (day to day). The monoclonal antibody used is directed against a native epitope which is fully exposed in the serum. Application of the SUMAL system further improves the practicability of the method. The assay should be suitable particularly for the recognition of epitopes of apo AI in serum HDL fractions associated with an atherogenic risk. The values obtained by the ELISA correspond to those measured turbidimetrically using polyclonal antibodies.

Antibodies, Monoclonal↗
PubMed1990

Development and regulation of hepatocellular fatty acid synthesis towards term: studies in isolated fetal rat hepatocytes.

De novo fatty acid synthesis was studied in hepatocytes freshly isolated from rat fetuses on day 20 and 22 (term) of gestation by using the 3H2O incorporation method. Within 1 h, about 4 nmoles palmitate equivalents/10(6) cells were synthesized on day 20 of gestation. Towards term this rate decreases to about two thirds of this value. Both on day 20 and 22 of gestation, fetal hepatocellular de novo fatty acid synthesis is stimulated by the addition of lactate and octanoate, whereas a stimulating effect by the addition of pyruvate was found on day 20 of gestation only. These data provide evidence that both cytosolic and mitochondrially derived NADH2 is an adequate source of reducing equivalents required for NADPH2-dependent fatty acid synthesis. Insulin stimulates fetal hepatocellular de novo fatty acid synthesis at term, whereas db-cAMP remains ineffective on both gestational days under study.

Animals↗