CD-fed Rats Research Articles

Essential pathogenic and metabolic differences in steatosis induced by choline or methione‐choline deficient diets in a rat model

Choline deficient (CD) and methione-choline deficient (MCD) diets are rodent models for steatosis, with potentially dissimilar biochemical backgrounds. The aim of this study was to assess the metabolic and pathological derangements in rats fed CD and MCD diets. Male Wistar rats received CD or MCD diet up to 7 weeks. Nutritional status, liver histopathology, Kupffer cell-mediated inflammation and injury, oxidative stress via thiobarbituric reactive species (TBARS), hepatic and plasma glutathione (GSH) and insulin homeostasis were assessed. In CD-fed rats, mainly microvesicular steatosis developed with occasional inflammatory cells. In MCD-fed rats, macrovesicular steatosis progressed to steatohepatitis (collagen deposition, activated stellate cells). Hepatic TBARS was increased and GSH decreased in the MCD-fed rats compared to no changes in the CD-fed rats. The CD-fed rats developed obesity, dyslipidemia and insulin resistance, in contrast to undetectable plasma lipids, unaffected insulin homeostasis and loss of body weight in the MCD-fed rats. The CD diet induced uncomplicated steatosis as compared to progressive inflammation and fibrinogenesis in the MCD diet. CD and MCD diets represent two pathogenically different models of steatosis. Although equivalence for the outcome of both diets can be found in clinical steatosis, the results of models using these diets should be compared with caution.

Fish Oil Affects Pancreatic Fat Storage, Pyruvate Dehydrogenase Complex Activity and Insulin Secretion in Rats Fed a Sucrose-Rich Diet

Rats fed a sucrose-rich diet (SRD) develop hypertriglyceridemia and a marked decline in beta cell function. The purpose of this study was to determine whether changes in triglyceride concentration and/or altered pyruvate dehydrogenase complex (PDHc) activity contribute to the beta cell dysfunction, and to analyze the effect of dietary fish oil on the altered patterns of insulin secretion and peripheral insulin resistance. Rats were fed an SRD for 210 d. One-half of the rats continued consuming the SRD until d 270. The other half received an SRD in which fish oil (FO) was partially substituted for corn oil until d 270. A group of rats was fed a control diet (CD) throughout the experiment. The islets of rats fed the SRD had a greater triglyceride concentration and lower PDHc activity than those fed the CD. Insulin secretion patterns under the stimulus of glucose, palmitate or L-arginine were impaired in SRD-fed compared with CD-fed rats. This was accompanied by peripheral insulin resistance, mild hyperglycemia, a sharp increase of plasma triglyceride and free fatty acid levels and greater epididymal and retroperitoneal fat weights. FO normalized and/or improved these variables. Our results indicate that the increased fat storage and decreased PDHc activity in the beta cells play a key role in the abnormal insulin secretion of rats chronically fed an SRD. This is consistent with the reversion of these alterations by dietary FO.

Effects of a partially hydrolyzed curdlan on serum and hepatic cholesterol concentration, and cecal fermentation in rats.

A significant reduction was observed for serum and hepatic cholesterol concentrations in the rats fed diet containing a 5% partially hydrolyzed curdlan (PHCD), whereas only the hepatic cholesterol concentration was decreased in the curdlan (CD)-fed rats. The cecal contents in the CD group contained a significantly larger amount of short-chain fatty acids, but not those in the PHCD group. CD, but not PHCD, significantly increased the population of cecal bifidobacteria. From the in vitro fermentation test with cecal contents from cellulose powder (CP) and CD-fed rats, PHCD proved to be easily fermented by both cecal contents; incidentally CD was more susceptible to the cecal contents from CD-fed rats than to those from CP-fed rats. These results suggest that PHCD is involved in the modulation of lipid metabolism and intestinal microflora through a different manner from the native CD in rats.

Dietary curdlan increases proliferation of bifidobacteria in the cecum of rats.

Significant increases in the amounts of short-chain fatty acids and lactate, and in numbers of bifidobacteria were observed in the cecum of curdlan (CD) -fed rats as compared with those of cellulose-fed ones. The in vitro proliferation of 5 species of bifidobacteria was markedly increased in the cultures containing the supernatant obtained from the cecal contents of CD-fed rats. These findings suggest that bifidus factors have been produced in the cecum of CD-fed rats.

Determination of trace cadmium and other elements in bone by epithermal neutron activation analysis

Epithermal neutron activation analysis (ENAA) was applied to measure quantitatively Cd and other elements in bone samples from control and Cd-fed rats. This method was found to be non-destructive to the bone samples, with no sign of “radiolytic charring” and was sensitive enough to detect and quantify Cd in bone samples at normal levels for mammals (viz. 0.5–1.0 μg/g) and higher. Two different thermal neutron shield materials were utilized, namely cadmium and boron. The boron shield resulted in a 27% improvement in the detection limit of Cd in bone. The accuracy of ENAA for Cd was assessed by intercomparison with electrothermal atomic absorption spectrophotometry (ETAAS), and the results were in fair agreement (±23%) with those from ENAA.

Protection from chlordecone-amplified carbon tetrachloride toxicity by cyanidanol: Biochemical and histological studies

Chlordecone (CD) pretreatment is well known to greatly potentiate CCl 4 toxicity. Previous work has shown that suppression of hepatocellular regeneration permits an ordinarily limited liver injury to progress in an irreversible manner. Insufficient hepatocellular energy has been proposed as a mechanism for suppressed hepatocellular regeneration. Since cyanidanol reportedly increases cellular ATP, this compound was employed to test the above hypothesis. The present study was designed to investigate the sequential biochemical and histological changes over a time course of 120 hr after CCl 4 administration. Male Sprague-Dawley rats (125–150 g) were maintained on 10 ppm CD diet for 15 days and were challenged with either a standard protocol dose (100 μl/kg) or a low (50 μ/kg, L) dose of CCl 4. Cyanidanol pretreatment at 48, 24, and 2 hr before CCl 4 administration to rats maintained on CD diet resulted in 100 or 70% animal survival, for CCl 4 (L) or the standard dose of CCl 4, respectively. Preliminary studies indicated that neither simultaneous nor subsequent administration of cyanidanol with CCl 4 challenge affords such protection. Prior treatment with cyanidanol and a latency period were found necessary for protection. Without cyanidanol, CD + CCl 4 combination caused 50 and 100% lethality after CCl 4 (L) and the standard dose, respectively, while the same doses of CCl 4 alone did not cause lethal effects. Plasma enzymes (alanine aminotransferase, asparatate aminotransferase, sorbitol dehydrogenase) in control rats showed only moderate and transient increases after CCl 4 challenge. The combination of CD + standard dose of CCl 4 resulted in progressive and marked elevations of all three serum enzymes at all time intervals until the death of animals. Cyanidanol pretreatment resulted in significant decline in the plasma enzyme elevations at later time points. Cyanidanol pretreatment increased hepatic ATP synthesis in control or CD rats. CCl 4 administration to control rats did not alter hepatic ATP levels, while in CD-fed rats hepatic ATP levels were significantly decreased. Cyanidanol pretreatment to CD + CCl 4 combination-treated rats did not significantly prevent the decline in hepatic ATP and glycogen levels. However, in the surviving rats a recovery in these parameters was observed. Light microscopic examination of livers from animals that received CCl 4 alone revealed only marginal cellular injury, at early time points only. However, CCl 4 challenge to rats maintained on CD resulted in progressive injury, characterized by the appearance of ballooned cells, necrotic cells, and cells with lipid droplets in the liver. Cyanidanol pretreatment to these rats caused decreased vacuolation and significantly reduced the progression of liver necrosis. These results show that cyanidanol protects against an irreversibly progressive phase of liver injury and the interactive lethality of CD + CCl 4. In conclusion, the cyanidanol protection of CD-amplified CCl 4 toxicity requires a latency period and appears to be through mechanisms other than interference with the injurious events such as free radical injury, bioactivation, or covalent binding of CCl 4. These findings are consistent with the possibility that protection might be through a facilitation of cellular supply of energy.

Effect of choline-deficiency and methotrexate administration on peroxisomal beta-oxidation, palmitoyl-CoA hydrolase activity and the glutathione content in rat liver.

Hepatic metabolism of long-chain fatty acids was studied in male rats fed a defined choline-deficient (CD) diet with and without choline and after methotrexate (MTX) administration. Peroxisomal beta-oxidation was increased approximately 4-fold in the peroxisome-enriched fraction of CD-fed animals, whereas the catalase activity was increased 1.3-fold. The urate oxidase activity was marginally affected. The CD-fed rats also revealed elevated capacity for hydrolysis of palmitoyl-CoA in the cytosolic fraction (2.0-fold), whereas the microsomal palmitoyl-CoA hydrolase activity was decreased. Notably, the increased peroxisomal beta-oxidation, the catalase activity and palmitoyl-CoA hydrolase activities (the membrane-bounded and cytosolic) were almost fully prevented by adding choline to the CD-diet. Thus, the change in these enzyme activities appears to be a consequence of a choline-deficiency provoked by the CD diet. MTX administration of normal fed rats (ND diet) had no effects on the peroxisomal beta-oxidation, catalase activity and urate oxidase activity. MTX treatment of the ND-fed animals, however, increased the mitochondrial palmitoyl-CoA hydrolase activity and decreased the microsomal enzyme activity. As choline-deficiency and MTX increased the hepatic lipid level, the overall results suggest that fat accumulation is not an 'induction signal' for increased peroxisomal beta-oxidation. The CD diet alone increased the reduced glutathione content in liver, whereas MTX did not significantly change this level. Whether the changes of H2O2-generating peroxisomal oxidation of long-chain fatty acids may be an important step in a chain of events, which eventually results in tumour formation by choline-deficiency, should be considered.

Modifying effects of supplemental selenium and sulfur on cadmium toxicity in rats.

Interactions among dietary supplements of selenium (Se), cadmium (Cd), and sulfur (S), and their effects on Cd toxicity in the cardiovascular system were explored in rats. Weanling male Sprague-Dawley rats were fed a Torula yeast-based diet in a 23 factorial design, which provided two levels each of supplemental Se (0 and 0.5 ppm-mg/kg diet—as Na2SeO3), and S (0 and 2% as K2SO2). Supplemental Cd (0 and 25 ppm-mg/L as Cd (C2H3O2)2 · 2H2O) was added to drinking water. Supplemental Cd induced cardiac hypertrophy only in rats not supplemented with Se. Selenium supplementation enhanced growth more markedly in Cd-fed rats than in rats not receiving Cd. Renal Cd content was increased by Cd feeding, but there was no significant effect of Cd feeding on hemoglobin or blood pressure. Supplemental Cd did not influence Se-GSH-Px activity in heart or kidney cytosol. Interactions between Se and Cd were not altered by S supplementation. These results suggest 1) that hypertension is not associated with increased kidney accumulation of Cd,per se; 2) a supplement of 0.5 ppm Se as selenite is sufficient to protect against Cd-induced reductions in Se-GSH-Px activity in heart and kidney cytosol; and 3) increased S intake does not compromise Se's protective effect against certain manifestations of Cd toxicity.

Cardiac physiologic-metabolic changes after chronic low-level heavy metal feeding.

Female Long-Evans hooded rats received 0 or 5 parts per million Cd, Pb, or Cd + Pb in the drinking water from weaning to 20 mo. Growth, measured as body weight, and organ weights were comparable among heavy metal and control animals. Indirect blood pressure responses measured trimonthly are discussed. Analysis of His bundle electrograms (HBE) and electrocardiograms recorded in sodium pentobarbital-anesthetized animals at 20 mo demonstrated that Cd selectively depressed conduction system excitability proximal to the common His bundle (atrioventricular node region), whereas Pb and Pb + Cd impaired conductivity distal to the common His bundle (His-Purkinje system). Perchloric acid extracts of liquid N2-freeze-clamped isolated perfused hearts, derived from a subpopulation of control and Cd-fed rats in which HBE analyses were not performed, were analyzed by phosphorus-31 nuclear magnetic resonance (31P-NMR) spectroscopic techniques to quantitate [31P]phosphate metabolite profiles. Cardiac active tension and atrioventricular node excitability were depressed in the Cd group. High-energy phosphate and glycerol 3-phosphorylcholine concentrations were depressed. Results demonstrate potentially significant pathophysiological changes within cardiovascular tissues that develop in the absence of overt heavy metal toxicity manifestations.

Potentiation of the hepatotoxicity of carbon tetrachloride following preexposure to chlordecone (Kepone) in the male rat

The effect of carbon tetrachloride (CCl 4) challenge on several parameters of hepatic injury was determined in chlordecone (CD)-pretreated male rats. Following a 15-day feeding of 0 or 10 ppm CD, rats received a single intraperitoneal challenge of 0, 25, 50, 100, or 200 μl CCl 4/kg. Twenty-four hours after CCl 4, biliary excretion of phenolphthalein glucuronide (PG), bile secretion, and serum transaminase (SGPT, SGOT) activities were examined as functional indices of hepatic injury. Feeding of 10 ppm CD alone resulted in decreased biliary excretion of PG (59% of control) but had no effect on bile flow or on serum transaminases. Twenty-four hours after CCl 4 alone, the two high doses caused decreased biliary excretion of PG in the absence of any effect on bile flow and doubled serum transaminases at the highest dose (200 μl/kg). A single challenge of CCl 4 to CD-fed rats resulted in a dose-dependent impairment of PG excretion at a dose of 50 μl/kg and higher. Bile secretory function was severely impaired in the CD-fed animals receiving CCl 4 at the doses of 100 and 200 μl/kg. Decreased bile flow was not seen in any other groups. Greatly potentiated hepatotoxicity was reflected in the form of elevated SGPT and SGOT activities which increased in excess of 30- and 10-fold, respectively, in CD-fed rats challenged with CCl 4 at 100 and 200 μl/kg. Parenchymal liposis (cytoplasmic sudanophilic droplets) developed in all CD-fed rats receiving CCl 4, while necrosis occurred after CCl 4 at 50 μl/kg and increased in a dosedependent manner. CCl 4 controls exhibited parenchymal liposis and limited centrilobular necrosis only at the two highest doses of CCl 4. These data indicate a great potential for production of severe liver damage resulting from interactions of CCl 4 and CD exposure at levels which may be independently nontoxic.