Physiological copper exposure in Jurkat cells induces changes in the expression of genes encoding cholesterol biosynthesis proteins

Copper is an essential micronutrient that functions as an enzymatic cofactor in a wide range of cellular processes. Although adequate Cu levels are essential for normal metabolism, excess Cu can be toxic to cells. Cellular responses to copper deficiency and overload involve changes in the expression of genes directly and indirectly involved in copper metabolism. However little is known on the effect of physiological copper concentration on gene expression changes. In the current study we aimed to establish whether the expression of genes encoding enzymes related to cholesterol (hmgcs1, hmgcr, fdft) and fatty acid biosynthesis and LDL receptor can be induced by an iso-physiological copper concentration. The iso-physiological copper concentration was determined as the bioavailable plasmatic copper in a healthy adult population. In doing so, two blood cell lines (Jurkat and THP-1) were exposed for 6 or 24 h to iso- or supraphysiological copper concentrations. Our results indicated that in cells exposed to an iso-physiological copper concentration the early induction of genes involved in lipid metabolism was not mediated by copper itself but by the modification of the cellular redox status. Thus our results contributed to understand the involvement of copper in the regulation of cholesterol metabolism under physiological conditions.     

Effect of prolonged ethanol ingestion on hepatic lipogenesis and related enzyme activities

1. Hepatic lipogenesis in vivo and the activities of enzymes associated with fatty acid synthesis in the liver were studied in rats fed for 21 days on liquid diets containing ethanol. 2. The ethanol-fed rats developed a moderate hepatic triacylglycerol accumulation during this period. When carbohydrate was replaced by ethanol in the diet, the rate of fatty acid synthesis was slower in the ethanol-fed rats on low-, medium- and high-fat diets than in the appropriate controls. However, when the fat/carbohydrate ratio was kept the same in the ethanol-fed and control rats, ethanol had no influence on the rate of fatty acid synthesis. 3. Glucose 6-phosphate dehydrogenase activity was lower in the ethanol-fed group. ;Malic' enzyme activity did not change during the ethanol treatment when the fat/carbohydrate ratio was kept unchanged. 4. The ATP citrate lyase activity was lower in the ethanol-fed rats on all diets, whereas acetyl-CoA synthetase activity was independent of the composition of the control diet, but was lower in the ethanol-fed rats, in which the concentration of the active form of pyruvate dehydrogenase was also lower. 5. It is concluded that hepatic fatty acid synthesis does not play any major role in ethanol-induced triacylglycerol accumulation. Careful design of the diets is necessary to reveal the specific effects of ethanol on the enzymes associated with lipogenesis.     

The Development of Enzymes of Energy Metabolism in the Brain of a Precocial (Guinea Pig) and Non-Precocial (Rat) Species

Abstract: Key enzymes of ketone body metabolism (3-hydroxybutyrate de-hydrogenase, 3-oxo-acid: CoA transferase, acetoacetyl-CoA thiolase) and glucose metabolism (hexokinase, lactate dehydrogenase, pyruvate dehydrogenase, citrate synthase) have been measured in the brains of foetal, neonatal and adult guinea pigs and compared to those in the brains of neonatal and adult rats. The activities of the guinea pig brain ketone-body-metabolising enzymes remain relatively low in activity throughout the foetal and neonatal periods, with only slight increases occurring at birth. This contrasts with the rat brain, where three- to fourfold increases in activity occur during the suckling period (0–21 days post partum), followed by a corresponding decrease in the adult. The activities of the hexokinase (mitochondrial and cytosolic), pyruvate dehydrogenase, lactate dehydrogenase and citrate synthase of guinea pig brain show marked increases in the last 10–15 days before birth, so that at birth the guinea pig possesses activities of these enzymes similar to the adult state. This contrasts with the rat brain where these enzymes develop during the late suckling period (10–15 days after birth). The development of the enzymes of aerobic glycolytic metabolism correlate with the onset of neurological competence in the two species, the guinea pig being a "precocial" species born neurologically competent and the rat being a "non-precocial" species born neurologically immature. The results are discussed with respect to the enzymatic activities required for the energy metabolism of a fully developed, neurologically competent mammalian brain and its relative sensitivity to hypoxia.     

Uteroplacental insufficiency alters hepatic fatty acid-metabolizing enzymes in juvenile and adult rats

Multiple adult morbidities are associated with intrauterine growth retardation (IUGR) including dyslipidemia. We hypothesized that uteroplacental insufficiency and subsequent IUGR in the rat would lead to altered hepatic fatty acid metabolism. To test this hypothesis, we quantified hepatic mRNA levels of acetyl-CoA carboxylase (ACC), carnitine palmitoyltransferase (CPTI), the beta-oxidation-trifunctional protein (HADH), fasting serum triglycerides, and hepatic malonyl-CoA levels at different ages in control and IUGR rats. Fetal gene expression of all three enzymes was decreased. Juvenile gene expression of CPTI and HADH continued to be decreased, whereas gene expression of ACC was increased. Serum triglycerides were unchanged. A sex-specific response was noted in the adult rats. In males, serum triglycerides, hepatic malonyl-CoA levels, and ACC mRNA levels were significantly increased, and CPTI and HADH mRNA levels were significantly decreased. In contrast, the female rats demonstrated no significant changes in these variables. These results suggest that uteroplacental insufficiency leads to altered hepatic fatty acid metabolism that may contribute to the adult dyslipidemia associated with low birth weight.     

Embryotoxicity of silver ions is diminished by ceruloplasmin—further evidence for its role in the transport of copper

The effect of alimentary administration of silver salts upon embryogenesis in rats was studied. Feeding of female rats throughout the term on a regular diet supplemented with AgCl did not cause alterations of their physiological functions, despite the fact that enzymatically active copper-containing ceruloplasmin (CP) was eliminated from the blood plasma. However, developmental abnormalities of embryos, their prenatal death or the 100% mortality of the newborns in the first 24 h of life was seen. Copper content in placenta and fetal tissues was strongly diminished. Cu, Zn-superoxide dismutase (SOD) activity decreased in cytoplasm of embryonic cells along with a drop, though less pronounced, in the tissues of the pregnant females. Embryotoxicity of AgCl was seriously diminished by repetitive injections of native CP to the pregnant rats. Such treatment resulted in an increase of SOD activity in placenta and embryonic tissues. The mortality of the newborns also became less. It is suggested that the embryotoxic effect of AgCl is caused by its ability to interfere with copper metabolism, in particular by altering the copper-transporting function of CP.     

Myeloid metaplasia in the spleen after acute haemolysis.

Copper sulphate, phenyl hydrazine and antiserum were administered to adult albino rats to produce equivalent haemolysis. Red pulp activity was studied for myeloid metaplasia after 24, 48 and 72 h, 5 and 7 days. Myeloid metaplasia was not related to the marginal zone proliferation. The activity rose rapidly with all the three agents and followed a similar curve despite the different patterns of haemolysis and erythrophagocytosis. Splenic erythrophagocytosis acts as a powerful feedback for splenic myeloid metaplasia. With copper, the release of reticulocytes and iron binding in haemoglobin is rapid as compared to the other agents, in spite of the initial lag phase, due to its effect on cellular metabolism.     

Silver (Ag+) reduces denitrification and induces enrichment of novel nirK genotypes in soil

The use of silver ions in industry to prevent microbial growth is increasing and silver is a new and an overlooked heavy-metal contaminant in sewage sludge-amended soil. The denitrifying community was the model used to assess the dose-dependent effects of silver ions on microorganisms overtime in soil microcosms. Silver caused a sigmoid dose-dependent reduction in denitrification activity, and no recovery was observed during 90 days. Dentrifiers with nirK, which encodes the copper nitrite reductase, were targeted to estimate abundance and community composition for some of the concentrations. The nirK copy number decreased by the highest addition (100 mg Ag kg(-1) soil), but the nirK diversity increased. Treatment-specific sequences not clustering with any deposited nirK sequences were found, indicating that silver induces enrichment of novel nirK denitrifiers.     

Involvement of reactive oxygen species in the bactericidal activity of activated carbon fibre supporting silver; Bactericidal activity of ACF(Ag) mediated by ROS

An activated carbon fibre supporting silver (ACF(Ag)) was tested for its antibacterial capacity against Escherichia coli (E. coli). Water that has passed through ACF(Ag) demonstrated strong bactericidal ability. This activity decreased over the time suggesting that generated bactericidal species were short lifespan. Since formation of reactive oxygen species (ROS) might be catalysed by silver impregnated and/or ACF itself, implication of ROS and silver was evaluated by the use of ROS scavengers and a silver ions neutralizing agent. The role of ROS in the E. coli mortality was confirmed by the use of a molecular approach which revealed a strong expression of oxidative stress genes.     

Dopamine and Norepinephrine in Discrete Areas of the Copper-Deficient Rat Brain1

Abstract: Copper deficiency was induced in post-weaning rats by feeding the dams a low copper diet during gestation and lactation. In confirmation of an earlier study, both dopamine and norepinephrine concentrations in the total brain were approximately 30% lower in deficient than in control rats. Doparnine in the corpus striaturm was depressed nearly 60%, but the concentration of norepinephrine in the hypothalamus was unchanged. Tyrosine concentrations in the striatum, hypothalamus, and total brain were not affected by copper deficiency, suggesting a catalytic defect rather than lack of substrate. Copper repletion restored norepinephrine level in total brain but did not affect the low level of dopamine. The results suggest that copper deficiency depresses a catalytic function of the adrenergic pathways and, further, adversely affects a structural component of the dopaminergic system during development.     

Tissue-specific alterations in lipoprotein lipase activity in copper-deficient rats

Copper deficiency results in alterations in lipid metabolism that include elevations in serum cholesterol and triglycerides and a decrease in whole-body respiratory quotient. Copper-deficient animals are also leaner even though electron micrographs of the myocardium present increased lipid droplet accumulation. To address whether a compromised copper status impacts triglyceride deposition in a tissue-specific manner, the activity of lipoprotein lipase was measured in adipose tissue and cardiac and skeletal muscle. Weanling rats fed a copper-restricted diet (<1 ppm) for 6 wk demonstrated a greater than twofold increase in cardiac lipoprotein lipase activity concomitant with a significant reduction in adipose tissue lipoprotein lipase activity. Skeletal muscle lipoprotein lipase activity was not altered by the copper-deficient state. The results of this study suggest that copper deficiency may induce a tissue-specific alteration in lipoprotein lipase activity in rats, which may contribute to the notable deposition of lipid substance in myocardium and the concomitant general body leanness.     

Absorption of 241Am in the gastrointestinal tract

In experiments with rats a study was made of a number of factors influencing the resorption of 241Am from the gastrointestinal tract (GIT). The resorption of 241Am from GIT was found to be 120-245 times more intensive in neonatal rats, during the first 21 days after birth (a milk diet), than in adult animals. A milk diet for adult rats produced a 5-fold increase in the resorption of 241Am from GIT. The additional administration of digestive enzymes, as a homogenate from pancreas and small intestine, produced a 7--9-fold increase in the rate of 241Am resorption from GIT.     

Fructose metabolizing enzymes in the rat liver and metabolic parameters: Interactions between dietary copper,type of carbohydrates,and gender

This study was conducted to determine the effects of nutrient interactions between dietary carbohydrates and copper levels on fructose-metabolizing hepatic enzymes in male and female rats. Male and female rats were fed diets for 5 weeks that were either adequate or deficient in copper that contained either starch or fructose. Rats of both sexes fed fructose as compared with those fed starch showed higher activity of hepatic fructose metabolizing enzymes. There were also significant differences in fructose metabolism of liver between the male and female rats. Female rats had lower hepatic ketohexokinase and triose kinase but higher triosephosphate isomerase activities compared with male rats. Male rats fed copper-deficient diets had lower aldolase B activity compared with those fed copper-adequate diets. Female rats fed copper-deficient diets had higher triosephosphate isomerase activity compared with rats fed copper-adequate diets. Our data suggest that gender differences in hepatic fructose metabolism may not be the primary reason for the severity of copper deficiency syndrome in male rats fed copper-deficient diet with fructose.     

Copper metabolism in analbuminaemic rats fed a high-copper diet

Copper metabolism in male Nagase analbuminaemic (NA) rats was compared with that in male Sprague Dawley (SD) rats fed purified diets containing either 5 or 100 mg Cu/kg diet. Dietary copper loading increased hepatic and kidney copper concentrations in both strains to the same extent, but baseline values were higher in the NA rats. There was no strain difference in true and apparent copper absorption nor in faecal endogenous and urinary copper excretion. NA rats had higher levels of radioactivity in kidneys at 2 hr after intraperitoneal administration of ⁶⁴Cu. As based on the distribution of added ⁶⁴Cu, about 70% of plasma copper appeared to be in the non-protein compartment in the NA rats, whereas in SD rats, it was only about 1%. It is concluded that the NA rats are able to maintain a relatively normal metabolism of copper, even after dietary copper challenge. In the NA rats, zinc concentrations in kidneys, liver and urinary zinc excretion were elevated when compared with SD rats. The high-copper diet did not affect tissue zinc concentrations and apparent zinc absorption in both strains of rats.     

Dietary beta- and gamma-cyclodextrins stimulation of hepatic metallothionein gene expression in rats

This study investigated whether hepatic metallothionein gene expression is affected by dietary cyclodextrins. Young male Wistar rats were fed a basal diet or cyclodextrin-supplemented (50 g of cyclodextrin per kg diet) diets for 7 d. Copper content in the liver did not show any significant changes among rats fed the basal, beta- and gamma-cyclodextrin diets. There were no differences in liver or serum zinc among groups. Copper content in serum was markedly decreased in rats fed the gamma-cyclodextrin-supplemented diet. Liver metallothionein mRNA levels were significantly elevated in both beta- and gamma-cyclodextrins-fed rats, but not in alpha-cyclodextrin-fed rats. Thus, the increase in hepatic metallothionein mRNA levels might be due to this mechanism except for the contents of copper and zinc in the liver.