Effect of copper deficiency and Sodium intake upon liver lipid and mineral composition in the rat

The effect of copper and sodium intake upon liver cholesterol concentrations, fatty acid profile, and mineral concentrations were studied in the Long-Evans rat. Forty-eight male weaning rats were divided into three groups of 16 each and fed a semipurified diet containing either 0, 3, or 8 mg of added copper/kg of diet. At 100 d of age, half of the animals in each group were given 1% NaCl as drinking water and the other half was given deionized-distilled water for 12 wk. Copper deficiency in rats produced elevations in liver palmitate and oleate concentrations, but decreases in linoleate concentrations. The ratio of oleate:stearate was higher in copper deficient rats. Liver copper levels were decreased, but liver iron concentrations were elevated in copper deficient rats. Sodium intake did not have an effect on any of the parameters studied. These results suggested that dietary copper deficiency alters both liver mineral and fatty acid composition.     

Dietary carbohydrate influences tissue fatty acid and lipid composition in the copper-deficient rat

Fructose and copper have been shown independently to influence long chain fatty acid metabolism. Since fructose feeding exacerbates copper deficiency, their possible interaction with respect to tissue long chain fatty acid and lipid composition was studied. Weanling male Sprague-Dawley rats were given diets containing 0.6 or 6 mg/kg copper. The carbohydrate source (627 g/kg) was either fructose or corn starch. After 3 wk, fatty acid profiles and total lipids in heart and liver were analyzed. Copper-deficient rats fed fructose had more severe signs of copper deficiency than those fed starch, according to heart/body wt ratio, hematocrit, and liver copper content. The fatty acid composition of heart and liver triacylglycerol was significantly different between groups, but the changes did not correlate with the severity of copper deficiency. In heart, phosphatidylinositol and phosphatidylserine, arachidonic acid and docosapentaenoic acid (n-6) were increased 193 and 217%, respectively, p<0.05) in rats given the copper-deficient diet containing fructose. Changes in the long chain fatty acids in heart phospholipids may be related to the higher mortality commonly observed in rats fed a copper-deficient diet containing fructose.     

Relative and combined effects of ethanol and protein deficiency on some hair trace elements

This study was performed in order to analyze the relative and combined effects of ethanol and protein deficiency on hair copper, zinc, manganese, and iron content in four groups of seven animals each which were pair-fed during 8 wk with (1) a nutritionally adequate diet, (2) a 36% (as energy) ethanol-containing isocaloric diet, (3) a 2% protein, isocaloric diet, and (4) a 36% ethanol, 2% protein isocaloric diet, respectively, following the Lieber-DeCarli model, and to analyze the relationship between hair copper, zinc, manganese, and iron content, and the liver and muscle content of these elements. Although there was a trend to higher levels of all the elements analyzed in the the hair of the low-protein fed animals, differences were statistically significant regarding copper and manganese, effects being solely attributable to the low protein diet, not to ethanol. Moreover, hair copper was significantly, inversely related with final weight and weight loss. There were significant relationship between liver zinc and muscle zinc (r=0.57, p=0.002), but not between liver or muscle zinc and hair zinc; no correlations were observed between muscle copper and hair copper, nor between liver manganese and hair manganese. An inverse, statistically significant correlation was observed between liver copper and hair copper (r=−0.39, p<0.05).     

Excess diet copper increases systolic blood pressure in rats

Twenty Wistar and 20 Spontaneously Hypertensive Rats were divided into two diet copper intake groups at 5 wk of age. One group was fed 18 mg Cu as cupric carbonate/kg feed and the other group 100 mg Cu/kg feed. The basal diet was formulated after the AIN-76 diet. Animals were fed their respective diets until 20 wk of age. At age 8, 14, and 20 wk rats were placed in metabolic cages and food and water consumption as well as urine output were quantitated. Systolic blood pressures were measured triweekly by the tail-cuff method. Rats fed 100 mg Cu/kg feed showed an increase in systolic pressure compared to rats fed 18 mg Cu/kg feed. The increase was more pronounced for the Wistar strain. Body weights, urine output, and feed and water consumption did not differ (p>0.05) by copper intake. Rats fed 100 mg Cu/kg feed did have greater hemoglobin levels. Heart: body and liver: body weight strain differences were apparent. This study demonstrated increased blood pressure in rats fed excess diet copper.     

Effect of protein deficiency on macroelement and trace element levels of weanling rats' small intestine and liver tissues

Protein energy malnutrition has become a major health issue in developing countries. In the present study, the effect of protein deficiency on the small intestine and liver tissue content of macroelements and trace elements was investigated in weanling rats. Forty-five male weanling Wistar albino rats were divided into three groups. The control group (C) was fed a standard diet containing 25% casein, whereas the two experimental groups E1 and E2 consumed 12% and 3% casein, respectively, over a period of 45 d. The tissue samples were analyzed for zinc, copper, iron, manganese, calcium, and magnesium by atomic absorption spectroscopy. The protein-deficient groups showed increased levels of iron in both tissues and decreased manganese in small intestine tissue from the E1 group. No other differences were found for the other elements. These results suggest that protein deficiency might cause iron accumulation in the liver and intestine and decreases of manganese in the small intestine.     

High arsenic intake raises kidney copper and lowers plasma copper concentrations in rats

The effect of high arsenic intake on copper metabolism was investigated. Male rats aged 6 wk had free access to purified diets containing either 0 or 100 mg As/kg diet and demineralized water for a period of 2 wk. Arsenic was added to the diet in the form of NaAsO₂. The high-arsenic diet decreased feed and water intake and body weight gain, but significantly increased liver weight. Kidney weight was not affected. Arsenic feeding drastically elevated kidney copper concentration, but significantly reduced copper concentration in plasma. Both true absorption and biliary excretion of copper were decreased significantly in rats fed the high-arsenic diet. True copper absorption was lowered essentially through the lower copper intake in the rats fed arsenic. It is speculated that arsenic feeding primarily leads to copper accumulation in the kidney, followed by a decrease in feed intake and thus in true, absolute copper absorption, a decrease in plasma copper concentration, and a decrease in biliary copper excretion.     

Dietary copper and dimethylhydrazine affect protein kinase C isozyme protein and mRNA expression and the formation of aberrant crypts in colon of rats.

Low dietary copper has been shown to decrease the expression of various protein kinase C (PKC) isozymes and increase the risk of colon cancer development in experimental animals. The purpose of this study was to investigate the relationship between dietary copper and carcinogen administration on PKC isozyme accumulation and aberrant crypt foci (ACF) formation in rats fed 0.9 and 7.7 microg Cu/g diet. After 24 and 31 d on the diets, the rats were injected with either dimethylhydrazine (DMH) (25 mg/kg i.p.) or saline and killed at two time points (2 wk and 8 wk after DMH). Rats fed low dietary copper had significantly lower (p<0.0001) hematocrits, hemoglobin, ceruloplasmin activity and plasma and liver copper concentrations than rats fed adequate dietary copper. Ingestion of low dietary copper significantly (p<0.005) increased the formation of DMH-induced ACF (116.8 vs 59.6). Low dietary copper significantly (p<0.05) decreased the concentration of PKC alpha, delta, and zeta in the colon at 2 wk but not at 8 wk. Thus, changes in PKC isoform protein concentration may be related to increased susceptibility of copper-deficient animals to colon cancer.     

Cadmium-induced alterations in ocular trace elements

The present report demonstrates, for the first time, that feeding rats 50 ppm cadmium for just 7 wk results in detectable levels of cadmium in the eye of rats. Furthermore, these ocular cadmium concentrations affect significant alterations in the levels of the essential trace elements selenium, calcium iron, and copper in the eye. Rats were fed a low-selenium (<0.02 ppm selenium), high-copper basal diet (50 ppm copper) supplemented with 0, 0.1, and 0.5 ppm selenium. The animals were either untreated or treated with 50 ppm cadmium admixed with their feed. Cadmium treatment resulted in significant reductions (up to 50%) in ocular selenium. Furthermore, rats fed the basal diet and given 100 ppm cadmium via their feed for 6 wk exhibited a 69% reduction in the activity of the selenoenzyme, glutathione peroxidase, in the eye. Cadmium treatment also resulted in reductions of up to 50% in ocular calcium, irrespective of dietary selenium supplementation. Iron levels were increased by 30% in rats fed the low-selenium diet and decreased by as much as 40% in rats fed the selenium-supplemented diets, compared to animals fed identical levels of selenium without cadmium. Ocular copper levels were significantly increased only in rats fed the low-selenium diet and treated with cadmium. Ocular zinc levels were not significantly affected by dietary cadmium or selenium.     

cis-acting determinants of basal and lipid-regulated apolipoprotein A-IV expression in mice

The levels of apolipoprotein A-IV (apoA-IV) mRNA are regulated by dietary lipid in the liver of both the mouse and rat. Thirteen different inbred mouse strains were fed a high lipid diet, and the effect on apoA-IV liver mRNA levels was examined. It was found that each strain responded in one of two ways. Mice of four strains had higher liver apoA-IV mRNA levels as compared with syngeneic mice fed a normal chow diet. Mice of the other nine strains had decreased liver apoA-IV mRNA levels as compared with syngeneic mice fed a normal chow diet. Using F1 hybrids between mice from BALB/c, C3H, and C57BL/6 and between 129 and C57BL/6, as well as recombinant inbred strains derived from a cross between BALB/c and C57BL/6, we have shown that both the normal level of liver apoA-IV mRNA in the chow-fed mice and the lipid-dependent regulation of apoA-IV mRNA levels are controlled by cis-acting genetic elements. The apoA-IV mRNA levels in mice fed a normal diet varied dramatically among strains, with the largest difference (90-fold) being between the 129/J inbred strain and the C57BL/6J strain. In addition, we have examined the expression of apoA-IV during mouse development. ApoA-IV mRNA is expressed early in mouse liver (16 days postcoitum), whereas others have shown previously that rat liver apoA-IV mRNA is undetectable until 14 days after birth. ApoA-IV mRNA levels in the intestine and apoA-I mRNA levels in the liver and intestine, by contrast, mirror the pattern seen in the rat.     

Mineral supplementation of white wheat flour is necessary to maintain adequate mineral status and bone characteristics in rats

This experiment was designed to compare the effect of ingestion of a wheat flours on mineral status and bone characteristics in rats. White flour was tested either without further mineral supplementation or with Mg, Fe, Zn and Cu supplementation. The flour diets were compared to a control purified diet. Four groups of 10 male Wistar rats each were fed one of the experimental diets for 6 wk and mineral status and tissue retention as well as bone characteristics were determined. As expected, mineral intake, except for calcium, was significantly lesser in rats fed the white flour diet than in the other groups. The rats fed the white flour diet had the lowest food intake, weight gain, fecal excretion and intestinal fermentation. The most important result was that Mg and Fe status were drastically lower in rats fed the white flour diet than in those fed whole flour or control diets. The status of these both elements were significantly improved by the mineral supplementation of white flour. There were no major significant differences between mineral-supplemented white flour and whole flour groups in mineral status. Furthermore, bone mineral densities (total, metaphyseal and diphyseal) were significantly lower in rats fed white flour diet compared to the other diet groups, while no significant difference was observed between the mineral-supplemented white flour, whole flour or control diet groups. In conclusion, the present work shows clearly the importance of mineral-supplementation of white wheat flour to sustain an adequate intake of minerals. Our results indicate also that the whole wheat flour did not negatively alter mineral bioavailability, in comparison to mineral supplemented white flour. Clinical studies are still needed to confirm these rat results in human.     

Copper modulates sex-specific fructose hepatoxicity in nonalcoholic fatty liver disease (NALFD) Wistar rat models

This study aimed to characterize the impact of dietary copper on the biochemical and hepatic metabolite changes associated with fructose toxicity in a Wistar rat model of fructose-induced liver disease. Twenty-four male and 24 female, 6-week-old, Wister rats were separated into four experimental dietary treatment groups (6 males and 6 females per group), as follows: (1) a control diet: containing no fructose with adequate copper (i.e., CuA/0% Fruct); (2) a diet regimen identical to the control and supplemented with 30% w/v fructose in the animals' drinking water (CuA/30% Fruct); (3) a diet identical to the control diet but deficient in copper content (CuD/0% Fruct) and (4) a diet identical to the control diet but deficient in copper content and supplemented with 30% w/v fructose in the drinking water (CuD/30% Fruct). The animals were fed the four diet regimens for 5 weeks, followed by euthanization and assessment of histology, elemental profiles and identification and quantitation of liver metabolites. Results from ¹H nuclear magnetic resonance metabolomics revealed mechanistic insights into copper modulation of fructose hepatotoxicity through identification of distinct metabolic phenotypes that were highly correlated with diet and sex. This study also identified previously unknown sex-specific responses to both fructose supplementation and restricted copper intake, while the presence of adequate dietary copper promoted most pronounced fructose-induced metabolite changes.     

Analysis of whole blood manganese by flameless atomic absorption spectrophotometry and its use as an indicator of manganese status in animals

To investigate the use of whole blood manganese (Mn) as an indicator of total body Mn, we measured Mn in whole blood and liver of rats fed purified diets containing adequate (45 micrograms Mn/g diet) or deficient (1 microgram Mn/g diet) Mn. The mean hepatic Mn concentration was significantly lower (P less than 0.001) in the Mn-deficient group compared to the control group, 0.36, microgram Mn/g and 1.73 micrograms Mn/g, respectively. Furthermore, whole blood Mn was significantly reduced (P less than 0.001) in the deficient group when compared to the control group, 4.0 ng Mn/ml and 8.6 ng Mn/ml, respectively. Hepatic Mn linearly regressed against whole blood Mn yielded a statistically significant (P less than 0.001) correlation coefficient of 0.775. These data suggest that whole blood Mn is a valid indicator of body Mn status and thus may be useful, in addition to the measurements of serum copper and zinc, for the diagnosis and prognosis of diseases in which the metabolism of trace elements is affected. In addition, this paper describes and delineates operational parameters for the measurement of whole blood Mn using the IL 551 atomic absorption spectrophotometer and the IL 555 B flameless atomizer.     

Interaction between nickel and iron in the rat

The interaction between nickel and iron was confirmed in rat metabolism. In a fully-crossed, two-way, three by four, factorially designed experiment, female weanling rats were fed a basal diet supplemented with iron at 0, 25, 50, and 100 μg/g and with nickel at 0, 5, and 50 μg/g. The basal diet contained about 10 ng of nickel and 2.3 μg of iron/g. After nine weeks, dietary iron affected growth, hematocrit, hemoglobin, plasma cholesterol, and in liver affected total lipids, phospholipids, and the contents of copper, iron, manganese, and zinc. By manipulating the iron content of the diet, effects of dietary nickel were shown in rats that were not from dams fed a nickel-deprived diet. Nickel affected growth, hematocrit, hemoglobin, plasma alkaline phosphatase activity, plasma total lipids, and in liver affected total lipids, and the contents of copper, manganese, and nickel. The interaction between nickel and iron affected hematocrit, hemoglobin, plasma alkaline phosphatase activity, and plasma phospholipids, and in liver affected size, content of copper, and perhaps of manganese and nickel. In severely iron-deficient rats, the high level of dietary nickel partially alleviated the drastic depression of hematocrit and hemoglobin, and the elevation of copper in liver. Simultaneously, high dietary nickel did not increase the iron level in liver and was detrimental to growth and appearance of severely iron-deficient rats. In nickel-deprived rats fed the borderline iron-deficient diet (25 μg/g) hematocrit and hemoglobin also were depressed. However, 5 μg Ni/g of diet were just as effective as 50 μg Ni/g of diet in preventing those signs of nickel deprivation. The findings in the present study suggested that nickel and iron interact with each other at more than one locus.     

Hepatic iron accumulation is not directly associated with induction of DNA strand breaks in the liver cells of Long-Evans Cinnamon (LEC) rats.

Effects of accumulation of copper and iron on induction of DNA strand breaks were investigated in Long-Evans Cinnamon (LEC) rats that spontaneously develop fulminant hepatitis. Copper and iron accumulated in the liver of LEC rats in an age-dependent manner from 4 to 15 weeks. Low-iron diet prevented the accumulation of iron in the liver, but did not prevent accumulation of copper. The amounts of DNA strand breaks that were estimated by comet assay in the liver cells of rats fed standard diet increased with age from 4 to 15 weeks. No significant differences were observed in the proportions of LEC rat liver cells without tail and the average lengths of tail momentum in the comet images between LEC rats that had been fed standard MF diet and low-iron diet. These results support the idea that accumulation of iron is not directly associated with the induction of DNA damage in the liver cells of LEC rats.     

Metallothionein mRNA levels are influenced by dietary cyclodextrins in rats

The relationship between metallothionein mRNA levels and the amounts of copper and zinc in liver, kidney and small intestine by feeding dietary cyclodextrin was examined in growing Wistar rats. alpha-, beta- or gamma-cyclodextrin was fed at 50 g/kg of diet for a 7-days period (ad libitum). After feeding, the liver zinc of rats fed beta-cyclodextrin was greater than those of rats fed the other three diets. Copper accumulated in kidney of rats fed alpha- or beta-cyclodextrin. Copper content in the small intestine did not show any alterations among rats fed all kinds of diets. The cyclodextrin-supplemented diets were ineffective in zinc content in every organ. There was the greatest level of copper in serum of rats fed beta-cyclodextrin, whereas the highest level of serum zinc was observed in rats fed gamma-cyclodextrin diet. Northern blot analysis demonstrated that dietary beta- and gamma-cyclodextrins, but not alpha-cyclodextrin markedly increased the metallothionein mRNA in the liver, whereas small intestinal metallothionein mRNA levels were markedly decreased. Kidney metallothionien mRNA levels were raised appreciably by all dietary cyclodextrin intakes. Metallothionein gene expressions in liver, kidney and small intestine were not proportional to liver and serum copper or zinc levels in those tissues. These results suggest that regulation of the metallothionein mRNA levels may at least partly involved with the accumulation of metals as copper in liver and kidney of rats fed cyclodextrins.     

Levels and distribution of zinc,copper, magnesium,and calcium in rats fed different levels of dietary zinc

Effects of altered dietary zinc on levels of zinc, copper, magnesium, and calcium in organ and peripheral tissues were studied. When rats fed a zinc-deficient diet (1.3 μg Zn/g) for 28 d were compared with rats fed a control diet (37.5 μg Zn/g), levels of zinc were slightly lower in plasma, hair, and skin and 50% lower in femur and pancreas, whereas the levels of copper were higher in all tissue except plasma. Magnesium levels were higher than controls in the heart and lower in the spleen, whereas the calcium levels were lower in plasma, lung, spleen, kidney, and skin and strikingly higher in brain, hair, and femur. When rats fed a zinc-supplemented diet (1.0 mg Zn/g) were compared to the same conrols, levels of zinc in these were higher in all organs and peripheral tissues studied, except heart, lung, and liver; copper levels were higher in liver, kidney, and spleen; magnesium levels were significantly higher in the spleen, but were little affected in other tissues, although calcium levels were higher in pancreas, spleen, kidney, and skin and lower in plasma and hair. These data indicate that overall copper organ and peripheral tissue levels are affected inversely, and zinc and calcium levels directly, by zinc nutriture.     

Effect of zinc supplementation on metallothionein,copper, and zinc concentration in various tissues of copper-loaded rats

The present study was designed to investigate the effects of Zn administration on metallothionein concentrations in the liver, kidney, and intestine of copper-loaded rats. Male CD rats were fed a diet containing 12 mg Cu and 67 mg Zn/kg body wt. They were divided into either acute or chronic experimental protocols. Rats undergoing acute experiments received daily ip injections of either Cu (3 mg/kg body wt) or Zn (10 mg/kg body wt) for 3 d. Chronic experiments were carried out on rats receiving Cu ip injections on d 1, 2, 3, 10, 17, and 24, Cu injections plus a Zn-supplemented diet containing 5 g Zn/kg solid diet, or a Zn-supplemented diet alone. Rats injected Zn or Cu had increased MT concentrations in liver and kidney. Zn produced the most important effects and the liver was the most responsive organ. Rats fed a Zn-supplemented diet had significantly higher MT concentrations in liver and intestine with respect to controls. Increased MT synthesis in the liver may contribute to copper detoxification; the hypothesis of copper entrapment in enterocytes cannot be confirmed.     

Effect of 2,3,7,8-Tetrachlorodibenzo-p-dioxin on the Hepatic Distribution of Iron,Copper, Zinc,and Magnesium in Rats

The distribution of iron, copper, zinc, and magnesium in hepatic subcellular fractions of male and female rats treated with 2,3,7,8-tetrachloro-dibenzo-p-dioxin (TCDD) was determined. Animals received 40 μg TCDD per kilogram per day for three days by mouth (PO) or the vehicle and were killed seven or nine days posttreatment. Iron, copper, zinc, and magnesium were determined by atomic absorption spectroscopy. The iron content of liver from female animals was twofold higher than male animals. The administration of TCDD increased the iron content of mitochondria in female and male rats and decreased iron content of microsomes of both sexes. Significant increases occurred in the copper content of whole liver, mitochondria, and cytosol of male rats and in whole liver and cytosol of female rats. Decreases in the copper content of the microsomes of male rats were observed following TCDD treatment; however, TCDD produced no changes in the zinc content of hepatic subcellular fractions of either sex. The magnesium content of female TCDD-treated rats increased in whole liver, mitochondria, and cytosol, while the magnesium content of microsomes was not altered. With respect to the subcellular distribution of iron, copper, zinc, and magnesium, TCDD produces differential effects. The altered distribution of some cations may contribute to the broad range of effects of TCDD.     

Effect of low dietary rubidium on plasma biochemical parameters and mineral levels in rats

The effects of low dietary rubidium on plasma biochemical parameters and mineral levels in tissues in rats were studied. Eighteen male Wistar rats, weighing about 40 g, were divided into two groups and fed the diets with or without supplemental rubidium (0.54 vs 8.12 mg/kg diet) for 11 wk. Compared to the rats fed the diet with supplemental rubidium, the animals fed the diet without rubidium supplementation had higher urea nitrogen in plasma; lower rubidium concentration in tissues; lower sodium in muscle; higher potassium in plasma, kidney and tibia, and lower potassium in testis; lower phosphorus in heart and spleen; lower calcium in spleen; higher magnesium in muscle and tibia; higher iron in muscle; lower zinc in plasma and testis; and lower copper in heart, liver, and spleen, and higher copper in kidney. These results suggest that rubidium concentration in tissues reflects rubidium intake, and that rubidium depletion affects mineral (sodium, potassium, phosphorus, calcium, magnesium, iron, zinc, and copper) status.     

Liver copper content of rats hypo- or hyperresponsive to dietary cholesterol

The question addressed is whether cholesterol intake reduces the hepatic copper content in rats. For this purpose we have compared the hepatic copper content of two selected rat inbred strains after feeding the animals a control or a high fat, high cholesterol diet. One strain was dietary cholesterol resistant (SHR/OlaIpcv), whereas the other strain was susceptible to dietary cholesterol (BN-Lx/Cub). Dietary cholesterol-susceptible rats have a lower baseline hepatic copper content when compared with their resistant counterparts. The consumption of a hypercholesterolemic diet decreased the liver copper concentration (expressed in microg/g dry weight) to about the same extent in both strains. However, dietary cholesterol did not reduce the absolute (expressed as microg/whole liver) and relative (expressed as microg/whole liver/100 g body weight) copper store of rats. The decrease of liver copper concentration after the high fat, high cholesterol diet is probably not caused by a decrease in whole hepatic copper content, but rather due to dietary-induced hepatomegaly.