Effect of experimental azotemia on intestinal transport of butyric acid

Earlier studies have revealed an impairment of jejunal absorption of long chain fatty acids in experimental uremia. We investigated the intestinal absorption of butyric acid which is a short chain fatty acid in experimental renal failure (RF). Sprague-Dawley rats were randomized into the RF group which had subtotal nephrectomy, a sham-operated control group, and a pair-fed group. In vivo recirculating perfusion (n = 5) and in vitro everted sac incubation (n = 8) were employed. The in vitro experiments were repeated substituting the serosal buffer by either predialysis or postdialysis sera from uremic individuals, or normal serum (n = 10). The rate of in vivo butyric acid absorption was significantly lower while the in vitro absorption was significantly higher in the RF group than those observed in the sham-operated and pair-fed groups which showed comparable values. The normality of butyric acid absorption in the pair-fed animals despite comparable weight loss with the RF group tends to exclude anorexia and weight loss as a cause of altered butyric acid transport in RF animals. The disparity between the in vivo and in vitro data is suggestive of an inhibitory influence of uremic environment which is present in vivo and absent in vitro. This viewpoint was corroborated by the observed fall in butyric acid absorption by sacs containing predialysis uremic serum as compared with those containing normal or postdialysis sera. The latter further suggests that the inhibitory factor(s) is dialyzable.     

Intestinal absorption of L-ascorbic acid in rats with renal failure

We studied L-ascorbic acid absorption in rats subjected to subtotal nephrectomy (renal failure (RF) group) and compared the results with those obtained in sham-operated normal animals and those pair-fed with their azotemic counterparts (PF group). In vivo recirculating perfusion and in vitro everted sac techniques were employed. The in vitro experiments were repeated substituting buffer within the serosal compartment with pooled sera from uremic and normal individuals. L-Ascorbic acid absorption in vivo in the RF group was significantly lower than those found in normal control and PF groups. In contrast, the in vitro mucosal to serosal transport was increased in the RF and PF groups when compared with the normal control group, suggesting increased permeability to L-ascorbic acid in the former groups. The disparity between in vivo and in vitro results in the RF animals is indicative of some inhibitory influence present in the intact uremic animals. However, experiments comparing the effect of uremic with normal sera on in vitro transport failed to reveal any suppressive effect of uremic chemical environment. In addition, serum ascorbic acid was reduced in PF and RF groups when compared with the normal control animals, thereby excluding elevated blood level as a cause of impaired absorption in intact animals with RF. In conclusion, in vivo jejunal absorption of L-ascorbic acid is impaired in rats with RF despite evidence of increased in vitro permeability. The latter appears to be mediated by reduced nutrient intake and weight loss. The inhibitory influence present in vivo could not be reproduced by incubation with uremic sera in vitro.     

Intestinal transport of pyridoxine in experimental renal failure

Renal failure (RF) has been shown to alter intestinal transport of a number of nutrients. We studied jejunal absorption of pyridoxine (B6) in rats rendered azotemic by subtotal nephrectomy (RF group) and compared the results with those obtained in normal rats subjected to sham operation (controls) and animals pair-fed (PF) with their RF counterparts. In vivo recirculating perfusion and in vitro everted sac techniques were employed. The in vitro experiments were repeated using sera from uremic and normal individuals to assess the possible effect of uremic chemical environment. The results showed significant reduction in B6 absorption in vivo in the RF group as compared to the control and PF groups. Paradoxically, the rate of in vitro B6 absorption determined for a wide range of concentrations was increased in the RF and PF groups as compared to the control group. The observed increase in B6 absorption in vitro suggests enhanced permeability in the RF and PF groups due probably to reduced nutrient intake which was common to both groups. The disparity between the in vivo and in vitro results is indicative of some inhibitory factor(s) present in the RF animals. Sacs containing uremic serum showed significantly suppressed B6 absorption in vitro as compared to those containing normal serum. These observations suggest that the uremic chemical environment may be, in part, responsible for the observed impairment of B6 transport in RF animals despite in vitro evidence of hyperpermeability.     

Vitamin D absorption, plasma concentration and urinary excretion of 25-hydroxyvitamin D in nephrotic syndrome

Nephrotic syndrome (NS) is commonly associated with vitamin D deficiency. Urinary losses of the protein-bound intermediary metabolite of this vitamin is thought to contribute to the deficiency state. The role of possible changes, if any, of vitamin D absorption has not been investigated previously in NS. We determined intestinal absorption of vitamin D3 as well as plasma concentration and urinary excretion of 25-hydroxyvitamin D3 in rats with puromycin aminonucleoside-induced NS. In vivo recirculating perfusion technique was employed at 100 and 600 nM perfusate concentrations. The results were compared with those obtained in animals receiving placebo injections provided with either free access to food (normal controls) or those pair-fed with their NS counterparts (pair-fed group). The NS group showed heavy proteinuria and hypoalbuminemia. In addition, the NS group exhibited marked urinary losses and significantly reduced plasma concentration of 25-hydroxyvitamin D. The rate of vitamin D3 absorption (given as nmol/100 cm/min) at 100 nM perfusate concentration in the NS group (0.161 +/- 0.029) was not significantly different from those obtained in the pair-fed group (0.202 +/- 0.058) and the normal control group (0.143 +/- 0.053). Likewise, no significant difference was found in the rats of vitamin D absorption at 600 nM concentration among the NS (1.073 +/- 0.383), pair-fed (0.955 +/- 0.229), and normal control (0.756 +/- 0.314) groups. Accordingly, intestinal absorption of vitamin D appears to be unaffected by the presence of experimental NS and as such the associated vitamin D deficiency can be managed by enteral supplementation.     

Effect of excessive vitamin A intake on muscle protein turnover in the rat.

3-Methylhistidine excretion in vivo and in vitro was monitored in hypervitaminotic and pair-fed control rats. Feeding with excess of retinyl palmitate (40 000 i.u./day per 100 g body wt.) significantly increased urinary 3-methylhistidine and creatinine output during a 4-day treatment interval. 3-Methylhistidine release from perfused rat hindquarters was also elevated after 5 days of vitamin treatment. To determine whether the adrenals were involved in mediating the above response, a study was conducted on adrenalectomized and sham-operated rats. Excessive vitamin A intake stimulated 3-methylhistidine excretion in vivo and in vitro in both adrenalectomized and sham-operated animals, thus suggesting that the vitamin A-induced acceleration in myofibrillar protein breakdown was not mediated by the adrenals. In both groups of rats, vitamin A treatment had no effect on the rate of protein synthesis, on the basis of incorporation in vitro of [3H]phenylalanine into muscle protein. Additional studies revealed that the addition of excess retinol to the perfusion medium (10 i.u./ml) had no significant effect on the rates of 3-methylhistidine release or [3H]phenylalanine incorporation in vitro. Finally, high doses of cortisol (7 mg/day per 100g body wt.) administered to intact rats for 5 days significantly increased rates of 3-methylhistidine excretion, both in vivo and in vitro.     

Dietary zinc deficiency decreases plasma concentrations of vitamin E

Experiments were conducted to examine the effects of dietary zinc (Zn) upon plasma vitamin E (E) concentrations to test the hypothesis that there may be a significant dietary interaction between these two nutrients. Weanling female Sprague-Dawley rats were fed diets that were (i) Zn-deficient (less than 0.9 micrograms Zn/g diet) ad libitum; (ii) Zn-adequate (50.9 micrograms Zn/g diet), pair-fed to the Zn-deficient group; and (iii) Zn-adequate (50.9 micrograms Zn/g diet) ad libitum. Plasma E in Zn-deficient animals (4.02 +/- 1.20 micrograms/ml) was significantly reduced (P less than or equal to 0.05) compared with results in both Zn-adequate pair-fed (9.21 +/- 0.70 micrograms/ml) and Zn-adequate ad libitum-fed (9.47 +/- 0.90 micrograms/ml) animals. Zn deficiency in this model system also resulted in significant (P less than or equal to 0.05) reductions in femur and plasma Zn concentrations as well as in plasma retinol, plasma triglyceride, and plasma cholesterol concentrations. Plasma albumin and total plasma protein concentrations were normal in Zn-deficient animals. With dietary Zn deficiency, the decrease in plasma E appeared to be out of proportion to associated decreases in plasma triglyceride and plasma cholesterol concentrations. Since E is associated with plasma lipoproteins, these data suggest that lipid and/or E malabsorption may be a consequence of Zn deficiency. In response to increased dietary intake of E, increments of plasma E were lower in Zn-depleted than in Zn-adequate, pair-fed animals. These findings suggest that dietary Zn deficiency possibly may increase the nutritional requirement for E necessary to maintain adequate plasma concentrations.     

Prolactin is an important regulator of intestinal calcium transport

Prolactin has been shown to stimulate intestinal calcium absorption, increase bone turnover, and reduce renal calcium excretion. The small intestine, which is the sole organ supplying new calcium to the body, intensely expresses mRNAs and proteins of prolactin receptors, especially in the duodenum and jejunum, indicating the intestine as a target tissue of prolactin. A number of investigations show that prolactin is able to stimulate the intestinal calcium transport both in vitro and in vivo, whereas bromocriptine, which inhibits pituitary prolactin secretion, antagonizes its actions. In female rats, acute and long-term exposure to high prolactin levels significantly enhances the (i) transcellular active, (ii) solvent drag-induced, and (iii) passive calcium transport occurring in the small intestine. These effects are seen not only in pregnant and lactating animals, but are also observed in non-pregnant and non-lactating animals. Interestingly, young animals are more responsive to prolactin than adults. Prolactin-enhanced calcium absorption gradually diminishes with age, thus suggesting it has an age-dependent mode of action. Although prolactin's effects on calcium absorption are not directly vitamin D-dependent; a certain level of circulating vitamin D may be required for the basal expression of genes related to calcium transport. The aforementioned body of evidence supports the hypothesis that prolactin acts as a regulator of calcium homeostasis by controlling the intestinal calcium absorption. Cellular and molecular signal transductions of prolactin in the enterocytes are largely unknown, however, and still require investigation.     

Impact of vitamin E supplement in standard laboratory animal diet on microvascular manifestation of ischemia/reperfusion injury

Aimed at improving animal fertility and health, diets for farm and laboratory animals have over the last few years been supplemented with increasing amounts of the antioxidant vitamin E. We now demonstrate by intravital microscopy that feeding hamsters with a vitamin E-supplemented “standard” rodent diet (60 ppm vitamin E) significantly reduces the microvascular manifestations of ischemia/reperfusion injury when compared to animals fed a nonsupplemented diet. Postischemic leukocyte adhesion to venular endothelium was reduced from 770 ± 204 cells/mm² at 24 h after reperfusion in control animals on the nonsupplemented diet to 403 ± 105 cells/mm² in animals on the “standard” rodent diet (means ± SD, N = 7 animals per group, p < 0.01). Animals on the nonsupplemented diet showed a dramatic loss of capillary perfusion density until 7 days after reperfusion (to 21 ± 13% of preischemic baseline values), whereas this loss was significantly attenuated (to 71 ± 12% of preischemic values, p < 0.01) in animals on the “standard” rodent diet. No difference in the extent of reperfusion injury was seen between animals on the “standard” rodent diet and animals on diets with substantially higher vitamin E supplements (300 ppm–30.000 ppm). Besides underscoring the benefit of vitamin E in reducing the extent of ischemia/reperfusion injury, this study raises the concern that vitamin E supplements in “standard” laboratory animal diets may have a far-reaching impact on biomedical research by jeopardizing established animal models of disease.     

Impairment of pancreatic acinar function by reserpine in vivo and in vitro

Summary Chronic reserpine treatment of animals, an experimental model for cystic fibrosis (CF), results in generalized exocrinopathy, impaired pancreatic secretion, and decreased pancreatic content of amylase. The mechanisms of altered acinar function and decreased amylase content in both CF and the reserpine-treated rat are unknown. To examine this alteration, the rate of [³H]phenylalanine (phe) incorporation into cellular protein was determined in pancreatic acinar cells after reserpine treatment of rats in vivo (7 d) and of cells in vitro (1 to 24 h). Acinar cells isolated from control, chronic reserpine-treated, and pair-fed rats were incubated in vitro with 0, 30, 50, or 100 μM reserpine. Reserpine treatment in vitro for 24 h of acinar cells from control rats significantly decreased amylase activity (20 to 70%), an effect similar to that of reserpine treatment in vivo. In vivo, reserpine treatment decreased [³H]phe incorporation (disintegrations per minute per milligram protein) 56% in freshly isolated cells, but did not alter intracellular specific activity (disintegrations per minute per nanomole phe, SA) of [³H]phe. Reserpine treatment (30 and 50 μM) in vitro for 1 h also decreased [³H]phe incorporation by freshly isolated cells from control (53 to 85%) and pair-fed (40 to 68%) rats. Reserpine treatment for 24 h in vitro significantly decreased [³H]phe incorporation by cells from control (82 and 98%), pair-fed (80 and 95%), and chronic reserpine-treated (90 and 97%) rats as compared with cells from respective in vivo treatments cultured with no reserpine. In vitro reserpine treatment also decreased the intracellular SA of [³H]phe in freshly isolated cells from control (14 and 36%) and pair-fed (35 and 39%) rats and in cultured cells from control (11 and 86%), pair-fed (60 and 88%), and chronic reserpine-treated (49 and 76%) rats. However, these alterations of SA by reserpine did not account for the decreased incorporation of [³H]phe into acinar protein, which remained significantly lower (70 to 88%) when expressed as total phe incorporation. These results suggest (a) that reserpine acts directly on acinar cells to alter function and (b) that the ability of the pancreas to synthesize digestive enzymes may be impaired in this model of cystic fibrosis. This study was supported in part by the Cystic Fibrosis Foundation, Bethesda, MD.     

Kinetics of intestinal sugar transport, in vivo

Sugar absorption by the small intestine has been studied in rat and hamster in vivo, with luminal perfusion, during 1 minute successive periods. Transport is calculated as the difference between absorption and diffusion. The diffusion component is evaluated in the presence of phlorizin or as absorption of sorbose. The resulting KT values for glucose and galactose (rat: 7.7 and 10 mM; hamster: 10 and 14 mM) and 3-0-methyl-glucose (hamster: 25-33 mM) are quite lower than those previously obtained in vivo, but still higher than those in vitro. The physiological levels of glucose in the intestine of normally fed animals imply that the diffusion component plays an important role in the proximal regions of the small intestine, especially in rat.     

Vitamin absorption in the in vivo intestine of normal and infected (Hymenolepis diminuta: Cestoda) rats

Uptake and serosal transfer of the vitamins thiamine, riboflavin and folic acid have been studied in vivo in normal and parasitized rats infected with Hymenolepis diminuta (Cestoda). Regional differences in intestinal uptake of all three vitamins in both uninfected and parasitized animals were not satistically significant. In the parasitized intestine mucosal uptake and serosal transfer of thiamine were significantly inhibited, with increased mucosal accumulation of the vitamin as luminal thiamine concentration increased. Apparent increased riboflavin mucosal uptake in parasitized animals, was not matched by the reduced serosal transfer, suggesting adsorption of the vitamin in the unstirred aqueous layers. Mucosal uptake of folic acid increased in the parasitized gut; serosal transfer and mucosal accumulation were not affected. These results, indicating vitamin malabsorption associated with infection by H. diminuta, are consistent with the parasite inhibiting mucosal passive transport mechanisms. This conclusion is supported by the changes in net water fluxes associated with vitamin uptake in the parasitized intestine.     

Vitamin B6 deficiency alters tissue iron concentrations in the Wistar rat

PurposeWe investigated the effect of a vitamin B6 deficiency and pair-feeding on tissue trace element status.MethodTissue zinc, copper and iron concentrations were measured in 3 groups of young, male Wistar rats receiving a diet of 3.5 mg/kg (control group), 0 mg/kg (deficient group) and a pair-fed group over 8 weeks. The pair-fed group received the same diet consumed by the control. Tissue trace element analysis was performed using atomic absorption spectrophotometry and plasma vitamin B6 status was determined using HPLC.ResultsDeficiency resulted in elevation in liver iron concentration and reduction in muscle iron concentration. Muscle copper concentrations were reduced in the pair-fed and deficient groups vs. the control group. Tissue zinc concentrations remained unaffected by the deficiency. Kidney iron and heart copper levels were elevated in the pair-fed group.ConclusionsThe liver and muscle iron changes were due to the deficiency and not to reduced calorie intake and the latter may be due to impaired heme synthesis. The differences in copper between the groups were due to reduced food intake. Zinc seems to form a fixed pool in these animals. A dietary deficiency of vitamin B6 impacts on the trace element status of certain tissues in key metabolic tissues and hence needs to be factored into the amelioration of the condition.     

The isoelectric fractionation of hen''s-egg ovotransferrin

1. ATP sulphurylase was assayed in various organs from vitamin A-deficient and pair-fed control rats at different stages of deficiency. Activity decreased slightly in the liver and markedly in the adrenal gland. Striking differences in liver activity were observed between pair-fed control and ad libitum-fed animals. This observation suggested that diet (apart from vitamin A) strongly influenced the activity of ATP sulphurylase. 2. Total starvation caused a severe decrease in activity in liver within 48hr. This was due to a lack of protein intake. 3. By feeding groups of vitamin A-deficient and pair-fed control rats on a diet containing 80% protein, the specific activity of the liver ATP sulphurylase was maintained in the pair-fed control group at the normal level of an ad libitum-fed rat, whereas it decreased by 25% (statistically significant at P<0.01) in the deficient rat. On a 20%-protein diet, there were no significant differences between vitamin A-deficient and pair-fed control rats. These relationships held also for enzyme activity expressed per g. of liver, per total liver and per g. of DNA. There were no differences in liver protein or DNA concentration between vitamin A-deficient and control rats on either protein intake. 4. Control rats on a 20%-protein diet had liver specific enzyme activities about one-half of those in control rats on an 80%-protein diet, as well as lower liver protein concentrations. 5. It is concluded that, when the effect of protein deprivation on ATP sulphurylase is separated from the effect of vitamin A deficiency, a lowering of the enzyme activity caused by the vitamin deficiency is demonstrable.     

Effects of Perinatal Vitamin B6 Deficiency on Dopaminergic Neurochemistry

Long-Evans dams were fed either a vitamin B6-deficient or a control diet from day 13-14 of gestation and throughout lactation. A control pair-fed group was also included because of differences in food intake between vitamin B6-deficient and control ad libitum dams. The progeny of vitamin B6-deficient dams had all the classic symptoms of B6 deficiency. These included weight loss, ataxia, tremor, and epileptic seizures. Concentrations of the neurotransmitter dopamine (DA), and its metabolites 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA), as well as D-2 dopamine receptor binding, 3,4-dihydroxyphenylalanine (DOPA) decarboxylase activity, and vitamin B6 levels were measured in the corpus striatum of progeny at 7, 14, and 18 days after birth. Striatal DA and HVA levels were significantly decreased in B6-deficient animals when compared to ad libitum or pair-fed controls. Daily injections of vitamin B6 to deprived animals from the 14th to 18th day after birth improved the abnormal movement and normalized the concentration of DA but not of HVA in corpus striatum. Striatal D-2 dopamine receptor binding using [3H]spiperone as ligand was significantly reduced in 18-day-old animals as compared to ad libitum and pair-fed controls. No significant differences were found at 14 days. The administration of vitamin B6 to deprived animals did not raise the level of D-2 receptor binding during the period of observation. Scatchard plots indicated that the differences in binding were due to changes in receptor number and not in KD. Corpus striatum DOPA decarboxylase activity with and without the addition of exogenous pyridoxal phosphate was significantly reduced in 14- and 18-day-old animals when compared to pair-fed controls.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effects of vitamin E and carotenoid status on oxidative stress in health and disease. Evidence obtained from human intervention studies

Vitamin E and carotenoids are known to act as antioxidants both in vitro and in vivo. In this review we present a series of studies in healthy subjects and in patients who exhibit either acute or chronic oxidative stress. In the EU-Commission funded VITAGE project we investigated the status and effects of vitamin E and carotenoids on oxidative stress in 300 healthy volunteers. Depletion studies limiting dietary vitamin E or carotenoid intake to 25% of the dietary reference intakes and subsequent repletion by supplementation with either large doses of vitamin E or intermediate doses of carotenoids showed significant changes in ex vivo LDL oxidizability, total plasma peroxide concentrations and urinary 8-oxo-7,8-dihydro-2^′-deoxyguanosine excretion. Patients on chronic hemodialysis present with oxidative stress in the presence of normal vitamin E but impaired vitamin C status and, due to anemia, need to be treated with parenteral iron. We studied the effects of a single oral dose of vitamin E taken 6 h prior to intravenous infusion of 100 mg iron, which exceeded the iron-binding capacity of transferrin. Vitamin E significantly reduced and in combination with a single dose of vitamin C completely abrogated acute oxidative stress induced by the iron load. Patients with cystic fibrosis are exposed to chronic oxidative stress due to an overproduction of reactive oxygen species as a result of neutrophil-dominated lung inflammation and impaired antioxidant status. Biochemical vitamin E and carotenoid deficiencies could be fully corrected even in the presence of fat malabsorption using intermediate doses of either RRR -tocopherol or all-rac -tocopheryl acetate and water-miscible all-trans β-carotene. Long-term supplementation reduced ex vivo LDL oxidizability, in vivo lipid peroxidation and lung inflammation.     

Role of endothelin-1 in regulation of the postnatal intestinal circulation

Newborn intestine is uniquely prone to vasoconstriction in response to a wide variety of perturbations. To test the hypothesis that endothelin (ET)-1 is an important factor in this process, we determined the effects of exogenous ET-1 administration and blockade of endogenous ET-1 in vivo and in vitro in 3- and 35-day-old swine. Intramesenteric artery administration of exogenous ET-1 to vascularly isolated in vivo gut loops (10(-9) M/kg bolus) caused vasoconstriction and reduced gut O(2) uptake similarly in these age groups. Selective blockade of ET(A) or ET(B) receptors with BQ-610 or BQ-788, respectively, in vascularly isolated in vivo gut loops had no effect on gut vascular resistance or O(2) uptake in either age group; within in vitro gut loops, BQ-610 significantly increased vasoconstriction when perfusion pressure was reduced below baseline, but only in 3-day-old animals; i.e., it impaired the autoregulatory response to perfusion pressure reduction. Exogenous ET-1 significantly decreased capillary perfusion within in vitro gut loops, as evidenced by a decrease in capillary filtration coefficient, but only in 3-day-old animals; furthermore, blockade of endogenous ET-1 activity with BQ-610 significantly enhanced capillary filtration coefficient in 3-day-old animals and increased O(2) extraction ratio. ET-1 did not depress intestinal metabolic rate, as evidenced by its effect on the O(2) uptake-blood flow relationship; it did compromise tissue oxygenation because of its effects on intestinal O(2) transport. ET-1 concentration in mesenteric venous effluent exceeded arterial concentration, but only in 3-day-old intestine, suggesting production of ET-1 by newborn intestine. We conclude that ET-1 exerts an age-dependent effect on intestinal hemodynamics in postnatal intestine, having a greater impact in 3- than in 35-day-old intestine.     

Effect of vitamin A deficiency on hepatic and intestinal drug metabolizing enzymes in rats

Feeding of vitamin A-deficient diet to male weanling rats for 10 weeks caused significant reduction in the hepatic cytochrome P-450, cytochrome b5, aminopyrine N-demethylase and arylhydrocarbon hydroxylase activities. Contrary to this, the levels of these Phase I enzymes were found to be significantly elevated in all the 3 portions (proximal, middle and distal) of the intestine in deficient animals as compared to corresponding pair-fed controls. Of the Phase II enzymes studied, UDP-glucuronyltransferase showed a significant decrease whereas glutathione S-transferase showed a significant increase in vitamin A-deficient rat liver and small intestine. The study suggests that vitamin A deficiency causes an imbalance between the Phase I and phase II drug metabolizing enzyme systems which may decrease the capacity of the organism to withstand the neoplastic effects of chemical carcinogens in vitamin A deficiency.     

Effect of vitamin A dietary intake on in vitro and in vivo activation of aflatoxin B1.

The mechanism by which vitamin A prevents or delays in chemical carcinogenesis remains unclear. In the present study, we assess the suggestive role of vitamin A in the initiation phase of carcinogenesis. We have conducted a dose-effect relationship between vitamin A dietary intake and aflatoxin B1 (AFB1) genotoxicity measured both in vitro and in vivo. Thus AFB1-induced mutagenesis in Salmonella typhimurium TA98 was investigated and compared to AFB1-induced single-strand breaks (SSBs) in DNA of rat hepatocytes. Rats were fed ad libitum with diet containing 0, 5, 50 or 500 IU of retinyl palmitate for 8 weeks. The AFB1-treated rats were injected i.p. with 1 mg/kg body weight. In the Ames test conditions TA98 back-reversion was negatively correlated with the log of vitamin A concentration in liver S9 fractions from experimental groups. However, the activities of metabolizing enzymes which specifically activate or deactivate AFB1 were found to be significantly decreased in vitamin A-deficient animals and weakly modified in vitamin A-supplemented animals. For in vivo experiments, the DNA elution rate of both AFB1-treated and untreated rats was increased in vitamin A deficiency condition (+79% and +17% respectively) and was reduced with the higher vitamin A dietary level (-44% and -53% respectively). DNA damage measured in vivo showed a significant positive correlation with mutagenic activity measured in the Ames test. These results confirm that the vitamin A status of animals can influence AFB1 genotoxic activity in vitro and indicate that this phenomenon also occurs in vivo. Thus a similar mechanism may be considered for the protective action of vitamin A both in vitro and in vivo. However, this mechanism is unlikely to involve modulation of the microsomal enzyme system responsible for AFB1 metabolism. Therefore a protective mechanism at the cytosolic or nuclear levels may be suggested.     

Protective effects of vitamin E on carbon tetrachloride-induced liver damage in rats

In this study we investigated whether the increase of hepatic vitamin E content by intraperitoneal administration, influences chronic liver damage induced by carbon tetrachloride (CCl(4)) in rats. Thirty adult male Wistar rats were divided into three groups. The first group was used as a control and the rats in the second group were administered CCl(4) in olive oil subcutaneously. Rats in the third group were administered intraperitoneally vitamin E (dl-alpha-tocopherol acetate, 100 mg kg(-1)). This administration was performed three times per week for five weeks. Liver samples were used for the determination of vitamin E levels, glutathione peroxidase (GSHPx) activities and histological examination. Serum levels of alanine aminotransferase, lactate dehydrogenase, alkaline phosphatase, aspartate aminotransferase, gamma-glutamyltranspeptidase, total and conjugated bilirubin were significantly (p<0.05, p<0.01, p<0.001) higher in animals treated with CCl(4) than in the controls and had returned to normal values by the administration of vitamin E + CCl(4 ). Liver vitamin E levels were significantly (p<0.05) lower in the CCl(4) group than in the control group. However, the liver vitamin E content was significantly (p<0.01, p<0.001) increased in the vitamin E + CCl(4) injected group. On the other hand, liver GSHPx activity was not statistically different among the groups. On histological examination, vitamin E administered animals showed incomplete, but significant, prevention of liver necrosis and cirrhosis induced by CCl(4 ). these data indicate that intraperitoneally administered vitamin E has protective effects against CCl(4)-induced chronic liver damage and cirrhosis as evidenced by biochemical data and conventional histological examination.     

Gamma-tocopherol supplementation inhibits protein nitration and ascorbate oxidation in rats with inflammation

Gamma-tocopherol (gammaT) complements alpha-tocopherol (alphaT) by trapping reactive nitrogen oxides to form a stable adduct, 5-nitro-gammaT [Christen et al., PNAS 94:3217-3222; 1997]. This observation led to the current investigation in which we studied the effects of gammaT supplementation on plasma and tissue vitamin C, vitamin E, and protein nitration before and after zymosan-induced acute peritonitis. Male Fischer 344 rats were fed for 4 weeks with either a normal chow diet with basal 32 mg alphaT/kg, or the same diet supplemented with approximately 90 mg d-gammaT/kg. Supplementation resulted in significantly higher levels of gammaT in plasma, liver, and kidney of control animals without affecting alphaT, total alphaT+gammaT or vitamin C. Intraperitoneal injection of zymosan caused a marked increase in 3-nitrotyrosine and a profound decline in vitamin C in all tissues examined. Supplementation with gammaT significantly inhibited protein nitration and ascorbate oxidation in the kidney, as indicated by the 29% and 56% reduction of kidney 3-nitrotyrosine and dehydroascorbate, respectively. Supplementation significantly attenuated inflammation-induced loss of vitamin C in the plasma (38%) and kidney (20%). Zymosan-treated animals had significantly higher plasma and tissue gammaT than nontreated pair-fed controls, and the elevation of gammaT was strongly accentuated by the supplementation. In contrast, alphaT did not significantly change in response to zymosan treatment. In untreated control animals, gammaT supplementation lowered basal levels of 3-nitrotyrosine in the kidney and buffered the starvation-induced changes in vitamin C in all tissues examined. Our study provides the first in vivo evidence that in rats with high basal amounts of alphaT, a moderate gammaT supplementation attenuates inflammation-mediated damage, and spares vitamin C during starvation-induced stress without affecting alphaT.