Effect Of Chronic Ethanol Feeding Research Articles

Protein metabolism in the small intestine of the ethanol‐fed rat

The effects of chronic ethanol feeding on the small intestine were investigated in young rats. Rats were fed a nutritionally-adequate liquid diet, containing 36 per cent of total energy as ethanol (treated, n = 7), or isovolumetric amounts of the same diet in which ethanol was substituted by isocaloric glucose (controls, n = 7). After six weeks the wet weight and total tissue contents of protein, RNA and DNA were significantly reduced by 21 per cent, 23 per cent, 16 per cent and 28 per cent respectively, (p less than 0.014). Rates of protein synthesis were measured with L[4(3H)]phenylalanine and fractional rates (defined as the percentage of constituent tissue protein synthesised each hour, i.e. ks, % h-1) were calculated from the specific radioactivity of free phenylalanine in both tissue homogenates and plasma. Ethanol-feeding reduced ks by approx 10 per cent (p less than 0.181). The amount of protein synthesized unit-1 RNA was also reduced by approx 15 per cent (p less than 0.059) but the amount of protein synthesis unit-1 DNA was unaffected by ethanol-feeding (p less than 1.000). In contrast, the absolute rates of protein synthesis were reduced by approximately 30 per cent (p less than 0.022). It was concluded that, as the small intestine contributes to approx. 20-25 per cent of whole body synthesis these results may have an important effect on whole body nitrogen homeostasis and may have implications for the gastrointestinal effects of ethanol seen during chronic alcoholic abuse.

Effects of chronic ethanol feeding on rat hepatocytic glutathione. Relationship of cytosolic glutathione to efflux and mitochondrial sequestration.

Chronic ethanol feeding to rats increases the sinusoidal component of hepatic glutathione (GSH) efflux, despite a lower steady-state GSH pool size. In the present studies, no increase of biliary GSH efflux in vivo was found in chronic ethanol-fed cells. Studies were performed on ethanol-fed and pair-fed cells to identify the kinetic parameters of cellular GSH concentration-dependent efflux. The relationship between cytosolic GSH and the rate of efflux was modeled by the Hill equation, revealing a similar Vmax, 0.22 +/- 0.013 vs. 0.20 +/- 0.014 nmol/min per 10(6) cells for ethanol-fed and pair-fed cells, respectively, whereas the Km was significantly decreased (25.3 +/- 2.3 vs. 33.5 +/- 1.4 nmol/10(6) cells) in ethanol-fed cells. The difference in Km was larger when the data were corrected for the increased water content in ethanol-fed cells. We found a direct correlation between mitochondria and cytosolic GSH, revealing that mitochondria from ethanol-fed cells have less GSH at all cytosolic GSH values. The rate of resynthesis in depleted ethanol-fed cells in the presence of methionine and serine was similar to control cells and gamma-glutamylcysteine synthetase remained unaffected by chronic ethanol. However, the reaccumulation of mitochondrial GSH as the cytosolic pool increased was impaired in the ethanol cells. The earliest time change in GSH regulation was a 50% decrease in the mitochondrial GSH at 2 wk.

Impaired Hepatic Elimination of Paranitrophenol and Its Metabolites in the Rat following Chronic Ethanol Pretreatment

Chronic ethanol feeding has been shown to enhance hepatic microsomal drug oxidation in humans and in laboratory animals. However, the effects of chronic ethanol administration on drug conjugation are less conclusive. We have studied the effects of chronic ethanol feeding on (a) the conjugation and elimination of p-nitrophenol (PNP) by the isolated perfused rat liver, (b) the formation of PNP glucuronide by hepatic microsomal PNP-glucuronyltransferase in vitro and (c) the hepatic content of UDP-glucuronic acid (UDPGA). PNP elimination from the isolated perfused rat liver was best described as a combination of parallel saturable and first-order processes. Ethanol pretreatment did not influence the former but resulted in a 48% reduction in the rate of elimination by the latter. This was associated with a significant reduction in recovery of PNP-glucuronide from bile, but no change in concentrations of PNP glucuronide or sulfate in perfusate. Michaelis constants and Vmax for PNP-glucuronyltransferase in native and solubilized microsomes and UDPGA concentrations in liver were not influenced by ethanol pretreatment. These results suggest that chronic ethanol treatment reduces PNP elimination in the intact liver primarily via a reduction in the biliary excretion of PNP glucuronide without altering glucuronidation per se.

The effect of chronic ethanol ingestion on protein metabolism in type-I- and type-II-fibre-rich skeletal muscles of the rat

1. The effects of chronic ethanol feeding on muscles containing a predominance of either Type I (aerobic, slow-twitch) or Type II (anaerobic, fast-twitch) fibres were studied. Male Wistar rats, weighing approx. 90 g or 280 g, were pair-fed on a nutritionally complete liquid diet containing 36% of total energy as ethanol, or isovolumetric amounts of the same diet in which ethanol was replaced by isoenergetic glucose. After 6 weeks feeding, fractional rates of protein synthesis were measured with a flooding dose of L-[4-(3)H]-phenylalanine and muscles were analysed for protein, RNA and DNA. 2. Ethanol feeding decreased muscle weight, protein, RNA and DNA contents in both small and large rats. Type-II-fibre-rich muscles showed greater changes than did Type-I-fibre-rich muscles. Changes in protein paralleled decreases in DNA. 3. The capacity for protein synthesis (RNA/protein), fractional rates of protein synthesis and absolute rates of protein synthesis were decreased by ethanol feeding in both small and large rats. The amounts of protein synthesized relative to RNA and DNA were also decreased. Changes were less marked in Type-I than in Type-II-fibre-rich muscles. Loss of protein, RNA and DNA was greater in small rats, but protein synthesis was more markedly affected in large rats. 4. It was concluded that chronic ethanol feeding adversely affects protein metabolism in skeletal muscle. Fibre composition and animal size are also important factors in determining the pattern of response.

Effect of chronic ethanol ingestion on pancreatic protein synthesis

The effect of chronic ethanol feeding on pancreatic synthesis protein synthesis was assessed by studying the rate of incorporation of [ 3H]leucine into proteins in isolated rat pancreatic acini in vitro. Chronic ethanol feeding increased the rate of protein synthesis (2–3-fold) compared to controls fed an isocaloric diet. The onset of the increase in protein synthesis was detectable 2 days after the beginning of ethanol feeding, reached a maximum after 7 days and remained constant for up to 4 months. The increased incorporation of [ 3H]leucine was not due to an increased turnover of proteins as measured in pulse-chase experiments. After separation of individual digestive enzymes by SDS-polyacrylamide gel electrophoresis and determination of the distribution of radioactivity in different proteins, a general increase in the rate of incorporation of the label into all of the protein was observed. In contrast to the observations made with isolated acini, there was no significant difference between the control and ethanol-fed groups when the rate of pancreatic protein synthesis was measured in vivo. However, overnight withdrawal of ethanol led to an increase of approx. 70% in protein synthesis in the ethanol-fed group. These results suggest that chronic ethanol ingestion modifies the control of pancreatic protein synthesis; the enhanced protein synthesis is expressed in isolated acini, i.e., in the absence of physiological factor during chronic ethanol ingestion and vivo after ethanol withdrawal.

Effect of chronic ethanol feeding on the hepatic secretion of very-low-density lipoproteins

Using Triton WR-1339 to inhibit plasma metabolism of VLDL, hepatic secretion of VLDL was measured in vivo in control and alcohol-fed rats. The composition of VLDL secreted from livers of chronically alcohol-fed rats was compared with pair-fed controls. The results show that the cholesterol, esterified and nonesterified, as well as the triacylglycerol content of VLDL secreted by the alcohol-fed rats was significantly reduced; protein and phospholipid constituents of the VLDL were unaltered. The apoprotein components of the VLDL showed no apparent difference between the two groups. However, fatty-acid composition of VLDL triacylglycerol and phospholipids show a significant reduction in 20:4 and an increase in 18:2 fatty acids in the alcohol-fed rats. VLDL secretion rates by chronic alcohol-fed rats was reduced significantly compared to the control animals, and this observation has been implicated in the pathogenesis of alcoholic fatty liver.

The effect of chronic ethanol feeding on ornithine decarboxylase activity and liver regeneration.

The effects of ethanol on liver regeneration are poorly understood. Acute and chronic exposure to ethanol have been found to exert opposite effects on the induction of ornithine decarboxylase, the rate-limiting enzyme for polyamine biosynthesis. Polyamines are necessary for DNA synthesis and liver regeneration after chemical or surgical liver injury. Short-term exposure to ethanol, which inhibits ornithine decarboxylase has been shown to inhibit DNA synthesis and liver regeneration, whereas more chronic exposure to ethanol increases ornithine decarboxylase activity and therefore could conceivably stimulate DNA synthesis and regeneration. To explore this later possibility, the effects of chronic ethanol consumption on ornithine decarboxylase activity, DNA synthesis and liver regeneration were studied in rats after sham laparotomy and partial hepatectomy. Chronic ethanol feeding failed to inhibit the induction of ornithine decarboxylase that occurred after partial hepatectomy and yet significantly inhibited posthepatectomy DNA synthesis and restitution of liver mass. These data suggest that the induction of hepatic polyamine biosynthesis is dissociated from DNA synthesis and liver regeneration after chronic consumption of ethanol.

The Effect of Chronic Ethanol Feeding on Acetyl COA Carboxylase (EC 6.4.1.2) and Fatty Acid Synthase (EC 2.3.1.85) Activities in Rat Liver

The Effect of Chronic Ethanol Feeding on Acetyl COA Carboxylase (EC 6.4.1.2) and Fatty Acid Synthase (EC 2.3.1.85) Activities in Rat Liver

Effect of chronic ethanol feeding on glutathione and glutathione-related enzyme activities in rat liver

The effects of chronic ethanol consumption on liver glutathione concentrations and glutathione-related enzyme activities were studied in rats over a period of 1–9 weeks. The animals received a liquid diet containing 36% of calories as ethanol or isocaloric carbohydrate. Glutathione concentrations were significantly enhanced following ethanol intake with increases of 99% after 3 weeks and a progressive decrease thereafter. Glutathione S-transferase activity reached a maximum increase of 36% after 2 weeks of ethanol feeding. Glutathione peroxidase activity remained unchanged for the first 6 weeks of treatment, with a tendency to decrease in the last weeks of ethanol consumption. Our findings indicate that chronic ethanol administration profoundly modifies the hepatic metabolism of glutathione and may thus have important effects on the detoxification of xenobiotics by the liver.

Alcohol and cancer.

Alcohol and cancer.

Effect of chronic ethanol and food deprivation on intestinal villus morphology and brush border membrane content of lipid and marker enzymes.

Brush border membranes (BBM) were isolated from the jejunum and ileum of control, ad libitum (CAL); control, food-restricted (CFR); control, weight gain (CWG); and ethanol-fed (EF) rabbits. Jejunal alkaline phosphatase activity was similar among control groups, but higher in CAL than EF animals. Sucrase activity was higher in EF and CWG animals than in CAL and CFR. The alkaline phosphatase/sucrase ratio was lower in EF than control animals. Ileal enzyme marker activity was similar among EF and control animals. Sucrase (S) activity was lower in the ileum than in the jejunum. Jejunal free fatty acid and phospholipid/cholesterol (PL/C) were lower in EF than control animals, whereas ileal lipid content was generally similar among all animal groups. Total phospholipid content was similar between sites, but the cholesterol and free fatty acid content were lower in the ileum than the jejunum. The phospholipid/cholesterol ratio was increased only in the ileum of EF animals. The amount of lecithin was decreased in the jejunal BBM of EF animals resulting in a decreased choline/amine phospholipid ratio as compared with control animals. The ileal phospholipid composition was similar among all groups. A large increase in villus height is observed in the jejunum of EF animals. Villus surface area and mucosal surface area are altered with ethanol feeding and food deprivation. Thus, (i) there is a gradient of S and cholesterol between the BBM of jejunum and ileum; (ii) changes in food intake are associated with changes in the morphology as well as the enzyme marker and lipid content of BBM.(ABSTRACT TRUNCATED AT 250 WORDS)

Effect of chronic ethanol feeding upon the catabolism of protein and lipid moieties of chylomicrons and very low density lipoproteins in vivo and in the perfused heart system.

Effect of chronic ethanol feeding for 6 weeks to male Wistar rats upon the catabolism of rat chylomicrons and very low density lipoproteins was investigated both in vivo and in the perfused heart system. The exponential decay curves in the plasma compartment or in the perfused heart system of these lipoproteins labeled in the protein or triacylglycerol or cholesterol moieties were determined. It was found that chronic ethanol feeding inhibited the catabolism of both protein and triacylglycerol moieties by 26-35%, whereas that of the cholesterol moiety was inhibited by 67-71%. Since the catabolism of the triacylglycerol moiety takes place essentially in the extrahepatic tissues while that of the cholesterol moiety occurs in the liver, it is concluded that ethanol affects the catabolism of triacylglycerol-rich lipoproteins in the liver more than in the extrahepatic tissues.

Effects of chronic ethanol feeding on glutathione turnover in the rat

Glutathione (GSH) is important in protection of cells against electrophilic drug injury and against reactive oxygen species. Both steady-state concentrations and turnover of GSH are important determinants of susceptibility of the hepatocyte to injury. Chronic ethanol administration is known to enhance susceptibility to electrophilic drug injury. We have examined the effects of chronic ethanol feeding on GSH turnover and the hepatic activities of GSH peroxidase and enzymes of the γ-glutamyl cycle in the rat. Turnover of GSH was measuremed in individual animals by measuring the decrease in specific activity of GSH in bile over time after i.v. administration of [ 35S]cysteine. Rats fed ethanol had significantly increased rates of GSH turnover, 0.287 ± 0.050 hr −1 vs 0.131 ± 0.041 hr −1 (P < 0.001), as well as steady-state GSH levels, 6.59 ± 1.55 vs 4.30 ± 1.28 μmoles/g liver (P < 0.01). The activities of gamma-glutamyltransferase (GGT) and GSH-synthesizing enzymes were corresponding increased significantly. By contrast, GSH peroxidase activity was decreased in ethanol-fed rats, 194 ± 20.8 vs 311 ± 89.9 nmoles NADPH oxidized/min/mg protein (P < 0.001). Biliary output and concentrations of GSH and GSSG were similar in both groups. The increase in turnover of GSH was not due to an increase in oxidation of GSH. There was, however, an association between GSH turnover and the activity of hepatic GGT in ethanol-fed but not in control rats.

Effects of Chronic Ethanol Feeding on Sympathetic Innervated Organs: Temporal Sequence of Biochemical, Functional, and Trophic Changes

In noradrenergic nerves as well as in effector organs of the sympathetic nervous system, chronic ethanol feeding induced biochemical, functional, and trophic changes. The time sequence of modifications in the activities of tyrosine hydroxylase, phenylethanolamine-N-methyl transferase, monoamine oxidase, endogenous norepinephrine levels, as well as the neuronal uptake mechanism and secretory responses to norepinephrine in the submaxillary gland of the rat, confirms that the effects of chronic ethanol administration are related to an increase in sympathetic tone.

Effects of chronic ethanol feeding and acetaldehyde metabolism on calcium transport by rat liver mitochondria

The prolonged feeding of ethanol to rats alters in vitro mitochondrial transport of calcium. Hepatic mitochondria isolated from rats fed ethanol for 7 weeks exhibited decreased retention of calcium in the presence of 4mM-Pi. This defect was associated with enhanced efflux of calcium when mitochondria were incubated with EGTA. Acetaldehyde at low, “physiological” concentrations (100μM) enhanced calcium retention by mitochondria but this response was blunted after chronic ethanol administration. The in vitro actions of acetaldehyde appear to be mediated, in part, by its metabolism in mitochondria since pretreatment of rats with cyanamide (an aldehyde dehydrogenase inhibitor) prevents this effect.

Effects of Chronic Ethanol Feeding on the Metabolism of Tryptophan and Nicotinamide in Rats

Oxidation of tryptophan and urinary excretion of N1-methylnicotinamide were studied in rats fed a modified DeCarli-Lieber liquid diet of low (1.1 mg/liter or 5 mg/kg dry diet) and adequate (5.5 mg/liter or 25 mg/kg dry diet) nicotinamide content with and without ethanol replacing carbohydrate to 36% of total calories. Protein was restricted to 12% of calories so that the tryptophan content was about 0.16% of dry weight. Rats were fed for 30 or 40 days and feeding was restricted to 7 or 12 hours per day. N1-Methylnicotinamide was measured in urine collected during the last 2 days of feeding. At the end of the experimental period, fasted rats were injected intraperitoneally with 0.8 µCi of L-[ring-2-14]tryptophan (0.12 or 0.11 µCi/mg tryptophan) or 0.7 µCi of[14C]formate (0.44 µCi/mg formate) per 100 g body weight. Expired CO2 was collected every 30 minutes or hourly for 5 or 6 hours. N1-Methylnicotinamide excretion was increased in ethanol-fed rats (P < 0.05). The percent of injected [14C]tryptophan recovered as 14CO2 was increased in rats fed ethanol in a low niacin diet as compared to controls (P < 0.01). Recovery of 14CO2 from [14C]formate oxidation was not affected by ethanol feeding. These findings suggest that the initial oxidation of tryptophan by tryptophan 2,3-dioxygenase is enhanced following chronic ethanol feeding.

Effects of chronic ethanol administration on hepatic glycoprotein secretion in the rat

The effects of chronic ethanol feeding on protein and glycoprotein synthesis and secretion were studied in rat liver slices. Liver slices from rats fed ethanol for 4–5 wk showed a decreased ability to incorporate [14C]glucosamine into medium trichloracetic acidprecipitable proteins when compared to the pair-fed controls; however, the labeling of hepatocellular glycoproteins was unaffected by chronic ethanol treatment. Immunoprecipitation of radiolabeled secretory (serum) glycoproteins with antiserum against rat serum proteins showed a similar marked inhibition in the appearance of glucosamine-labeled proteins in the medium of slices from ethanol-fed rats. Minimal effects, however, were noted in the labeling of intracellular secretory glycoproteins. Protein synthesis, as determined by measuring [14C]leucine incorporation into medium and liver proteins, was decreased in liver slices from ethanol-fed rats as compared to the pair-fed controls. This was the case for both total proteins as well as immunoprecipitable secretory proteins, although the labeling of secretory proteins retained in the liver slices was reduced to a lesser extent than total radiolabeled hepatic proteins. When the terminal sugar, [14C]fucose, was employed as a precursor in order to more closely focus on the final steps of hepatic glycoprotein secretion, liver slices obtained from chronic ethanol-fed rats exhibited impaired secretion of fucose-labeled proteins into the medium. When ethanol (5 or 10 mM) was added to the incubation medium containing liver slices from the ethanol-fed rats, the alterations in protein and glycoprotein synthesis and secretion caused by the chronic ethanol treatment were further potentiated. The results of this study indicate that liver slices prepared from chronic ethanol-fed rats exhibit both impaired synthesis and secretion of proteins and glycoproteins, and these defects are further potentiated by acute ethanol administration.

Effect of chronic ethanol feeding on pancreatic enzyme secretion in rats in vitro.

The effect of long-term ethanol intake on pancreatic digestive enzyme secretion was determined in Sprague-Dawley rats. Weight-matched triplets were fed Lieber-DeCarli diet containing 5% w/v concentration of ethanol, isocaloric amounts of Lieber-DeCarli diet, or rat chow ad libitum for 6, 12, and 18 months. Basal and bethanechol-stimulated secretion of amylase, lipase, trypsinogen, chymotrypsinogen, and pancreatic secretory trypsin inhibitor (PSTI) was determined. In the ethanol-fed group, basal secretion of trypsinogen and chymotrypsinogen was increased at 6, 12, and 18 months. In addition, basal secretion of amylase and lipase was increased and that of PSTI was decreased at 12 and 18 months. Secretion of PSTI was stimulated by bethanechol (10(-4)M), whereas the secretion of digestive enzymes was not stimulated in the ethanol-fed versus two control groups. At 12 months the dose-response curve of amylase and lipase secretion was shifted upwards in the ethanol-fed group with increase in ED50. These data are suggestive of membrane perturbations leading to increased basal secretion and a subsensitivity of the cholinergic receptors in the ethanol-fed group. Increased basal secretion of proteases in the presence of diminished trypsin inhibitor indicates that premature activation of proenzymes could occur resulting in pancreatitis.

Effect of chronic ethanol feeding on hepatic mitochondria in the monkey.

The effect of chronic ethanol feeding on hepatic mitochondrial morphology and histochemically measured succinic dehydrogenase activity was assessed. Five monkeys of the species Macaca radiata received a nutritionally adequate diet containing 50% of the calories as ethanol, while five others were pair-fed the same diet except that ethanol was isocalorically substituted by carbohydrate. Liver morphology was assessed at 12 and 24 months and at sacrifice after 40 to 48 months of ethanol feeding. The ethanol-fed animals developed mild to moderate fatty liver as did some of the controls. No necrosis or fibrosis developed. All ethanol-fed animals developed centrilobolar megamitochondria and centrilobular "shift" in histochemically assayed succinic dehydrogenase activity characteristic of animals fed ethanol for prolonged periods. These mitochondrial changes persisted throughout the 48-month test period without progressive increase in severity or accompanying pathology. It is concluded that the morphologic and histochemically assessed mitochondrial changes do not necessarily represent a progressive destructive effect of ethanol.

Effect of chronic ethanol feeding on hepatic collagen in the monkey.

The effect of chronic ethanol feeding was determined on parameters of hepatic collagen metabolism in the monkey. Four monkeys of the species Macaca radiata received a nutritionally adequate diet containing 50% of the calories as ethanol, while four others were pair-fed a diet in which ethanol was isocalorically substituted by carbohydrate. Liver biopsies were obtained at 3, 12, and 24 months, and the animals were killed between 40 and 48 months after initiation of the diets. The ethanol-fed animals developed various degrees of fatty infiltration, but no necrosis, inflammation, or fibrosis. The amount and distribution of collagen Types I, III, and IV demonstrated by immunohistochemical techniques was not altered after ethanol feeding. No changes were found in hepatic protein-bound hydroxyproline or in collagen prolyl hydroxylase activity at the various time intervals. Liver free proline and the incorporation of labeled proline into protein-bound hydroxyproline by liver slices were not altered after 40 to 48 months of ethanol feeding. This study shows that prolonged feeding of ethanol together with an adequate diet results in fatty infiltration but not deposition or alteration of hepatic collagen metabolism.