bluuebunny/crossref_metadata_2025 huggingface dataset:
Sensitivity to Ethanol‐Induced Ataxia in HOT and COLD Selected Lines of Mice
Authors: Schafer Gwen L. , Crabbe John C.
Published: April 2006

Studies with inbred strains of mice have suggested that there may be a genetic correlation between strain sensitivities to the ataxic and hypothermic responses to ethanol (EtOH), which would suggest that some genes influence both responses. To test this hypothesis, EtOH sensitivity was determined in replicate lines of mice selectively bred for sensitivity (COLD) or resistance (HOT) to acute ethanol hypothermia. Several tests were used to index ataxia, related traits such as muscle strength, and locomotor activity. The screen test yielded a dose‐dependent EtOH‐induced decrease in performance that did not differ between the selected lines. Based on the dose—response characteristics of this task, 2.5 g/kg of EtOH was used as the test dose for the remaining experiments. Results from the fixed‐speed rotarod and the grid test of motor incoordination also indicated no significant differences between HOT and COLD mice in sensitivity to EtOH impairment. When the selected lines were tested on an accelerating rotarod, COLD mice were impaired by the acute EtOH injection, but HOT mice were unaffected. COLD mice were more sensitive to EtOH‐induced decrements in grip strength and locomotor activity. Overall, the results indicated that HOT and COLD mice were only differentially sensitive to EtOH in some tasks related to ataxia, suggesting that some genes must be associated uniquely with EtOH induced hypothermia or ataxia. The mixed results from the various tests indicate that ataxia can best be conceived as a group of related complex behaviors that cannot be assessed adequately by the use of a single task and that ataxia‐related behaviors are influenced by different groups of genes.
Alcohol and Cytokine‐inducible Transcription Factors
Authors: Zeldin Gillian , Yang Shi Qi , Yin Ming , Lin Hui Zhi , Rai Rudra , Diehl Anna Mae
Published: April 2006

Cytokines, such as TNFα, modulate the behavior of many cells by regulating the expression of a wide array of genes. When a cytokine binds to its receptor on the cell surface, the receptor becomes activated and activates signal transduction cascades. These cascades typically involve a series of phosphorylation reactions that lead to sequential activation of various kinases. The targets of these kinases include DNA binding proteins that regulate the transcription of target genes. The activity of DNA binding proteins, such as c‐Jun and NF‐kappa B, titrates the transcriptional activity of cytokine‐regulated genes. Both acute and chronic alcohol consumption of ethanol increase hepatic expression of TNFα. After acute ethanol consumption, this is associated with increased induction of several TNF‐dependent regenerative events, including the activation of c‐Jun and increased binding activity of NF‐kappa B. However, chronic consumption of ethanol appears to impede TNFα signaling in the liver because it attenuates the increases in c‐JUN activity and NF‐kappa B binding, which normally follow partial hepatectomy. These results suggest that one mechanism by which ethanol influences liver cell behavior is by influencing local expression of TNFα and changing the activity of TNF‐regulated transcription factors.
Ethanol Ingestion Impairs Neutrophil Bactericidal Mechanisms against Streptococcus pneumoniae
Authors: Jareo Patti W. , Preheim Laurel C. , Gentry Martha J.
Published: April 2006

Ethanol ingestion impairs both killing of selected pneumococcal strains by rat polymorphonuclear leukocytes (PMNL) in vitro and clearance of these same strains from experimentally infected rat lungs. To determine the mechanism(s) of this impairment, we isolated neutrophils (PMNL) by a magnetic cell sorting technique from the peripheral blood of chow‐fed rats (C‐PMNL) or rats pair fed for 7 days with a liquid diet providing 36% of its calories as either ethanol (E‐PMNL) or dextrin‐maltose (P‐PMNL). Phagocytosis of fluorochrome‐labeled bacteria and oxygen radical production, as determined by oxidation of dihydrorhodamine 123, were measured by flow cytometry. Degranulation, as determined by lysozyrne release, was measured as lysis of a suspension of Micrococcus lysodeikticus. E‐PMNL, P‐PMNL, and C‐PMNL were equivalent in their ability to phagocytose pneumococci and to produce an oxidative burst in response to stimulation by opsonized zymosan. However, E‐PMNL produced fewer oxygen radicals and released less lysozyme than either P‐PMNL or C‐PMNL when stimulated by exposure to S. pneumoniae. There was no difference in oxygen radical production by E‐PMNL and P‐PMNL when stimulated with phorbol myristate acetate, but both cell types mounted a significantly reduced response in comparison to C‐PMNL. These data suggest that ingestion of ethanol for 7 days significantly reduces both the oxidative burst and degranulation of rat PMNL in response to S. pneumoniae, thereby compromising anti‐pneumococcal activity.
Acute Hemodynamic, Pituitary, and Adrenocortical Responses to Alcohol in Adult Female Sheep
Authors: Cudd Timothy A. , Chen Wei‐Jung A. , West James R.
Published: April 2006

Alcohol was infused intravenously Into chronically cannulated adult female sheep as a 40% solution (w/v) at doses of 0.5,1.0, 1.5, or 2.0 g/kg over 1 hr. Saline infusions, equal in volume to the highest dose, served as a control. Dose‐dependent peak blood alcohol concentrations (BACs) were attained 60 min after the beginning of alcohol infusion for all doses (88.5 ± 3.7, 213.2 ± 11.0, 373.2 ± 14.3, and 494.1 ± 34.5 mg/dl ± SE, respectively). Plasma cortisol concentrations Increased in response to the 0.5 g/kg infusions (BACs less than 100 mg/dl), whereas both ACTH and cortisol concentrations increased in the 1.0 and 2.0 g/kg dose groups. Mean arterial pressure, heart rate, and Pao2, increased, whereas Pao2, decreased in response to the 1.5 and 2.0 g/kg infusions. Arterial pH declined in the highest dose group. Respiratory rate was lower in all groups receiving alcohol compared with that of the control group. Hematocrit did not change. We conclude that BACs in adult female below 100 mg/dl (levels easily achieved by social drinkers) result in activation of the hypothalamus‐pituitaty‐adrenal axis. At high BACs (>350 mg/dl), pituitary adrenal responses are accompanied by increase in heart rate, blood pressure, and Pao2, and decreases in Pao2, and arterial pH. These findings support the hypothesis that alcohol acts directly on the brain to mediate pituitary adrenal responses and that the additional responses to high BACs (the blood gas and hemodynamic responses), might be mediated by direct actions of alcohol on the brain, by cerebral Ischemia, or by alcohol‐mediated suppression of vemtilatory drive and hypoxemia.
Effect of the Cytochrome P‐1450IIE1 Genotype on Ethanol Elimination Rate in Alcoholics and Control Subjects
Authors: Ueno Yasuhiro , Adachi Junko , Imamichi Hiroyuki , Nishimura Akiyoshi , Tatsuno Yoshitsugu
Published: April 2006

We studied an influence of genetic polymorphisms in the cytochrome P‐450IIE1 (CYP2E1) gene on ethanol elimination rate in alcoholic patients and healthy subjects. The CYP2E1 genotype was determined by polymerase chain reaction‐restriction fragment length polymorphism method for 124 alcoholics and 54 healthy subjects. There was no significant difference in the gene frequency of CYP2E1 between alcoholics and healthy control subjects. Blood ethanol concentrations in the 65 alcoholics on admission ranged from 0.32 to 4.22 mg/ml. In the patients with the c1/c2 genotype, the elimination rate was significantly correlated with blood ethanol concentration. In each of the three genotypes of CYP2E1, the patients were divided into three groups based on ethanol concentrations. The average of the ethanol elimination rate in the patients with c1/c2 having blood ethanol levels of ≧2.5 mg/ml was significantly higher than the rates in the two other groups of c1/c2. When blood ethanol levels were ≧2.5 mg/ml, the elimination rate in the patients with c1/c2 was significantly higher than that in those with c1/cl. Regardless of the CYP2E1 genotype, the elimination rate in the alcoholics was higher than that in the control subjects when blood ethanol levels were < 1.0 mg/ml. These results suggest the possibility that the c2 allele of CYP2E1 influences the rate of ethanol elimination at high ethanol levels. The rate of ethanol elimination was independent of liver disorder judged by serum total bilirubin values.
Association of Restriction Fragment‐Length Polymorphisms in the Alcohol Dehydrogenase 2 Gene with Alcoholic Brain Atrophy
Authors: Maezawa Yoshihiko , Yamauchi Masayoshi , Searashi Yasuyuki , Takeda Kunihiko , Mizuhara Yuuji , Kimura Taketo , Toda Gotaro , Suzuki Hitoshi , Sakurai Susumu
Published: April 2006

Alcohol abuse can induce brain atrophy, but it only occurs in some alcoholics. To investigate whether genetic polymorphism of alcoholmetabolizing enzymes [including alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH)] was related to alcoholic brain atrophy, we determined restriction fragment‐length polymorphisms of the ADH2 and ALDH2 genes in 77 male alcoholics. Computed tomography was used to determine the severity of brain atrophy. Digestion with Mae III and Mbo II after polymerase chain reaction amplification showed that the ADH21 gene frequency was significantly higher in patients with brain atrophy than in those without brain atrophy (x2= 9.274, p < 0.01), whereas no significant association was observed between brain atrophy and the ALDH2 gene. Multivariate analysis (including age, total alcohol intake, liver cirrhosis, and ADH2 genotype) showed that the ADH21/ADH21 genotype was associated with alcoholic brain atrophy. These findings suggest that the ADH21 allele may be associated with alcoholic brain atrophy.
Changes in GABAA Receptor Function and Cross‐Tolerance to Ethanol in Diazepam‐Dependent Rats
Authors: Toki Sadamu , Saito Toshikazu , Nabeshima Akemi , Hatta Shinichi , Watanabe Masayuki , Takahata Naohiko
Published: April 2006

Changes in γ‐aminobutyric acidA (GABAA) receptor function and their relation to cross‐tolerance to ethanol (EtOH) were studied in diazepam (DZP)‐dependent rats. Physical dependence on DZP was induced in male Fischer rats by using the drug‐admixed food method. The 36CI– influx into cerebral cortical synaptoneurosomes induced by 10 μM GABA in DZP‐withdrawn rats was significantly increased, compared with control and DZP‐tolerant rats. Although enhancement of GABA‐dependent 36CI– influx by the addition of EtOH and flunitrazepam (FZ) was recognized in the control, there was no such effect of EtOH or FZ in the DZP‐tolerant animals. On the other hand, GABA‐dependent 36CI– influx was enhanced by FZ in the withdrawn group. The addition of picrotoxin and bicuculline inhibited GABA‐dependent 36CI– influx in each group. The stimulatory effect of FZ on GABA‐dependent 36CI– influx was inhibited by the addition of Ro 15–1788 in the control group. However, such an inhibitory effect was not observed in the withdrawn group. The antagonistic effect of Ro 15–4513 on EtOH stimulation of GABA‐dependent 36CI– influx observed in the control was not recognized in the withdrawn group. In a [3H]FZ assay of binding to benzodiazepine (BZ) receptors, Bmax values were significantly increased in DZP‐withdrawn animals, but decreased in the DZP‐tolerant group, compared with the control. When [3H]muscimol binding was examined, the Kd of high‐affinity sites of the GABAA receptor in withdrawn rats was significantly lower than in the control. In low‐affinity binding sites, the values of Kd and Bmax were significantly decreased, compared with those in the control. The present study indicates that GABAergic transmission involving the regulation of GABA‐dependent chloride channels is altered in DZP‐dependent rats. Alterations of the GABAA/BZ/chloride channel complex function may be related to the cross‐tolerance between BZ and EtOH.
Evaluation of Ethanol on Gastric Epithelial Restoration in Vitro
Authors: Murai Toshio , Watanabe Sumio , Hirose Miyoko , Kobayashi Osamu , Maehiro Kouhei , Ohkura Ryuichi , Miwa Hiroto , Kitamura Tsuneo , Ogihara Tatsuo , Oide Hirosumi , Miyazaki Akihisa , Sato Nobuhiro
Published: April 2006

Ethanol exerts damaging effects on gastric mucosa and delays ulcer healing. To investigate the effect of ethanol on the wound repairing process, we used a wound repair model using primary cultured gastric mucosal cells. A confluent monolayer gastric mucosal cell sheet consisting mainly of mucous cells was wounded to make a cell‐free area of constant size. Cell‐free area was restored with time after wounding and monitored every 12 hr using a computer image analyzer to observe epithelial cell restoration quantitatively in the presence and absence of ethanol (2.0%). It was found that, although the control wound was completely repaired in 36 to 48 hr, the group treated with 2.0% ethanol showed a significant delay of repair. In the control, 5‐bromodeoxyuridine‐positive cells appeared around the wound in 24 to 36 hr. In contrast, the group treated with 2.0% ethanol showed no 5‐bromodeoxyuridine‐positive cells during the experiment. In conclusion, 2.0% ethanol retarded the repair of gastric mucosal restoration by inhibiting the initial gastric cell migration, followed by inhibition of proliferation of cells.
Effects of Long‐Term Alcohol Intake on ADP Ribosylation in Rat Liver Plasma Membranes
Authors: Nomura Fumio , Noda Masatoshi , Nakai Toshiaki
Published: April 2006

Mono‐ADP‐ribosylation, in which the ADP‐ribosyl moiety is transferred from NAD to an acceptor protein, is one of the important posttranslational modifications of cellular proteins. Because mounting evidence suggests significant biological roles of this reaction in transmembrane signal transduction and other cell metabolic reactions, we assessed how long‐term alcohol intake alters toxin catalyzed‐ and endogenous mono‐ADP‐ribosylation in the liver of a rat model. We first found that thiol‐preactivated cholera toxin‐catalyzed ADP‐ribosylation of the α‐subunit of the stimulatory GTP‐binding protein was enhanced after long‐term alcohol intake. Unexpectedly, but interestingly, this enhancement was not accompanied by a concomitant increase of cholera toxin‐catalyzed stimulation of the adenylate cyclase activity. We also found that long‐term alcohol intake remarkably enhanced endogenous mono‐ADP‐ribosylation of a 58 kDa protein in plasma membranes. Thus, long‐term alcohol intake stimulated endogenous, as well as, toxin‐catalyzed mono‐ADP‐ribo‐sylations. Characterization of the 58 kDa protein may uncover pathophysiological roles of this interesting phenomenon in alcohol‐induced liver damage.
Effect of Chronic Ethanol Feeding on Kupffer Cell‐Mediated Antitumor Cell Activity
Authors: Kato Shinzo , Kurose Iwao , Higuchi Hajime , Ebinuma Hirotoshi , Saito Hidetsugu , Miura Soichiro , Ishii Hiromasa
Published: April 2006

We have previously reported that the Kupffer cell has antitumor activity through mitochondrial damage to tumor cells by nitric oxide production. In this study, the effect of chronic ethanol feeding on antihepatoma cell activity of the Kupffer cell was examined in rats. Male rats of the Wistar strain were fed ethanol chronically for 8 weeks by liquid diets. Kupffer cells were isolated from the control rat or the ethanol‐fed rat, and cocultured with AH 70 cells, a rat hepatoma cell line. Fluorescence of rhodamine 123 or propidium iodide was observed as indicators of the mitochondrial damage or cell membrane injury, respectively, by a laser scanning confocal microscopy. Mitochondrial damage of AH 70 cells as indicated by reduction of rhodamine 123 fluorescence was smaller by the coculture with Kupffer cell from the ethanol rat than that from the control. Cell membrane barrier dysfunction of AH 70 cell was less frequently observed with the Kupffer cell from ethanol‐fed rats. A metabolite of nitric oxide (nitrite and nitrate) was less in the cultured medium with the ethanol Kupffer cell than with the control Kupffer cell. Ca2+mobilization, which induces inducible nitric oxide synthase and observed by the fluorescence of fluo‐3, in Kupffer cells cocultured with AH 70 cells was suppressed in ethanol‐fed rats. These result suggests that chronic ethanol feeding suppresses antitumor cell activity of Kupffer cell through the impairment of Ca2+mobilization and nitric oxide production.
Effect of Chronic Ethanol Feeding on Nitric Oxide Synthesis by Rat Kupffer Cells
Authors: Kimura Hiroyuki , Miura Soichiro , Higuchi Hajime , Kurose Iwao , Tsuzuki Yoshikazu , Shigematsu Takeharu , Ebinuma Hirotoshi , Kato Shinzou , Ishii Hiromasa
Published: April 2006

Kupffer cells contribute to the important role of the liver defense mechanism through nitric oxide (NO) production. In this study, the effect of chronic ethanol administration on the ability of Kupffer cells to synthesize and release NO was investigated after stimulation with lipopolysaccharide (LPS). Male Wistar rats were chronically fed ethanol for 8 weeks according to the method described by DeCarli and Lieber et al. (J Nutr. 91:331–336, 1967). Kupffer cells were isolated and cultured with LPS (1 μg/ml) for 24 hr. The levels of nitrite and nitrate, metabolites of NO, were determined in the culture medium. NO synthase (NOS) activity in Kupffer cells was determined by the method that measures conversion of [14C]arginine into [14C]citrul‐line. In control rats, a significant increase of nitrite and nitrate levels in culture medium was observed after LPS treatment. The magnitude of this increase was significantly smaller in chronic ethanol‐fed rats. When the activity of NOS was determined, inducible NOS (iNOS) activity was higher than that of constitutive NOS, and LPS administration produced a significant elevation of iNOS activity in both control and chronic ethanol‐fed rats. However, the elevation of iNOS activity by LPS stimulation was diminished by chronic ethanol administration. Distribution of iNOS in Kupffer cells as determined by an immunofluorescence method using a laser scanning confocal image system showed a lower expression of iNOS in chronic ethanol‐fed rats even in the presence of LPS. These results demonstrate that the excessive production of NO by increased iNOS activity in Kupffer cells is diminished by chronic ethanol administration.
Ethanol‐Induced Oxidative Stress in the Liver
Authors: Kurose Iwao , Higuchi Hajime , Kato Shinzo , Miura Soiohiro , Ishii Hiromasa
Published: April 2006

Oxygen stress is well recognized to be a key step in the pathogenesis of ethanol‐associated liver injury. Ethanol administration induces an increase in lipid peroxidation either by enhancing the production of oxygen‐reactive species and/or by decreasing the level of endogenous antioxidants. Numerous experimental studies have emphasized the role of the ethanol‐inducible cytochrome P‐450 in the microsomes, as well as the molybdo‐flavoenzymes xanthine oxidase in the cytosol. This review shows the putative role of ethanol‐induced disturbances in iron metabolism in relation to iron as a prooxidant factor. Ethanol administration also affects the mitochondrial free radical generation. Although many previous studies suggest a role for active oxygens in ethanol‐induced mitochondrial dysfunction in hepatocytes, the detailed mechanism of ethanol‐induced oxidative stress on mitochondria remains to be clarified further. Studies of our laboratory using a confocal laser scanning microscopic system strongly suggest that active oxidants produced during ethanol metabolism modulate mitochondrial energy synthesis in isolated and cultured hepatocytes. In addition, our investigations implicate endogenous glutathione‐glutathione peroxidase system and catalase as important antioxidants and cytoprotective machinery in the hepatocyte mitochondria exposed to ethanol. The fluorographic investigations using the confocal laser scanning microscopy may be useful to extend our knowledge and provide a new view about ethanol‐associated oxidative stress and metabolic changes in hepatocytes.
Progression of Type C Chronic Hepatitis to Liver Cirrhosis and Hepatocellular Carcinoma—Its Relationship to Alcohol Drinking and the Age of Transfusion
Authors: Noda Katsuhisa , Yoshihara Harumasa , Suzuki Kunio , Yamada Yukinori , Kasahara Akinori , Hayashi Norio , Fusamoto Hideyuki , Kamada Takenobu
Published: April 2006

Alcohol drinking has been reported to be an important factor that modulates the development and prognosis of chronic hepatitis B; however, little is known about an interrelationship between alcohol intake and the progression of chronic hepatitis C to liver cirrhosis (LC) and hepatocellular carcinoma (HCC). We attempted to clarify this interrelationship in patients with hepatitis C and history of blood transfusion. Thirty LC and 85 HCC patients were enrolled. In patients with LC, no significant correlation was observed between the amount of alcohol intake and the period from transfusion to diagnosis. The period from transfusion to diagnosis in HCC patients with alcohol intake ≥46 g/day and < 46 g/day were 26 ± 6 and 31 ± 9 years, respectively, resulting in a significant difference (p < 0.05). The period from transfusion to diagnosis of LC and/or HCC showed significant negative correlation with the age of transfusion (r= 0.82, Y = ‐0.67X+ 48.0, p < 0.01; r= 0.76, Y= ‐0.70X+ 54.1, p < 0.001, respectively). This correlation was also observed in patients with HCC, regardless of the amount of alcohol intake. In conclusion, these data suggest that alcohol drinking might be an important factor that promotes an occurrence of HCC in patients with hepatitis C, and that hepatitis C virus infection in the elderly promotes development of liver disease via LC to HCC.
Noncompetitive‐Like Inhibition of Ethanol Elimination by Cyanamide Treatment: Pharmacokinetic Study
Authors: Fujimiya Tatsuya , Li Yu‐Jiang , Uemura Koichi , Ohbora Yumiko , Komura Setsuo
Published: April 2006

The effect of acetaldehyde accumulation of ethanol elimination is of interest in medico‐legal practice in Japan. We examined the pharmacokinetic mechanism of the inhibition of ethanol metabolism by cyanamide, an inhibitor of mitochondrial aldehyde dehydrogenase. An ethanol solution (0.25‐2.0 g/kg body weight) was injected intravenously into male rabbits with or without administration of cyanamide. Cyanamide was injected intraperitoneally (25 mg/kg body weight) to the cyanamide‐treated group 2 hr before ethanol injection. Blood ethanol and acetaldehyde concentrations were measured periodically by head‐space gas chromatography. The MULTI(RUNGE) computer program was applied for the pharmacokinetic analysis. One‐ or two‐compartment open models with Michaelis‐Menten elimination kinetics were used for simultaneous multi‐line fitting. The ethanol elimination rate decreased by cyanamide treatment. The border‐point concentration between pseudolinear and curvilinear phases was not affected by cyanamide treatment. The estimated Vmax value decreased by cyanamide treatment, whereas the Km value did not change. Our results correspond to a noncompetitive‐like inhibition of ethanol metabolism. Km is related to the border point between pseudolinear and curvilinear phases. Thus, our findings in the blood ethanol concentration‐time curve suggest adequate curve‐fitting. The product, or competitive, inhibition of alcohol dehydrogenase by acetaldehyde had been reported in enzymological study. The pharmacokinetic manner of inhibition in vivo was different from the enzymologic mechanism in vitro. Other metabolic factors related to ethanol metabolism are thought to be more important than acetaldehyde accumulation itself.
Genotypes of Alcohol‐Metabolizing Enzymes and the Risk for Alcoholic Chronic Pancreatitis in Japanese Alcoholics
Authors: Matsumoto Michinaga , Takahashi Hisao , Maruyama Katsuya , Higuchi Susumu , Matsushita Sachio , Muramatsu Taro , Okuyama Keiji , Yokoyama Akira , Nakano Masayuki , Ishii Hiromasa
Published: April 2006

Genetic predisposition to alcoholism and alcoholic liver disease has been reported. However, genetic susceptibility to alcoholic pancreatitis is still a matter of debate. To determine it, we examined genotype patterns of aldehyde dehydrogenase (ALDH2), alcohol dehydrogenase (ADH2 and ADH3), and cytochrome P‐4502E1 (CYP2E1) in alcoholic pancreatitis patients. In 296 alcoholic patients, 52 cases showed findings of chronic pancreatitis by ultrasonography and x‐ray computed tomography and/or had a history of pancreatitis (P+). The remaining 244 patients had neither abnormal findings of the image examinations nor a history of pancreatitis (P‐). As for the ADH2 genotype, distribution of 21/21, 21/22, and 22/22 was 22,37, and 42% in P+ patients, whereas 34,35, and 30% in P‐ patients, respectively. The frequency of ADH22/22 genotype was significantly higher in P+ patients, compared with that in P patients. There were no significant differences in the distribution of ADH3, ALDH2, and CYP2E1 genotypes between P+ and P‐ patients. In 14 alcoholic patients who showed low contents of fecal chymotrypsin, which suggests dysfunction of pancreatic exocrine, the rate of ADH22/22 genotype also tended to be higher (50%) than in 74 controls who showed normal contents of the fecal chymotrypsin (28%). No differences were observed in genotypes of ADH3, ALDH2, and CYP2E1. Moreover, the frequency of ADH22/22 genotype was significantly higher in autopsy cases with interlobular fibrosis in the pancreas, which suggests alcoholic pancreatic damage, than in cases with only intralobular pancreatic fibrosis. These data suggest that the risk of alcoholic pancreatitis seems to be associated with the presence of ADH22/22 genotype.
Investigation of Genetic Risk Factors Associated with Alcoholism
Authors: Harada Shoji , Okubo Takehito , Tsutsumi Mikihiro , Takase Shujiro , Muramatsu Taro
Published: April 2006

Alcoholism is a multifactorial disease influenced by genetic‐environmental interaction. Genetic variation of the receptor may be associated with alcohol dependence due to its modified function in behavioral and physiological responses. In the present study, polymorphic alleles of cholecystokinin B receptor (CCKBR), serotonin 1A receptor (HT1AR) genes, and mitochondrial DNA were analyzed. DNAs were isolated from the blood samples of 112 healthy controls and 106 alcoholics. Genetic variation was detected by SSCP analysis, followed by direct sequencing of polymerase chain reaction product as well as restriction fragment‐length polymorphism. Three different mutations were found in the exon 3 sequence of CCKBR: His (CAT) at aa207 → His (CAC) (5.4%), Arg (CGC) at aa215 → His (CAC) (4.5%), and Val (GTG) at aa138 → Met (ATG) (0.9%) in controls. Genotypic distribution of alcoholics was not significantly different with that in controls. A proline (CCG) to leucine (CTG) substitution at amino acid 16 of HT1AR was found in alcoholics (4.5%) and in controls (4.7%). This mutation site of HT1AR was different in comparison with the variants reported by Nakhai et al. (Biochem Biophys. Res.Commun. 210:530 ‐536,1995). Analysis of the mitochondrial DNA showed that a 491 bp deletion in the sequence of ATPase exists as heteroplasmy in 58% of alcoholics, but not in controls. Heteroplasmic deletion of mitochondrial DNA may be a useful marker for alcohol abuse. Further study is undergoing to elucidate the cause and significance of this deletion in alcoholics.
Effects of Ethyl Alcohol Administration to THA Rat Dams During Their Gestation Period on Learning Behavior and on Levels of Monoamines and Metabolites in the Brain of Pups after Birth
Authors: Furuya Hiroyuki , Aikawa Hiroyuki , Yoshida Takahiko , Okazaki Isao
Published: April 2006

Children born from chronic alcoholic mothers have shown behavioral teratogenic effects more frequently than morphological malformations. To investigate the possible mechanisms and evaluate maternal alcohol dosage levels to induce behavioral dysfunctions, we gave pregnant Tokai High Avoider (THA) rats 0, 5,10, and 20% ethanol (EtOH) as drinking water during the gestation period. We evaluated the brain function of pups born of alcohol‐administered dams. Sidman avoidance behavior test and the levels of monoamines (nor‐adrenalin, dopamine, and serotonin) and metabolites (3,4‐dihydroxy‐phenyl acetic acid, homovanillic acid, and 5‐hydroxyindole acetic acid) in whole brain were examined for neurobehavioral and neurochemical effects. EtOH‐exposed THA offsprings showed high pre‐and postnatal mortality, growth deficits, and brain weight reductions. Compared with the results of the same conditioning experiment using Wistar rats, the THA rat may have higher susceptibility to the effects of in utero EtOH exposure than Wistar rats. EtOH‐exposed THA pups exhibited deficits in avoidance operant learning that were not shown in Wistar rats. We also observed the increased levels of all monoamines that were assumed to be related with the deficit of learning, the decreased levels of 3,4‐dihydroxyphenyl acetic acid and homovanillic acid, and the unchanged levels of 5‐hydroxyindole acetic acid in pups from dams administered 10 and 20% EtOH. However, contrary alteration of monoamines and their metabolites were shown in pups from 5% EtOH‐administered dams.
Interaction of Protein Phosphatases and Ethanol on Phospholipase C‐Mediated Intracellular Signal Transduction Processes in Rat Hepatocytes: Role of Protein Kinase A
Authors: Higashi Katsuyoshi , Hoshino Makoto , Nomura Tomoyuki , Saso Katsuhisa , Ito Makoto , Hoek Jan B.
Published: April 2006

Phospholipase C (PLC)‐mediated signal transduction processes in rat hepatocytes are subject to modulation by protein phosphatases (PPases) and protein kinases, including protein kinase A (PKA) and protein kinase C. Ethanol (EtOH) stimulates PLC activity in liver cells in the absence of hormones, and EtOH pretreatment inhibits the subsequent stimulation of PLC by hormonal stimuli. There is evidence that protein kinase activities are involved in these actions of EtOH. We investigated the effects of okadaic acid (OKA), a PPase inhibitor, and 8‐(4‐chlorophenylthio)adenosine 3′:5′‐cyclic monophosphate (cpt‐cAMP), a cell permeant cAMP analog that activates PKA, on EtOH‐induced PLC activation. In addition, we studied the combined effects of cpt‐cAMP and EtOH/OKA on vasopressin‐induced PLC activation. PLC activation (cytosolic Ca2+ mobilization and inositol trisphosphate accumulation) induced by EtOH and vasopressin was inhibited by treatment with OKA, and was potentiated by cpt‐cAMP. OKA treatment prevented the effect of cpt‐cAMP. Pretreatment with EtOH caused inhibition of vasopressin‐induced PLC activation. EtOH also decreased the enhancing effect of cpt‐cAMP on the responses to vasopressin. The susceptibility to enhancement by cpt‐cAMP plotted as a function of the initial rate of vasopressin‐induced Ca2+ mobilization in EtOH‐treated cells was similar to the pattern observed in OKA‐treated cells. These data suggest that interactions of OKA and PKA on EtOH‐induced PLC activation occurred at the level of G‐protein, and indicate that EtOH may act as an inhibitory agent of PPase.
Inhibitory Effect of Ethanol on Hepatocyte Growth Factor‐Induced DNA Synthesis and Ca2+ Mobilization in Rat Hepatocytes
Authors: Saso Katsuhisa , Higashi Katsuyoshi , Nomura Tomoyuki , Hoshino Makoto , Ito Makoto , Moehren Gisela , Hoek Jan B.
Published: April 2006

Hepatocyte growth factor (HGF) is the most potent mitogen identified for hepatocytes and is thought to be an important growth factor in the regulation of liver regeneration. Its effects are mediated through a tyrosine kinase receptor, the product of c‐met proto‐on‐cogene. One of the downstream signaling processes activated by HGF is phospholipase C‐γ. HGF stimulation of liver cells causes formation of inositol 1,4,5‐triphosphate, which releases Ca2+ from intracellular Ca2+ ([Ca2+]i) stores, and causes elevation of cytosolic Ca2+ levels. It is known that liver regeneration is inhibited by both acute and chronic ethanol (EtOH) treatment. We investigated the effect of EtOH on HGF‐induced DNA synthesis and mobilization of [Ca24], in rat hepatocytes in primary culture. DNA synthesis was monitored by [3H]thymidine incorporation in primary cultures of hepatocytes 42 hr after stimulation with HGF. HGF concentration required for maximum DNA synthesis was 0.3 to 1 ng/ml, and DNA synthesis was inhibited by 100 mM EtOH at HGF concentrations in the range of 0.1 to 5 ng/ml. This inhibition was strongest (45 to 47% inhibition) at a low concentration of HGF (0.1 to 0.3 ng/ml) and decreased at an HGF concentration >1 ng/ml. HGF‐induced changes in [Ca2+], were measured in single fura 2‐loaded hepatocytes by fluorescence imaging techniques. The Ca2+ response induced by HGF (0.3 to 5 ng/ml) was inhibited by EtOH, with an EC50, of ‐50 mM. Analysis of Ca2+ response patterns in individual cells indicated that EtOH suppressed the number of responsive cells and made Ca2+ responses more transient, but did not affect peak [Ca2+]j elevation; thus suggesting an inhibition at the level of phospholipase C‐y‐activation. These data indicate that inhibition by EtOH of the response of liver cells to HGF may contribute to the inhibitory effect of EtOH on liver regeneration.
Microheterogeneity with Concanavalin A Affinity of Serum Transferrin in Patients with Alcoholic Liver Disease
Authors: Inoue Takahiro , Yamauchi Masayoshi , Toda Gotaro , Ohkawa Kiyoshi
Published: April 2006

Microheterogeneity of transferrin (Tf) with concanavalin A (ConA) affinity was investigated by sensitive lectin‐affinity electrophoresis and antibody‐affinity blotting technique of sera obtained from patients with alcoholic liver disease (ALD) and normal subjects. Serum Tf was separated by ConA into three bands–a strongly ConA‐reac‐tive major band (C1), a weakly reactive minor band (C2), and a non‐reactive trace band (C3). The C3 fraction was significantly increased in patients with ALD before alcohol abstinence, compared with normal subjects and patients with ALD after 4 weeks of abstinence. Furthermore, a significant correlation was found between the C3 fraction and serum carbohydrate‐deficient transferrin or γ‐glutamyl‐transpeptidase. These results indicate that the microheterogeneity of serum Tf in patients with ALD may be a more complex abnormality of elongation and processing on the glycans than merely a loss of terminal sialic acids. Determination of the C3 fraction is a useful marker for ALD.
Correlation Between Intrasinusoidal Neutrophilic Infiltration and Ceroid‐Lipofuscinosis in Alcoholic Liver Fibrosis With or Without Fatty Change: Clinicopathological Comparison with Nutritional Fatty Liver
Authors: Kishi Masaya , Maeyama Shiro , Koike Junki , Aida Yoshio , Yoshida Hideki , Uchikoshi Toshiyuki
Published: April 2006

A histopathological study was conducted on alcoholic liver fibrosis with fatty change (21 cases) and alcoholic liver fibrosis without fatty change (18 cases) in comparison with nutritional fatty liver (27 cases). The diagnoses of alcoholic liver fibrosis groups were clinically fulfilled according to the criteria established by the Alcohol and Liver Research Group (Chief: Professor Takeuchi) of the Ministry of Education of Japan. Histological diagnosis of alcoholic liver fibrosis with fatty change was based on moderate and/or greater fatty metamorphosis of the hepatic lobules, alcoholic liver fibrosis without fatty change on a lesser degree of fatty metamorphosis than alcoholic liver fibrosis with fatty change, and nutritional fatty liver on clinicopathological features. All 66 cases were negative for viral markers of hepatitis B surface antigen and anti‐hepatitis C virus in serum. Intrasinusoidal neutrophil infiltrations were significant in cases of alcoholic liver fibrosis groups more often than in cases of nutritional fatty liver. The degree of intrasinusoidal neutrophil infiltration in cases of alcoholic liver fibrosis groups was higher in cases who had last consumed alcohol recently, compared with those with longer abstinence. In alcoholic liver fibrosis with fatty change and nutritional fatty liver groups, mild‐to‐moderate degrees of ceroid‐lipofuscinosis were recognized, but both fatty change and ceroid‐lipofuscinosis were decreased according to the deterioration of fibrotic changes in alcoholic liver fibrosis with fatty change cases. On the other hand, it is significant that the frequency of ceroid‐lipofuscinosis in alcoholic liver fibrosis without the fatty change group was lower than those of the alcoholic liver fibrosis with fatty change and nutritional fatty liver groups. Distribution of ceroid‐lipofuscinosis has a tendency to be recognized around the central zone (zone III) of alcoholic liver fibrosis with fatty change cases with mild fibrosis, as in nutritional fatty liver cases, and the ceroid‐lipofuscinosis disperses with the progression of fibrosis. These results suggest that fibrosis and fatty droplet deposition lead to microvascular heterogeneity. Therefore, the degree and distribution of fatty droplets, ceroid‐lipofuscinosis, and intrasinusoidal neutrophil infiltration differ, depending on the etiology of fatty liver, and are an important histopathological barometer in cases of alcoholic liver fibrosis with fatty change and alcoholic liver fibrosis without fatty change, thus indicating the degree of fibrosis and the period since last alcohol intake.
Hepatocellular Carcinoma Associated with AlcoholicLiver Disease: A Clinicopathological Study and Genetic Polymorphism of Aldehyde Dehydrogenase 2
Authors: Ohhira Motoyuki , Fujimoto Yoshinori , Matsumoto Akinori , Ohtake Takaaki , Ono Minoru , Kohgo Yutaka
Published: April 2006

In this paper, we describe a clinicopathological study of primary hepatocellular carcinoma (HCC) associated with alcoholic liver disease without hepatitis virus infection. In 180 HCC patients who were admitted to Asahikawa Medical College Hospital from 1987 to 1995, 10 patients (6%) had HCC associated with pure alcoholic liver disease (AI‐HCC), whereas the HCC in 165 patients was associated with chronic viral liver diseases, in 2 with primary biliary cirrhosis, in 1 each with coexistence of the hepatitis C virus infection and hemochromatosis, and in 2 with cirrhosis of unknown origin. In the AI‐HCC group, all patients were male. The diagnosis of HCC was obtained at the age of 54 to 67 years old, and the duration of ethanol intake was 33 to 40 years. Four cases had a history of temperance. As an underlying liver disease, liver fibrosis was found in three cases and liver cirrhosis in seven cases. HCC was diagnosed histologically in all cases. Serum α‐fetoprotein and PIVKA‐II were positive in patients with advanced HCC. In cases with small HCC, the tumor was resected surgically in three cases and percutaneous ethanol injection was performed in two cases. In four cases with small HCC, the patients were alive without tumor recurrence during the observation period. In advanced HCC, transcatheter arterial chemolipiodolization was performed. In the analysis of genetic polymorphism of ALDH 2, all AI‐HCC had ALDH 21/21.
Two Survival Cases of Alcoholic Lactic Acidosis Complicated with Diabetes Mellitus and Alcoholic Liver Disease
Authors: Ishii Kunihide , Kumashiro Ryukichi , Koga Yuriko , Tanikawa Kyuichi , Furudera Shigeki , Mitsuoka Masazumi , Sakamoto Teruo , Kaku Nobuo
Published: April 2006

We have experienced two patients with alcoholic lactic acidosis complicated with liver disease and diabetes mellitus who were successfully treated. They developed hypoglycemia, dehydration, lactic acidosis, and renal failure after drinking a large volume of alcohol without eating for 1 week before onset. Acidosis was thought to be directly related to excessive alcoholic intake, because it was not associated with severe liver failure and rhabdomyolysis. During monitoring of respiratory and circulatory functions, a rapid infusion of fluids adjusting to water and electrolyte imbalance was performed. A mixture of physiological saline and 5% glucose solution was thought to be effective in these cases. Patients recovered from renal failure and lactic acidosis without hemodialysis. Our experience will hopefully provide a key to successful treatment of fatal alcoholic lactic acidosis.
In Vitro Alcohol Dehydrogenase‐Mediated Acetaldehyde Production by Aerobic Bacteria Representing the Normal Colonic Flora in Man
Authors: Jokelainen K. , Siitonen A. , Jousimies‐Somer H. , Nosova T. , Heine R. , Salaspuro M.
Published: April 2006

Excessive ethanol consumption has been related with the development of liver cirrhosis, as well as with rapid intestinal transit time and diarrhea. Moreover, heavy drinking is associated with an increased incidence of cancer of the oropharynx, larynx, esophagus, and colorectum. Acetaldehyde of microbial origin has recently been suggested as a possible pathogenetic factor behind this alcohol‐associated gastrointestinal morbidity. The present in vitro study was aimed to investigate alcohol dehydrogenase activity and acetaldehyde formation capacity of some major aerobic bacteria representing the normal colonic flora in man. Cytosolic alcohol dehydrogenase activity and cytosolic protein concentration were determined spectrophotometrically. Alcohol dehydrogenase activity was then calculated as nmoles of reduced substrate produced by milligrams of protein per minute. The ability of different bacteria to produce acetaldehyde was determined by incubating the intact bacterial suspension in closed vials containing ethanol (final concentration 22 mM)for 1 hr at 37°C. The acetaldehyde formed during the incubation was analyzed by headspace gas chromatography. Marked differences in the alcohol dehydrogenase activity and acetaldehyde forming capacity were found among the strains tested. The alcohol dehydrogenase activity varied from 606 ± 91 nmol/min/mg protein (Escherichia coli IH 50546) to 1 ± 0.2 nmol/min/mg protein (E. coli IH 50817), and acetaldehyde formation varied from 1,717 ± 2 nmol acetaldehyde/109 colony‐forming units (Klebsiella oxytoca IH 35403) to 5 ± 2 nmol acetaldehyde/109 colony‐forming units (Pseudomonas aeruginosa ATCC 27853). There was a statistically significant correlation (r= 0.77; p < 0.001) between alcohol dehydrogenase activity and acetaldehyde production from ethanol, strongly suggesting the catalytic role of bacterial alcohol dehydrogenase in this reaction.
Long‐Term Ethanol and Nicotine Treatment Elicit Tolerance to Ethanol
Authors: Collins Allan C. , Wilkins Lincoln H. , Slobe Brian S. , Cao Jian‐Zhe , Bullock Amy E.
Published: April 2006

Several previous studies have shown that 1 to 2 weeks of treatment with ethanol elicits tolerance to several effects produced by ethanol and cross‐tolerance to nicotine‐induced hypothermia. Similarly, short‐term, high‐dose nicotine treatment produces tolerance to nicotine and cross‐tolerance to ethanol‐induced hypothermia. In the studies reported here, C57BL/6 mice were force‐fed ethanol, nicotine, or an ethanol/nicotine combination in the drinking water for 6 months. All of the chronic drug‐treated mice developed tolerance to ethanol as measured by open‐field activity, body temperature, and sleep‐time tests. Ethanol tolerance is due, in part, to enhanced metabolism and reduced CNS sensitivity in the two ethanol‐treated groups but only to reduced CNS sensitivity in the nicotine‐treated group. Similar levels of tolerance to nicotine developed in those two groups that were nicotine‐treated, but no tolerance to nicotine was seen in those animals treated with ethanol alone. The tolerance to nicotine may be related to an upregulation of brain (cortex, hippocampus, and hypothalamus) [3H]‐nicotine binding, but ethanol tolerance is not readily explained by changes in the number of the brain high affinity nicotine binding sites.
Role of Catalase in Rat Gastric Mucosal Ethanol Metabolism In Vitro
Authors: Salmela Katja S. , Kaihovaara Pertti , Salaspuro Mikko , Roine Risto P.
Published: April 2006

To evaluate the possible role of catalase in gastric ethanol metabolism in rats, we studied acetaldehyde formation from ethanol by gastric mucosal homogenate under various in vitro conditions. Homogenized rat gastric mucosa produced significant amounts of acetaldehyde in a time and ethanol concentration‐dependent manner, even in the absence of added NAD. Both acetaldehyde formation and catalase activity peaked around the physiological pH, whereas alcohol dehydrogenase (ADH) activity was in that pH range low and reached peak values only at a higher pH of 9 to 10. Catalase inhibitors sodium azide (SA) and 3‐amino‐1,2,4‐triazole (3‐AT) had little effect on ADH activity but markedly decreased catalase activity and acetaldehyde formation (1 mM of SA to 56 ± 13% of control, 5 mM of 3‐AT to 67 ± 3% of control; mean ± SE). 4‐Methylpyrazole decreased ADH activity significantly, but did not affect acetaldehyde formation. Heating of the homogenate at 60°C for 5 min decreased ADH activity only slightly, but totally abolished catalase activity and reduced acetaldehyde formation to 39 ± 3% of control. Addition of a H2O2 generating system [β‐d(+)‐glucose + glucose oxidase] increased acetaldehyde formation in a concentration‐dependent manner up to 8‐fold of the control value. Our results strongly suggest that, in addition to ADH, catalase may play a significant role in gastric ethanol metabolism in rats.
Mitochondrial Dysfunction after Fetal Alcohol Exposure
Authors: Marin‐Garcia Jose , Ananthakrishnan Radha , Goldenthal Michael J.
Published: April 2006

Specific mitochondrial enzyme activities and mRNA levels were measured in the heart, brain, and liver tissues of a group of 1‐day‐old neonatal rats whose mothers were alcohol‐fed during pregnancy and compared with a control group. The results show a significant decrease in mitochondrial ATP synthase activity in both the brain and liver, as well as a decrease in complex III activity in the liver of rats exposed to alcohol. Other mitochondrial enzymes activities (e.g., citrate synthase, cytochrome c oxidase, and complex I), as well as specific mitochondrial transcript levels, were not significantly affected. Heart mitochondrial enzyme activities were not significantly affected. These data reveal that a tissue‐specific response occurs after fetal exposure to alcohol and may explain some of the cellular events occurring in fetal alcohol syndrome resulting in abnormal growth and neurological development.
Serotonin2C Receptors and Serotonin2C Receptor‐Mediated Phosphoinositide Hydrolysis in the Brain of Alcohol‐Preferring and Alcohol‐Nonpreferring Rats
Authors: Pandey Subhash C. , Lumeng Lawrence , Li Ting‐Kai
Published: April 2006

To examine the role of serotonin2C (5HT2C) receptors in alcohol drinking behavior, the binding indices of 5HT2C receptors were determined in various brain regions of alcohol‐preferring (P) and alcohol‐nonpreferring (NP) rats. 5HT2C receptor‐mediated phosphoinositide hydrolysis in the choroid plexus of P and NP rats was also determined. It was observed that the densities of 5HT2C receptors are significantly higher in the hippocampus, the amygdala, and the choroid plexus, but not in the cortex of P rats compared with NP rats. The Kd values of [3H]mesulergine binding to 5HT2C receptors were not different in these brain regions of P rats compared with NP rats. It was also observed that 5HT‐stimulated [3H]inositol 1‐phosphate formation was significantly higher in the choroid plexus of P rats compared with NP rats. The results of this study indicate that the numbers of 5HT2C receptors are higher in the hippocampus, the amygdala, and the choroid plexus, and that 5HT2C receptor‐mediated phosphoinositide hydrolysis is more elevated in the choroid plexus of P rats compared with NP rats. Thus, it seems from these results that increased 5HT2C receptors may be involved in the genetic vulnerability to alcohol drinking behavior.