bluuebunny/crossref_metadata_2025 huggingface dataset:
Role of newly synthesized cholesterol or its metabolites on the regulation of bile acid biosynthesis after short-term biliary diversion in the rat
Authors: Vlahcevic Reno Z. , Pandak William M. , Hylemon Philip B. , Heuman Douglas M.
Published: March 2007

Cholesterol 7αhydroxylase, the rate-limiting enzyme in the bile acid biosynthetic pathway, is thought to be regulated by hydrophobic bile acids through negative feedback control. The role of cholesterol in the regulation of cholesterol 7αhydroxylase is more controversial, in part because of incomplete understanding of the relationship between the pathways of cholesterol synthesis and degradation. The main objective of this study was to define the interaction between these two pathways in an experimental model in which the supply of newly synthesized cholesterol was interrupted by sustained infusion of mevinolin (lovastatin), an inhibitor of 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMG-CoA reductase) or accelerated by a continuous infusion of mevalonate, a cholesterol precursor. The study was carried out in rats subjected to short-term bile fistula. In one set of experiments, rats were treated postoperatively with mevinolin (5 mg/kg loading dose followed by 2 mg/kg/hr infusion), mevalonate (180 μmol/hr infusion) or both for up to 96 hr. In a separate set of experiments, rats were infused intraduodenally with taurocholate (36 μmol/100 gm/hr for up to 96 hr). We determined cholesterol 7αhydroxylase- and HMG-CoA reductase specific activities at those time intervals, whereas bile acid synthesis rates were determined throughout the study. Compared with rats not subjected to surgery, rats with short-term biiary diversion had increases in cholesterol 7αhydroxylase activity of 259% and 827% at 48 and 96 hr, respectively. The increase in bile acid biosynthesis was less pronounced. Continuous infusion of mevinolin completely prevented increases in cholesterol 7α-hydroxylase specific activity and bile acid biosynthesis at both time intervals. Intraduodenal infusion of taurocholate alone also suppressed cholesterol 7α-hydroxylase activity. In contrast, infusion of mevalonate led to rapid up-regulation of cholesterol 7α-hydroxylase, with activity 526% greater than basal levels at 48 hr. We conclude that regulation of cholesterol 7α-hydroxylase is under multivalent control in volving a number of regulatory factors. Up-regulation of cholesterol 7α-hydroxylase after short-term biliary diversion occurs as a result of diversion of bile acids away from the liver and the presence of a normally functioning cholesterol biosynthetic pathway. Thus regulation of cholesterol 7α-hydroxylase appears to be under the dual control of bile acids and newly synthesized cholesterol or its metabolites. Lipoprotein cholesterol, a substrate for cholesterol 7α-hydroxylase and bile acid synthesis, does not compensate for the lack of endogenous production of cholesterol and, at least in this model, does not appear to be regulatory. (HEPATOLOGY 1993;18:660-668.)
Cryoglobulinemia and hepatatrophic viruses
Authors: Baker Alfred L.
Published: March 2007

Essential mixed cryoglobulinemia is frequently associated with chronic hepatitis. This report presents four cases of cryoglobulinemia with vasculitis associated with chronic hepatitis related to hepatitis C virus infection. Hepatitis C virus infection was ascertained in the four patients by both the presence in the serum of anti-HCV antibodies detected by the fourantigen recombinant immunoblot assay and of HCV RNA detected by polymerase chain reaction. In two patients tested, anti-HCV antibodies were not detected after centrifugation in the purified cryoglobulin but were detected in the supernatant. HCV RNA was detected in the purified cryoglobulin in all four patients and was detected in the supernatant in three patients. In one patient receiving recombinant interferon alfa, serum aminotransferases normalized and cryoglobulin disappeared; in another patient receiving recombinant interferon alfa, serum aminotransferases remained high and the cryoglobulin persisted. The presence of HCV RNA in the cryoglobulin and the parallelism of the changes of the cryoglobulinemia and of the serum aminotransferases during recombinant interferon alfa administration suggest that HCV infection is responsible for the production of cryoglobulinemia and vasculitis. It is proposed that HCV infection is a cause of cryoglobulinemia associated with chronic hepatitis.

Background.

Type II cryoglobulinemia is a vasculitis characterized by cryoglobulins consisting of complexes of polyclonal IgG and monoclonal IgM rheumatoid factors. The cause of these immune complexes is unknown, though both the hepatitis B (HBV) and C (HCV) viruses have been suspected.

Methods.

We studied 19 patients with Type II cryoglobulinemia for markers of HCV and HBV infection. Quantitative HCV antibody and RNA studies were performed on whole serum, cryoprecipitates, and supernatants.

Results.

Eight patients (42 percent) had HCV antibodies, and 16 (84 percent) had HCV RNA. Of the 19 patients, 5 (26 percent) had HBV markers, but only 1 had evidence of active HBV infection. Control serum samples from nine patients with Type I cryoglobulinemia were negative for HCV antibody and HCVRNA. There was a close, although not exclusive, association of one type of rheumatoid factor (WA) with HCV RNA. HCV antibody and HCV RNA were concentrated approximately 10-fold and 1000-fold, respectively, in the Type II cryoglobulins examined.

Conclusions.

Type II cryoglobulinemia is strongly associated with concomitant HCV infection and a high rate of false negative serologic tests. HCV virions and HCV antigen-antibody complexes are concentrated in the cryoprecipitates, most commonly in association with the WA type of rheumatoid factor, suggesting a role for HCV in the pathogenesis of mixed cryoglobulinemia.
Amino acid transport and glutathione homeostasis: What is the mechanism for cysteine uptake from bile?
Authors: Mailliard Mark E.
Published: March 2007

Transport of L-cysteine and a cysteine S-conjugate, S-(1,2-dichlorovinyl)-L-cysteine (DCVC) was investigated in rat liver canalicular plasma membrane (cLPM) vesicles. Cysteine uptake into an osmotically active intravesicular space was temperature sensitive and further enhanced by an inwardly directed Na+ gradient. Na+ -dependent and -independent L-cysteine uptake exhibited saturation kinetics with apparent K
m of 53 ± 0.7 and 1300 ± 300 μM and Vmax of 95 ± 21 and 1600 ± 200 pmol ± mg protein−1; 10 sec−1 for the Na+ dependent components, and an apparent K
m of 207 ± 48 μM and a Vmax of 355 ± 71 pmol ± mg protein−1 ± 10 sec−1 for the Na+ -independent component. Na+-dependent uptake was inhibited by L-alanine, glycine, L-phenylalanine and L-leucine, whereas Na+-independent uptake was inhibited by L-phenylalanine, L-leucine and 2-amino-2-norbornanecarboxylic acid. Both Na+ -dependent and -independent L-cysteine transport processes were inhibited by several cysteine S-conjugates, with DCVC having the strongest effect. Inhibition of [35S]-cysteine uptake by DCVC was noncompetitive with a K, of 1.2 ± 0.1 mM. On the other hand, uptake of [35 S IDCVC by the rat cLPM vesicles was not stimulated by a Na+-gradient, but was inhibited by several other amino acids, including L-cysteine. Further investigation of [35SlDCVC uptake in rat cLPM vesicles indicated a saturable Na+ -independent process with an apparent K, of 155 f 42 μM, and a V
max of 393 f 53 pmol mg protein −1 5 sec −1. Time-dependent nonspecific binding of DCVC to rat cLPM vesicles was diminished by addition of aminooxyacetic acid, an inhibitor of cysteine conjugate β-lyase activity. These findings demonstrate that cysteine and DCVC uptake into cLPM vesicles occurs by both saturable and nonsaturable transport mechanisms. These transport pathways may function in vivo for uptake and recycling of biliary L-cysteine for glutathione turnover and some cysteine S-conjugates for intrahepatic mercapturic acid biosynthesis.
Deletion of the E-cadherin gene in hepatitis B virus-positive chinese hepatocellular carcinomas
Authors: Slagle Betty L. , Zhou Yi-Zhong , Birchmeier Walter , Scorsone Kathleen A.
Published: March 2007

Frequent allele loss from chromosome 16q was recently described for human tumors of the breast, prostate gland and liver, indicating the possible presence of a tumor-suppressor gene on that chromosome arm. In this study, the chromosome 16 allele status of 38 hepatocellular carcinomas in Chinese patients was determined with restriction-fragmentlength polymorphism analysis. Tumor-specific allele loss was detected in 14(74%) of 19 patients informative for 16p markers and in 22 (85%) of 26 patients informative for 16p markers. Quantitative densitometric analysis revealed reduction to hemizygosity of the E-cadherin cell adhesion gene (localized to 16q22.1) in 18 (64%) of the 28 patients for whom quantitative data were available. Reduced expression of E-cadherin has been associated with invasion and metastasis in several human cell lines and primary tumors, and our results suggest that one mechanism of reduced E-cadherin expression is the loss of one copy of the E-cadherin gene. (HEPATOLOGY 1993;18:757-762).
Five-year follow-up of a prospective randomized trial of hepatitis B recombinant DNA yeast vaccine vs. plasma-derived vaccine in children: Immunogenicity and anamnestic responses
Authors: Lai Ching-Lung , Wong Benjamin Chun-Yu , Yeoh Eng-Kiong , Lim Wai-Ling , Chang Wai-Kwan , Lin Hsiang-Ju
Published: March 2007

In a prospective randomized trial, 318 children aged between 3 mo and 11 yr who were negative for all hepatitis B markers were randomized to receive two 5-μg doses of hepatitis B recombinant DNA yeast vaccine at 0 and 1 mo (group 1), three 5-μg doses of hepatitis B recombinant DNA yeast vaccine at 0, 1 and 6 mo (group 2) or three 10-μg doses of plasma-derived hepatitis B vaccine (group 3). The HBs antibody response rate at 8 mo was between 93% and 99%; it was still 75% to 87% at 5 yr in all three groups. Geometric mean titers at 1 yr were 83, 1,085 and 858 mIU/ml in groups 1, 2 and 3, respectively. These values had decreased after 5 yr to 47, 131 and 250 mIU/ml. Subjects in group 1 showed a significantly less proportional drop in geometric mean titer at the fifth year than did subjects in group 2 (p = 0.05) or group 3 (p = 0.015). None of the children developed HBc antibody, even after 5 yr of follow-up. We noted 42 episodes of significantly increased HBs antibody titers, probably due to anamnestic response, even when the titers had dropped to low levels. The mean age at which anamnestic response occurred was 8.7 yr. We conclude that (a) the recombinant vaccine and plasma-derived vaccine are comparable in safety and immunogenicity; (b) two doses of vaccine was as effective in protecting hepatitis B infection as three doses, despite lower HBs antibody titers; (c) anamnestic responses occurred most frequently around 4 yr after a child began attending school; and (d) a booster dose was not necessary at 5 yr, probably because of ability of healthy child vaccinees to mount good anamnestic responses. (HEPATOLOGY 1993;18:763-767).
Pilot study of recombinant human granulocyte-macrophage colony-stimulating factor in the treatment of chronic hepatitis B
Authors: Martin Julio , Bosch Orencio , Moraleda Gloria , Bartolome Javier , Quiroga Juan Antonio , Carreño Vicente
Published: March 2007

Recombinant human granulocyte-macrophage colony-stimulating factor is being used to improve the immunological function of patients with various diseases and to ameliorate hematological disorders. We investigated the tolerance and possible antiviral effect of the administration of daily doses of recombinant human granulocyte-macrophage colony-stimulating factor (3, 1 or 0.5 μg/kg body wt) to nine patients with chronic hepatitis B, alone or in combination with 5 MU interferon-α2b. Recombinant human GM-CSF reduced significantly (p < 0.02) hepatitis B virus DNA levels. The three doses used were equally effective. Of the eight patients who completed the study, four became negative for HBV DNA and HBeAg; two of them seroconverted to HBe antibody. These four patients showed improvement in the histological activity of their liver disease. Ultimately, two patients regained normal ALT values. 2′,5′-Oligoadenylate synthetase activity increased significantly (p < 0.01) in cell lysates of mononuclear cells cultured in vitro, coinciding with the reductions in hepatitis B virus DNA levels. Recombinant human granulocyte-macrophage colony-stimulating factor was well tolerated but produced a dose-dependent increase in white blood cell counts. It became intolerable at doses of 3 μg (and was reduced to 1.5 μg); this effect was reversible after cessation of recombinant human granulocyte-macrophage colonystimulating factor treatment. No remarkable variations occurred in other parameters. In conclusion, recombinant human granulocyte-macrophage colonystimulating factor administration is safe and tolerable at doses of 0.5 to 1 μg/kg body wt and may exert an antiviral effect in chronic hepatitis B. The efficacy of granulocyte-macrophage colony-stimulating factor should be considered in the prevention of hematological disorders and in the increased susceptibility to bacterial infections, during interferon therapy in chronic hepatitis B virus infection. (HEPATOLOGY 1993;18:775-780).
Hepatitis B virus replication in diverse cell types during chronic hepatitis B virus infection
Authors: Mason Andrew , Wick Mark , White Heather , Perrillo Robert
Published: March 2007

Hepatitis B virus-specific nucleic acid sequences and proteins have been detected in extrahepatic tissues of acutely and chronically infected patients. However, apart from peripheral blood mononuclear cells and bone marrow cells, little is known about the specific cell types that permit viral replication. In this study, we assessed the extrahepatic tissues of four patients who died with chronic hepatitis B virus infection and two uninfected controls by means of in situ hybridization and immunohistochemical study. Three of these patients had diffuse extrahepatic distribution of the virus. Hepatitis B virus nucleic acid sequences and proteins were detected in the lymph nodes, spleen, bone marrow, kidney, skin, colon, stomach, testes and periadrenal ganglia. The following cell types were found to be positive for hepatitis B virus: endothelial cells, macrophages/monocytes, hematopoietic precursors, basal keratinocytes, mucosal epithelial cells, stromal fibroblasts and sustentacular and neuronal cells. It is probable that these cells could support viral replication because hepatitis B virus DNA replicative intermediates, viral transcripts and HBsAg and HBcAg proteins were detected in most. These findings may be relevant to the initiation of extrahepatic syndromes associated with chronic hepatitis B virus infection such as vasculitis, glomerulonephropathy, neuropathy and dermatitis. (HEPATOLOGY 1993;18:781-789).
Intercellular adhesion molecule-1 concentration in sera of patients with acute and chronic liver disease: Relationship to disease activity and cirrhosis
Authors: Zöhrens Gregor , Armbrust Thomas , Pirzer Ursula , zum Büschenfelde Karl-Hermann Meyer , Ramadori Giuliano
Published: March 2007

To study the influence of chronic hepatitis on intercellular adhesion molecule-1 serum concentration, we measured intercellular adhesion molecular-1 in the serum of 84 patients with chronic liver disease (17 chronic persistent hepatitis, 42 chronic active hepatitis and 25 active cirrhosis) caused by hepatitis B virus (n = 46), hepatitis C virus (n = 10) and autoimmunity (n = 28). Furthermore, 20 patients with acute viral hepatitis (16 hepatitis B virus and 4 hepatitis A virus) and 6 patients with acute drug-induced hepatitis were included. Sera from 20 healthy persons were used as control. Follow-up examinations were performed during immunosuppressive therapy in 20 patients with autoimmune chronic liver disease (13 chronic active hepatitis and 7 active cirrhosis). Intercellular adhesion molecule-1 serum concentration was significantly increased in patients with acute viral hepatitis, drug-induced hepatitis, chronic active hepatitis and active cirrhosis compared with healthy controls and with patients with chronic persistent hepatitis. Intercellular adhesion molecule-1 was also significantly increased in severe chronic active hepatitis and active cirrhosis compared with moderate chronic active hepatitis and moderate active cirrhosis. Serum concentration of intercellular adhesion molecule-1 decreased significantly in patients with autoimmune chronic liver disease after 2 mo of immunosuppression when remission was present. A close correlation between aspartate aminotransferase and intercellular adhesion molecule-1 serum levels was found. We conclude the following: (a) in chronic liver disease intercellular adhesion molecule-1 serum concentration may represent, at least in part, hepatocellular damage; and (b) intercellular adhesion molecule-1 serum level does not differentiate between chronic autoimmune and chronic viral hepatitis. (HEPATOLOGY 1993;18:798-802).
Changes of serum 7S fragment of type IV collagen and N -terminal propeptide of type III procollagen after transcatheter arterial embolization as a model of acute liver injury
Authors: Suou Takeaki , Yamada Sadako , Kobayashi Junko , Hoshino Ushio , Nishimuki Eiji , Kawasaki Hironaka
Published: March 2007

To evaluate the mechanism of increased serum concentrations of the N-terminal propeptide of type III procollagen and the 7S fragment of type IV collagen in acute liver injury, we serially measured serum levels of these substances after transcatheter arterial embolization in patients with hepatocellular carcinoma. The results demonstrated marked increases in serum levels of the 7S fragment of type IV collagen in accordance with increased serum levels of transaminase after transcatheter arterial embolization. In contrast, a transient decrease in serum levels of the N-terminal propeptide of type III procollagen was found after transcatheter arterial embolization and transcatheter arterial infusion, although no significant changes in serum levels of the N-terminal propeptide and 7S fragment were seen after hepatic arteriography alone. The absolute change in serum levels of the 7S fragment after treatment showed positive correlation with serum transaminase levels and negative correlation with the decrease in serum levels of α-fetoprotein, although serum levels of N-terminal peptide did not demonstrate such a correlation. The results suggest that in acute liver injury induced by transcatheter arterial embolization, at least in part, increased in serum levels of the 7S fragment of type IV collagen reflect accelerated liberation of type IV collagen from the destroyed surrounding nontumorous tissues and that decreased serum N-terminal propeptide of type III procollagen reflects diminished synthesis of type III collagen caused by anticancer drugs. (HEPATOLOGY 1993;18:809-815).
Clearance by the liver in cirrhosis. II. Characterization of propranolol uptake with the multiple-indicator dilution technique
Authors: Gariépy Louise , Fenyves Daphna , Kassissia Ibrahim , Villeneuve Jean-Pierre
Published: March 2007

We studied the steady-state hepatic extraction and single-pass hepatic uptake of propranolol in isolated perfused livers from normal rats and compared these values with those of rats with carbon tetrachloride-induced cirrhosis, rats treated with chlorpromazine (an inhibitor of propranolol metabolism) and rats with acute liver injury. The kinetics of propranolol transport in the liver were characterized by means of the multiple-indicator dilution technique, and estimates of cellular influx, efflux and sequestration rate constants were obtained with a computer fit to the model of Goresky. The outflow pattern of propranolol in the hepatic veins was then resolved into throughput material, which had swept past the hepatoyctes along with albumin, and returning material, which had entered the cells but returned in the outflow after escaping metabolic sequestration. The steady-state extraction of propranolol was significantly decreased in the three experimental groups compared with that in controls, but the outflow profile differed within each group. In cirrhotic animals, influx was markedly decreased and the sequestration rate constant remained unchanged; most of the propranolol in the outflow consisted of throughput material. In rats treated with chlorpromazine, the sequestration rate constant was decreased, and propranolol in the outflow was mainly returning material. In rats with acute liver injury, both influx and sequestration rate constants were decreased. Indicator dilution curves for nonsequestered tracers showed a decreased transit time for red blood cells and abnormal diffusion of albumin and sucrose into the space of Disse in cirrhotic rats compared with the other groups. These results demonstrate that the decreased hepatic extraction of propranolol in cirrhotic rats is due to impaired cellular influx rather than to reduced metabolism. We speculate that the limitation of propranolol cellular entry in cirrhosis is related to restriction of diffusion of protein-bound propranolol into the space of Disse, to the presence of small intrahepatic shunts or to both (HEPATOLOGY 1993;18:823-831).
Stimulation of mono-ADP ribosylation in rat liver plasma membranes after long-term alcohol intake
Authors: Nomura Fumio , Noda Masatoshi
Published: March 2007

ADP ribosylation is considered one of the important covalent modifications of cellular proteins catalyzed by ADP ribosyltransferase, which transfers ADP ribose moiety of NAD to an acceptor protein. Because a growing body of evidence has suggested significant biological roles for mono-ADP ribosylations in transmembrane signal transduction and other cell metabolism, how alcohol intake alters them is of interest. Cholera toxin and pertussis toxin have been widely used as probes to investigate the roles of GTP-binding proteins (G-proteins) in the transduction of hormonal and sensory signals. We first tested effects of long-term alcohol intake on these toxin-catalyzed ADP ribosylations of G-proteins in rat liver plasma membranes. Treatment of rat liver plasma membrane with [32P]NAD and thiol-preactivated cholera toxin resulted in the labeling of a 44-kD band, most likely an α-subunit of the stimulatory GTP-binding protein, the extent of which was much greater in alcohol-fed rats than in pair-fed controls. Analogous experiments with pertussis toxin also demonstrated enhancement of toxin-catalyzed ADP ribosylation of the inhibitory GTP-binding protein after long-term alcohol intake. More interesting was that long-term alcohol intake remarkably stimulated endogenous mono-ADP ribosylation of a 58-kD protein in a GTP-dependent manner. In vitro, ethanol (50 mmol/L) or a single load of ethanol (3 gm/kg) did not stimulate the reaction. Thus long-term alcohol intake stimulated both toxin-catalyzed and endogenous mono-ADP ribosylations of proteins in rat liver plasma membranes. Pursuit of alcohol interaction with mono-ADP ribosylation may provide an interesting approach to the study of alcohol's effects on the liver. (HEPATOLOGY 1993;18:870-873).
Regulation of iron metabolism in HepG2 cells: A possible role for cytokines in the hepatic deposition of iron
Authors: Hirayama Michiaki , Kohgo Yutaka , Kondo Hitoshi , Shintani Naoaki , Fujikawa Koshi , Sasaki Katsunori , Kato Junji , Nhtsu Yoshiro
Published: March 2007

In chronic inflammation it is reported that serum iron is depleted and hepatic iron is increased because of reticuloendothelial system iron blockade. However, recent studies indicate that hepatic parenchymal cells increase the uptake of transferrin-bound iron after in vivo stimulation with bacterial lipopolysaccharide, suggesting that endotoxemia itself or lipopolysaccharide-induced production of inflammation-related cytokines may also be responsible for this phenomenon. In this study the actions of inflammation-related cytokines on the synthesis of iron-binding proteins (transferrin and ferritin) and transferrin receptor and the uptake of transferrin-bound iron were investigated in a human hepatoblastoma cell line, HepG2, which is the most commonly used cell line for examining the regulation of hepatic protein synthesis by cytokines. The cells were exposed to interleukin-1β, interleukin-6 or tumor necrosis factor-α separately for 24 hr. In each cytokine treatment group, the level of transferrin, which is secreted into the conditioned medium, was found to be decreased compared with that of untreated cells. On the other hand, the biosynthesis of ferritin was markedly elevated after the same treatment. This increase in ferritin by cytokine treatment was diminished when deferoxiamine was used concomitantly to deplete intracellular chelatable iron. After stimulation with interleukin-β, interleukin-6 or tumor necrosis factor-α, 59Fe-labeled transferrin uptake into the cells was increased by 36%, 48%, or 18%, respectively, and this uptake was inhibited by the addition of excess unlabeled transferrin. A binding study with 125I-labeled diferric transferrin revealed that the three cytokines increased the number of transferrin receptors on the cell surface by 1.15-fold to 1.35-fold. A metabolic labeling study using 35S-methionine showed that the increase of transferrin-receptor expression was not caused by the redistribution between the cell surface and internal pool but by de novo synthesis. These findings suggest that not only reticuloendothelial system blockade of iron release, but also enhanced liver cell uptake of transferrin-bound iron by cytokine stimulation as well as endotoxin is responsible for the hypoferremia seen during inflammation. (HEPATOLOGY 1993;18:874-880).
Oxidative stress produced by suprahepatic occlusion and reperfusion
Authors: Gonzalez-Flecha Beatriz , Reides Claudia , Cutrin Juan C. , Llesuy Susana F. , Boveris Alberto
Published: March 2007

In this article the spontaneous chemiluminescence and the steady-state concentration of hydrogen peroxide were determined in rat liver as indicators of oxidative stress in the tissue. Hydroperoxide-initiated chemiluminescence and the activity of antioxidant enzymes (catalase, superoxide dismutase and glutathione peroxidase) were also measured to evaluate antioxidant defenses and serum activity of lactate dehydrogenase and aspartate aminotransferase. Mitochondrial morphology and mitochondrial respiratory control ratio were measured as indicators of cell and mitochondrial damage. Xanthine dehydrogenase and xanthine oxidase activities were determined as a possible source of oxyradicals. No significant changes were observed after 10 or 30 min of vena cava occlusion in any of the measured parameters. In contrast, 10 min of occlusion followed by 10 min of reperfusion increased chemiluminescence (from 18 ± 3 to 32 ± 5 cps/cm 2), hydrogen peroxide (from 0.10 ± 0.01 to 0.17 ± 0.01 μmol/L), lactate dehydrogenase (from 80 ± 2 to 330 ± 30 U/L), and aspartate aminotransferase (from 42 ± 2 to 100 ± 10 U/L). Liver reperfusion was also associated with mitochondrial swelling and decreased mitochondrial respiratory control (from 5.6 ± 0.3 to 2.6 ± 0.1). The activity of the antioxidant enzymes and xanthine oxidase was instead without change. After 30 min of vena cava occlusion and 10 min of reperfusion a more marked increase in chemiluminescence (37 ± 5 cps/cm 2), hydrogen peroxide (0.30 ± 0.01 μmol/L), lactate dehydrogenase (730 ± 10 U/L) and aspartate aminotransferase (140 ± 10 U/L) was observed. No further changes were found in either mitochondrial morphology or respiratory control (2.4 ± 0.1) in isolated mitochondria. A parallel decrease in the activity of cytosolic (36%) and mitochondrial (57%) superoxide dismutase, catalase (34%) and glutathione peroxidase (34%) was also observed without any change in xanthine oxidase activity. Chain-breaker antioxidants were also diminished as indicated by an increase in the hydroperoxide-initiated chemiluminescence (40%). The results indicate the occurrence of an oxidative stress in association with a sequence of 10 or 30 min of occlusion followed by 10 min of reperfusion. The unchanged activity of xanthine oxidase, the modifications in mitochondrial morphology, the decrease in mitochondrial respiratory control and the relatively high inactivation of the mitochondrial superoxide dismutase indicate that the mitochondria are the main source of oxyradicals during reperfusion. An increased rate of superoxide radical and hydrogen peroxide generation by mitochondria, associated with a decreased activity of antioxidant enzymes, would be the major causes of oxidative stress and the related cell damage. (HEPATOLOGY 1993;18:881-889).
Metabolic adaptation of the kidney to hyperammonemia during chronic liver insufficiency in the rat
Authors: Dejong Cornelis H. C. , Deutz Nicolaas E. P. , Soeters Peter B.
Published: March 2007

The aim of this study was to evaluate the role of renal ammonia and glutamine metabolism in the metabolic adaptation to chronic liver insufficiency-induced hyperammonemia in the rat. To this purpose, urinary excretion, renal net exchange and tissue concentrations of ammonia and amino acids were measured in anesthetized, normal control rats that did not undergo surgery, in control rats that underwent sham surgery, in rats that underwent portacaval shunting and in rats that underwent both portacaval shunting and bile duct ligation. Rats that underwent sham surgery and portacaval shunting were pair-fed with rats that underwent portacaval shunting and biliary obstruction, to correct for anorexia in that group, and all rats that were operated on were studied 7 and 14 days after surgery. Arterial ammonia and glutamine levels were elevated in groups that underwent portacaval shunting and portacaval shunting plus biliary obstruction at all time points. At days 7 and 14, total renal ammonia production decreased in rats that underwent portacaval shunting and in rats that underwent portacaval shunting plus biliary obstruction, associated with a 50% decrease in net renal glutamine uptake and strongly diminished net ammonia release into the renal vein, which was most prominent in the group that underwent portacaval shunting plus biliary obstruction. Urinary ammonia excretion was similar in rats that underwent portacaval shunting and in those that underwent sham surgery but was increased more than 200% at days 7 and 14 in rats that underwent portacaval shunting plus biliary obstruction. In this group, in contrast to portacaval-shunted rats, the kidney appeared to be an organ of net ammonia disposal from the body. In separate experiments in unanesthetized, unrestrained rats, similar changes in urinary ammonia excretion were observed without changes in arterial pH, excluding an effect of anesthesia or pH on the obtained results. These results indicate that the kidney plays an important role in the metabolic adaptation to hyperammonemia during chronic liver insufficiency in the rat. (HEPATOLOGY 1993;18:890-902).
Relationship between genomic DNA ploidy and parameters of liver damage during necrosis and regeneration induced by thioacetamide
Authors: Díez-Fernández Carmen , Boscá Lisardo , Fernández-Simón Lourdes , Alvarez Alberto , Cascales María
Published: March 2007

Thioacetamide proved to be a potent necrogenic agent when a single dose of 6.6 mmol/kg was administered intraperitoneally to rats. Its necrogenic ability was assessed on the basis of morphological and biochemical changes. The injury of centrilobular hepatocytes showed a peak of cell death 24 hr after thioacetamide administration; it was followed immediately by the regenerative response. Parallel increases of serum aminotransferases, isocitrate dehydrogenase and γ-glutamyl transferase activities were observed. Severe liver damage was also evident at 24 hr on the basis of glutathione depletion (29% of control), malondialdehyde production (169%), cytochrome P-450 level decrease (26%) and increased activity of glutathione S-transferase (160%). We checked the regenerative response by determining nuclear DNA content in isolated hepatocytes 0, 6, 12, 18, 24, 36, 48 and 72 hr after thioacetamide administration. Changes in DNA cell distribution between G0-G1, S and G2 + M phases of the cell cycle were observed. The sharp decrease in the percentage of the tetraploid cell population (G2 + M phases) and the abrupt increase of the S-phase cells at 36 and 48 hr suggest transition from adult to fetal in hepatocyte populations obtained 24 and 36 hr after thioacetamide treatment. At 72 hr of treatment, hepatocyte populations showed recovery to adult state. In the shift from the adult to fetal, registered at 24, 36 and 48 hr after thioacetamide administration, mitosis seemed to precede the synthesis of DNA. (HEPATOLOGY 1993;18:912-918).
Quantitative detection of hemoglobin saturation in the liver with near-infrared spectroscopy
Authors: Kitai Toshiyuki , Tanaka Akira , Tokuka Atsuo , Tanaka Koichi , Yamaoka Yoshio , Ozawa Kazue , Hirao Konomu
Published: March 2007

To quantify the changes in oxygen saturation of hemoglobin in the liver in hypoxia and liver transplantation, we applied a novel method using near-infrared spectroscopy. Instead of the conventional two-wavelength method, we obtained near-infrared data from a wide spectral range of 700 nm to 1,000 nm with continuous-wave spectroscopy. To correct the flattened spectral shape caused by photon scattering in living tissue, we then applied an equation taking into account the relationship between absorber concentration and actual absorption in scattering materials as assessed with time-resolved spectroscopy. Hepatic hemoglobin oxygen saturation was calculated with multicomponent analysis. In room air, hepatic hemoglobin oxygen saturation of rabbit was calculated as 57.4% ± 2.5% (mean ± S.E.M., n = 7). Arterial, portal and hepatic venous hemoglobin oxygen saturation were simultaneously measured as 97.5% ± 0.7%, 77.1% ± 3.4% and 55.5% ± 4.6%, respectively. The changes in hepatic hemoglobin oxygen saturation seen in graded hypoxia were also close to those in hepatic venous hemoglobin oxygen saturation, suggesting that the average oxygenation state of sinusoidal blood approximates that in the central vein. We tested the clinical applicability of this method in a case of liver transplantation. It was determined that the hepatic hemoglobin oxygen saturation of the graft liver was heterogeneously distributed and that the initially low level of hepatic hemoglobin oxygen saturation was increased by the ligation of portal-systemic shunts. (HEPATOLOGY 1993;18:926-936).
Contribution of sinusoidal endothelial liver cells to liver fibrosis: Expression of transforming growth factor-β1 receptors and modulation of plasmin-generating enzymes by transforming growth factor-β1
Authors: Rieder Hartmut , Armbrust Thomas , zum Büschenfelde Karl-Hermann Meyer , Ramadori Giuliano
Published: March 2007

Transforming growth factor-βH1 is an important cytokine in the pathophysiology of liver fibrosis, stimulating the production of extracellular matrix. Whether this cytokine can also control the degradation of matrix proteins in liver cells has not been investigated. Because plasmin is an important protease for the degradation of matrix glycoproteins, we investigated whether sinusoidal endothelial liver cells could contribute to fibrosing liver disease through the modulation of plasmin-generating enzymes in response to transforming growth factor-β1. Sinusoidal endothelial cells from guinea pig liver were investigated in pure monolayer culture. Using 125I-labelled transforming growth factor-β, we demonstrated high-affinity binding sites on sinusoidal endothelial cells at a density of 9.3 × 102 per cell, and a dissociation constant of about 5.5 × 10−11 mol/L. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed the known three classes of membrane receptors for transforming growth factor-β. Using biosynthetic labeling of proteins with 35S-methionine, immunoprecipitation and sodium dodecyl sulfate-polyacrylamide gel electrophoresis, we showed that sinusoidal endothelial cells produce and secrete plasminogen activator inhibitor type 1 from the beginning of culture. Treatment of confluent cell cultures for 24 hr with transforming growth factor-β1 increased synthesis and release of plasminogen activator inhibitor type 1. The response was almost maximal at a concentration of 1 ng transforming growth factor-β/ml and paralleled the increased synthesis of fibronectin. On reverse fibrin autography we proved that transforming growth factor-β1 stimulated the release of functionally active plasminogen activator inhibitor type 1. On the other hand, transforming growth factor-β1 decreased in a dose-dependent manner the release of plasminogen activators as shown by fibrin autography. From these studies it is evident that sinusoidal endothelial liver cells respond to transforming growth factor-β1 in a fibrogenic manner, reducing the plasmin-generating potential, resulting in decreased proteolysis of extra-cellular matrix glycoproteins. Therefore we postulated that sinusoidal endothelial cells may participate in early stages of fibrotic liver disease such as capillarization and collagenization of liver sinusoids. (HEPATOLOGY 1993;18:937-944).
Induction of apoptosis by transforming growth factor-β1 in the rat hepatoma cell line MCA-RH7777: A possible association with tissue transglutaminase expression
Authors: Fukuda Kazunori , Kojiro Masamichi , Chiu Jen-Fu
Published: March 2007

We report here that transforming growth factor-β1 induces cell death in the Morris hepatoma cell line McA-RH7777. We assessed the type of cell death induced by transforming growth factor-β1 in this hepatoma cell line on the basis of morphological and biochemical characteristics. Dying cells, which detached from the cell monolayer, showed morphological characteristics of apoptosis (programmed cell death) such as chromatin condensation, nuclear disintegration and cellular fragmentation into clusters of eosinophilic globules. DNA isolated from these cells showed a ladder pattern consisting of multimers of 180 to 190 bp, indicating extensive DNA cleavage into oligonucleosomal units by an endogenous endonuclease. Treatment of the dead cells with detergents and chaotropic agents resulted in formation of insoluble shells, so-called apoptotic bodies, suggesting extensive cross-linking of cell proteins by tissue transglutaminase. Furthermore, increased amounts of cytosolic tissue transglutaminase, which has been recognized as a possible marker of apoptosis, and extensive crosslinking of cytokeratin polypeptides was demonstrated in TGF-β1-treated hepatoma cells on immunoblot analysis. These results provide strong evidence that the cell death induced by TGF-β1 in McA-RH7777 hepatoma cells is mainly apoptotic. It also suggests that a specific induction of the cytosolic tissue transglutaminase may be involved in the TGF-β1-induced pathways of apoptotic cell death in McA-RH7777 hepatoma cells. (HEPATOLOGY 1993;18:945-953).
Endothelins 1 and 3: Potent cholestatic agents secreted and excreted by the liver that interact with cyclosporine
Authors: Bluhm Renata E. , Frazer Marshall G. , Vore Mary , Pinson Wright C. , Badr Kamal F.
Published: March 2007

Autoradiographic studies have shown that the liver accumulates endothelin. High-affinity binding sites for endothelin have been identified on rat liver plasma membranes. We investigated the role of endothelin isopeptides as mediators of cholestasis with isolated rat liver perfused by a recirculating solution of buffer and blood. These studies demonstrated that endothelin-1, as measured by means of radioimmunoassay, was cleared from the perfusate by the liver and that the liver concentrated both endothelin-1 and endothelin-3 in bile. Addition of endothelin-1 to the liver perfusate solution increased the concentration of endothelin-3 measured in the perfusate, suggesting that endothelin-1 caused release or secretion of endothelin-3. Both endothelin-1 and endothelin-3 at 5 nmol/L caused almost complete cessation of bile flow, but this effect was more prolonged after endothelin-1 than after endothelin-3 administration. Because it has been reported that cyclosporine increases endothelin levels, we studied the interaction of these two compounds. Cyclosporine (100 μmol/L) also produced cholestasis. Endothelin-3 secretion in bile, however, was decreased in livers perfused with cyclosporine compared with secretion in controls. Simultaneous addition of endothelin-1 and cyclosporine that on their own were not significantly cholestatic produced cholestasis. In conclusion, endothelin is a potent cholestatic agent secreted and excreted by the liver. It may potentiate the cholestatic action of cyclosporine. (HEPATOLOGY 1993;18:961-968.)
Hepatic blood volume responses and compliance in cats with long-term bile duct ligation
Authors: Schafer Joshua , D'Almeida Mark S. , Weisman Harvey , Lautt Wayne W.
Published: March 2007

Hepatic capacitance responses were compared in sham-operated and 14-day bile duct-ligated cats under pentobarbital anesthesia. Both groups were subjected to splenectomy and had the anterior hepatic nerve plexus sectioned to allow stimulation; the posterior plexus was intact. Blood volume compensation for hemorrhage was reduced in the bile duct ligation group compared with the control group: The liver compensated for 20.1% and 10.6% of blood loss, respectively. Portal hypertension did not exist and hepatic compliance was unaltered despite the presence of severe biliary hyperplasia, portal tract distortion and fibrosis. The capacitance responses to hemorrhage in the bile duct ligation group were accounted for entirely by a passive compliant response to reduced portal pressure, whereas the sham surgery group showed an additional active component. Responses of liver volume and venous resistance to norepinephrine were normal, but responses to nerve stimulation were reduced. The nerve dysfunction was not universal; reflex arterial blood pressure response to carotid occlusion was normal. The data suggest that hepatic blood volume response to hemorrhage is mediated by passive compliant responses to reduced portal pressure (stressed volume) and active responses (unstressed volume) to nerve stimulation but that long-term bile duct ligation produces selective hepatic neuropathy resulting in loss of the active component. (HEPATOLOGY 1993;18:969-977).
Ito cell contraction in response to endothelin-1 and substance P
Authors: Sakamoto Masaharu , Ueno Takato , Kin Motoaki , Ohira Hiromasa , Torimura Takuji , Inuzuka Sadataka , Sata Michio , Tanikawa Kyuichi
Published: March 2007

The contractile response of cultured Ito cells to endothelin-1 and substance P was examined. Ito cells were obtained from rat liver by perfusion with collagenase, followed by separation through centrifugal elutriation, and were cultured for 24 hr. The area of the Ito cells was measured after treatment with endothelin-1 or substance P at various concentrations in the culture medium. The area of the cells decreased dose dependently after treatment with endothelin-1 or substance P. The area of Ito cells before addition of interleukin-1 or substance P was defined as 100%. The area of the cells after treatment with endothelin-1 or substance P medium was expressed as the percentage against the area before treatment with endothelin-1 or substance P. The percentage in area after treatment with 200 nmol/L endothelin-1 was as follows: 81% ± 13% at 30 min, 77% ± 15% at 60 min, 87% ± 15% at 120 min and 99% ± 18% at 180 min. The maximal decrease in area occurred at 60 min after treatment. The percentage values for 200 nmol/L substance P were as follows: 88% ± 15% at 10 min, 95% ± 17% at 30 min and 101% ± 17% at 60 min. The maximal decrease in area was noted at 10 min. Thus Ito cells contracted in response to treatment with endothelin-1 or substance P. The mode of the extent and onset of the contraction was different for the two peptides. These findings suggest that Ito cells are involved in the regulation of the hepatic sinusoidal microcirculation. (HEPATOLOGY 1993;18:978-983).
The effect of ethanol on one-carbon metabolism: Increased methionine catabolism and lipotrope methyl-group wastage
Authors: Trimble Kenneth C. , Molloy Anne M. , Scot John M. , Weir Donald G.
Published: March 2007

Deficiency of choline and methionine produces hepatic steatosis similar to that seen with ethanol, and supplementation with these lipotropes can prevent ethanol-induced fatty liver. These effects are thought to occur through alterations in membrane phospholipid metabolism, but the mechanism whereby this occurs and the precise nature of the changes brought about by ethanol in the interactions of choline and methionine metabolism remain unclear. Through the known effects on hepatic glutathione (which requires as a precursor a product of methionine catabolism), ethanol might affect hepatic one-carbon metabolism, which requires the participation of both methionine and choline in the transfer of methyl groups. This has been investigated with a radiorespirometric technique to assess the in vivo oxidation of the methyl groups of lipotropes and their intermediates in ethanol- and control-fed rats. Enzyme activities of one-carbon transfer reactions and the hepatic levels of methionine and α-aminobutyrate, an end product of methionine catabolism, have been measured. The effect of ethanol feeding on hepatic S-adenosylmethionine and S-adenosylhomocysteine has also been assessed. Ethanol increases the oxidation to carbon dioxide of the methyl group of methionine by a factor of 2.9 (p = 0.002) and produces a 3.6-fold (p = 0.0001) accumulation of α-aminobutyrate, indicating a marked increase in methionine catabolism. Hepatic methionine levels are unchanged by ethanol, however, and this may be explained by a dramatic increase in the turnover of the methyl groups of choline and betaine in response to ethanol (times 3.6 and 4.2, respectively, p < 0.003), suggesting greatly increased use of the choline oxidation pathway to remethylate homocysteine through betaine homocysteine methyltransferase. The activity of this enzyme is more than doubled (p = 0.002) in ethanol-fed animals, and that of methionine synthase is significantly decreased. Hepatic S-adenosylmethionine is decreased by ethanol (59.6 vs. 100.9 nmol/gm, p < 0.001), possibly representing increased consumption, and S-adenosylhomocysteine is increased. We propose that ethanol stimulates catabolism of methionine to generate cysteine and replenish glutatione, but at the same time, the cell attempts to conserve methionine through a futile cycle of enhanced choline oxidation. The result is a dramatic wastage of methyl groups. (HEPATOLOGY 1993;18:984-989).
Lessons from the repression of the α-fetoprotein gene in the adult liver
Authors: Bilir Bahri M.
Published: March 2007

The developmental regulation of the α-fetoprotein (AFP) gene in liver results in high-level expression in the fetus, followed by dramatic transcriptional repression after birth. We have examined the mouse AFP gene for transcriptional control sequences that may be involved in its postnatal repression in liver. We showed previously that removal of a DNA region between positions −250 base pairs (bp) and −838 bp of the AFP gene resulted in the persistence of expression of an AFP minigene in the postpartum liver of transgenic mice (Vacher and Tilghman, Science 250;1732-1735, 1990). This study examines the distribution of these transgene transcripts in liver using in situ hybridization. We show that there is a zonal distribution of minigene transcripts in the adult livers of these animals. Hepatocytes surrounding the central veins express high levels of minigene transcripts, while hepatocytes in the intermediate and portal areas contain few, if any, transcripts. Quantitative RNAsc protection analysis shows a decrease in transgene RNA levels after birth, consistent with repression in all but a small subset of hepatocytes. These results indicate that repression in the pericentral hepatocytes is dependent upon the presence of a cis-acting, negative-regulatory domain, which is located between the enhancers and the proximal promoter of the AFP gene. In contrast, this domain is not essential for complete repression of AFP transgenes in the intermediate zone and periportal hepatocytes.