bluuebunny/crossref_metadata_2025 huggingface dataset:
Up-regulation of heme-binding protein 23 (HBP23) gene expression by lipopolysaccharide is mediated via a nitric oxide-dependent signaling pathway in rat Kupffer cells
Authors: Immenschuh Stephan , Stritzke Janina , Iwahara Shin-ichiro , Ramadori Giuliano
Published: July 2004

Heme-binding protein 23 (HBP23) is a cytosolic protein that binds the prooxidant heme with high affinity and has been implicated in the cellular protection against reactive oxygen species (ROS). Because lipopolysaccharide (LPS) stimulates macrophages to produce large amounts of ROS the gene expression of HBP23 was analyzed during treatment with LPS in cultured rat Kupffer cells (KC). HBP23 was constitutively expressed in KC and up-regulated on the protein and messenger RNA (mRNA) level by LPS with a time response distinct from that of TNFα, but in coordination with that of heme oxygenase-1 (HO-1), which is the inducible isoform of the rate-limiting enzyme of heme degradation. A parallel up-regulation of HBP23 and HO-1 mRNA by LPS was also observed in cultured peritoneal macrophages and peripheral blood monocytes. HBP23 mRNA induction by LPS occurred on the transcriptional level as indicated by blocking with actinomycin D. The induction of HBP23 mRNA expression by LPS was preceded by that of the inducible nitric oxide synthase (iNOS) and the production of nitrite in KC. Treatment with the NOS inhibitor NG-monomethyl l -arginine prevented HBP23 mRNA induction by LPS, which was reversed by an excess of l -arginine. Both the nitric oxide (NO)-donor S-nitroso-N-acetylpenicillamine and the peroxynitrite donor SIN-1 increased HBP23 mRNA expression. HBP23 mRNA induction by LPS was down-regulated by interleukin 10 and transforming growth factor β1 with a NO-independent mechanism. LPS-stimulated KC exhibited marked protection against the cytotoxicity mediated by H2 O2. The data suggest that NO and peroxynitrite are major mediators of the LPS-dependent up-regulation of HBP23 in KC.
Acute sporadic non-A, non-B hepatitis in Northeastern Brazil: Etiology and natural history
Authors: Paraná Raymundo , Vitvitski Ludmila , Andrade Zilton , Trepo Christian , Cotrim Helma , Bertillon Pascale , Silva Fernanda , Silva Luciana , de Oliveira Irismar R. , Lyra Luis
Published: July 2004

In a 4-year follow-up study, patients with acute sporadic non-A, non-B (NANB) hepatitis were evaluated to determine the etiology and natural history of the disease. Acute hepatitis C virus (HCV) was detected in 13 of 43 (30%) of patients, anti-hepatitis E virus (HEV) IgG in 5 (12%), and 25 (58%) were considered non-A-E. The HCV RNA was detected in all HCV patients but none of the non-A-E cases. The initial clinical and biochemical presentation of the HCV and non-A-E cases was quite similar, although 2 of the non-A-E patients had severe disease. The 5 patients who were found to be anti-HEV IgG-reactive recovered within 6 months of follow-up. Of the 13 HCV cases, alanine transaminase (ALT) levels returned to normal in 7 (53.8%), while 6 (46.2%) continued to show abnormal ALT after 6 months of follow-up. However, 9 (69.2%) of them remained HCV-RNA-positive, denoting virological/biochemical dissociation. Long-term follow-up showed a reappearance of HCV RNA in 2 of the 4 patients who were in virological remission performing 84% of chronicity rate. Acute non-A-E hepatitis patients were less likely to evolve toward chronicity, as compared with acute HCV cases (16% vs. 84%; P = .0001). Only 4 (16%) of the non-A-E patients were hepatitis G virus (HGV)-RNA-positive. Concerning risk factors for acquiring parenterally transmitted viruses, tattooing was the only one that could be associated with HCV transmission (P = .002). No risk factors could be identified for putative non-A-E virus transmission. Liver biopsies performed for chronic HCV patients showed a variable degree of inflammation, while the non-A-E patients presented less severe histological disease.
In situ nucleic acid detection of PDC-E2, BCOADC-E2, OGDC-E2, PDC-E1α, BCOADC-E1α, OGDC-E1,and the E3 binding protein (protein X) in primary biliary cirrhosis
Authors: Harada Kenichi , Sudo Yoshiko , Kono Naoko , Ozaki Satoru , Tsuneyama Koichi , Gershwin Eric M. , Nakanuma Yasuni
Published: July 2004

The characteristic serological feature of primary biliary cirrhosis (PBC) is the presence of antimitochondrial antibodies (AMAs), and the major proteins recognized by AMAs are subunits of the 2-oxo acid dehydrogenase complexes (2-OADC), including the E2 components of the pyruvate dehydrogenase complex (PDC), the 2-oxo-glutarate dehydrogenase complex (OGDC), the branched-chain 2-oxoacid dehydrogenase complex (BCOADC), the E3 binding protein (E3BP or protein X) and the E1a component of mammalian PDC. Previous work has postulated that either E3BP, or a molecule cross-reactive with the PDC-E2 molecule, is uniquely expressed on the surface of biliary epithelial cells in PBC. To address this issue, we performed in situ hybridization for all of the major 2-OADC components at the mRNA level, including PDC-E2, BCOADC-E2, OGDC-E2, PDC-E1a, BCOADC-E1a, OGDC-E1, and E3BP using 13 PBC and 9 control livers using 7 mitochondrial antisense probes. In both PBC and controls, the expression of all 2-OADC component mRNA studied herein were found in hepatocytes and infiltrating mononuclear cells, without significant differences. Interestingly, however, despite published data on immunohistochemical staining, interlobular bile ducts including the injured bile ducts in PBC were generally negative or only faintly positive, with the exception of 1 bile duct in 1 of 13 cases of PBC and 1 of 9 control liver specimens. Moreover, confocal microscopic examination and image analysis revealed that the mRNA signal intensity of each of the 2-OADC components in the bile ducts of PBC was relatively lower in comparison with control liver diseases. These data suggest that continuous enhanced synthesis of the 2-OADC components is not likely to be occurring in the biliary epithelial cells in PBC, and that an increase of PDC-E2 or E3BP immunoreactivity in PBC is caused by exogenous imported or cross-reactive molecules
Ceramide induces caspase-independent apoptosis in rat hepatocytes sensitized by inhibition of RNA synthesis
Authors: Jones Brett E. , Lo Chau R. , Srinivasan Anu , Valentino Karen L. , Czaja Mark J.
Published: July 2004

Ceramide has been implicated as a second messenger in intracellular signaling pathways leading to apoptosis in nonhepatic cells. To determine whether ceramide can mediate hepatocyte apoptosis, the cytotoxicity of ceramide was determined in rat hepatocytes. The rat hepatocyte cell line, RALA255-10G, and primary rat hepatocytes were completely resistant to toxicity from 10 to 100 μmol/L C2 ceramide. Resistance was not the result of a failure to take up ceramide, because ceramide treatment did cause nuclear factor-κB (NF-κB) activation. Because ceramide may mediate cell death from tumor necrosis factor α (TNF-α), the ability of RNA synthesis inhibition and NF-κB inactivation to sensitize hepatocytes to ceramide toxicity was examined. RALA hepatocytes were sensitized to ceramide toxicity by coadministration of actinomycin D (ActD). Cell death occurred by apoptosis as determined by the presence of morphological evidence of apoptosis, caspase activation, poly(ADP-ribose) polymerase (PARP) degradation, and DNA hypoploidy. Despite the induction of apoptosis associated with caspase activation, cell death from ActD/ceramide was not blocked by caspase inhibition. Inhibition of NF-κB activation also sensitized RALA hepatocytes to ceramide toxicity, but to a lesser extent than for TNF-α. Thus, unlike many nonhepatic cell types, rat hepatocytes are resistant to cell death from ceramide because of the transcriptionally dependent up-regulation of a protective gene(s). The ability of ActD and NF-κB inactivation to sensitize RALA hepatocytes to ceramide toxicity suggests that ceramide may act as a downstream mediator of TNF-α toxicity.
Chronic hepatitis C virus patients with breakthroughs during interferon treatment can successfully be retreated with consensus interferon
Authors: Heathcote Jenny E. , James Stephen , Mullen Kevin D. , Hauser S. C. , Rosenblate H. , Albert Donald G.
Published: July 2004

Patients with chronic hepatitis C who have not had a sustained hepatitis C virus (HCV)-RNA response or serum alanine transaminase (ALT) response to a 6-month course of interferon (IFN) may respond to higher dose retreatment with consensus interferon (CIFN). Some nonresponders to initial IFN treatment have a transient response defined as undetectable HCV RNA or normalization of ALT during treatment, but subsequently have a “breakthrough” while still on treatment. The aim of this study was to determine if nonresponders who had breakthroughs responded differently to CIFN retreatment than nonresponders without breakthroughs using data from a large, multicenter trial. ALT and HCV RNA were monitored frequently during initial IFN therapy (either 9 mcg CIFN or 3 MU IFN-α2b 3 times per week). HCV-RNA breakthroughs were observed in 86 of 467 (18%) of all treated patients, and ALT breakthroughs were observed in 90 of 467 (19%) of all treated patients. There was no association between breakthroughs and the presence of either binding or neutralizing anti-IFN antibodies. When the patients who were nonresponders to initial IFN treatment were retreated with CIFN (15 mcg) for 12 months, 27% of those with viral breakthroughs had a sustained viral response compared with 8% in prior nonresponders without breakthroughs (P = .102). Sustained ALT responses were observed in 39% with breakthroughs compared with 10% in those without breakthroughs (P = .014). The data suggest that prior nonresponders with breakthroughs have a greater chance of responding to retreatment than do nonresponders without breakthroughs. However, most breakthrough patients would be missed unless repeated HCV-RNA testing were conducted during therapy.
A pilot study of the CY-1899 T-cell vaccine in subjects chronically infected with hepatitis B virus
Authors: Heathcote Jenny , McHutchison John , Lee Samuel , Tong Myron , Benner Kent , Minuk Gerald , Wright Teresa , Fikes John , Livingston Brian , Sette Alex , Chestnut Robert
Published: July 2004

Clinical observations suggest that eradication of the hepatitis B virus (HBV) is immune-mediated. Vigorous cytotoxic T lymphocyte (CTL) activity directed at HLA class I-bound viral epitopes are detected during acute hepatitis B, but not in chronic hepatitis B carriers. A CTL epitope derived from the hepatitis B core protein amino acids 18-27 has been incorporated into a vaccine also comprised of a T-helper cell epitope and 2 palmitic acid residues (CY-1899). The aim of this study was to determine whether repeated doses of CY-1899 given to patients with chronic hepatitis B could initiate in vivo CTL activity and viral clearance. Patients with chronic hepatitis B received up to 4 doses (ranging from 0.05 mg to 15 mg) 6 weeks apart. Following vaccination, patients were monitored for hepatitis B surface antigen and “e” status, HBV-DNA levels, liver biochemistry, CTL activity, and any adverse events. Ninety patients with chronic hepatitis B infection received CY-1899. Mean CTL responses were all low but were maximal following vaccination with 5 mg CY-1899. Peak CTL responses never exceeded 10 lytic units (LU) regardless of vaccine dose, this value being well below that seen following resolution of acute hepatitis B. No significant changes in liver biochemistry or viral serology were observed during follow-up. No serious adverse events were noted. Administration of the single-epitope vaccine, CY-1899, initiated CTL activity, but of a magnitude lower than that observed during spontaneous HBV clearance. This low-level CTL activity was not associated with viral clearance.
Inhibition of system A amino acid transport and hepatocyte proliferation following partial hepatectomy in the rat
Authors: Freeman Thomas L. , Ngo Hao Q. , Mailliard Mark E.
Published: July 2004

System A, the sodium-dependent neutral amino acid transport activity, has a 3-fold increase in its initial uptake velocity into hepatocytes following partial hepatectomy (PH) in the rat. The purpose of this study was to examine the effect of inhibition of System A-mediated amino acid transport on hepatocyte proliferation and liver regeneration. We describe thein vivocompetitive inhibition of System A activity following PH by the nonmetabolizable, System A-specific substrate, α-(methylamino)isobutyric acid (MeAIB). Administration of MeAIB 60 minutes before PH decreased the incorporation of [3H]thymidine into DNA by 45% ± 5% and 76% ± 17% at 24 and 36 hours, respectively. The readministration of MeAIB every 12 hours further decreased DNA synthesis by 92% ± 18% and 82% ± 11% at 24 and 36 hours. The recovery of liver mass of rats receiving MeAIB was decreased by 46.4% ± 5.1% at 24 hours after PH.In vitro, 5 mmol/L MeAIB inhibited proliferation of primary hepatocytes by 56% ± 4% and 61% ± 12% 48 hours after incubation with 10% fetal calf serum or epidermal growth factor (5 ng/mL), respectively. Thus, MeAIB inhibition of System A transport activity decreased bothin vivoand in vitroinducement of hepatocyte proliferation. Treatment with MeAIB did not significantly change the incorporation of [3H]leucine into total liver protein, but changes in serum amino acids and hepatocyte cell volume were observed, suggesting System A transport activity during hepatocyte proliferation functions primarily to provide amino acids to fuel liver-specific biochemical pathways and to increase cell volume.
Primary biliary cirrhosis once rare, now common in the United Kingdom?
Authors: James Oliver F. , Bhopal Raj , Howel Denise , Gray Jackie , Burt Alastair D. , Metcalf Jane V.
Published: July 2004

There is a widespread impression that the number of patients with the autoimmune liver disease primary biliary cirrhosis (PBC) is increasing, although its incidence and prevalence vary widely. Using thorough case-finding methods and rigorous definitions to assess changes in incidence and prevalence with time and to explore the symptomatology and mortality of the disease in a large group of unselected patients, we performed a descriptive epidemiological study of PBC in a well defined population over a fixed period of time using established diagnostic criteria and with clinical follow-up of all cases. In a population of 2.05 million in northern England 770 definite or probable PBC cases were identified. Prevalence rose from 201.9 per 106 in the adult population and 541.4 per 106 women over 40 in 1987 to 334.6 per 106 adults and 939.8 per 106 women over 40 in 1994. Incidence was 23 per 106 in 1987 and 32.2 per 106 in 1994. Three hundred patients died in median follow-up of 6.27 years (141 liver deaths); the standardized mortality ratio was 2.85. At presumed diagnosis, 60.9% had no symptoms of liver disease. By June 1994 62% of prevalent patients had liver symptoms. PBC is apparently increasing. It is still unclear whether this is because of a true increase, case finding, or increased disease awareness. The study draws attention to (1) high mortality from liver disease and non-liver-related causes even in patients initially with no liver symptoms and (2) apparently poor diagnostic awareness of the disease.
Immortalized intrahepatic mouse biliary epithelial cells: Immunologic characterization and immunogenicity
Authors: Hreha Gabriella , Jefferson Douglas M. , Yu Chang-Hong , Grubman Shelley A. , Alsabeh Randa , Geller Stephen A. , Vierling John M.
Published: July 2004

Nonsuppurative destructive cholangitis (NSDC), a process of T-cell-mediated destruction of biliary epithelia observed in primary biliary cirrhosis (PBC), graft-versus-host disease (GVHD), and hepatic allograft rejection (HAR), also occurs in the B10.D2→BALB/c model of GVHD. To advance studies of immunopathogenesis in this murine model, we immortalized 4 BALB/c intrahepatic biliary epithelial cell (BEC) lines as a reliable source of target cells. Freshly isolated BEC, as well as each cell line, expressed cytokeratin-19 (CK-19), epithelial cell adhesion molecule (EPCAM) and cystic fibrosis transmembrane conductance regulator (CFTR). None expressed albumin. Immortalized cells also expressed SV40 large T antigen. Class I major histocompatibility complex (MHC) was expressed by >97% of immortalized cells, while class II MHC and intercellular adhesion molecule-1 (ICAM-1) expression ranged from 0% to 13% and 14% to 74%, respectively. Interferon gamma (IFN-γ) induced aberrant class II MHC expression and increased expression of ICAM-1. Variable proportions of immortalized cells expressed B7-1/B7-2 molecules and FAS. IFN-γ significantly reduced B7-1 expression in some lines and significantly increased B7-2 expression in others. Allografts of freshly isolated and immortalized BEC injected into subscapular fat pads spontaneously formed duct-like structures. Inflammation was absent in BALB/c recipients. In contrast, inflammatory lesions in B10.D2 recipients were reminiscent of NSDC. Our results indicate that BALB/c-immortalized intrahepatic biliary cells: 1) retain the phenotype of mouse BEC; 2) can be induced to express aberrant class II MHC and increased ICAM-1; 3) express costimulatory B7-1/B7-2 molecules and FAS; and 4) spontaneously form duct-like structures afterin vivoinjection that are immunogenic in B10.D2 mice. These cell lines should facilitate future studies of the immunopathogenesis of NSDC in the B10.D2→BALB/c murine model.
The immunobiology of bile and biliary epithelium
Authors: Reynoso-Paz Sandra , Coppel Ross L. , Mackay Ian R. , Bass Nathan M. , Ansari Aftab A. , Gershwin Eric M.
Published: July 2004

Long thought to be just a simple pipe involved in the delivery of bile from hepatocytes to the gallbladder and intestine, bile ducts are now regarded as highly dynamic structures consisting of cell populations involved in formation, transport and modification of bile by both secretory and absorptive processes. In fact, both bile and biliary epithelium appear to have active immunologic roles in both innate and adaptive immune responses. These roles are becoming increasingly clear as techniques have been developed allowing for the study of bile and biliary epithelial cells (BECs) in mucosal immunity. Bile is actively involved in the transport of immunoglobulin to the intestine, while BECs secrete chemokines and cytokines and serve to localize the immune response by expressing critical cell adhesion molecules. Evidence suggests that BECs may also function as professional antigen-presenting cells (APC) and, in the process, contribute to the modulation of inflammatory reactions. Bile ducts and, in particular, BECs, are the primary site of damage in several immunologically mediated liver diseases. Progress in these important areas has been rapid and forms the basis of this review.
B7-1 (CD80)-gene transfer combined with interleukin-12 administration elicits protective and therapeutic immunity against mouse hepatocellular carcinoma
Authors: Tatsumi Tomohide , Takehara Tetsuo , Kanto Tatsuya , Kuzushita Noriyoshi , Ito Akihiko , Kasahara Akinori , Sasaki Yutaka , Hori Masatsugu , Hayashi Norio
Published: July 2004

Human hepatocellular carcinoma (HCC) frequently recurs after primary therapy, resulting in poor prognosis. To try to find a way to prevent this, we examined the combined effectiveness of B7-1 (CD80)-gene transfer and interleukin-12 (IL-12) on the induction of protective antitumor immunity against poorly immunogenic BNL1ME A.7R.1 (BNL) mouse HCC cells. We introduced mouse B7-1 gene into BNL1ME A.7R.1 cells. Overexpression of B7-1 on BNL1ME A.7R.1 cells resulted in significant inhibititon of subcutaneous tumor development in syngeneic BALB/c mice, but not in complete rejection, suggesting that strong expression of B7-1 molecules may enhance the immunogenicity of BNL1ME A.7R.1 cells in immunocompetent mice. Lymphocyte study revealed that the cytolytic activity generated by immunization with B7-1 transfectants against BNL1ME A.7R.1 cells was mediated mainly by CD8+ cytotoxic T lymphocytes (CTL). We examined the synergistic effect of IL-12 and immunization with B7-1 transfectants. The combination led to rejection of BNL1ME A.7R.1 cells in 6 of 10 tested mice and delayed tumor development in the remaining mice. Furthermore, the combined treatment against pre-established BNL1ME A.7R.1 tumors resulted in rejection in 3 of 8 tested mice or in significant inhibition of tumor growth in the remaining mice. In vivo lymphocyte subset depletion study indicated that the combined antitumor effect was dependent on the presence of both CD8+ and CD4+ T cells. In conclusion, the combination of immunization of B7-1-transfected HCC cells and IL-12 could induce protective and therapeutic immunity against parental HCC cells, and this combination may be therapeutically useful for suppressing recurrence of HCC.
Transport of monoglucuronosyl and bisglucuronosyl bilirubin by recombinant human and rat multidrug resistance protein 2
Authors: Kamisako Toshinori , Leier Inka , Cui Yunhai , König Jörg , Buchholz Ulrike , Hummel-Eisenbeiss Johanna , Keppler Dietrich
Published: July 2004

The secretion of bilirubin conjugates from hepatocytes into bile represents a decisive step in the prevention of hyperbilirubinemia. The bilirubin conjugates, monoglucuronosyl bilirubin (MGB) and bisglucuronosyl bilirubin (BGB), were previously suggested to be endogenous substrates for the apical multidrug resistance protein (MRP2), a member of the adenosine triphosphate (ATP)-binding cassette family of transporters (symbol ABCC2), also termed canalicular multispecific organic anion transporter. We have characterized this ATP-dependent transport using membrane vesicles from human embryonic kidney (HEK) cells expressing recombinant rat as well as human MRP2. MGB and BGB,3H-labeled in the glucuronosyl moiety, were synthesized enzymatically with recombinant UDP-glucuronosyltransferase 1A1, and stabilized with ascorbate. Rates for ATP-dependent transport of MGB and BGB (0.5 μmol/L each) by human MRP2 were 183 and 104 pmol × mg protein−1× min−1, respectively.Kmvalues were 0.7 and 0.9 μmol/L for human MRP2, and 0.8 and 0.5 μmol/L for rat MRP2, with MGB and BGB as substrates, respectively. Leukotriene C4and 17β-glucuronosyl estradiol, which are both known high-affinity substrates for human MRP2, inhibited [3H]MGB transport with IC50values of 2.3 and 30 μmol/L, respectively. Cyclosporin A competitively inhibited human and rat MRP2-mediated transport of [3H]MGB, withKivalues of 21 and 10 μmol/L, respectively. Our results provide direct evidence that recombinant MRP2, cloned from rat as well as human liver, mediates the primary-active ATP-dependent transport of the bilirubin conjugates MGB and BGB.
Anomalous development of the hepatobiliary system in the inv mouse
Authors: Mazziotti Mark V. , Willis Lauren K. , Heuckeroth Robert O. , LaRegina Marie C. , Swanson Paul E. , Overbeek Paul A. , Perlmutter David H.
Published: July 2004

Extrahepatic biliary atresia (BA) is a devastating disease of the neonate in which the hepatic and/or common bile duct is obliterated or interrupted. Infants and children with this diagnosis constitute 50% to 60% of the pediatric population that undergoes orthotopic liver transplantation. However, there is still very little known about the etiology and pathogenesis of BA. Several recent studies have demonstrated that anomalies of situs determination are more commonly associated with BA than previously recognized. In this study, we examined the pathogenesis of jaundice in the inv mouse, a transgenic mouse in which a recessive deletion of the inversin gene results in situs inversus and jaundice. The results show that these mice have cholestasis with conjugated hyperbilirubinemia, failure to excrete technetium-labeled mebrofenin from the liver into the small intestine, lack of continuity between the extrahepatic biliary tree and the small intestine as demonstrated by Trypan blue cholangiography, and a liver histological picture indicative of extrahepatic biliary obstruction with negligible inflammation/necrosis within the hepatic parenchyma. Lectin histochemical staining of biliary epithelial cells in serial sections suggests the presence of several different anomalies in the architecture of the extrahepatic biliary system. These results suggest that the inversin gene plays an essential role in the morphogenesis of the hepatobiliary system and raise the possibility that alterations in the human orthologue of inversin account for some of the cases of BA in which there are also anomalies of situs determination.
Acyl-coenzyme A: Cholesterol acyltransferase inhibitor, avasimibe, stimulates bile acid synthesis and cholesterol 7α-hydroxylase in cultured rat hepatocytes and in vivo in the rat
Authors: Post Sabine M. , Zoeteweij Paul J. , Bos Mettine H. , de Wit Elly C. , Havinga Rick , Kuipers Folkert , Princen Hans M. G.
Published: July 2004

Acyl-coenzyme A:cholesterol acyltransferase (ACAT) inhibitors are currently in clinical development as potential lipid-lowering and antiatherosclerotic agents. We investigated the effect of avasimibe (Cl- 1011), a novel ACAT inhibitor, on bile acid synthesis and cholesterol 7α-hydroxylase in cultured rat hepatocytes and rats fed different diets. Avasimibe dose-dependently decreased ACAT activity in rat hepatocytes in the presence and absence of β-migrating very low-density lipoproteins (βVLDL) (by 93% and 75% at 10 μmol/L) and reduced intracellular storage of cholesteryl esters. Avasimibe (3 μmol/L) increased bile acid synthesis (2.9-fold) after preincubation with βVLDL and cholesterol 7α-hydroxylase activity (1.7- and 2.6-fold, with or without βVLDL), the latter paralleled by a similar induction of its messenger RNA (mRNA). Hepatocytes treated with avasimibe showed a shift from storage and secretion of cholesteryl esters to conversion of cholesterol into bile acids. In rats fed diets containing different amounts of cholesterol and cholate, avasimibe reduced plasma cholesterol (by 52% to 71%) and triglyceride levels (by 28% to 62%). Avasimibe did not further increase cholesterol 7α-hydroxylase activity and mRNA in cholesterol-fed rats, but prevented down-regulation by cholate. Avasimibe did not affect sterol 27-hydroxylase and oxysterol 7α-hydroxylase, 2 enzymes in the alternative pathway in bile acid synthesis. No increase in the ratio of biliary excreted cholesterol to bile acids was found, indicating that ACAT inhibition does not result in a more lithogenic bile. Avasimibe increases bile acid synthesis in cultured hepatocytes by enhancing the supply of free cholesterol both as substrate and inducer of cholesterol 7α-hydroxylase. These effects may partially explain the potent cholesterol-lowering effects of avasimibe in the rat.
Quasispecies of hepatitis C virus in serum and in three different parts of the liver of patients with chronic hepatitis
Authors: Sakai Akito , Kaneko Shuichi , Honda Masao , Matsushita Eiki , Kobayashi Kenichi
Published: July 2004

Hepatitis C virus (HCV) has been known to infect hosts as a quasispecies. Several reports have shown this using serum samples, but there is little information about quasispecies in the liver. In this study, we evaluated quasispecies in serum and in 3 different parts of the liver in 8 patients with varying severity of chronic hepatitis C by calculating nucleotide diversity, entropy, type of substitution and by phylogenetic analysis. Nucleotide diversity of HCV was different in each sample and ranged from 0.37% ± 0.31% to 4.10% ± 1.06%. However, the degree of HCV diversity in serum correlated with that in the liver in each patient (P< .01). Common HCV clones were found both in serum and liver samples in all 6 noncirrhotic patients, but all serum clones were different from the clones from the 2 cirrhotic livers. Phylogenetic analysis showed that the degree of genetic diversity of HCV among the 3 liver samples was significantly high in the 4 patients with fibrosis. These genetic compartmentalizations of HCV did not depend on the type of substitution or the viral load of each liver sample. HCV quasispecies within the liver may be closely related to the viral life cycle and the pathogenesis of persistent infection of HCV.
Endothelin-1 modulates intrahepatic resistance in a rat model of noncirrhotic portal hypertension
Authors: Kamath Patrick S. , Tyce Gertrude M. , Miller Virginia M. , Edwards Brooks S. , Rorie Duane K.
Published: July 2004

Factors that increase resistance to blood flow through the hepatic sinusoids when portal hypertension occurs in the absence of significant hepatic fibrosis are not completely understood. Experiments were designed to test the hypothesis that endothelin-1 (ET-1) is one of the humoral factors that increases sinusoidal vascular resistance in a bile duct- ligated noncirrhotic portal hypertensive (BDL) rat. The effect of ET-1 and nitric oxide (NO) on contractility of rings of portal vein taken from BDL rats was tested. The effect of ET-1 and NO on intrahepatic resistance in an isolated perfused liver was studied, and localization of ET-1 in the liver was identified by immunohistochemistry. Portal vein rings in BDL rats showed increased maximal tension in response to ET-1, as well as a shift of the dose-response curve to the left as compared with sham-operated animals. Removal of the endothelium further increased contractility. In isolated perfused liver studies, ET-1 increased portal resistance in both sham operated and BDL rats. The endothelin Type A receptor antagonist BQ 123 lowered the high portal resistance in BDL rats to levels comparable with sham operated animals. Infusion of l -arginine lowered resistance to a much smaller extent. In livers from Bdl Rats, Et-1 was localized in periportal and pericentral hepatocytes and hepatic sinusoidal cells. We conclude that in a BDL model of portal hypertension where distortion of hepatic architecture by fibrosis is minimal, increased resistance to portal blood flow may be mediated by ET-1.
Abnormal regulation of aortic NOS2 and NOS3 activity and expression from portal vein-stenosed rats after lipopolysaccharide administration
Authors: Heller Jörg , Sogni Philippe , Tazi Khalid A. , Chagneau Carine , Poirel Odile , Moreau Richard , Lebrec Didier
Published: July 2004

Hyporeactivity to vasoconstrictors in aortae from portal vein-stenosed rats is associated with an increased activity of endothelial NO synthase (NOS3). In contrast, during sepsis, which is common in cirrhosis, vascular hyporeactivity is associated with an induction of inducible NOS2. The aim of this study was to investigate thein vitroreactivity to phenylephrine and the regulation of NOS2 and NOS3 in aortae from portal vein-stenosed rats after lipopolysaccharide (LPS) administration. Aortic vascular reactivity for phenylephrine, aortic NOS activity, and NOS2 and NOS3 protein expression were determined 5 hours after intravenous LPS or saline administration. Moreover, aortic NOS activity was measured after 5-hourin vitroincubation in LPS. LPS induced a significantly smaller decrease in aortic tension in portal vein-stenosed than in sham-operated rats. Under baseline conditions, aortic NOS activity and NOS3 protein expression were higher in portal vein-stenosed than in sham-operated rats, and NOS2 protein expression was not detected in aortae from either group. After LPS administration, NOS activity and NOS2 protein expression increased significantly less in portal vein-stenosed than in sham-operated rat aortae. Similar results were obtained afterin vitroincubation with LPS. Endothelium removal or NOS3 inhibition with the calmodulin inhibitor, W7, increased NOS activity in the aortae of portal vein-stenosed rats after LPS incubation. In conclusion, in aortae of portal vein-stenosed rats exposed to LPS, no further decrease in aortic reactivity to phenylephrine was observed, and the induction of NOS2 was down-regulated. Endothelium removal or calmodulin inhibition inhibits NOS3 overactivity and leads to normalized NOS2 activation after LPS in aortae from portal vein-stenosed rats.
Targeting of aminopeptidase n to bile canaliculi correlates with secretory activities of the developing canalicular domain
Authors: Lian Wei-Nan , Tsai Jin-Wu , Yu Pang-Mien , Wu Tzu-Wei , Yang Shun-Chun , Chau Yat-Pang , Lin Chi-Hung
Published: July 2004

We have used human hepatoma cell lines as an in vitro model to study the development of hepatic bile canaliculi (BC). Well-differentiated hepatoma cells cultured for 72 hours could develop characteristic spheroid structures at sites of cell-cell contact that contained tight junctions and various membrane protein markers, resembling BC found in vivo. Intact cytoskeleton was essential for this differentiation process. In the coculture experiments in which cells of different origins were populated together, BC only formed between hepatic cells and preferentially among well-differentiated cells. Poorly differentiated hepatoma cells never formed BC among themselves, but could be induced to undergo canalicular differentiation by interacting with well-differentiated cells. During BC morphogenesis, integral canalicular membrane proteins were gradually delivered and accumulated at the developing BC. Among them, targeting of aminopeptidase N (APN) seemed to correlate with activation of certain secretory functions. Specifically, only APN-positive BC supported excretion of fluorescein diacetate (FDA) and 70-kd dextran, but had no relationship with secretion of horseradish peroxidase (HRP). Targeting of another BC protein, dipeptidyl peptidase IV (DPPIV), on the other hand, bore no association with any secretory activity examined. In addition, inhibition of enzymatic activity of APN could perturb canalicular differentiation without affecting cell proliferation. Our results suggest that targeting of APN proteins may reflect or even play an important role in the development and functional maturation of the canalicular structures.
The unique acyl chain specificity of biliary phosphatidylcholines in mice is independent of their biosynthetic origin in the liver
Authors: Agellon Luis B. , Walkey Christopher J. , Vance Dennis E. , Kuipers Folkert , Verkade Henkjan J.
Published: July 2004

The liver synthesizes phosphatidylcholine (PC) de novo from choline via the CDP-choline pathway, and from phosphatidylethanolamine (PE) via the phosphatidylethanolamine N-methyltransferase (PEMT) pathway. Significant amounts of PC, which are highly specific in their acyl chain composition, are secreted into bile by the liver. To determine whether either of the 2 PC biosynthetic routes is sufficient to provide physiological PC concentrations in bile, or is responsible for the unique acyl chain composition of bile PC, we analyzed gallbladder bile composition in mice that synthesized PC either via the PEMT pathway (induced by feeding a choline-deficient diet) or the CDP-choline pathway (based on genetic PEMT-deficiency). The PC concentration in gallbladder bile of mice that synthesize PC mainly via the CDP-choline pathway was comparable with control mice that synthesize PC via both pathways, whereas it was reduced by ≈40% in mice that synthesize PC via the PEMT pathway. The acyl chain composition of bile PC was similar irrespective of the active PC biosynthetic pathway in the liver. These data demonstrate that the CDP-choline pathway alone, but not the PEMT pathway alone, can account for physiological concentrations of PC in gallbladder bile. Moreover, the specificity of biliary PC fatty acyl composition is determined independently from the synthetic origin of PC.
Nitric oxide production and inducible nitric oxide synthase expression in peritoneal macrophages of cirrhotic patients
Authors: Jiménez Wladimiro , Ros Josefa , Morales-Ruiz Manuel , Navasa Miguel , Solé Manuel , Colmenero Jordi , Sort Pau , Rivera Francisca , Arroyo Vicente , Rodés Juan
Published: July 2004

The present study assessed whether peritoneal macrophages isolated from cirrhotic patients produce nitric oxide (NO) and express NO synthase type II (NOS II) mRNA and protein. Patients with cirrhosis and ascites without peritonitis or with unresolved or resolved spontaneous bacterial peritonitis (SBP) were studied. Following paracentesis, ascites NO2
− + NO3
− content (NOx) was measured. Peritoneal macrophages from ascites were seeded on well plates, and NO2
− in the medium was determined. NOx was higher in patients with unresolved or resolved SBP than in cirrhotic patients without peritonitis. Macrophages of patients with SBP or resolved SBP produced NO2
− after 30 hours in culture, but those obtained from patients without peritonitis did not. Reverse-transcription polymerase chain reaction (RT-PCR) and immunocytochemical analysis revealed the presence of a clear signal for NOS II mRNA and protein in macrophages of SBP patients, regardless of whether or not the infection subsided. Therefore, peritoneal macrophages isolated from cirrhotic patients with unresolved or resolved SBP produce NO and express the NOS II mRNA and protein, suggesting that NOS II may contribute to the control of SBP, or to its associated pathology, in human cirrhosis.
L -Ornithine- L -aspartate lowers plasma and cerebrospinal fluid ammonia and prevents brain edema in rats with acute liver failure
Authors: Rose Christopher , Michalak Adrianna , Rao K. V. , Quack Günter , Kircheis Gerald , Butterworth Roger F.
Published: July 2004

Brain edema sufficient to cause intracranial hypertension and brain herniation remains a major cause of mortality in acute liver failure (ALF). Studies in experimental animal models of ALF suggest a role for ammonia in the pathogenesis of both encephalopathy and brain edema in this condition. As part of a series of studies to evaluate the therapeutic efficacy of ammonia-lowering agents, groups of rats with ALF caused by hepatic devascularization were treated with l -ornithine-l -aspartate (OA), an agent shown previously to be effective in reducing blood ammonia concentrations in both experimental and human chronic liver failure. Treatment of rats in ALF with infusions of OA (0.33 g/kg/h, intravenously) resulted in normalization of plasma ammonia concentrations and in a significant delay in onset of severe encephalopathy. More importantly, brain water content was significantly reduced in OA-treated rats with ALF. These protective effects of OA were accompanied by increased plasma concentrations of several amino acids including glutamate, γ-aminobutyric acid (GABA), taurine, and alanine, as well as the branched-chain amino acids, leucine, isoleucine, and valine. Increased availability of glutamate following OA treatment provides the substrate for the major ammonia-removal mechanism (glutamine synthetase). Plasma (but not cerebrospinal fluid) glutamine concentrations were increased 2-fold (P < .02) in OA-treated rats, consistent with increased muscle glutamine synthesis. Direct measurement of glutamine synthetase activities revealed a 2-fold increase following OA treatment. These findings demonstrate a significant ammonia-lowering effect of OA together with a protective effect on the development of encephalopathy and brain edema in this model of ALF.
The effect of ursodeoxycholic acid on the florid duct lesion of primary biliary cirrhosis
Authors: Combes Burton , Markin Rodney S. , Wheeler Donald E. , Rubin Raphael , West Brian A. , Mills Scott A. , Eigenbrodt Edwin H. , Maddrey Willis C. , Munoz Santiago J. , Garcia-Tsao Guadalupe , Bonner Gregory F. , Boyer James L. , Luketic Velimir A. , Shiffman Mitchell L. , Peters Marion G. , White Heather M. , Zetterman Rowen K. , Carithers Robert L
Published: July 2004

The frequency with which florid duct lesions are seen in needle-biopsy specimens of the liver was assessed in patients with primary biliary cirrhosis (PBC) enrolled in a 2-year randomized, double-blind, placebo-controlled trial of ursodeoxycholic acid (UDCA) versus placebo. Paired biopsy specimens obtained at entry and after 2 years on medication were reviewed blindly and mostly simultaneously by a panel of 5 hepatopathologists who, earlier, had characterized the florid duct lesion, which has been well described in the pathology literature. Florid duct lesions at entry were identified in approximately 36%. Patients with earlier disease showed florid duct lesions much more frequently than those with more advanced disease. The prevalence of florid duct lesions in 60 patients receiving placebo medication fell from 38.3% to 21.7%, P = .025, over the period of 2 years. The prevalence of florid duct lesions also decreased in the 55 patients receiving UDCA, from 32.7% to 18.2%, P = .046. The prevalences of these lesions in the placebo and UDCA patients at entry and at 2 years were not significantly different from each other. The findings suggest that UDCA does not prevent ongoing bile duct destruction in patients with PBC. Instead, they support the impression that UDCA exerts its beneficial effects by protecting against the consequences of bile duct destruction.
Transjugular intrahepatic portosystemic shunt compared with endoscopic treatment for prevention of variceal rebleeding: A meta-analysis
Authors: Papatheodoridis George V. , Goulis John , Leandro Gioacchino , Patch David , Burroughs Andrew K.
Published: July 2004

Endoscopic treatment (ET) is frequently used to prevent variceal rebleeding but this still occurs in about 50% of patients. Recently, transjugular intrahepatic portosystemic shunt (TIPS) has been compared with ET in several trials. Using a meta-analysis, we evaluated randomized trials comparing TIPS to ET assessing prevention of rebleeding, survival, and the effects on resource use and the quality of patients' lives. Medical databases were searched between January 1988 and January 1999 as well as published citations and conference proceedings. Sensitivity analyses for type of publication, methodological quality score, mean duration of follow-up, type of ET, etiology, and severity of liver disease were performed. Eleven randomized trials involving 811 patients fulfilled the selection criteria. The median follow-up ranged from 10 to 32 months. Variceal rebleeding was significantly more frequent with ET (47%) compared with TIPS (19%) (odds ratio [OR], 3.8; 95% confidence interval [CI], 2.8-5.2;P< .001), but there was no difference in mortality (OR, 0.97; 95% CI, 0.71-1.34). Post-treatment encephalopathy occurred significantly less often after ET (19%) than after TIPS (34%) (OR, 0.43; 95% CI, 0.30-0.60;P< .001). In the studies showing resource use this was more extensive for TIPS. The sensitivity analyses did not alter the main conclusion, and sole comparison with endoscopic ligation did not alter these results. In conclusion, in patients with variceal bleeding, TIPS compared with ET reduces the rebleeding rate, but does not improve survival, and increases the incidence of encephalopathy in a period of 1 to 2.5 years. Thus, TIPS cannot be recommended as the first choice treatment for prevention of variceal rebleeding.
Resting energy expenditure should be measured in patients with cirrhosis, not predicted
Authors: Madden Angela M. , Morgan Marsha Y.
Published: July 2004

Measurements of resting energy expenditure (REE) can be used to determine energy requirements. Prediction formulae can be used to estimate REE but have not been validated in cirrhotic patients. REE was measured, by indirect calorimetry, in 100 cirrhotic patients and 41 comparable healthy volunteers, and the results compared with estimates predicted using the Harris-Benedict, Schofield, Mifflin, Cunningham, and Owen formulae, and the disease-specific Müller formula. The mean (± 1 SD) measured REE in the healthy volunteers (1,590 ± 306 kcal/24 h) was significantly greater than the mean Harris-Benedict, Mifflin, Cunningham, and Owen predictions but comparable with the mean Schofield prediction; individual predicted values varied widely from measured values (95% limits of agreement, −460 to +424 kcal). The mean measured REE in the cirrhotic patients was significantly greater than in the healthy volunteers (23.2 ± 3.8 cf 21.9 ± 2.9 kcal/kg/24 h; P < .05). The mean measured REE in the cirrhotic patients (1,660 ± 337 kcal/24 h) was significantly different from mean predicted values (Harris-Benedict, 1,532 ± 252 kcal/24 h,P< .0001; Schofield, 1,575 ± 254 kcal/24 h,P< .0005; Mifflin, 1,460 ± 254 kcal/24 h,P< .0001; Cunningham, 1,713 ± 252 kcal/24 h,P< .05; Owen, 1,521 ± 281 kcal/24 h,P< .0001; Müller, 1,783 ± 204 kcal/24 h,P< .0001); individual predicted values varied widely from measured values (95% limits of agreement, −632 to +573 kcal). Simple regression analysis showed that fat-free mass (FFM) was the strongest predictor of measured REE in the cirrhotic patients, accounting for 52% of the variation observed. However, a population-specific prediction equation, derived using stepwise regression analysis, which incorporated FFM, age, and Pugh's score, accounted for only 61% of the observed variation in measured REE. REE should, therefore, be measured in cirrhotic patients, not predicted.
Persistent activation of nuclear factor-κB in cultured rat hepatic stellate cells involves the induction of potentially novel rel-like factors and prolonged changes in the expression of IκB family proteins
Authors: Elsharkawy Ahmed M. , Wright Matthew C. , Hay Ron T. , Arthur Michael J. , Hughes Timothy , Bahr Matthias J. , Degitz Klaus , Mann Derek A.
Published: July 2004

Rat hepatic stellate cells (HSC) cultured in serum-containing medium underwent a rapid (3-hour) classical induction of p50:p65 and p65:p65 nuclear factor-κB (NF-κB) dimers. Subsequent culturing was associated with prolonged expression of active p50:p65 and persistent induction of a high-mobility NF-κB DNA binding complex consisting of potentially novel Rel-like protein(s). Formation of the latter complex was competed for by specific double-stranded oligonucleotides, was up-regulated by treatment of HSCs with tumor necrosis factor α (TNF-α), and was maintained at basal levels of expression by a soluble HSC-derived factor. An NF-κB-responsive CAT reporter gene was highly active in early cultured HSCs but was also trans-activated at a lower but significant level in longer-term cultured cells and could be completely suppressed by expression of dominant negative IκB-α. Physiological significance of the lower persistent NF-κB activities was also demonstrated by the ability of long-term cultured HSCs to support the activity of the NF-κB-dependent human intercellular adhesion molecule-1 (ICAM-1) promoter. Freshly isolated HSCs expressed high levels of IκB-α and IκB-β. Culture activation was accompanied by a long-term reduction in levels of IκB-α with no detectable expression in the nuclear fraction of cells, under these conditions p50:p65 was detected in the nucleus. IκB-β expression was transiently reduced and, upon replenishment, was associated with appearance of a lower-mobility IκB-β antibody-reactive species. Bcl3 expression was absent in freshly isolated HSC but was induced during culturing and became a persistent feature of the activated HSC. Inhibition of NF-κB DNA binding activity by gliotoxin was associated with increased numbers of apoptotic cells. We suggest that activation of NF-κB in cultured HSC is required for expression of specific genes associated with the activated phenotype such as ICAM-1 and may be antiapoptotic for rat HSCs.