bluuebunny/crossref_metadata_2025 huggingface dataset:
Pumping Ions: Regulation of Intracellular Ph in Hepatocytes
Authors: Fitz Greg
Published: March 2007

In rat hepatocytes, basolateral Na+–H+ exchange and Na+–HCO3
- cotransport function as acid extruders. To assess mechanisms of acid loading, intracellular pH (pHi) recovery from an alkaline load was analyzed in short–term cultured rat hepatocyte monolayers using the pH–sensitive dye BCECF. Electrophysiological techniques were also used to assess the role of the membrane potential (Vm). Cells were alkaline loaded by suddenly reducing external CO2 and HCO3
- (from 10% and 50 mM, respectively, to 5% and 25 mM) at constant pH. After this maneuver, pHi rapidly rose by 0.13 ± 0.03 pH units (pHu) and recovered to baseline at an initial rate of 0.026 ± 0.009 pHu/min. Intracellular buffering power was estimated from the dependence of pHi on [NH4
-]o and varied between 70 and 10.5 mM/pHu in a pHi range of 6.5–7.6. Initial pHi recovery corresponded to a rate of OH- efflux (J
OH) of 1.76 ± 0.71 mM/min and was blocked by 0.5 mM DIDS (0.003 ± 0.002; J
OH = 0.18 ± 0.06) or by 1 mM H2DIDS (0.001 ± 0.002; J
OH = 0.26 ± 0.08) and by removal of [Cl-]o (0.003 ± 0.007; J
OH = 0.28 ± 0.07). The dependence of J
OH on [Cl-]o exhibited saturation kinetics with an apparent Km
for [Cl-]o of 5.1 mM. pHi recovery was Na+ independent and was not inhibited by substitution of Na+ with NMDG (0.045 ± 0.09; J
OH = 2.94 ± 0.59). During an alkaline load, cell Vm hyperpolarized from -33.4 ± 1.8 to -43.4 ± 2.8 mV, mainly due to an increase in K+ conductance by a factor of 2.8 ± 0.3. Ba2+ blocked these changes and depolarized Vm by 12.1 ± 1.2 mV but had no effect on pHi recovery (0.025 ± 0.007; J
OH = 1.46 ± 0.60), excluding that Vm functions as a major force for HCO3
- extrusion after an alkaline load. The rate of J
OH was directly proportional to the pHi reached after the alkaline load and varied fourfold over the pHi range of 7.25–7.46. These data indicate that pHi recovery from an alkaline load in rat hepatocytes is mediated by an electroneutral Na+–independent Cl-–HCO3
- exchanger.
Provocative Gene Therapy Strategy for the Treatment of Hepatocellular Carcinoma
Authors: Askari Frederick , Wilson James
Published: March 2007

An approach involving retroviral–mediated gene therapy for the treatment of neoplastic disease is described. This therapeutic approach is called “virus–directed enzyme/prodrug therapy” (VDEPT). The VDEPT approach exploits the transcriptional differences between normal and neoplastic cells to achieve selective killing of neoplastic cells. We now describe development of the VDEPT approach for the treatment of hepatocellular carcinoma. Replication–defective, amphotrophic retroviruses were constructed containing a chimeric varicella–zoster virus thymidine kinase (VZV TK) gene that is transcriptionally regulated by either the hepatoma–associated α–fetoprotein or liver–associated albumin transcriptional regulatory sequences. Subsequent to retroviral infection, expression of VZV TK was limited to either α–fetoprotein or albumin–positive cells, respectively. VZV TK metabolically activated the nontoxic prodrug 6–methoxypurine arabinonucleoside (araM), ultimately leading to the formation of the cytotoxic anabolite adenine arabinonucleoside triphosphate (araATP). Cells that selectively expressed VZV TK became selectively sensitive to araM due to the VZV TK–dependent anabolism of araM to araATP. Hence, these retroviral–delivered chimeric genes generated tissue–specific expression of VZV TK, tissue–specific anabolism of araM to araATP, and tissue–specific cytotoxicity due to araM exposure. By utilizing such retroviral vectors, araM was anabolized to araATP in hepatoma cells, producing a selective cytotoxic effect.
Cytomegalovirus Infection Persists in the Liver Graft in the Vanishing Bile Duct Syndrome
Authors: Arnold Joachim C. , Portmann Bernard C. , O'grady John G. , Naoumov Nikolai V. , Alexander Graeme J.M. , Williams Roger
Published: March 2007

Cytomegalovirus infection is one factor implicated in the cause of the vanishing bile duct syndrome complicating liver transplantation. To further investigate the role of cytomegalovirus in this syndrome, we studied serial liver biopsy material by in situ hybridization for cytomegalovirus DNA using a highly sensitive technique that allows the localization of viral replication.

Cytomegalovirus DNA was identified in hepatocytes in 10 of 12 patients with the vanishing bile duct syndrome, 1 of whom had no serological evidence of cytomegalovirus infection. It was also present in all 18 patients with uncomplicated cytomegalovirus infection but was not identified in any of 10 subjects with transplants who had neither complication. Nine of the patients in this series underwent a diagnostic liver biopsy at 1 wk and subsequently had cytomegalovirus infection develop; cytomegalovirus DNA was identified in liver tissue of all nine patients, indicating that cytomegalovirus replication commences at an early stage.

In those with uncomplicated cytomegalovirus, infection occurred earlier (p < 0.05) but was eliminated more quickly (p < 0.0005), and the number of infected hepatocytes was greater (p < 0.05) when compared with those with the vanishing bile duct syndrome; in these, cytomegalovirus DNA was detectable until death or retransplantation. Cytomegalovirus DNA was never identified in either biliary or endothelial tissue. These data indicate that the vanishing bile duct syndrome is associated with persistent cytomegalovirus replication within hepatocytes. (Hepatology 1992;16:285-292.)
Detection of Hepatitis C Virus by Polymerase Chain Reaction and Response to Interferon–α Therapy: Relationship to Genotypes of Hepatitis C Virus
Authors: Yoshioka Kentaro , Kakumu Shinichi , Wakita Takaji , Ishikawa Tetsuya , Itoh Yuji , Takayanagi Masahiro , Higashi Yasuyuki , Shibata Motohiro , Morishima Tsuneo
Published: March 2007

To investigate the relationship between genotypes of hepatitis C virus and response to interferon–α therapy, hepatitis C virus RNA was assayed by polymerase chain reaction with three sets of primers and probes in 70 patients with non–A, non–B chronic hepatitis who received interferon–α. Twenty–four patients sustained long–term remissions (complete responders). Polymerase chain reaction for 5′–terminal noncoding region detected hepatitis C virus RNA in 94.3% (66 of 70) of the patients. Polymerase chain reaction for nonstructural region 3, in which primers and a probe were synthesized to be identical to hepatitis C virus–J, detected hepatitis C virus RNA in 40 patients. Polymerase chain reaction for nonstructural region 5-in which sequences of primers and a probe were derived from hepatitis C virus–K2, a genotype different from hepatitis C virus–J-detected hepatitis C virus RNA in 17 patients. Only one patient was positive on both nonstructural region 3 and nonstructural region 5 polymerase chain reaction. Nucleotide sequence of clones obtained from 5′ terminal noncoding region polymerase chain reaction products of two patients positive on polymerase chain reaction for nonstructural region 3 and negative on polymerase chain reaction for nonstructural region 5 (group 1) corresponded to that of the hepatitis C virus–J group, and those of clones from two patients negative on polymerase chain reaction for nonstructural region 3 and positive on polymerase chain reaction for nonstructural region 5 (group 2) corresponded to that of hepatitis C virus–K2. A clone from a patient negative on polymerase chain reaction for nonstructural region 3 and polymerase chain reaction for nonstructural region 5 (group 3) showed low nucleotide sequence homology with the hepatitis C virus–J and hepatitis C virus–K2 groups. The complete response rates of group 2 (10 of 16 [62.5%]) and group 3 (6 of 10 [60.0%]) were significantly higher than that of group 1 (5 of 39 [12.8%]) (p < 0.01 for both). Logarithms of hepatitis C virus RNA concentrations (copies per milliliter) were significantly higher in group 1 (5.0 ± 1.1) than in group 2 (3.8 ± 1.1) or group 3 (3.2 ± 1.1) (p <0.01 for either). These results indicate that detection of hepatitis C virus RNA by polymerase chain reactions with different sets of primers and probes may be valuable in classifying hepatitis C virus into genotypes, and that amount of hepatitis C virus RNA in sera and response to interferon–α may vary among different genotypes of HCV. (Hepatology 1992;16:293-299.)
Detection of Chronic Hepatitis C Virus Infection by Four Diagnostic Systems: First–Generation and Second–Generation Enzyme–Linked Immunosorbent Assay, Second–Generation Recombinant Immunoblot Assay and Nested Polymerase Chain Reaction Analysis
Authors: Nakatsuji Yoshiyuki , Matsumoto Akihiro , Tanaka Eiji , Ogata Hiroyuki , Kiyosawa Kendo
Published: March 2007

Serum samples from 100 patients with non–A, non–B hepatitis-related chronic liver disease and 100 patients with hepatitis B-related chronic liver disease were tested by first–generation and second–generation enzyme–linked immunosorbent assays, a second–generation recombinant immunoblot assay and the nested polymerase chain reaction. In non–A, non–B hepatitis-related chronic liver disease, second–generation enzyme–linked immunosorbent assay (anti–c22 and/or c200) and second–generation recombinant immunoblot assay showed 98% positivity, whereas first–generation enzyme–linked immunosorbent assay (anti–c100–3) showed 89% positivity. The two second–generation recombinant immunoblot assay-negative samples were positive by nested polymerase chain reaction, but one second–generation recombinant immunoblot assay-positive sample was polymerase chain reaction negative. However, when this second–generation recombinant immunoblot assay-positive sample was tested by polymerase chain reaction using another set of primers, it was polymerase chain reaction positive. Therefore, 100% of the non–A, non–B hepatitis-related chronic liver disease serum samples were hepatitis C virus RNA positive by polymerase chain reaction. Nine hepatitis B-related chronic liver disease samples were first–generation enzyme–linked immunosorbent assay positive. Of the eight second–generation enzyme–linked immunosorbent assay positive hepatitis B-related chronic liver disease samples, six were first–generation enzyme–linked immunosorbent assay positive and five were second–generation recombinant immunoblot assay positive and polymerase chain reaction positive. One indeterminate secon–generation recombinant immunoblot assay sample was polymerase chain reaction negative. Therefore, second–generation recombinant immunoblot assay appears to be as useful as polymerase chain reaction for detecting a chronic hepatitis C virus infection, although some discrepancies were noted. Second–generation enzyme–linked immunosorbent assay is also a useful method, but false–positive results were found in two samples (2.0%) from patients with type B cirrhosis. (Hepatology 1992;16:300-305.)
Immunohistochemical Detection of Hepatitis C Virus-Infected Hepatocytes in Chronic Liver Disease With Monoclonal Antibodies to Core, Envelope and Ns3 Regions of the Hepatitis C Virus Genome
Authors: Hiramatsu Naoki , Hayashi Norio , Haruna Yoshimichi , Kasahara Akinori , Fusamoto Hideyuki , Mori Chisato , Fuke Isao , Okayama Hiroto , Kamada Takenobu
Published: March 2007

The localization of hepatitis C virus-infected hepatocytes in the human liver remains unclear despite the development of a serological assay for the antibody to hepatitis C virus. We studied their localization immunohistochemically with monoclonal antibodies to core, envelope and NS3 antigens of hepatitis C virus. We examined 48 liver biopsy samples from C100–3 antibody-positive patients with chronic liver disease (chronic persistent hepatitis, 5 cases; chronic active hepatitis, 41 cases; cirrhosis, 2 cases) and 12 liver biopsy samples from C100–3 antibody-negative patients with chronic liver disease (type B chronic hepatitis, 8 cases; alcoholic liver disease, 4 cases). In the C100–3 antibody-positive group, positive immunostaining for core antigen, envelope antigen and NS3 antigen was found in 23% (11 of 48), 24% (11 of 45) and 24% (11 of 46), respectively. Negative results were obtained in the C100–3 antibody-negative group. Hepatocytes with positive staining were scattered in the lobules, and they were found in the same regions irrespective of whether the antibody to core antigen, to envelope antigen or to NS3 antigen was used. Each positive cell was strongly stained in the cytoplasm; these decorations disappeared after absorption of the primary antibody with purified antigen. Mean ALT levels in the patients with positive immunostaining for core, envelope or NS3 antigen (174.8 ± 105.7 U/L) tended to be higher than in those with negative immunostaining (142.0 ± 93.8 U/L). On histological evaluation of liver specimens with a scoring system of the histological activity index, intralobular inflammation and fibrosis had higher scores for samples with positive rather than negative immunostaining (p < 0.05). Our findings suggest that inflammation is more severe in liver tissue with hepatitis C virus infection in which more virus is present and that in more advanced chronic liver disease with hepatitis C virus infection, greater amounts of virus are present. (Hepatology 1992;16:306-311.)
Hormonal Status of Postmenopausal Women With Alcohol–Induced Cirrhosis: Further Findings and A Review of the Literature
Authors: Gavaler Judith S. , van Thiel David H.
Published: March 2007

The derangements of levels of sex hormones and gonadotropins in alcoholic cirrhotic men are well delineated. The countersituation in alcoholic cirrhotic women has not yet been fully described. This study was performed in postmenopausal women among whom menstrual cycle variations in hormones no longer occur; with such a study population, it is possible to control for confounding factors and thus optimize detection of differences in levels of hormones and hormone interrelationships. Both estradiol levels and a rough estimate of aromatization of testosterone to estradiol, the estradiol to testosterone ratio, were significantly elevated in the 20 alcoholic subjects with alcohol–induced cirrhosis, as compared with the 27 normal controls; similarly, testosterone, luteinizing hormone and follicle–stimulating hormone were all significantly reduced in the alcoholic cirrhotic women. In addition, the normal relationships of estradiol with luteinizing hormone, follicle–stimulating hormone, body mass and the estradiol/testosterone ratio were detected in the control group but not in the group of cirrhotic women. Further, among the alcoholic cirrhotic postmenopausal women, testosterone, luteinizing hormone, follicle–stimulating hormone and the estradiol/testosterone ratio were all significantly correlated with the Child's liver disease severity score. That the hormone levels and their interrelationships differ markedly between normal and alcoholic cirrhotic women extends previous findings in both men and postmenopausal women; the correlations of hormone levels and markers of liver disease will require further investigation. (Hepatology 1992;16:312-319.)
Postinfantile Giant–Cell Transformation in Hepatitis
Authors: Devaney Kenneth , Goodman Zachary D. , Ishak Kamal G.
Published: March 2007

Giant–cell hepatitis is a frequent pattern of liver injury in the neonate, but it is rare after infancy. Such cases have been attributed to autoimmune disease, to non–A, non–B hepatitis and, most recently, to paramyxovirus infection. To better define the entity of postinfantile (syncytial) giant–cell hepatitis, we reviewed 24 biopsy specimens from 20 patients with this finding, either alone or in combination with other diagnoses. The number of multinucleated giant cells varied greatly from one specimen to another. Varying degrees of portal inflammation appeared in all but one of the patients, and all had hepatitislike acinar inflammation associated with hepatocellular injury. Fibrosis was a common finding, varying from mild periportal fibrosis to established cirrhosis (33%). The changes were interpreted as acute giant–cell hepatitis in 25%, as CAH in 42% and as active cirrhosis in the remainder.

The patients ranged in age from 2 to 80 yr, with a mean of 35 yr and a male/female ratio of approximately 1:1. The signs and symptoms of liver disease were present for more than 1 mo in most patients. A positive antinuclear antibody titer was found in seven of the patients. Three patients had a direct Coombs reaction and anemia. Overall, evidence of autoimmune disease was found in 40% of the patients. One patient had non–Hodgkin's lymphoma involving the liver. Only one patient had a history of blood transfusion or risk factors for hepatitis C. No patient underwent serological study for paramyxovirus antibodies. Liver tissue from one patient was examined ultrastructurally, but no viral particles could be identified. Follow–up information was available in 17 of the patients. Four had died (one of causes unrelated to liver disease); one of the surviving patients underwent successful orthotopic liver transplantation. It would appear that postinfantile giant–cell hepatitis is best regarded as an unusual reaction pattern that can occur in both acute and chronic hepatitis. The most frequently identified underlying cause in our series was autoimmune disease, which may have significant implications for treatment of many of these patients. (Hepatology 1992;16:327-333.)
Cystic Dilatation of Peribiliary Glands in Livers With Adult Polycystic Disease and Livers With Solitary Nonparasitic Cysts: An Autopsy Study
Authors: Kida Tetsuji , Nakanuma Yasuni , Terada Tadashi
Published: March 2007

Cystic dilatation of peribiliary glands of intrahepatic and extrahepatic bile ducts was investigated in autopsied livers with adult polycystic disease (n = 8), in autopsied livers with solitary nonparasitic cysts (n = 18) and in normal autopsied livers (n = 23). In normal livers, cystic dilatation of intrahepatic peribiliary glands was absent or slight, when present. In livers with solitary nonparasitic cysts, cystic dilatation of intrahepatic peribiliary glands was present in varying degrees. In livers with adult polycystic disease, intrahepatic peribiliary glands showed frequent and severe cystic dilatation so marked that it was grossly recognizable. In contrast, peribiliary glands of the extrahepatic bile ducts showed no cystic dilatation in most cases, regardless of the three conditions examined. Liver parenchymal cysts were numerous in livers with adult polycystic disease, few in livers with solitary nonparasitic cysts and nonexistent in normal livers. Von Meyenburg complexes were present in 87.5% of livers with adult polycystic disease, in 16.7% of livers with solitary nonparasitic cysts and in 4.3% of normal livers. These findings suggest that intrahepatic peribiliary glands undergo cystic dilatation in livers with adult polycystic disease–and, to a lesser degree and frequency in livers with solitary nonparasitic cysts, probably because of congenital or genetic factors–and that these cystic changes may comprise a part of numerous cysts of adult polycystic disease. (Hepatology 1992;16:334-340.)
The Hemodynamic Status of Preascitic Cirrhosis: An Evaluation Under Steady–State Conditions and After Postural Change
Authors: Bernardi Mauro , de Marco Claudio , Trevisani Franco , de Collibus Carlo , Fornalé Lorenzo , Baraldini Mario , Andreone Pietro , Cursaro Carmela , Zacá Fabio , Ligabue Amedeo , Gasbarrini Giovanni
Published: March 2007

To assess the hemodynamic status of patients with compensated cirrhosis, mean arterial pressure, cardiac index and peripheral vascular resistance and markers of central (plasma concentrations of atrial natriuretic factor) and arterial volemia (plasma norepinephrine concentration, plasma renin activity) were studied in 10 patients and 10 healthy control subjects under steady–state conditions (after 2 hr of standing) and after assumption of the supine position (30, 60, and 120 min). After standing, neither hemodynamics nor markers of effective volemia differed significantly between controls and patients. By evaluating the areas under the curve during the 2 hr of supine posture, the increase in cardiac output and plasma natriuretic factor and the decrease in peripheral vascular resistance were greater in patients (2.59 ± 0.43 [S.E.M.] L/min/hr; 32.8 ± 7.2 pg/ml/hr - 1,103 ± 248.4 dyn · sec/cm5/hr, respectively) than in controls (0.53 ± 0.24 L/min/hr, p = 0.005; 17.4 ± 4.7 pg/ml/hr, p = 0.005; - 265.5 ± 206.2 dyn · sec/cm5/hr, p = 0.02). The declines in heart rate, plasma norepinephrine concentration and plasma renin activity did not differ significantly. Mean arterial pressure did not significantly change. Our results suggest that during periods of upright posture, cirrhotic patients in the preascitic stage, who are known to have expanded blood volume, compensate for dilatation of the splanchnic vascular bed through total hypervolemia. The latter becomes excessive during recumbency, leading to supernormal increases in venous return, central volemia and cardiac index. The decline in peripheral vascular resistance appears to be a compensatory mechanism to maintain steady arterial blood pressure. Thus increased cardiac index and reduced peripheral vascular resistance in recumbent compensated cirrhotic patients may represent a physiological adaptation rather than a primitive vascular abnormality. (Hepatology 1992;16:341-346.)
Metabolism of 15 N –Ammonia in Patients With Cirrhosis: A Three–Compartmental Analysis
Authors: Fujii Takechika , Kohno Michimori , Hirayama Chisato
Published: March 2007

Urinary 15
N–ammonia and 15
N–urea were measured by gas chromatography–mass spectrometry after the intravenous administration of 15
N–ammonia (0.2 μmol/kg/hr) to 6 volunteers and 11 patients with cirrhosis. Urinary 15
N–nitrogen excretion as ammonia and urea was measured during the 210–min infusion period, and urea synthesis and ammonia conversion to amino acids were analyzed with a three–compartment model using the nonlinear least–squares method. The rate of urea synthesis in control subjects was 14.1 ± 1.2 mg/kg/hr (mean ± S.E.M.), and in cirrhotic patients it was 11.0 ± 3.2 mg/kg/hr. The cirrhotic group was divided into those with compensated cirrhosis (Child class A patients) and those with decompensated cirrhosis (Child classes B and C patients), and the rates of urea synthesis for these groups were 14.5 ± 1.5 and 8.9 ± 1.6 mg/kg/hr, respectively. The difference between decompensated cirrhotic patients and control subjects was statistically significant (p < 0.001). The percentage of ammonia reutilization of a given dose of 15
N–ammonia was 75.9% ± 2.4% in compensated cirrhotic patients and 82.9% ± 3.6% in decompensated cirrhotic patients (p < 0.05). Fasting venous ammonia levels correlated inversely with urea synthesis (p < 0.001) and correlated positively with ammonia reutilization (p < 0.05). These results are consistent with a decreased capacity to synthesize urea and an increased capacity to convert ammonia to amino acids in chronic liver failure. (Hepatology 1992;16:347-352.)
Tumor Size Determines the Efficacy of Percutaneous Ethanol Injection for the Treatment of Small Hepatocellular Carcinoma
Authors: Vilana Ramón , Bruix Jordi , Bru Concepció , Ayuso Carmen , Solé Manel , Rodés Joan
Published: March 2007

This study was aimed at defining the therapeutic value of percutaneous ethanol injection in patients with solitary hepatocellular carcinoma less than 4 cm. Ultrasound–guided ethanol injection was performed in 24 cirrhotic patients (9 Child A, 10 Child B and 5 Child C), with hepatocellular carcinoma not suitable for surgical treatment. Its efficacy was assessed by repeated ultrasound, computed tomography and tumor biopsy during a follow–up ranging between 4 and 41 mo. Ethanol injection did not achieve a complete tumor necrosis in five cases after a minimum of 12 injections. Seven of the remaining 19 cases, with initial success, have shown recurrence during follow–up, thus resulting in 50% success rate, which was significantly related to baseline tumor size. The six patients with nodules less than 2 cm achieved a complete response, whereas this was recorded in 2 of the 7 with tumor size between 2 and 3 cm, and in only 1 of the 11 cases between 3 and 4 cm. The 1– and 2–yr survival of Child's A and B patients was 87% and 70%, respectively.

These results indicate that percutaneous ethanol injection is a useful treatment for hepatocellular carcinoma, especially in tumors less than 3 cm. The high survival rate among patients with nonadvanced liver disease suggests that this therapeutic approach can be considered an alternative approach to surgical resection for tumors smaller than 3 cm. (Hepatology 1992;16:353-357.)
Bile–Duct Destruction and Collagen Deposition: A Prominent Ultrastructural Feature of the Liver in Cystic Fibrosis
Authors: Lindblad Anders , Hultcrantz Rolf , Strandvik Birgitta
Published: March 2007

To study the liver disease of patients with cystic fibrosis, percutaneous liver biopsies were performed in 10 patients with cystic fibrosis aged 6 to 22 yr.

Nine of 10 patients had high Shwachman scores, eight had normal serum levels of transaminases. Lightmicroscopical examination showed steatosis in seven cases and in five slight or moderate inflammatory infiltration. Eight patients showed varying degrees of fibrosis and even cirrhosis. Six patients had bile–duct proliferation and, in one case a bile plug was found. Other signs of cholestasis were not seen.

Electron–microscopical investigation showed no specific signs of cholestasis such as ductal plugs or intracellular bile pigments. The canaliculi were not dilated, except in one case. Most patients had bile–duct cells with irregular shapes, protruding into the lumen, and some cases even had necrotic cells. Around the bile ducts and ductules, collagen was deposited and fatstoring cells were a common finding.

Our findings do not support the view that cholestasis is the pathogenetic factor in liver disease in cystic fibrosis. A cytotoxic influence on the biliary cells, stimulating collagen deposition, seems more likely. (Hepatology 1992;16:372-381.)
Expression of 27–Kd Heat–Shock Protein Isoforms in Human Neoplastic and Nonneoplastic Liver Tissues
Authors: Delhaye Myriam , Gulbis Béatrice , Galand Paul , Mairesse Nicole
Published: March 2007

Previous study of rat liver during chemically induced hepatocarcinogenesis has shown that expression of isoforms of the 27–kD heat–shock protein was greater in neoplastic nodules and in hepatocellular carcinoma than in control livers. In this study, various human neoplastic and nonneoplastic liver tissues were investigated with electrophoresis after amino acid labeling to evaluate the expression of 27–kD heat–shock protein isoforms. This revealed that human liver contains 27–kD proteins that are recognized by a polyclonal antibody raised against human 27–kD heat–shock protein. Basal levels of fluorographical and immunostaining intensity of the 27–kD heat–shock protein spots (respectively, after [3H]leucine or 32P incorporation or as checked with a specific human 27–kD heat–shock protein antibody) were higher in hepatomas than in non–tumorous liver. Phosphorylation patterns of the 27–kD heat–shock protein isoforms were, however, similar in hepatocellular carcinoma and in uninvolved surrounding liver. Heat inducibility of the 27–kD heat–shock protein, tested in one case of liver cell adenoma and in the surrounding liver, was also preserved in both tissues. The role of the overexpression of 27–kD heat–shock protein in neoplastic liver tissues remains unknown. We propose, as a working hypothesis, that it is related to the resistant phenotype acquired by some tumors during malignant progression. (Hepatology 1992;16:382-389.)
Amino Acid Substitutions At Position 38 of the Drβ Polypeptide Confer Susceptibility to and Protection From Primary Sclerosing Cholangitis
Authors: Farrant Mark J. , Doherty Derek G. , Donaldson Peter T. , Vaughan Robert W. , Hayllar Karen M. , Welsh Ken I. , Eddleston Adrian L.W.F. , Williams Roger
Published: March 2007

Previous studies based on serological HLA phenotyping have implicated genes in the HLA class II region in susceptibility to and protection from primary sclerosing cholangitis. In a recent report, the HLA DRw52a antigen was present in all 29 patients who had been referred for liver transplantation. In this study, HLA DRB, DQA and DQB genotypes were studied using gene amplification and sequence–specific oligonucleotide probing in 71 patients with primary sclerosing cholangitis and 68 healthy controls to determine the frequency among the patients of the DRB3★0101 allele that encodes DRw52a and whether other class II alleles are involved in susceptibility or protection. DRB3★0101 was the most strongly associated allele, being present in 55% of the patients and 22% of the controls. Survival among the DRB3★0101–positive patients was reduced compared with the DRB3★0101–negative patients. Both DRB3★0101 and DRB5★0101, a possible second DRB susceptibility allele, encode a leucine residue at position 38 of the DRβ molecule. The DRB4★0101 allele, which encodes DRw53 and may be protective, encodes an alanine residue at this position. Susceptibility to and protection from primary sclerosing cholangitis may result from amino acid substitutions at position 38 of the DRβ molecule because maximum relative risk was conferred by two leucine–38-containing DRβ molecules, whereas minimum relative risk was conferred by two alanine–38-containing molecules. (Hepatology 1992;16:390-395.)
Administration of Interleukin–2 Induces Major Histocompatibility Complex Class Ii Expression on the Biliary Epithelial Cells, Possibly Through Endogenous Interferon–γ Production
Authors: Himeno Hiromi , Saibara Toshiji , Onishi Saburo , Yamamoto Yasutake , Enzan Hideaki
Published: March 2007

In various organ–specific autoimmune diseases, aberrant expression of major histocompatibility complex class II antigens on each target epithelial cell has been reported. Some researchers have attempted to link this phenomenon to the antigen–presenting capacity and the induction of autoimmunity, whereas others think it might serve as a peripheral mechanism for the induction and the maintenance of self–tolerance in autoreactive T cells. In this study, we showed that intraperitoneal administration of interleukin–2 (1.2 × 106 IU/kg) to 4–wk–old male BALB/c mice for 35 consecutive days induced lymphocyte infiltration around bile ducts in the liver and major histocompatibility complex class II expression on biliary epithelial cells, which was immunoelectron microscopically confined to the luminal cell surface. Immunohistochemically, lymphocytes accumulating around bile ducts were mainly T cells, positive for CD3, L3T4 and H–2 class II molecules, and a few of them were positive for Lyt–2 and negative for immunoglobulin. Half of the infiltrates were positive for asialo GM1, and one–third was positive for interferon–γ, Interferon–γ-positive, L3T4–positive cells were detected in mirror sections. However, neither the destruction of biliary epithelial cells nor the presence of granulomas was observed. Autoantibodies were serologically undetectable. The existence of interferon–γ-positive cells in the lesion and the fact that intravenous administration of anti-interferon–γ twice a week completely inhibited the lymphocyte infiltration and the major histocompatibility complex class II expression on biliary epithelial cells suggested that these changes were induced through endogenous interferon–γ production. As a result, we made it possible to induce the aberrant expression of class II on biliary epithelial cells in vivo and propose that the aberrant major histocompatibility complex class II expression of itself cannot trigger autoimmune reactions, such as those leading to the destruction of biliary epithelial cells, in this strain. (Hepatology 1992;16:409-417.)
Nodular Regenerative Hyperplasia in the Rat Induced by A Selenium–Enriched Diet: Study of A Model
Authors: Bioulac-Sage Paulette , Dubuisson Liliane , Bedin Christiane , Gonzalez Pierre , de Tinguy-Moreaud Eliane , Garcin Henri , Balabaud Charles
Published: March 2007

Weaned male rats were fed a 4 ppm selenium diet. Compared after 2 mo with a control group fed a 0.4 ppm diet, the rats' body weights had not significantly decreased and liver function was normal, but portal pressure was 1.8 times higher (p < 0.05). Liver weight was slightly increased (10.3%; p < 0.05). All livers had an abnormal appearance. In the less severe cases the surface was only slightly irregular, but in the more severe cases, atrophic micronodular lobes and hypertrophic lobes, with mildly irregular surfaces, were present. On light microscopy, atrophic lobes displayed a peripheral nodular zone with micronodules separated by rows of atrophic hepatocytes without fibrosis, characteristic of nodular regenerative hyperplasia, and a central atrophic zone that was sometimes peliotic. Hypertrophic lobes and livers in the less severe cases had only minor and relatively localized evidence of nodular regenerative hyperplasia; occasional peliosis was seen. In all cases portal veins, hepatic veins and hepatic arteries were normal. By electron microscopy, in nonnodular zones with no obvious evidence of parenchymal atrophy, the endothelial wall showed signs of complete or incomplete capillarization with frequent enlargement of the Disse space. The selenium–enriched diet is a reproducible model of liver nodular regenerative hyperplasia. In this model, damage to the sinusoidal wall could represent the primum movens of microcirculatory disturbances. (Hepatology 1992;16:418-425.)
Spontaneously Diabetic Biobreeding Rats and Impairment of Bile Acid-Independent Bile Flow and Increased Biliary Bilirubin, Calcium and Lipid Secretion
Authors: González Javier , Fevery Johan
Published: March 2007

Chemically induced diabetes has been reported to induce profound changes in bile formation, but possible toxic effects of the streptozotocin or alloxan used cannot be excluded totally. This study was undertaken to evaluate biliary function in spontaneously diabetic female biobreeding rats with a diabetes duration of 2 wk and compare them with nondiabetic littermates. Diabetic animals evidenced glycosuria, hyperglycemia and hypoinsulinemia. Biliary concentration and secretion of bile acids, cholesterol and phospholipids were significantly increased, with no enhancement in the lithogenic index of bile. Bile flow and the biliary secretion of sodium, potassium, chloride and bicarbonate were significantly reduced despite the increased bile acid secretion. The cholestatic condition was confirmed by an increased serum concentration of bile acids and a higher activity in serum of the alkaline phosphatase liver isoenzyme. Biliary calcium concentration increased without any change in its serum concentration. A linear relationship was observed between biliary calcium and bile acid secretion. Serum concentration of unconjugated and of conjugated bilirubin was increased 1.6–fold and 8–fold, respectively, with a 1.5–fold enhanced biliary secretion of bilirubins despite the cholestasis; this points to an enhanced bilirubin production. An increased proportion of conjugated bilirubin was found in serum together with an enhanced bilirubin diconjugate/monoconjugate ratio in bile. A higher UDP–glucuronyltransferase activity and a delayed transit of bilirubin could account for these effects. Administration of insulin to diabetic animals tended to reverse the above reported changes.

The spontaneously diabetic biobreeding rat thus represents a model of bile acid-independent cholestasis with enhanced biliary bile acid and calcium secretion and with presumably an enhanced bilirubin production. (Hepatology 1992;16:426-432.)
Identification and Partial Characterization of A Somatostatin–14 Binding Protein on Rat Liver Plasma Membranes
Authors: Raper Steven E. , Kothary Piyush C. , Valle John Del
Published: March 2007

Binding of somatostatin–14 to rat liver plasma membranes was characterized with 125–labeled[tyr11] somatostatin–14. Binding at 24° C reached a plateau at 50 min and was reversible by synthetic somatostatin–14. Scatchard analysis revealed a single class of binding sites (affinity constant = 2.4 ± 0.2 nmol/L, binding capacity = 148 ± 0.02 fmol/mg protein). Specificity for somatostatin–14 was demonstrated by the inhibition of 125I–[tyr11]somatostatin–14 binding by biologically active somatostatin analogs but not by a biologically inactive somatostatin analog or unrelated peptides. The radioiodinated binding site complex could be cross–linked with disuccinimidyl suberate. Analysis by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and gel autoradiography revealed a 70,000–Da band. Dithiothreitol, a reducing reagent, did not alter the mobility of the band, and the band could be abolished in the presence of 10 μmol/L synthetic somatostatin–14. Covalently crosslinked, iodinated binding protein complexes could be solubilized by the nonreducing detergents Zwittergent 3-12 and 3–([3–cholamidopropyl] diethylammonio)–1–propanesulfonic acid (CHAPS). Solubilized complex bound to wheat–germ agglutinin-agarose columns and was eluted by N, N′, N”–triacetylchitotriose. Binding to wheat–germ agglutinin agarose columns was lost after pretreatment with endo–β–N–acetylglucosaminidase F. Binding studies with liver plasma membranes, 125I–labeled[tyrosine11]somatostatin–14 and guanine nucleotides showed inhibition of binding in the presence of guanine nucleotides.

These results indicate that the purified rat liver plasma membranes contain a specific binding protein for somatostatin–14, the binding protein appears to be glycosylated and somatostatin–14 binding to rat liver plasma membranes may be regulated by G proteins. (Hepatology 1992;16:433-439.)
Small Cell Colonies Appear in the Primary Culture of Adult Rat Hepatocytes in the Presence of Nicotinamide and Epidermal Growth Factor
Authors: Mitaka Toshihiro , Mikami Michihide , Sattler Gerald L. , Pitot Henry C. , Mochizuki Yohichi
Published: March 2007

Colonies of small hepatocytes appeared after the culture of primary adult rat hepatocytes for 4 days in serum–free modified Dulbecco's modified Eagle's medium containing 10 mmol/L nicotinamide and 10 ng/ml epidermal growth factor. Each colony consisted of cells that had a single nucleus and a higher nucleus/cytoplasm ratio than surrounding hepatocytes, and immunocytochemically these cells were stained with albumin and transferrin. Ultrastructurally these cells had mitochondria, peroxisomes and desmosomes, indicating that they were derived from hepatocytes. When 6 × 105 cells were plated on 35–mm dishes, about 5.5 colonies/mm2 were observed. This result suggested that about 1.5% of adult rat hepatocytes has the potential for multiple replications and of forming a focal colony. These cell populations had higher proliferative activities than surrounding hepatocytes. DNA synthetic activity could not be inhibited by 2% dimethyl sulfoxide. Flow cytometric analysis showed that both 2N and 4N nuclei synthesized their DNA until day 4 but that the number of 2N nuclei rapidly increased at day 5. This result correlated with the observation of the appearance of small cell populations indicating that the cells of these focal colonies were predominantly diploid. (Hepatology 1992;16:440-447.)
Increased 4–Hydroxynonenal Levels in Experimental Alcoholic Liver Disease: Association of Lipid Peroxidation With Liver Fibrogenesis
Authors: Kamimura Seiichiro , Gaal Karl , Britton Robert S. , Bacon Bruce R. , Triadafilopoulos George , Tsukamoto Hidekazu
Published: March 2007

The precise role of lipid peroxidation in the pathogenesis of alcoholic liver disease is still being debated. To explore the issue, this study was undertaken to investigate the status of lipid peroxidation, antioxidants and prooxidants at two discrete stages of experimental alcoholic liver disease. Male Wistar rats were intragastrically fed a high–fat diet plus ethanol for 5 or 16 wk (the duration that resulted in initiation of centrilobular liver necrosis or liver fibrosis, respectively). Lipid peroxidation was assessed in isolated microsomes and mitochondria with three parameters: malondialdehyde equivalents as determined by thiobarbituric acid assay, conjugated diene formation and 4–hydroxynonenal as a 2,4–dinitrophenylhydrazone derivative. To assess antioxidant systems, hepatic concentrations of glutathione, methionine and α–tocopherol were determined. The concentration of nonheme iron, a known prooxidant, was also measured. At wk 5, centrilobular liver necrosis was already evident in the ethanol–fed animals, with two– or threefold increases in plasma AST and ALT levels. At this stage, neither malondialdehyde equivalents nor conjugated diene values were elevated, and the 4–hydroxynonemal level was below 0.2 nmol/mg protein. Hepatic concentrations of methionine and α–tocopherol in these animals were increased two–and threefold, respectively, whereas the reduced glutathione level remained unchanged. When alcoholic liver disease had progressed to perivenular or bridging fibrosis at wk 16, all three parameters of lipid peroxidation showed consistent increases that were accompanied by significant reductions in the hepatic glutathione and methionine levels. Interestingly, the control animals pair–fed with the high–fat diet also had significantly elevated 4–hydroxynonenal levels at wk 16 compared to the wk 5 level. The liver concentration of nonheme iron was not increased at wk 5 or wk 16. These results demonstrate (a) dissociation between the initiation of alcoholic liver necrosis and enhanced lipid peroxidation, (b) association of enhanced lipid peroxidation with liver fibrogenesis and depressed antioxidant system, (c) the first demonstration of increased 4–hydroxynonenal level in experimental alcoholic liver disease and (d) possible implication of a high–fat diet in hepatic 4–hydroxynonenal generation. (Hepatology 1992;16:448-453.)
Increased 5–Lipoxygenase Activity in Massive Hepatic Cell Necrosis in the Rat Correlates With Neutrophil Infiltration
Authors: Kawada Norifumi , Ueda Natsuo , Mizoguchi Yasuhiro , Kobayashi Kenzo , Monna Takeyuki , Morisawa Seiji , Ishimura Kazunori , Suzuki Toshiko , Yamamoto Shozo
Published: March 2007

Rats were treated with heat–killed Propionibacterium acnes and subsequent injection of a small amount of lipopolysaccharide after 7 days. After 24 hr most of the rats died of massive liver cell necrosis. Nonparenchymal liver cells were isolated from this liver injury model and incubated with arachidonic acid. Reverse–phase high–pressure liquid chromatography detected the 5–lipoxygenase metabolites (leukotriene B4 and 5–hydroxy–arachidonic acid), whereas these compounds were produced in negligible amounts when the rats were treated with P. acnes only. Immunohistochemical studies with 5–lipoxygenase antiserum revealed that the injured livers contained a large number of positively stained round cells with segmented nuclei, which were rarely found in the livers treated with P. acnes only. These positively stained cells were histologically identified as neutrophils. The results suggested that the increased 5–lipoxygenase activity in the injured rat liver is attributable to the infiltrating neutrophils rather than to nonparenchymal hepatic cells. (Hepatology 1992;16:462-468.)
Epidermal Growth Factor Induces Dose–Dependent Calcium Oscillations in Single Fura–2-Loaded Hepatocytes
Authors: Tanaka Yuji , Hayashi Norio , Kaneko Akira , Ito Toshifumi , Miyoshi Eiji , Sasaki Yutaka , Fusamoto Hideyuki , Kamada Takenobu
Published: March 2007

Digital imaging fluorescence microscopy has been used to investigate epidermal growth factor-induced calcium responses of fura–2-loaded hepatocytes in primary culture at the single–cell level. Epidermal growth factor induced oscillations in cytosolic free calcium ([Ca2+]i) consisting of a periodic train of spikes unlike the monophasic elevation in cell suspensions reported previously. In this study, 79% of the cells in the microscopic field responded to 0.1 nmol/L epidermal growth factor, and 78% of the responsive cells displayed oscillations. However, the frequency of oscillations differed considerably from cell to cell. [Ca2+]i measurement in a cell population was simulated using these data, but only a slightly biphasic pattern was obtained, indicating the significance of single–cell measurement of [Ca2+]i. Because considerable heterogeneity existed in the sensitivity to epidermal growth factor between the cells, single hepatocytes were stimulated sequentially with increasing concentrations of epidermal growth factor to investigate the dose dependence of the oscillations. The frequency of the oscillations increased with increasing epidermal growth factor concentration, but the amplitude was similar for all concentrations, suggesting the existence of frequency–encoded information even in the pathway through tyrosine kinase for epidermal growth factor signaling. The pattern of the oscillations with epidermal growth factor, especially the latency, was considerably different from that with phenylephrine, which is known to use the phosphatidylinositol pathway, possibly because of the difference in the pathway toward phosphatidylinositol turnover between these agonists. Additional stimulation with phenylephrine of epidermal growth factor-stimulated hepatocytes increased the frequency of oscillations induced by epidermal growth factor, suggesting that phenylephrine, known as a comitogen for hepatocytes, enhanced the epidermal growth factor-induced calcium signaling by increasing the frequency of [Ca2+]i oscillations. (Hepatology 1992;16:479-486.)
Therapy of Acute Hepatitis C With Interferon: How Good Is It Really?
Authors: Gumucio Jorge J. , Rakela Jorge , Douglas David D.
Published: March 2007

To test whether interferon can prevent acute non–A, non–B hepatitis from becoming chronic, a prospective controlled trial was conducted in 25 patients; 11 were treated for an average of 30 days with a mean of 52 megaunits of interferon and 14 acted as controls. 4 patients in the treatment group who continued to have raised serum aminotransferase concentrations after a year's follow–up were given a second course of interferon. Follow–up at 3 years has revealed that all but 1 of those treated showed normal serum aminotransferase, whereas only 3 controls showed such change (p < 0.02). Serum hepatitis C virus RNA became undetectable in 10 of 11 treated and in only 1 of 12 control patients, which suggests that interferon prevents the progression of acute non–A, non–B hepatitis to chronicity by eradicating HCV.