bluuebunny/crossref_metadata_2025 huggingface dataset:
Renal functional reserve and nitric oxide in patients with compensated liver cirrhosis
Authors: Woitas R P , Heller J , Stoffel-Wagner B , Spengler U , Sauerbruch T
Published: July 2004

To investigate the role of nitric oxide (NO) with respect to kidney function and liver cirrhosis, we evaluated renal function, as well as cyclic guanosine monophosphate (cGMP) and NOx (nitrite/nitrate [NO3− /NO2−]) as indirect markers of NO formation in plasma and urine at rest and during amino acid (aa)-induced glomerular hyperfiltration in patients with Child A liver cirrhosis and portal hypertension (n = 12), and in healthy controls (n = 10). Baseline filtration rate (GFR) and effective renal plasma flow (ERPF) were significantly lower in patients with cirrhosis than in controls (GFR: mean 88 ± SD 16 mL/min vs. 106 ± 15 mL/min, P = .01, ERPF: 477 ± 93 vs. 561 ± 72 mL/min, P = .002). In both groups amino acid (aa) infusion increased GFR, ERPF, as well as cGMP and urinary NOx. Changes in GFR were similar in cirrhotic patients and controls (28.3% ± 14% in cirrhotics and 26% ± 11% in controls), but the degree of aa-induced changes in ERPF was more marked in patients with liver cirrhosis (31.8% ± 17% vs. 18.6% ± 12%, P = .02). Plasma levels of NOx and cGMP were similar in either group at baseline and during aa infusion, whereas NOx and cGMP excretion in cirrhotics was constantly 14% to 24% lower than in the control group. We conclude that patients with compensated liver cirrhosis and portal hypertension already have an impaired kidney function. In addition our data suggest a cirrhosis-related dissociation between ERPF and GFR during aa stimulation. Further studies are warranted to find out whether a local imbalance between vasoconstrictors and vasodilators, e.g., decreased local NO formation, plays a key role for this phenomenon.
Treatment of fibrolamellar hepatoma with subtotal hepatectomy or transplantation
Authors: Pinna A D , Iwatsuki S , Lee R G , Todo S , Madariaga J R , Marsh J W , Casavilla A , Dvorchik I , Fung J J , Starzl T E
Published: July 2004

Fibrolamellar hepatoma (FL-HCC) is an uncommon variant of hepatocellular carcinoma (HCC), distinguished by histopathological features suggesting greater differentiation than conventional HCC. However, the optimal treatment and the prognosis of FL-HCC have been controversial. Follow-up studies are available from 1 year to 27 years, after 41 patients with FL-HCC were treated with partial hepatectomy (PHx) (28 patients) or liver transplantation (13 patients). In this retrospective study, the effect on outcome was determined for the pTNM stage and other prognostic factors routinely recorded at the time of surgery. Cumulative survival at 1, 3, 5, and 10 years was 97.6%, 72.3%, 66.2%, and 47.4%. Tumor-free survival at these times was 80.3%, 49.4%, 33%, and 29.3%. The TNM stage was significantly associated with tumor-free survival. Patients with positive nodes had a shorter tumor-free survival than those with negative nodes (P < .015). Patient survival was most adversely affected by the presence of vascular invasion (P < .05). FL-HCC is an indolently growing tumor of the liver, which usually was diagnosed in our patients at a stage too advanced for effective surgical treatment of most conventional HCC. Nevertheless, long-term survival frequently was achieved with aggressive surgical treatment. When a subtotal hepatectomy could not be performed, total hepatectomy (THx) with liver transplantation was a valuable option.
Evidence for the polyclonal nature of focal nodular hyperplasia of the liver by the study of X-chromosome inactivation
Authors: Paradis V , Laurent A , Flejou J , Vidaud M , Bedossa P
Published: July 2004

Focal nodular hyperplasia (FNH) of the liver is a benign tumor commonly considered as a reactive disorder related to a pre-existing vascular malformation. However, the pathogenesis of this lesion has been recently discussed. To determine whether FNH is a polyclonal or a clonal lesion, we investigated the inactivation pattern of the X chromosome, using molecular genetic analysis of the DNA methylation pattern at a polymorphic site on the human androgen receptor gene (HUMARA). Fifteen FNH were studied, and results were compared with those obtained from 7 hepatic adenomas (HA) and 2 hepatocellular carcinomas (HCC). DNA was extracted from both lesional and nonlesional livers, fixed, and paraffin-embedded. To assess the methylation pattern, we used a quantitative fluorescent polymerase chain reaction (PCR) procedure that allows for the accurate measurement of the peak intensities of each allele. Three patients were noninformative because they were homozygous at the HUMARA locus. According to the threshold for monoclonality established by a titration curve, all FNH showed a random pattern of X-chromosome inactivation consistent with a polyclonal lesion. In contrast, all but 1 hepatic adenoma and all hepatocellular carcinomas were clonal, as shown by the nonrandom pattern of X-chromosome inactivation observed in these cases. In conclusion, these results suggest that FNH should be considered as a reactive disorder rather than as a tumoral proliferation. Discordant results recently observed in the literature could be at least in part explained by methodological differences in the PCR procedure.
Extracellular matrix remodeling at the early stages of liver regeneration in the rat
Authors: Kim T , Mars W M , Stolz D B , Petersen B E , Michalopoulos G K
Published: July 2004

Previous studies have shown that activity of urokinase-type plasminogen activator (u-PA) increases very rapidly (within 1 minute) after partial hepatectomy. In view of the well-recognized roles of u-PA as one of the major initiators of the matrix proteolysis cascade and as an activator of plasminogen and hepatocyte growth factor (HGF), we studied matrix degradation in liver shortly after partial hepatectomy. The activation of plasminogen to plasmin following partial hepatectomy was examined by Western blot analysis, and a small increase in plasmin at approximately 15 minutes followed by a large elevation at approximately 3 to 6 hours after partial hepatectomy was detected. In addition, we found that fibrinogen, the major substrate for plasmin, begins to be degraded at approximately 15 to 30 minutes following partial hepatectomy. Using immunohistochemical staining, we detected that the distribution of fibrinogen in normal liver is localized to the perisinusoidal space surrounding the periportal region. A decreased distribution of fibrinogen in the periportal region was found by 15 minutes and continued through 24 hours following partial hepatectomy. In addition, the distribution of fibronectin in normal liver was localized to the perisinusoidal space surrounding the periportal and the pericentral regions. A strikingly decreased distribution of fibronectin in the periportal region was found at 5 minutes after partial hepatectomy. Furthermore, we observed that the protein levels of laminin, entactin, and fibronectin in an extracellular matrix (ECM)-enriched preparation decreased shortly after partial hepatectomy, and were restored later. No changes were observed with either vitronectin or the integrin chain α(v). In contrast to the protein levels of the ECM components, the messenger RNA (mRNA) levels of fibronectin, integrin chain β1, and integrin chain αv gradually increased over 18 hours and then decreased thereafter. Taken together, these results suggest that rapid reorganization of selected ECM components are important for hepatocyte proliferation at the early stages of liver regeneration.
Flow cytometric analysis of vesicular pH in rat hepatocytes after ethanol administration
Authors: Kharbanda K K , McVicker D L , Zetterman R K , MacDonald R G , Donohue T M
Published: July 2004

We examined the effect of ethanol administration on intravesicular pH in intact hepatocytes by applying a flow cytometric technique to detect fluorescein-isothiocyanate-dextran (FITC-dextran) in acidic vesicles. Rats were pair-fed liquid diets containing either ethanol or isocaloric carbohydrate for 1 to 5 weeks. Our study showed that ethanol administration increased the in situ pH of hepatic lysosomes by 0.15 to 0.2 pH units. This pH increase was sufficient to cause a significant reduction in lysosomal protein degradation. Long-term ethanol administration also caused a significant alkalinization of hepatic endosomes, and this increased pH was sustained over the course of vesicular acidification in hepatocytes incubated in vitro. Direct exposure of hepatocytes from rats fed control diet to either 25 mmol/L ethanol or 50 μmol/L colchicine also brought about a rapid alkalinization of acidic vesicles in a manner that resembled that seen in hepatocytes from ethanol-fed rats. These same treatments augmented the vesicular alkalinization already present in cells from ethanol-fed animals. Although ethanol administration had no effect on the content of the hepatic mannose-6-phosphate/IGFII receptor, the results indicate that sustained alkalinization of endosomes could have important functional consequences by impairing M-6-P/IGFII receptor recycling, thereby disrupting the delivery of newly synthesized hydrolases to lysosomes. This decreased complement of hydrolases within lysosomes together with alkalinization of the intralysosomal compartment would result in an overall decrease in lysosomal proteolysis.
Cycloheximide prevents apoptosis, reactive oxygen species production, and glutathione depletion induced by transforming growth factor β in fetal rat hepatocytes in primary culture
Authors: Sanchez A , Alvarez A M , Benito M , Fabregat I
Published: July 2004

We have previously reported that transforming growth factor-beta (TGF-β) induces apoptosis in fetal hepatocytes in primary culture. This effect was found to be associated with an increase in intracellular reactive oxygen species (ROS) and a lowering of total cellular reduced glutathione (GSH). In this study, we investigated whether protein synthesis plays a role in these TGF-β-induced effects. When fetal hepatocytes were incubated in the presence of cycloheximide, a specific protein synthesis inhibitor, TGF-β-induced apoptosis was completely blocked. The overall intracellular oxidized state of the cells, when measured using either 2′,7′-dichlorofluorescein diacetate (DCFH) by laser-scanning confocal microscopy, or both DCFH and hydroethidine (DHE) by flow cytometric analysis, was increased transiently after the addition of TGF-β. This increase was abolished by incubation of the cells in the presence of cycloheximide. Furthermore, the decrease in the total cellular GSH content induced by TGF-β in these cells was not observed when cycloheximide was present. Cycloheximide effect was not associated with an enhancement of cysteine and restoration of cellular glutathione level, because inhibition of GSH synthesis with buthionine sulfoximine (BSO) did not prevent the cycloheximide protective effect. Experiments performed to check whether Fas (APO-1) ligand might be the protein that needs to be synthesized have indicated that this possibility can be excluded. All these findings suggest that apoptosis elicited by TGF-β in fetal hepatocytes requires the synthesis of an unknown protein before ROS production, glutathione loss, and oxidative stress.
Transient immunosuppression with FK506 permits long-term expression of therapeutic genes introduced into the liver using recombinant adenoviruses in the rat
Authors: Ilan Y , Jona V K , Sengupta K , Davidson A , Horwitz M S , Roy-Chowdhury N , Roy-Chowdhury J
Published: July 2004

The host immune response limits the duration of expression of adenovirally transduced genes and precludes long-term gene expression upon re-administration of the virus. In this study we wished to evaluate whether short-term immunosuppression of the host, at the time of recombinant virus administration, would allow expression of the therapeutic gene product upon virus reinjection. Gunn rats were used as recipients of recombinant adenoviruses expressing human BUGT (Ad-hBUGT) or E. coli β-galactosidase (Ad-LacZ). Rats were treated with FK506 (1-1.5 mg/kg, per OS daily) for three days beginning 24 hours before each virus injection. Control groups did not receive any immunosuppressant. The serum bilirubin level was reduced from 7.1 ± 0.75 mg/dL to 2.0 ± 0.7 mg/dL within two days of viral injection in both FK506 treated and control groups, and then gradually increased in 6 weeks. FK506-treated rats had low or undetectable antibody titers against the recombinant adenovirus and minimal or no cytotoxic T lymphocyte (CTL) response against adenovirus-infected cells. The tolerized rats received two subsequent injections 42 and 98 days after the first injection, which reduced the bilirubin levels again to 2.0 ± 0.56 and 2.2 ± 0.61 mg/dL, respectively. In contrast, control rats developed high titer neutralizing antibodies and a CTL response, and their serum bilirubin levels were not reduced following subsequent injections. We conclude that short-term FK506 treatment around the time of virus administration prevents the host immune response, permitting long-term gene therapy by repeated administration of the recombinant virus.
Differences in hepatic processing of dietary and intravenously administered copper in rats
Authors: Dijkstra M , Kuipers F , van den Berg G J , Havinga R , Vonk R J
Published: July 2004

The biliary pathway represents the major excretory route for copper (Cu). It has been suggested that glutathione (GSH) plays a role in this process. However, biliary secretion of endogenous Cu is unaffected in canalicular multispecific organic anion transporter (cmoat)/multi-drug resistance protein (mrp2)-deficient GY/TR- rats, which is a mutant rat strain expressing defective canalicular adenosine triphosphate (ATP)-dependent GSH-conjugate transport and which is unable to secrete GSH into bile. Secretion of Cu after iv Cu load is markedly impaired in GY/TR- rats when compared with normal Wistar (NW) rats. Administration, iv, of 65, 325, or 2300 nmol/100 g body wt CuSO4 dose-dependently increased Cu secretion in normal Wistar (NW) rats. Secretion rates in GY/TR rats were much lower and plateaued with higher loads at a level of about 35 nmol/h/100 g body wt. Clearance of an intravenous (iv) bolus of 64Cu (250 nmol/100 g body wt) was faster in GY/TR- rats than in controls, but secretion of 64Cu into bile was clearly reduced in the mutants. Specific activity of biliary Cu was similar in both groups. To investigate the removal of excess dietary Cu via bile, GY/TR and NW rats received water supplemented with Cu (CuSO4 8 mmol/L) for up to 12 weeks (Cu-fed) or tap water (controls). Cu feeding resulted in an increase of biliary Cu secretion from ˜ 6 to ˜ 30 nmol/h/100 g body wt within two weeks, both in NW and GY/TR- rats; Cu secretion also did not further increase during the course of the experiment. Hepatic Cu content was similar in NW and GY/TR- rats and progressively increased during Cu feeding. Our data indicate that biliary secretion of diet-derived Cu proceeds exclusively via a saturable Cu transporting system, which is distinct from cmoat/mrp2 and which is independent of biliary GSH. This transport may be mediated by the recently identified Cu-ATPase. In contrast, excess hepatic Cu after iv Cu load depends on cmoat/mrp2 activity for rapid removal. It is concluded that iv administered and dietary (endogenous) Cu is, in part, processed differently by rat liver, which might be related to differences in Cu redox state.
Additive inhibitory effect of hydrocortisone and cyclosporine on low-density lipoprotein receptor activity in cultured HepG2 cells
Authors: Al Rayyes O A , Wallmark A , Floren C
Published: July 2004

Both glucocorticoids and cyclosporine are used to prevent rejection in organ transplant recipients. However, long-term treatment with these drugs is known to induce hyperlipidemia and premature development of atherosclerosis. In previous studies, we have shown that the immunosuppressive drug cyclosporine inhibits catabolism of low-density lipoproteins (LDL) mainly by reducing the expression of LDL-receptor messenger RNA (mRNA), thus explaining the increased plasma levels of LDL cholesterol observed in patients treated with cyclosporine. In the present study, our objective was to investigate the mechanism by which glucocorticoids increase plasma levels of LDL cholesterol. We studied the catabolism of LDL in the human hepatoma cell line HepG2. Our results show that hydrocortisone at physiologically relevant concentrations inhibits LDL binding, uptake, and degradation in a dose-dependent way. Moreover, hydrocortisone also reduces the expression of LDL-receptor mRNA in a dose-dependent way. Cyclosporine also has an additive inhibitory effect on hydrocortisone in the catabolism of LDL. The 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitor fluvastatin reverses the inhibitory effect of both hydrocortisone and cyclosporine. We conclude that treatment with hydrocortisone and/or cyclosporine induces increased plasma levels of LDL cholesterol because of reduced hepatic LDL receptor activity. HMG-CoA reductase inhibitors reverse this undesirable effect and thus reduce the risk of the development of atherosclerosis in patients subjected to immunosuppressive treatment.
Effect of moderate exercise on insulin sensitivity and substrate metabolism during post-exercise recovery in cirrhosis
Authors: Petrides A S , Matthews D E , Eβer U
Published: July 2004

We examined whether a single bout of moderate exercise has a beneficial effect on insulin sensitivity and fuel homeostasis in cirrhosis. Clinically stable cirrhotic patients and age-, sex-, and weight-matched controls participated in insulin clamp studies (either euglycemic hyperinsulinemic or hyperglycemic hyperinsulinemic) in combination with indirect calorimetry and [6,6-2H2]glucose. Three to seven days later, studies were repeated following a single bout of exercise (30 minutes of treadmill exercise at 60% of maximal aerobic capacity). After an overnight fast, following exercise, both cirrhotic and control individuals showed a shift in fuel utilization to enhanced lipid oxidation, decreased glucose oxidation, and increased nonoxidative glucose disposal rates (i.e., glycogen synthesis in muscle) when compared with pre-exercise rates but differences were statistically significant only in the patient group. During euglycemic hyperinsulinemia, insulin-mediated glucose disposal was significantly reduced in cirrhotic patients (3.43 ± 0.26 vs. 7.36 ± 0.48 mg/kg/min, P < .01). Following exercise, glucose uptake increased significantly in cirrhotic patients when compared with pre-exercise levels (P < .05) but remained unchanged in the control group. The increase in total body glucose disposal in cirrhotic patients was entirely accounted for by an increase in nonoxidative glucose disposal (0.81 ± 0.20 vs. 0.51 ± 0.15 mg/kg/min, P < .05). During combined hyperglycemia/hyperinsulinemia, however, insulin sensitivity was unaffected by exercise in both patients and control individuals. In summary, in cirrhotic patients, a single bout of moderate exercise 1) causes a shift in substrate utilization with an increase in lipid oxidation in the postexercise period that is significantly more pronounced than in controls, and 2) increases insulin sensitivity only during euglycemia but not during the more physiological condition of hyperglycemia. Single bouts of moderate exercise therefore may not have a beneficial effect on the metabolic status of patients with chronic liver disease.
Induction of cMrp/cMoat gene expression by cisplatin, 2- acetylaminofluorene, or cycloheximide in rat hepatocytes
Authors: Kauffmann H , Keppler D , Kartenbeck J , Schrenk D
Published: July 2004

The human multidrug-resistance-associated protein (MRP), a member of the adenosine triphosphate (ATP)-binding cassette transporter superfamily, is frequently overexpressed in tumor cells resistant to antineoplastic drugs. In the rat, two Mrp isoforms have been identified, Mrp and cMrp. cMrp, also called Mrp2 or cMoat (canalicular multispecific organic anion transporter), is expressed in the canalicular membrane of rat hepatocytes and mediates the excretion of glucuronate, sulfate, and glutathione conjugates into bile. We investigated the expression of cMrp and Mrp in rat hepatocytes in primary culture. Treatment with the chemical carcinogen 2-acetylaminofluorene (2-AAF), the antineoplastic drug cisplatin, and the protein-synthesis inhibitor cycloheximide led to a dose-dependent and time-dependent increase in cmrp gene expression. A 347-base pair cmrp complementary DNA (cDNA) probe served to demonstrate the induction of cmrp messenger RNA (mRNA) with 40 μmol/L 2-AAF, 5 μmol/L cisplatin, or 5 μmol/L cycloheximide. An analogous response was obtained for the increase in cMrp protein. Mrp mRNA was below the detection limit in Northern blots of RNA from liver and hepatocyte cultures, in contrast to rat testis mRNA which served as a positive control. Immunofluorescence microscopy of cultured hepatocytes was used to visualize cMrp in the plasma membrane. Treatment with 2-AAF led to a marked increase in the immunofluorescence signal confirming the cMrp-inducing potency of 2-AAF. In conclusion, the inducing effect of the compounds studied may reflect a general inducibility of hepatic cMrp by a variety of cytotoxic, carcinogenic, and chemotherapeutic agents which is likely to be of relevance for the acquisition of multidrug resistance during chemotherapy and in the process of chemical carcinogenesis in the liver.
Clinical and family studies in genetic hemochromatosis: Microsatellite and HFE studies in five atypical families
Authors: Adams P C , Campion M L , Gandon G , LeGall J , David V , Jouanolle A M
Published: July 2004

A candidate gene (HFE) has been described for hereditary hemochromatosis on chromosome 6. The study of well-defined atypical hemochromatosis families using genetic markers may increase our understanding of the sensitivity and the specificity of genotyping in hemochromatosis. One hundred and thirteen Canadian families with genetic hemochromatosis were surveyed to find atypical families as possible examples of people with genetic recombinations. All families underwent clinical investigations including iron studies and HLA typing. Each individual was typed at three polymorphic microsatellite loci (D6S105, D6S1260, and D6S299) on chromosome 6. Sixteen subjects were studied for the two missense mutations described for the candidate gene for hemochromatosis (C282Y, H63D). There were eight HLA-identical siblings found in four different families (five men, three women; age range 30-72) with normal transferrin saturation and ferritin levels. There were two patients identified who were homozygous for the C282Y mutation without biochemical evidence of iron overload, and two patients with no evidence of the mutation with significant iron overload. Our conclusions are as follows: 1) finding HLA-identical siblings without iron overload does not confirm a genetic recombination, 2) difficulties in phenotypic definition of disease and the description of new iron overload syndromes that may differ from classical genetic HC cause complicated genetic studies, and 3) finding iron-loaded patients without a C282Y mutation and patients that are homozygous for the C282Y mutation without evidence of iron overload may limit the use of genotyping in population screening for hemochromatosis.
Identification and functional characterization of the promoter region of the human organic anion transporting polypeptide gene
Authors: Kullak-Ublick G , Beuers U , Fahney C , Hagenbuch B , Meier P J , Paumgartner G
Published: July 2004

The organic anion transporting polypeptide (OATP) of the basolateral hepatocyte membrane mediates multispecific uptake of anionic and other amphipathic substrates from sinusoidal blood plasma. To investigate the mechanisms controlling OATP expression, the 5′-flanking region of the human OATP gene was isolated from a P1-derived artificial chromosome genomic clone. Sequence analysis of the OATP promoter showed a number of consensus binding sites for both ubiquitous and liver-enriched transcription factors. Transfection of HepG2 cells with a series of 5′-deleted promoter-luciferase constructs identified the minimal promoter region within 91 base pairs relative to the transcription initiation site. A putative silencer element was localized in the -662/-440 region. The minimal promoter was also active in Chang liver, Madin-Darby canine kidney, and Chinese hamster ovary cells, indicating that basal promoter function is independent of liver-specific regulatory mechanisms. In transfected HepG2 cells, taurocholate (100 μmol/L) stimulated and triiodothyronine (1 μmol/L) inhibited OATP promoter activity, whereas hydrocortisone, dexamethasone, β-estradiol, estrone-3-sulfate, and testosterone had no significant effect. Reverse-transcription polymerase chain reaction analysis showed an increase in OATP messenger RNA in the livers of four patients with chronic cholestatic liver disease compared with three noncholestatic controls. The up-regulation of OATP expression by taurocholate could serve to enhance the sinusoidal efflux of toxic intracellular compounds during chronic cholestasis.
Spontaneous and iatrogenic spreading of liver-derived cells into peripheral blood of patients with primary liver cancer
Authors: Louha M , Poussin K , Ganne N , Zylberberg H , Nalpas B , Nicolet J , Capron F , Soubrane O , Vons C , Pol S , Beaugrand M , Berthelot P , Franco D , Trinchet J C , Brechot C , Paterlini P
Published: July 2004

The prognosis for patients with primary liver cancer (PLC) often depends on tumor recurrence and the development of extrahepatic metastases, particularly after liver transplantation. We have developed a sensitive test to detect both spontaneous circulation of tumor cells and the spread of liver cells due to chemoembolization and alcoholization. Reverse-transcription polymerase chain reaction was used to search for cells expressing α-fetoprotein (AFP) messenger RNA in the peripheral blood of 84 patients with PLC and 102 controls (55 patients with chronic hepatitis and/or cirrhosis, 10 patients with benign liver tumors or liver metastases from intestinal cancers, and 37 healthy individuals). By spiking the blood of healthy volunteers with HepG2 cells, we assessed the sensitivity limit: one HepG2 cell mixed with 107 leukocytes. All 102 controls tested negative. In contrast, 28 patients (33.3%) with PLC tested positive. Positivity for the test was significantly associated with portal thrombosis, tumor size, intravascular tumor emboli, serum AFP level, and extrahepatic metastases. Patients were followed up for a mean period of 39 ± 51 weeks: the probability of developing extrahepatic metastases was significantly higher in positive than in negative patients. Eighteen negative patients with PLC were tested before, 1 hour after, and 24 hours after locoregional therapy: 9 tested positive either 1 or 24 hours after alcoholization or chemoembolization. In conclusion, we have developed a highly specific and sensitive test to detect circulating tumor cells in patients with PLC. This test is likely to be clinically useful in evaluating the risk of developing extrahepatic metastases and the possibility of iatrogenic spreading of liver-derived, possibly tumorous, cells.
Prevalence, risk factors, and genotype distribution of hepatitis C virus infection in the general population: A community-based survey in southern Italy
Authors: Guadagnino V , Stroffolini T , Rapicetta M , Costantino A , Kondili L A , Menniti-Ippolito F , Caroleo B , Costa C , Griffo G , Loiacono L , Pisani V , Foca A , Piazza M
Published: July 2004

In 1996 the prevalence, risk factors, and genotype distribution of hepatitis C virus (HCV) infection were assessed in the general population of a town in southern Italy. The sample was selected from the census by a systematic 1:4 sampling procedure. The participation rate was 96.6%. Among the 1,352 subjects enrolled, 195 (14.4%) tested reactive to antibody to HCV (anti-HCV) with enzyme immunoassay (EIA 3). When further tested with recombinant immunoblot assay (RIBA 3), 170 subjects (87.2%) tested positive, 23 subjects (11.8%) had indeterminate results, and 2 subjects (1%) tested negative. Thus, the overall anti-HCV EIA-positive RIBA-confirmed prevalence was 12.6% (170 of 1,352 subjects) and increased from 1.3% in subjects younger than 30 years to 33.1% in those ≥ 60 years of age. This latter age group accounted for 72.3% of all anti-HCV-positive subjects. Females tested positive more frequently than males (14.1% vs. 10.5%; P < .05). Alanine transaminase (ALT) concentrations were abnormal in only 4.1% (7/170) of anti-HCV EIA-positive RIBA-confirmed subjects. This suggests that ALT screening is not useful in the detection of anti-HCV-positive subjects in a general population. The results of multiple logistic regression analysis showed that an age of less than 45 years, the use of glass syringes, and dental therapy were all independent predictors of anti-HCV positivity. HCV RNA was detected by polymerase chain reaction in 75.9% of the 195 anti-HCV EIA-positive subjects: in 84.7% (144/170) of the RIBA-confirmed subjects; in 17.4% (4/23) tested as RIBA indeterminate; and in neither of the two subjects who tested RIBA negative. HCV type 1b was detected in 75 subjects (50.7%), type 2b in 1 subject (0.7%), type 2c in 66 subjects (44.6%), type 3a in 4 subjects (2.7%), and type 4 in two subjects (1.3%). These figures differ from those of Italian patients with chronic liver disease in whom genotype 2 is more rare. None of the individuals was infected with more than one genotype. The distribution of the two most common HCV viral types (1b and 2c) was not statistically different in terms of mean age, sex, or risk factors and suggests that they may have had a parallel spread in this community. These findings provide one of the highest overall anti-HCV prevalence rates in a general population with a likely cohort effect, i.e., decreased risk of infection along generations. These observations may indicate an epidemic or focus of hepatitis C that occurred several years earlier. The majority of anti-HCV-positive subjects in the oldest age group and with no clinical evidence suggests that HCV infection is a very prolonged and indolent disease.
Liver failure in children with hepatitis A
Authors: Debray D , Cullufi P , Devictor D , Fabre M , Bernard O
Published: July 2004

There have been very few reports dealing with liver failure related to hepatitis A in children. Moreover, the criteria usually used for selecting patients with fulminant hepatitis A for liver transplantation have not been evaluated in children. Therefore, the current study was conducted retrospectively in a single French urban pediatric liver transplantation center to serve as a reminder of the potential severity of hepatits A in children and to identify predictors of outcome. Children were selected by chart review using a data base system and were grouped according to outcome for analyses purposes. Over a 15-year period, 24 children with hepatitis A showed evidence of liver failure, including 6 children who did not develop hepatic encephalopathy, 7 children in whom encephalopathy occurred but resolved spontaneously, and 11 children in whom death or liver transplantation was the outcome. The mean age at onset was 6.5 years. Those with the most rapid onset of liver failure from onset of jaundice had the best chance of recovery without developing encephalopathy. Otherwise, no predictive factors of outcome were found at onset of liver failure. Among the 18 children who developed encephalopathy, the best early prognostic indicator of a poor outcome irrespective of the grade of encephalopathy, appeared to be a prothrombin time level below 21% of normal combined with a serum bilirubin level above 400 μmol/L. Therefore, these two prognostic indicators may be helpful in deciding liver transplantation in children with hepatitis A-induced fulminant liver failure.
Molecular features of the hepatitis B virus nucleocapsid T-cell epitope 18-27: Interaction with HLA and T-cell receptor
Authors: Bertoletti A , Southwood S , Chesnut R , Sette A , Falco M , Ferrara G B , Penna A , Boni C , Fiaccadori F , Ferrari C
Published: July 2004

The strength of the cytotoxic T lymphocyte (CTL) response is believed to influence the final outcome of hepatitis B virus (HBV) infection. Among the different CTL epitopes so far identified, the sequence 18-27 of the HBV nucleocapsid antigen is widely recognized by CTL of HLA-A2-positive patients with acute self-limited HBV infection, and represents the main component of a peptide-based therapeutic vaccine aimed at stimulating the antiviral CTL response in patients with chronic hepatitis B. In the present study, we further analyzed the features of this important HBV region by the following: 1) defining the contribution of individual residues of the epitope to the interaction with the T-cell receptor (TCR) and with the HLA-A0201 molecule; 2) assessing the antigenicity of this viral region in the context of the different HLA-A2 subtypes; and 3) testing whether this sequence can stimulate not only HLA-class I but also HLA class II restricted T-cell responses. A clear hierarchy was observed in the ability of individual residues to act as TCR or HLA binding sites. Furthermore, the sequence HBc18-27 was able to be recognized by specific CTL when presented in the context of different HLA-A2 subtypes. Finally, this HBV region was also found to stimulate HLA class II restricted T-cell responses. These data further increase the potential coverage and efficacy of therapeutic vaccines based on the HBc18-27 sequence.
Isolation and molecular characterization of hepatitis B virus X-protein from a baculovirus expression system
Authors: Urban S , Hildt E , Eckerskorn C , Sirma H , Kekule A , Hofschneider P H
Published: July 2004

The X protein (HBx) of the human Hepatitis B Virus (HBV) is a regulatory protein that exercises a transcriptional activator function on a variety of regulatory elements and is therefore considered to be involved in the development of human hepatocellular carcinoma (HCC). So far, most attempts at elucidating HBx function have been undertaken at the genetic level, reflecting the difficulties in detecting the very low amounts of the protein in infected livers. Consequently, the questions of intracellular localization and posttranslational modification have not yet been completely answered. We therefore constructed recombinant baculoviruses that allowed expression of HBx and the hexa histidine HBx fusion protein HBxHis in insect cells. Cell fractionation experiments revealed that only a minor part of HBx is detectable in a soluble form in the cytosolic fraction, whereas most of the protein forms intracellular aggregates. These results could be confirmed by confocal laser immunofluorescence. The fusion of a hexa-histidine tag to the amino terminus of HBx allowed a rapid one-step purification by metal chelate affinity chromatography. The detailed analysis of purified HBxHis using electrospray ionization mass spectrometry uncovered two major components: the unmodified, monomeric, fully oxidized form with five intramolecular disulfide bridges, and its N-acetylated modification. Additionally, two minor peaks with mass differences of Δ m = +80 da suggested that a small fraction of HBx becomes posttranslationally phosphorylated in insect cells. No further modifications could be observed, indicating that only phosphorylation might play a role in a possible posttranslational regulation of this viral activator.
Evidence of functional and structural cardiac abnormalities in cirrhotic patients with and without ascites
Authors: Pozzi M , Carugo S , Boari G , Pecci V , de Ceglia S , Maggiolini S , Bolla G B , Roffi L , Failla M , Grassi G , Giannattasio C , Mancia G
Published: July 2004

Cirrhosis is associated with cardiovascular abnormalities. Scanty information is available as to whether these include left ventricle diastolic dysfunction and wall thickness increase. To this aim in 27 cirrhotic patients with tense ascites, 17 cirrhotic patients with previous episodes of ascites (not actual), and 11 controls we investigated by echocardiography and echocolor Doppler left ventricle diastolic function (E wave, A wave, E/A ratio, deceleration time of E wave), systolic function (ejection fraction), and wall thickness (left ventricle posterior wall thickness + interventricular septum thickness) along with neurohumoral variables. All measurements (supine position) were repeated after total paracentesis (10.7 ± 0.6 L of ascites) in ascitic patients. Both in patients with and without ascites E/A ratio was reduced as compared with controls (0.93 ± 0.07 and 0.97 ± 0.06 vs. 1.18 ± 0.08, P < .05) while left ventricle wall thickness was increased (18.6 ± 0.6 and 20.1 ± 0.8 vs. 17.2 ± 0.7, P < .05 and P < .01, respectively), irrespective of the postviral or alcoholic cause of liver disease. In all cirrhotics both right and left atrial and right ventricle diameters were significantly greater. Ejection fraction was slightly but significantly (P < .01) reduced in ascitic patients. Paracentesis induced a reduction of the highly increased basal plasma renin activity, aldosterone, norepinephrine (P < .01), and epinephrine (P < .05) and improved diastolic function (E/A, P < .05). Systolic function was unaffected. Thus, irrespective of ascites and cause, advanced cirrhosis is associated with left ventricle diastolic dysfunction and wall thickness increase. We can speculate that neurohumoral overactivity, known to stimulate cardiac tissue growth, may challenge the heart, promoting fibrosis and exerting a further hindrance to ventricular relaxation in patients with cirrhosis experiencing episodes of ascites.
Hepatocellular carcinoma in primary biliary cirrhosis and its impact on outcomes
Authors: Jones D E , Metcalf J V , Collier J D , Bassendine M F , James O F
Published: July 2004

In this study we have determined the incidence of hepatocellular carcinoma (HCC) development in primary biliary cirrhosis (PBC) and its effects on patient survival. Six hundred and sixty seven patients with liver histology compatible with or diagnostic of PBC were seen over a 20-year period. Two hundred and seventy three patients who had stage III or IV disease on their last biopsy and who had been followed up for at least 1 year following that biopsy (total follow-up with advanced disease 2,010 patient years) were identified (243 female, 30 male). Patients who developed HCC were identified and their confounding risk factors were excluded. Mayo risk scores were calculated for each clinic attendance and expected survival for each time point was compared with subsequent actual survival. Sixteen cases of HCC were seen in the patients with stage III or IV disease on last biopsy, providing an overall incidence of 5.9% in this group. Fourteen of these patients had died of HCC related causes, and 2 patients were alive at the census point. The incidence of HCC was significantly higher in males with stage III/IV disease than in females (20% vs. 4.1%, P < .005). Nine of one hundred and eight (8.3%) total female deaths in this group was attributable to HCC compared with 5 of 11 (45.5%, P < .05) male deaths. HCC was not seen in any of the 394 patients with stage I and II PBC followed-up over the same time period. Throughout the disease course of all PBC patients with HCC, the Mayo prognostic model over-predicted survival. Whereas it is a relatively rare complication of cirrhotic PBC in women, HCC is a relatively common cause of death in male PBC patients with cirrhosis. HCC typically develops several years after the onset of cirrhosis, and is poorly predicted by prognostic models. In view of these findings, consideration should be given to careful screening for HCC in male PBC patients with cirrhosis. The risk of HCC development may be an additional reason to consider earlier transplantation in these patients.
The prophylactic effect of aprotinin on intraoperative bleeding in liver transplantation: A randomized clinical study
Authors: Garcia-Huete L , Domenech P , Sabate A , Martinez-Brotons F , Jaurrieta E , Figueras J
Published: July 2004

Fibrinolysis has been recognized as an important cause of intraoperative bleeding during orthotopic liver transplantation (OLT). Several investigators have used prophylactic administration of aprotinin in patients to inhibit fibrinolysis and to decrease transfusion requirements, morbidity, and mortality. Nevertheless, the role of aprotinin in this situation is not yet clear. The goal of this study was to determine the effects of prophylactic administration of aprotinin on intraoperative bleeding and blood requirements, and on hemostatic changes during OLT. Eighty consecutive patients were included in a double-blind, prospective study and were randomized in two groups. In group A (n = 39), an initial dose of 2 × 106 kallikrein inactivator units (KIU) of aprotinin was administered in the induction of anesthesia followed by infusion of 5 × 105 KIU/h until the end of the procedure. The control group (n = 41) received an identical volume of saline solution. The majority of the operations were performed with vena cava preservation (piggy-back technique) without venovenous bypass. During the anhepatic phase, a significant increase in levels of tissue plasminogen activator, thrombin- antithrombin complexes (TAT) and D-dimers (DD) was noted in both groups. A significant increment of TAT was observed in group A during reperfusion. The remaining hemostatic parameters were similar in both groups. Intraoperative requirements of packed red cells, fresh-frozen plasma (FFP), platelets, and cryoprecipitate were similar in the two groups. Our results suggest that prophylactic administration of aprotinin is not useful in reducing bleeding and blood product requirements during OLT.
Urinary sodium balance in patients with cirrhosis: Relationship to quantitative parameters of liver function
Authors: Wensing G , Lotterer E , Link I , Hahn E G , Fleig W E
Published: July 2004

The relationship between the impairment in hepatic and renal function in cirrhosis has not been well established. This study investigated urinary sodium excretion in comparison with quantitative parameters of liver function in 75 patients with various degrees of cirrhosis kept on a constant salt diet of 120 mmol/d for 5 days before the start of the study. The aminopyrine breath test (ABT), indocyanine green (ICG) elimination, galactose elimination capacity (GEC), and hepatic sorbitol elimination (HSE) served as quantitative parameters of liver function. Results for the quantitative tests were compared with those for the Child-Pugh score. Urinary sodium excretion showed a significant nonlinear relationship to ABT (r = .70; P < .0001). Less-significant correlations were observed for ICG (r = .60), the Child-Pugh score (r = -.57), GEC (r = .44), and HSE (r = .34). Because a number of significant correlations were observed between the different liver function tests, multivariate analysis was used to further elucidate the relationship between hepatic function and sodium excretion. Only one independent predictor of urinary sodium excretion could be identified, and that was the ABT (P < .02). More than half of the nonascitic patients showed a urinary sodium excretion of less than 80% of dietary sodium intake, indicating impaired renal sodium handling in preascitic cirrhosis. Based on the 95% confidence interval (CI) for ABT of nonascitic patients with normal (mean ABT 0.56% dose × kg/mmol CO2; 95% CI: 0.44 to 0.69) and reduced urinary sodium excretion (mean ABT 0.26% dose × kg/mmol CO2; 95% CI: 0.18 to 0.35), a threshold level of ABT of about 0.4 (% dose × kg/mmol CO2) for conservation of normal urinary sodium excretion in cirrhosis can be defined. This ABT value reflects an approximate 50% reduction in function compared with the mean of cirrhotic patients with normal liver and kidney function (0.81% dose × kg/mmol CO2). The presence of ascites was also associated with a reduction in ABT to below 0.4 (% dose × kg/mmol CO2), while, for all other parameters, either the cut-off point was close to the lower limit of normal or no cut-off level could be detected. In conclusion, the results of the present study provide further evidence that the impairment in urinary sodium excretion in cirrhosis is related to hepatic function. The data suggest a nonlinear relationship. Because ABT has been shown to reflect functional hepatocellular mass, the occurrence of sodium retention and ascites appears to be related to a threshold of an approximate 50% reduction in functional liver cell mass.
Fate of the human liver after hemihepatic portal vein embolization: Cell kinetic and morphometric study
Authors: Harada H , Imamura H , Miyagawa S , Kawasaki S
Published: July 2004

The favorable therapeutic effect of preoperative portal vein embolization (PVE) was analyzed by assessing various volumetric, cell kinetic and morphometric parameters and examining histologically the embolized and nonembolized lobes of 15 patients who underwent extended right lobectomy 2 to 3 weeks after PVE. Each lobar volume was calculated from computed tomography (CT) images, hepatocyte proliferation was evaluated by assessing proliferative cell nuclear antigen (PCNA) expression and mitosis, hepatocyte apoptosis was evaluated by the terminal deoxyribonucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) assay, and the hepatocyte numerical density as well as the sinusoidal volumetric ratio (Vvs) and the mean hepatocyte volume (Cv) were calculated using morphometric methods. PVE induced hepatocyte apoptosis and atrophy of the embolized lobe (from 798 ± 213 to 708 ± 222 cm3; P < .05). The increased sinusoidal volume in this lobe (17.7% ± 4.5% and 21.7% ± 5.7%, periportal and pericentral area, respectively) may have been attributable to hepatocyte deletion. Cells in the nonembolized lobe entered a highly active phase of proliferation within 2 weeks after PVE. Further evidence of cellular proliferation was provided by the increased nonembolized lobar volume (from 379 ± 132 to 545 ± 130 cm3; P < .01) and numerical density of hepatocyte nuclei (Nv) (5.38 ± 1.26 vs. 3.11 ± 0.85 × 105/mm3; P < .01, nonembolized vs. embolized lobe, respectively). In conclusion, these results indicate that the favorable effect of PVE is attributable to a net gain of functional hepatocyte mass and/or early induction of hepatocyte proliferation following hepatectomy.