bluuebunny/crossref_metadata_2025 huggingface dataset:
Hepatitis C in human immunodeficiency virus-coinfected patients: Increased variability in the hypervariable envelope coding domain
Authors: Sherman K E , Andreatta C , O’Brien J , Gutierrez A , Harris R
Published: November 2004

Patients coinfected with the hepatitis C virus (HCV) and the human immunodeficiency virus (HIV) were studied with regard to nucleotide sequence variability in the E2/NS1 first hypervariable region of the HCV genome. The nucleotide variability within individual patients was compared to patients infected only with HCV. The proportion of predicted synonymous and nonsynonymous amino acid changes, and the relationship to putative high-antigenicity sites, were evaluated in the hypervariable envelope domain. Ninety-one clones from 10 patients with HCV/HIV coinfection were sequenced, following polymerase chain reaction (PCR) amplification of the hypervariable region. The control HCV group included 53 clones from 7 patients. Sequence analysis encompassed the region coding for amino acids 384 to 414. Consensus sequences from each patient were used as the internal standard for nonsynonymous amino acid codon variability. Cumulative proportional comparison at each amino acid site revealed increased variability in HCV RNA from patients with HCV/HIV coinfection versus HCV alone (P < .05). The greatest variability was observed at amino acids 386, 397, 400, 402, 405, 407, and 414, with > l0 percent clonal variation at these sites. Jameson-Wolf plots were used to predict putative high-antigenicity domains. Nonsynonymous clonal variation resulted in alteration of putative antigenic sites within the hypervariable region. All clones had at least one high-probability site. Clones with unique predicted antigenic domains were observed more frequently in HIV/HCV coinfected patients, and, independent of viral titer, were consistent with increased sequence variability. These data suggest an accumulation of envelope variants in the HCV/HIV coinfected patients, which could be related to ineffective viral clearance, and may help explain prior reports of interferon (IFN) resistance in this patient group.
Behavior of various cholesterol crystals in bile from patients with gallstones
Authors: Portincasa P , van Erpecum K J , Jansen A , Renooij W , Gadellaa M , van Berge-Henegouwen G P
Published: November 2004

Besides classical plate-like cholesterol monohydrate crystals, a variety of crystal shapes have recently been described in model biles but their relevance for human gallstone formation is unknown. We therefore studied crystallization behavior in gallbladder bile from cholesterol stone patients (54 untreated, 13 ursodeoxycholate-treated) and 6 pigment stone patients. Bile preparation by ultrafiltration or ultracentrifugation left biliary lipid composition unchanged but plates and their aggregates, and arcs and needles crystallized more extensively while spirals and tubules crystallized less extensively in ultra-centrifuged bile than in ultrafiltered bile. Plates, aggregates, and arcs/needles were seen in 90%, 36%, and 18% of the cases respectively of fresh unfiltered biles of untreated cholesterol stone patients, while spirals and tubules were always absent. In ultrafiltered biles arcs/needles, plates and aggregates progressively developed as persistent forms. Spirals and tubules occurred transiently and were associated with increased deoxycholic acid (+41%, P = .039) and with more extensive cholesterol crystallization. Rate/extent of crystallization of all crystal forms was higher (P < .0001) for multiple than solitary cholesterol stone patients. Ursodeoxycholate-treated patients had atypical platelike cholesterol crystals in fresh unfiltered biles that decreased in size at prolonged observation and in 2 cases even dissolved after 15 and 20 days. No crystals ever developed in ultra-filtered bile of ursodeoxycholic acid (UDCA)-treated patients during 21 days. Pigment stone patients seldom developed crystals. Thus, plates, aggregates and arcs/needles are persistent forms with high crystallization rate in multiple cholesterol stone patients. Tubules and spirals are transient forms that are associated with more extensive crystallization. Patients treated with ursodeoxycholate often have atypical crystals in their fresh bile.
Inhibition of hereditary hepatitis and liver tumor development in Long-Evans cinnamon rats by the copper-chelating agent trientine dihydrochloride
Authors: Sone K , Maeda M , Wakabayashi K , Takeichi N , Mori M , Sugimura T , Nagao M
Published: November 2004

Trientine dihydrochloride (trientine) is an alternative medicinal copper chelating agent for patients with Wilson’s disease of penicillamine intolerance. We examined the effects of trientine on the spontaneous development of hepatitis and hepatic tumors, by its short- term and long-term administration to Long-Evans cinnamon (LEC) rats with an accumulation of copper in the liver, as animal models of Wilson’s disease. Male rats were given trientine in their drinking water at 1500 ppm for 18 weeks, from 6 weeks to 24 weeks of age in short-term experiment, and 1500 ppm for 27 weeks then 750 ppm for 52 weeks, from 8 to 87 weeks of age in the long-term experiment. Development of hepatitis was observed in the control LEC rats at 18 weeks of age. They had high levels of plasma transaminases (glutamic oxaloacetic transaminase [GOT], glutamic pyruvic transaminase [GPT]), and on pathological examination, hepatocyte destruction was observed. Histological findings revealed that short-term administration of trientine inhibited the development of hepatitis remarkably. The plasma GOT and GPT levels of treated animals were only slightly higher than those of normal LEA (Long-Evans with agouti coat color) rats, a sibling line of LEC rats. Copper levels in the liver were decreased by a maximum of 50 percent. In the long-term administration of trientine, the incidence of hepatic cell carcinoma (HCC) in the treated rats was 67 percent that of the untreated LEC rats, and the number of HCCs per rat in the treated group was 0.7 +/- 0.5, being significantly lower as compared with 4.7 +/- 3.5 in the untreated rats. Additionally, the development of cholangiofibrosis in LEC rats was completely prevented by long-term administration of the agent. The copper level in the liver of treated rats was reduced by 33 percent at 87 weeks of age. Development of HCC in LEC rats might be partly, but not totally, because of copper accumulation. No effects on the levels of copper, iron, or zinc in the liver of LEA rats was detected, and no adverse effects were detected in either LEC or LEA rats after both short- and long-term administration of trientine in drinking water.
Kupffer cell inactivation prevents lipopolysaccharide-induced structural changes in the rat liver sinusoid: An electron-microscopic study
Authors: Sarphie T G , D’Souza N B , Deaciuc I V
Published: November 2004

Scanning and transmission electron-microscopic examination of the rat liver sinusoid was performed in this study after in vivo treatment of rats with gram-negative bacterial lipopolysaccharide (LPS, 1 mg/Kg(-1) body weight), with or without pretreatment with gadolinium chloride (GdCl3 10 mg/Kg−1 body weight). Twenty-seven and 48 hours after GdCl3 administration, to inactivate/eliminate part of the Kupffer cell population, a decrease in the number of visualized Kupffer cells was observed, without evident effects on the sinusoidal endothelial cell or on the hepatocyte. Three and 24 hours after its administration, LPS produced ultrastructural changes in the sinusoid characterized by morphological evidence of Kupffer cell activation (i.e., swelling and expanded philopodia anchoring the Kupffer cell to the luminal surface of the sinusoidal wall), and a marked decrease in the population of endothelial cell fenestration. The reduction in the number of fenestrae was associated with a change in the diameter of fenestrae and can be interpreted as a component of the “capillarization” process of the hepatic sinusoid. Such ultrastructural changes were prevented by the administration of GdCl3 24 hours before LPS injection. Hence, these findings suggest that LPS-induced structural changes in the liver sinusoid are mediated by an LPS-induced Kupffer cell activation. Coupled with previous experimental data, showing similar effects of GdCl3 on one of the hepatic sinusoidal endothelial cell (SEC) functions, i.e., hyaluronan scavenging, the data presented in this study strongly support the view that Kupffer cells modulate both the hepatic SEC’s functional as well as ultrastructural properties.
Contribution of immune response to the hepatic fibrosis induced by porcine serum
Authors: Bhunchet E , Eishi Y , Wake K
Published: November 2004

To investigate whether hepatic fibrosis induced by porcine serum in rats is caused by an immune reaction to porcine serum, rats that were immunologically tolerant exclusively to porcine serum were subjected to the repeated injection of porcine serum over a long period. This porcine serum-tolerant group consisted of 15 Wistar rats that had been injected intraperitoneally with porcine serum twice a week from the first postnatal day for 18 weeks. The control group consisted of 16 Wistar rats, aged 8 weeks, that were injected intraperitoneally with porcine serum twice a week for 10 weeks. Livers were fixed and examined by light microscopy. The serum of each rat was subjected to indirect enzyme-linked immunosorbent assay (ELISA) to measure the level of antibody to porcine albumin. In addition, immunohistochemical staining for ED1 was performed on untreated normal and porcine serum-induced fibrotic rat livers to examine the distribution of macrophages and their precursors, the monocytes. All rats in the tolerant group showed an extremely low antibody level (x = 68.27 +/- 4.53), and none (0/15) developed hepatic fibrosis. The majority of rats in the control group showed a very high antibody level (x = 1242.19 +/- 201.15); 75 percent (12/16) developed hepatic fibrosis. Data indicate that, despite the prolonged, repeated injections of porcine serum, if an immune response to porcine serum does not occur, the rats do not develop hepatic fibrosis. The porcine serum-tolerant rats developed hepatic fibrosis after 4 weeks of CCl4 treatment, indicating that injection of porcine serum into neonatal rats did not cause anergy of fibrogenesis, thereby preventing the animal from developing hepatic fibrosis. In normal rat liver, ED1-positive cells, which include nearly all Kupffer cells, were located pre-dominantly in the periportal area. In fibrotic rat liver, ED1-positive cells aggregated prominently in the newly formed and advanced connective tissue septa developed mainly between the neighboring central veins, and in fibrotic parts of the liver capsule. Aggregation of ED1-positive cells was rarely observed in nonfibrotic parts of the liver capsule. The difference between normal and fibrotic rat liver in distribution of EDl-positive cells suggests an involvement of macrophages in fibrogenesis and septum formation. In conclusion, our study showed a significant contribution by the immune response to porcine serum antigens leading to porcine serum-induced rat hepatic fibrosis--processes in which macrophages may be important. This study may lead to an understanding of the mechanism responsible for this form of experimental hepatic fibrosis.
Frequent Expression of Muc1 Apomucin on Biliary Epithelial Cells of Damaged Small Bile Ducts in Primary Biliary Cirrhosis and Chronic Viral Hepatitis: An Immunohistochemical Study
Authors: Sasaki M , Nakanuma Y
Published: November 2004

MUC1 apomucin is a specific target tumor antigen recognized by cytotoxic T cells in a major histocompatibility complex (MHC) unrestricted fashion in patients with pancreatic and breast cancer. This T–cell–mediated immune mechanism against MUC1 apomucin expressing cells has not been evaluated in nonneoplastic immune–mediated diseases. Therefore, we immunohistochemically surveyed the expression of MUC1 apomucin on biliary epithelial cells of small bile ducts in various hepatobiliary diseases, including primary biliary cirrhosis (PBC). MUC1 apomucin was detected using the monoclonal antibody DF3 and the streptavidin–biotin complex, in livers from 31 patients with PBC, 67 with chronic viral hepatitis (CH) with or without cirrhosis, 31 with extrahepatic biliary obstruction (EBO), 30 with hepatolithiasis, and from 23 normal individuals. MUC1 apomucin was infrequently and focally expressed in the biliary epithelial cells of the small bile ducts in 3 of 23 normal livers. In contrast, MUC1 apomucin was frequently and strongly expressed on the luminal surface of biliary epithelia] cells of small bile duct, in 22 of 31 patients with PBC, and in 50 of 67 patients with CH. In particular, high levels of MUC1 apomucin were expressed in bile ducts showing chronic nonsuppurative destructive cholangitis (CNSDC) in PBC and hepatitic duct injuries in CH. In EBO and hepatolithiasis, MUC1 apomucin was focally and weakly expressed in 29% and 30% of livers examined, respectively. More MUC1 apomucin was expressed in PBC and CH than in EBO, hepatolithiasis, and normal liver (

P

< .01, respectively). Frequent and high luminal expression of MUC1 apomucin on biliary epithelial cells in damaged small bile ducts in PBC and CH may be related to T–cell–mediated immunologic mechanisms in these diseases, probably by an MHC–unrestricted recognition process.
Intensive Care Unit Admissions With Cirrhosis: Risk–Stratifying Patient Groups and Predicting Individual Survival
Authors: Zimmerman J E , Wagner D P , Seneff M G , Becker R B , Sun X , Knaus W A
Published: November 2004

Prognosis for acutely ill patients with cirrhosis is influenced by the severity of hepatic abnormalities and by dysfunction of other organ systems. The purpose of this study was to examine the usefulness of the Acute Physiology, Age, and Chronic Health Evaluation (APACHE III) prognostic system for risk–stratifying groups of intensive care unit (ICU) patients with cirrhosis and in predicting individual survival. We used data for 17,440 ICU admissions at 40 American hospitals to select 117 of the 537 patients with a history of cirrhosis who were ventilated on ICU day 1, a group known to have a high mortality rate. We then calculated each patient's probability of hospital death on ICU days 1 through 7, using seven previously validated multivariate equations. Hospital mortality was 63% for the 117 study patients. The most important determinants of risk for hospital death on ICU day 1 were the acute physiology score of APACHE III, ICU admission diagnosis, and operative status. Daily changes in the acute physiology score caused a rise or fall in the probability of hospital mortality and was useful in assessing individual response to therapy. APACHE III accurately risk stratifies critically ill patients with cirrhosis because it accounts for many of the factors known to influence prognosis. This capability can be used to assess severity of illness and risk–stratify patients with cirrhosis during clinical trials. Daily prognostic estimates based on physiological changes over time reflect patient response and can help physicians to assess the incremental benefit of therapy.
The Caffeine Breath Test Does Not Identify Patients Susceptible to Tacrine Hepatotoxicity
Authors: Fontana R J , Turgeon D K , Woolf T F , Knapp M J , Foster N L , Watkins P B
Published: November 2004

Therapy with tacrine, a promising new treatment for Alzheimer's disease, must be discontinued in up to 15% of patients because of hepatocellular toxicity. Recent studies using human liver microsomes have suggested that a single liver enzyme, cytochrome P450 1A2 (CYP1A2), catalyzes the major route of metabolism and elimination of tacrine, and also catalyzes the pathway(s) involved in the generation of reactive metabolites capable of covalent protein binding and cytotoxicity. Because CYP1A2 activity has been shown to vary up to 60– fold among patients, we proposed that a convenient measure of CYP1A2 activity, the [13C 3–methyl] caffeine breath test (CBT), might be clinically useful in identifying patients most susceptible to tacrine liver toxicity. To test this hypothesis, we administered the CBT to 37 patients with Alzheimer's disease before they began treatment with tacrine. Twenty patients received 2 mg/kg of [(13)C 3–methyl] caffeine. The remaining 17 patients received the commercially available CBT kit, which employs a constant 200–mg dose. The activities of two other major drug–metabolizing enzymes (cytochrome P450 3A4 and 2D6 [CYP3A4 and CYP2D6]) were also measured in these 17 patients. We found that the results obtained from the CBT protocol did not predict the peak serum alanine transaminase (ALT) observed in the patients. The measured CYP3A4 and CYP2D6 activities also failed to predict the susceptible patients. However, the result of the standardized–dose CBT correlated well with the logarithm of the steady–state plasma tacrine level obtained in 10 patients (R2 = .69,

P

= .003). We conclude that the CBT will not be clinically useful in determining the subset of patients most susceptible to tacrine hepatotoxicity. However, the correlation we observed between CBT results and tacrine blood levels is the first evidence supporting a critical role for CYP1A2 activity in the disposition of the drug

in vivo

.
Liver Abscess Formation After Local Treatment of Liver Tumors
Authors: de Baère T , Roche A , Amenabar J M , Lagrange C , Ducreux M , Rougier P , Elias D , Lasser P , Patriarche C
Published: November 2004

Our goal was to determine a subset of patients at high risk of developing liver abscesses after local treatment of liver tumors (LTLT) and establish guidelines for the conduct of LTLT in the safest conditions in such patients. Five hundred sixty–one LTLT, 489 transhepatic arterial chemoembolizations (TAC, 10 hepatic embolizations, and 62 percutaneous intratumor injections (PIT), were retrospectively reviewed for liver parenchyma necrosis and abscess formation. Four patients developed abscesses, three after TAC and one after PIT. Despite broad–spectrum antibiotherapy, percutaneous drainage, and surgery, two patients died. A left hepatectomy was required in the other two patients for cure. All four patients had a carcinoid or a neuroendocrine pancreatic tumor. Three out of four patients had bilioenteric anastomoses, and the fourth had recently undergone cholecystectomy and papillotomy. A Lipiodol/doxorubicin mixture without any particulate embolization was injected in the three patients who developed abscesses after TAC. LTLT in patients with bilio enteric anastomosis or papillotomy and/or neuroendocrine or carcinoid tumor should be performed with strict precautions during the procedure and for peri–procedural care.
Acute Reactant Cytokines and Neutrophil Adhesion After Warm Ischemia in Cirrhotic and Noncirrhotic Human Livers
Authors: Clavien P , Camargo C A , Gorczynski R , Washington M K , Levy G A , Langer B , Greig P D
Published: November 2004

Animal studies suggest that acute phase reactant cytokines and polymorphonuclear leukocytes (PMN) may play a critical role in ischemia– reperfusion injury. To evaluate whether similar mechanisms are operative in human liver, six cirrhotic and nine noncirrhotic patients undergoing right hepatectomy were randomized for utilization of hepatic vascular exclusion (HVE) as a model of ischemia–reperfusion injury. Portal and systemic levels of acute reactant cytokines (interleukin 6 [IL–6], interleukin 1 [IL–1], tumor necrosis factor α [TNF–α]) and neutrophil adhesion in serial liver biopsy specimens were studied. Correlations among mediators, leukocyte adhesion, and markers of liver injury were also evaluated. Hepatic vascular exclusion resulted in substantial and reproducible changes in portal and arterial IL–6 levels in both cirrhotic and noncirrhotic patients. Portal and systemic cytokine levels were comparable in most instances, whereas levels were usually higher in cirrhotic patients than in noncirrhotic patients. Negative correlations were found between IL–6 levels at the time of reperfusion and later TNF–α levels. IL–6 levels correlated negatively with numerous markers of hepatocellular injury and the number of postoperative complications. Hepatic vascular exclusion increased neutrophils adhesion after reperfusion in cirrhotic patients but not in noncirrhotic patients. In cirrhotic patients, the degree of leukocyte adhesion after reperfusion correlated with several postoperative markers of liver injury. This study in humans shows that acute reactant cytokines are released during liver ischemia and, interestingly, that IL–6 levels strongly correlate with clinical and laboratory measures of injury. Further studies to evaluate possible causal relationship with hepatic injury are warranted, with emphasis on the role of IL–6 and PMN adhesion.
Ursodeoxycholic Acid or Clofibrate in the Treatment of Non–Alcohol–Induced Steatohepatitis: A Pilot Study
Authors: Laurin J , Lindor K D , Crippin J S , Gossard A , Gores G J , Ludwig J , Rakela J , McGill D B
Published: November 2004

Non–alcohol–induced steatohepatitis (NASH) is characterized by elevated serum aminotransferase activities with hepatic steatosis, inflammation, and occasionally fibrosis that may progress to cirrhosis. No established treatment exists for this potentially serious disorder. Our aim was to conduct a pilot study to evaluate the safety and estimate the efficacy of ursodeoxycholic acid (UDCA) and clofibrate in the treatment of NASH. Forty patients were diagnosed with NASH based on a compatible liver biopsy with other causes of liver disease, including alcohol abuse, excluded by history, serum tests, and use of ultrasound. Twenty–four patients received 13 to 15 mg/kg/d of UDCA for 12 months. Sixteen patients with hypertriglyceridemia were placed on clofibrate, 2 g/day for 12 months. Twenty–five women and 15 men entered the study. Six of 40 patients (15%) withdrew because of side effects. Four additional patients were withdrawn because of noncompliance; one of them later required liver transplantation. In the UDCA group, the decreases in mean serum levels of alkaline phosphatase, alanine transaminase (ALT), and γ–glutamyl transpeptidase (GGT) as well as histological grade of steatosis were significant. Among the patients treated with clofibrate, no change from baseline was found in mean ALT, aspartate transaminase (AST), GGT, bilirubin, triglycerides, and cholesterol, or in histological grade of steatosis, inflammation, or fibrosis after 12 months of treatment as compared with entry. Alkaline phosphatase activities decreased significantly from baseline. Despite the known lipid–lowering effects of clofibrate, it did not appear to be of clinical benefit in the treatment of NASH in this 1–year pilot study. However, treatment of NASH with UDCA for 12 months resulted in significant improvement in alkaline phosphatase, ALT, GGT, and hepatic steatosis. The possible benefit of UDCA therapy should be further investigated in the context of a randomized, controlled trial.
Nonlinear Kinetics After High–Dose Omeprazole Caused by Saturation of Genetically Variable Cyp2c19
Authors: Rost K L , Roots I
Published: November 2004

Nonlinear kinetics of omeprazole and its metabolites were investigated after treatment with repeated high doses. Extensive metabolizers relating to cytochrome P450 2C19 (CYP2C19) activity received for 1 week either omeprazole at 40 mg/d (n = 14) or 60 mg/d omeprazole twice daily (n = 8). Five poor metabolizers (PMs) received 40 mg/d for 1 week. Comparison of omeprazole plasma kinetics between extensive metabolizers (EMs) and PMs after 40–mg treatment revealed a dominant role of CYP2C19 over cytochrome P450 3A CYP3A in omeprazole metabolism. Comparing the omeprazole doses of 40 mg and 60 mg in eight EMs on day 7 of treatment showed that CYP2C19–dependent plasma clearance of omeprazole and omeprazole sulfone was reduced from 19.0 to 8.4 L/h (

P

< .001) and from 19.8 to 9.2 L/h (

P

= .012), respectively. Similarly, formation half–life of 5′–hydroxyomeprazole increased from 0.58 to 1.45 hours (

P

= .025) with the higher dose. CYP3A–dependent metabolic routes remained unaffected. Thus, high–dose treatment with omeprazole uncovers saturation kinetics for CYP2C19 pathways in EMs, and CYP3A becomes the predominant enzyme of omeprazole elimination. Moreover, these individuals may be at risk for side effects due to high omeprazole concentrations if high–dose omeprazole treatment is combined with drugs inhibiting CYP3A activity.
Hepatic Sinusoidal Endothelium Heterogeneity With Respect to Mannose Receptor Activity Is Interleukin–1 Dependent
Authors: Asumendi A , Alvarez A , Martinez I , Smedsrød B , Vidal-Vanaclocha F
Published: November 2004

Using fluorescein isothiocyanate–conjugated ovalbumin (OVA–FITC), 125I–mannan, or 125I–invertase as specific ligands for the mannose receptor, we have quantified its activity in mouse and rat hepatic sinusoidal endothelium (HSE), under both basal conditions and after lipopolysaccharide (LPS) or human recombinant interleukin–1β (IL–1β) stimulations. Mouse treatment for 4 hours with 5 μg/kg IL–1β significantly increased OVA–FITC uptake by HSE. Ligand uptake exhibited a sublobular compartmentalization: In control mice as well as in IL–1β–stimulated mice, the ligand distributed preferentially in the periportal and septal areas; no OVA–FITC was observed in the perivenous sinusoids.

In vitro

exposure of mouse HSE to 100 pg/mL LPS or 1 ng/mL IL–1β for 6 hours significantly (

P

< .01) increased OVA–FITC uptake. Blocking IL–1 receptors in HSE by addition of 100 ng/mL IL–1 receptor antagonist (IL–1Ra) before stimulation with LPS or IL–1β abrogated the increase in mannose receptor–mediated uptake.

In vitro

endocytosis assays showed that rat HSE uptake of 125I–mannan or 125I–invertase progressively increased with both exposure time and concentration of added IL–1β. Upregulation of mannose receptor–mediated uptake in response to IL–1β or LPS was also blocked by previous addition of IL–1Ra to rat HSE. Flow cytometric analysis showed a significant HSE heterogeneity in mannose receptor–mediated endocytosis in response to IL–1β treatment: type I endothelial cells (EC–I, defined by their small size and high cytoplasmic density) significantly (

P

< .01) increased OVA–FITC uptake compared with type II endothelial cells (EC–II, defined by their large size and low cytoplasmic density). In addition, the subset of EC–I contained three times more IL–1β–binding cells than the EC–II subset. Because EC–I and EC–II are preferentially located in the periportal and perivenous segments of hepatic sinusoids, respectively, these results suggest that IL–1β, apart from upregulating mannose receptor activity, contributes to the sublobular compartmentalization of this endothelial cell function.
Modulation of Basal Hepatic Glycogenolysis by Nitric Oxide
Authors: Borgs M , Bollen M , Keppens S , Yap S H , Stalmans W , Vanstapel F
Published: November 2004

We perfused livers from fed rats with a balanced salt solution containing 1 mmol/L glucose. Under these conditions a low steady rate of glycogenolysis was observed (approximately 1.7 μmol glucose equivalents/g/min; 20% of the maximal glycogenolytic activity). Nitric oxide (NO) transiently stimulated hepatic glucose production. A maximal response (on average doubling basal glucose output) was observed with 34 μmol/L NO. The same concentration of nitrite (NO2
–) was ineffective. Half–maximal effects were seen at 8 to 10 μmol/L NO, irrespective of the flow direction (portocaval or retrograde). This glycogenolytic response to NO corresponded to a partial activation of phosphorylase. The NO effect was not additive to maximal stimulation of glycogenolysis (7.7 ± 0.2 μmol hexose equivalents/g/min; n = 4) by 100 μmol/L dibutyryl cyclic adenosine monophosphate (Bt2cAMP). The requirement for activation of phosphorylase was also evidenced by the ineffectiveness of NO in phosphorylase–kinase–deficient livers of

gsd/gsd

rats. The NO effect was blocked by co–administration of cyclooxygenase inhibitors (50 μmol/L ibuprofen, 50 μmol/L indomethacin, or 2 mmol/L aspirin), suggesting a mediatory role of prostanoids from nonparenchymal cells. This conclusion was confirmed by the fact that NO did not activate phosphorylase in isolated hepatocytes. Moreover, NO was no longer glycogenolytic in livers perfused with Ca2
+–free medium, in agreement with the known mediatory role of Ca2
+ in prostanoid–mediated responses. Surprisingly, in Ca2
+– free medium NO inhibited the basal glucose production. This coincided with an increased elution of cyclic guanosine monophosphate (cGMP). Inhibition of glycogenolysis by NO under these conditions was blocked by 1 mmol/L theophylline, suggestive for involvement of cGMP–stimulated cAMP phosphodiesterase. However, we could not confirm that an increase in cGMP resulted in a drop in cAMP. In conclusion, NO recruits opposing mechanisms with respect to modulation of basal hepatic glycogenolysis. In the presence of Ca2
+, activation of phosphorylase with stimulation of glycogenolysis dominates. Cyclooxygenase inhibitors abolish this effect. Activation by NO of the cyclooxygenase in nonparenchymal cells is a distinct possibility. In the absence of Ca2
+, inhibition of basal glycogenolysis becomes observable. It remains to be established whether this results from cGMP–mediated stimulation of hydrolysis of cAMP.
Thalidomide Inhibits Tumor Necrosis Factor α, Decreases Nitric Oxide Synthesis, and Ameliorates the Hyperdynamic Circulatory Syndrome in Portal–Hypertensive Rats
Authors: Lopez-Talavera J C , Cadelina G , Olchowski J , Merrill W , Groszmann R J
Published: November 2004

A hyperdynamic circulatory state frequently is observed in portal hypertension with liver failure or extensive portal–systemic shunting. Tumor necrosis factor α (TNF) causes marked hypotension in mammals by inducing nitric oxide synthesis and has been shown to play a role in the development of the hemodynamic changes observed in portal hypertension. Thalidomide selectively inhibits TNF production by enhancing messenger RNA degradation. We investigated the systemic and portal hemodynamic effects of thalidomide in a prehepatic model of portal hypertension and evaluated whether suppressing TNF synthesis decreases NO production. Portal hypertension was induced by partial ligation of the portal vein (PVL). Animals received thalidomide (T) (50 mg/kg/d) + water or water alone (W), orally, daily for 2 days before and 13 days after PVL operation, at which time hemodynamic studies were performed and TNF plasma levels were obtained. Sham–operated animals were studied identically. In an additional group of PVL animals, 24–hour urinary excretion of NO2
– and NO3
– was measured during treatment. PVL animals receiving T presented with a significantly higher mean arterial pressure and systemic vascular resistance and significantly lower portal pressure, TNF plasma levels, and 24–hour urinary excretion of NO2
– and NO3
–, in comparison with rats receiving W. A significant correlation (

r

= –0.61) was observed between TNF plasma levels and mean arterial pressure among PVL animals. Thalidomide did not have any significant effects on sham rats. Thalidomide inhibits TNF synthesis and reduces NO production, blunts the development of the hyperdynamic circulation, and decreases portal pressure in PVL–operated rats.
Overexpression of Matrix Metalloproteinase 9 Gene in Hepatocellular Carcinoma With Invasive Potential
Authors: Arii S , Mise M , Harada T , Furutani M , Ishigami S , Niwano M , Mizumoto M , Fukumoto M , Imamura M
Published: November 2004

The prognosis for hepatocellular carcinoma (HCC) depends mainly on the clinicopathological characteristic regarding invasion and metastasis. In addition, another distinguishing feature of HCC is the high incidence of concomitant liver cirrhosis, in which the extracellular matrix proliferates markedly. Therefore, the present study was designed to investigate the molecules responsible for the invasion potential of HCC by focusing on matrix metalloproteinase (MMP) in particular, MMP–2 and MMP–9 and the corresponding tissue inhibitor of metalloproteinase (TIMP–2 and TIMP–1), because these enzymes participate in the degradation of the extracellular matrix including the basement membrane. Tumorous and adjacent nontumorous liver samples were obtained from 23 HCC patients who underwent a partial hepatectomy. In 16 of the 23 HCC samples, transcripts for MMP–9 were detected in the tumorous tissues, and 15 of 16 of these samples showed stronger expression in the tumorous tissues than in the nontumorous tissues. On the other hand, MMP–2 messenger RNA (mRNA) was detected in 18 of the 23 cases. Eight of these 18 cases showed more intense expression in the tumorous tissues than in the nontumorous tissues, whereas the expression levels were lower in the tumorous tissues than in the nontumorous tissues in 7 of 18 samples. With respect to the correlation between the clinicopathological features and mRNAs expression, it was found that the expression of MMP–9 mRNA in HCC with capsular infiltration was significantly higher than in HCC without capsular infiltration. HCC with capsular infiltration also tended to have a higher ratio of MMP–9 mRNA expression to TIMP–1 mRNA expression. In addition, the expression of MMP–2 mRNA in nontumorous cirrhotic tissues was significantly higher than in nontumorous tissues from patients with chronic hepatitis. Immunohistochemical examinations revealed that MMP–9 immunoreactivity was the most intense in the HCC cells, particularly in those cells in the marginal areas of the tumorous tissues. In conclusion, the present study shows that MMP–9 is closely participated in capsular infiltration in HCC.
Effects of Epinephrine on Glucose Metabolism in Patients With Alcoholic Cirrhosis
Authors: Schricker T , Albuszies G , Weidenbach H , Beckh K , Ensinger H , Anhäupl T , Radermacher P , Vogt J , Adler G , Georgieff M
Published: November 2004

The cirrhotic liver has been shown to be resistant to the actions of various glucoregulatory hormones. The objective of this study was to investigate the effects of epinephrine on hepatic glucose metabolism in cirrhotic patients. Thirteen cirrhotic and eight healthy subjects were studied. Hepatic glucose production and turnover of alanine and glycerol were measured using stable isotope technique before and during 70 and 150 minutes of epinephrine infusion (0.1 microgram/kg/min). β–Adrenoreceptor binding sites and affinity in mononuclear leukocyte membranes also were determined. Hepatic glucose production and alanine turnover in normals significantly increased during epinephrine infusion, but did not change in cirrhotics. Glycerol turnover increased after 70 minutes of epinephrine infusion in both groups. Epinephrine induced a significant rise of high–affinity β–adrenoreceptor binding sites in normals, yielding a significant correlation between hepatic glucose production and receptor density (r = .94,

P

< .0001). In cirrhotic patients, similar changes in the number of high–affinity β– adrenoreceptors were observed, but no correlation with hepatic glucose production was detected. The cirrhotic liver did not respond normally to the stimulatory effect of epinephrine on hepatic glucose production. Because this blunted response was not related to changes of β– adrenoreceptors, our findings suggest that epinephrine resistance in cirrhosis was caused by a postreceptor defect.
Transforming Growth Factor β1–Regulated Gene Expression of Ito Cells
Authors: Knittel T , Janneck T , Muller L , Fellmer P , Ramadori G
Published: November 2004

During liver fibrogenesis, Ito cells are regarded as the principal matrix synthesizing cells and transforming growth factor β 1 (TGF–β 1) appears to be the main fibrogenic mediator. This study analyzed the effects of TGF–β 1 on Ito cell activation, proliferation, and on the expression of a set of matrix proteins, antiproteases, and TGF–β receptors both in “early cultured” and “culture–activated” Ito cells. Rat liver Ito cells at day 2 of primary culture (“early cultured” cells) were mainly smooth muscle α actin (SMA)–negative, whereas cells at day 6 were judged as “activated” cells (SMA–positive). Following 24–hour exposure to 1 ng/mL TGF–β1, total protein synthesis, cell proliferation, and expression of the “activation” marker SMA were not significantly changed. In addition to previously described stimulatory effects on collagen types I and III, fibronectin, undulin, and proteoglycan–gene expression, TGF–β also dose–dependently increased synthesis and secretion of tenascin, laminin, entactin, collagen type IV, and α2–macroglobulin, but decreased C1–esterase inhibitor production by Ito cells, as revealed by immunoprecipitation of endogenously labeled proteins and by Northern blot analysis. The stimulatory effect of TGF–β was evident both in “early cultured” as well as “culture–activated” Ito cells. By reverse– transcription polymerase chain reaction (RT–PCR) analysis, TGF–β type II, III, and TGF–β/activin type I receptors were present in Ito cells, and their expression pattern was not changed upon TGF–β exposure. Northern blot analysis demonstrated that type I TGF–β/activin receptor was induced during in vitro activation and that TGF–β exposure resulted in a slight increase of type I and III receptor messenger RNAs. In summary, the data illustrate that TGF–β is an important fibrogenic mediator acting both on “early cultured” as well as “culture–activated” Ito cells, rather than a mitogenic or morphogenic mediator. The differential regulation of TGF–β/activin receptors during

in vitro

activation and their up–regulation by TGF–β1 might represent a mechanism by which the receptor complex regulates TGF–β signalling in Ito cells.
Intravenous Administration of Follistatin: Delivery to the Liver and Effect on Liver Regeneration After Partial Hepatectomy
Authors: Kogure K , Zhang Y , Kanzaki M , Omata W , Mine T , Kojima I
Published: November 2004

When 1 μg 125I–follistatin was administered into a rat intravenously, radioactivity levels in serum decreased rapidly. Analysis with a biexponential equation showed that the initial half– life and the terminal half–life were 4.0 and 130.8 minutes, respectively. After 2 hours of infusion, approximately 9% of the follistatin infused remained in the liver, which was much more than that in kidney, spleen, pancreas, intestine, or lung. Autoradiography of the liver obtained at 24 hours of infusion revealed that numerous grains were located in parenchymal cells. Radioactivity of 125I– follistatin in the liver remained elevated until 72 hours and declined markedly thereafter. When a booster shot of 125I–follistatin was administered at 72 hours, radioactivity in the liver at 120 hours was markedly increased compared with that in rats that received a single shot of 125I–follistatin. We then examined the effect of intravenous infusion of follistatin on liver regeneration after hepatectomy of 70%. Immediately after the hepatectomy, either 1 microgram follistatin or saline was infused intravenously. In some rats, a booster shot was infused at 72 hours. After 120 hours of hepatectomy of 70%, remnant liver weight, liver regeneration rate, and DNA content were significantly (

P

< .05) higher in rats that received a booster shot of follistatin at 72 hours than those in control rats. These results indicate that follistatin administered intravenously accumulates in the liver and promotes liver regeneration after partial hepatectomy.
Interferon Gamma Protects Against Hepatic Tumor Growth in Rats by Increasing Kupffer Cell Tumoricidal Activity
Authors: Karpoff H M , Tung C , Ng B , Fong Y
Published: November 2004

An increasing number of hepatic resections are being performed as potentially curative surgery for malignant liver neoplasms. Hepatectomy and subsequent liver regeneration produce a local environment that enhances growth of microscopic residual tumor. To determine if pretreatment with murine interferon γ (IFN–γ) can protect against such enhanced tumor growth, Buffalo rats were randomized to receive a 3–day treatment of IFN–γ (50,000 U/qD intraperitoneally) or saline. Groups then underwent intrasplenic injection of 10(6) Morris hepatoma cells, followed 1 hour later by sham (control) or partial hepatectomy (PH) of 70%. PH significantly enhanced tumor growth within the liver control, 8 ± 3 nodules per liver; PH, 73 ± 12 nodules per liver;

P

< .001). This enhancement was attenuated by prior administration of IFN–γ IFN–γ/PH, 16 ± 3;

P

< .001 vs. PH). Growth factor release and liver regeneration were not affected significantly by pretreatment with IFN–γ. The effect of IFN–γ on tumor growth is associated with a significant enhancement of Kupffer cell (KC)–mediated tumoricidal activity (percentage of specific lysis, 55 ± 10% control, 78 ± 11% IFN–γ,

P

< .01) but not lymphocyte–mediated tumoricidal activity. Because microscopic residual disease may be present after hepatectomies for cancer, IFN–γ may be useful agent in retarding growth of residual tumors.
Population of Hepatic Macrophages and Response of Perfused Liver to Platelet–Activating Factor During Production of Thioacetamide–Induced Cirrhosis in Rats
Authors: Noda S , Masumi S , Moriyama M , Kannan Y , Ohta M , Sugano T , Yamate J
Published: November 2004

The response of hepatic macrophages and the effects of platelet– activating factor (PAF) in perfused liver were studied during production of experimental cirrhosis induced by thioacetamide (TAA) in female Sprague–Dawley rats. Within 4 weeks of TAA administration (300 mg/L drinking water), an increase in hepatic macrophage population and enlargement in cell size preceded the alterations characteristic of cirrhosis. During 12 weeks of TAA administration, the changes in hepatic macrophages were maintained and cirrhosis of the micronodular type developed with a marked increase in hydroxyproline content. Although TAA treatment for 4 weeks had no effect on oxygen consumption or hepatic portal pressure in the perfused liver, the increment in hepatic portal pressure and decrement in oxygen consumption induced by PAF in TAA–treated rats were double those in control rats. The amounts of prostaglandin D2 (PGD2) and thromboxane B2 (TxB2) in perfusate induced by PAF were seven– and fivefold greater, respectively, in TAA–treated rats than in control rats. Zymosan mimicked the effects of PAF. These results are consistent with the hypothesis that hepatic macrophages and PAF play important roles in the development of cirrhosis induced by TAA.
Inhibition of Experimentally Induced Cirrhosis in Rats by Hypothyroidism
Authors: Oren R , Dotan I , Papa M , Marravi Y , Aeed H , Barg J , Zeidel L , Bruck R , Halpern Z
Published: November 2004

The coexistence of hyperkinetic circulation, hypermetabolism, and hyperactivity of the sympathetic nervous system is encountered in both cirrhosis and hyperthyroidism. Several drugs, such as propylthiouracil and propranolol, that are beneficial for treating some patients with chronic liver diseases are also prescribed for the treatment of thyrotoxicosis. We investigated the effects of experimentally induced hypo– and hyperthyroidism on the development of cirrhosis induced in rats by thioacetamide (TAA). We specifically examined whether hypothyroidism could prevent and hyperthyroidism could aggravate liver damage. Hypothyroidism induced by methimazole (MMI, 0.04%), propylthiouracil (PTU 0.05%), and by thyroidectomy was confirmed by a significant elevation of thyroid–stimulating hormone (TSH) levels. Hyperthyroidism (decreased TSH levels) was induced by eltroxin (ELT: 50 micrograms/kg). Thirteen groups of 10 rats each were studied: euthyroid controls (3 groups: water, TAA 1.5 months, and TAA 3 months), hypothyroid (6 groups: MMI, PTU, surgical, MMI–TAA, PTU–TAA, surgical–TAA), and hyperthyroid (4 groups:ELT 1.5 months and 3 months, and ELT–TAA for 1.5 months and 3 months). Hepatic fibrosis (scored from 0 to 3) was significantly reduced (

P

< .0001) in hypothyroid rats as compared with euthyroid controls, and was aggravated in TAA–treated hyperthyroid rats (

P

< .0001). Quantitative microscopic analysis of liver biopsy specimens from all groups confirmed the semiquantitative histopathological scores (

P

< .001). Direct intrasplenic pressure measurement revealed a significant portal pressure elevation in the TAA and the ELT–treated rats (from 4.7 ± 0.1 in the euthyroid group to 8.1 ± 2.3 and 10.2 ± 2.1 and 12.5 ± 1.6 in the TAA, ELT and ELT– TAA groups, respectively). However, in the hypothyroid–TAA groups, the portal pressure was found to be within the euthyroid normal range (4.6 ± 1.2 and 5.8 ± 0.6 in the PTU–TAA and surgical–TAA, respectively). After 12 weeks, the mean spleen weight of rats receiving only TAA was significantly higher than the TAA–treated hypothyroid rats (

P

< .0001), indicating that the hypothyroid TAA–treated rats were less portal hypertensive. These results suggest that induced hypothyroidism can inhibit, whereas hyperthyroidism can aggravate, the development of cirrhosis in a rat model.
Isolation of Highly Purified, Functional Carboxy–Terminally Truncated Hepatitis B Virus Middle Surface Protein Activators From Eucaryotic Expression Systems
Authors: Hildt E , Urban S , Eckerskorn C , Hofschneider P H
Published: November 2004

Carboxy–terminally truncated hepatitis B virus (HBV) middle surface proteins (MHBst) show a transcriptional activator function. Two different subtypes of MHBst activators can be distinguished: an ER–localized type, represented here by MHBst76 (truncated at amino acid 76), and a cytosol–localized type, represented here by MHBst63. To characterize the MHBst activator on the protein level and to analyze posttranslational modifications, we established recombinant baculoviruses encoding for fusion proteins of MHBst76 or MHBst63 and of an amino terminal hexa–his tag. Both proteins could be obtained in high purity by affinity chromatography using Ni–nitrilo–tri–acetate agarose. In addition, 6H–MHBst76 was also isolated from transiently transfected HepG2 cells. Both the

Spodoptera frugiperda

(Sf9) cell–derived and the HepG2 cell–derived MHBst proteins were found to be unglycosylated. A detailed analysis of Sf9 cell–derived 6H–MHFBst76 by electrospray– ionization mass spectrometry showed that a fraction of this protein is N–terminally acetylated and phosphorylated or sulfated. Electric–field– mediated transfer of the highly purified proteins into reporter cells demonstrated that the isolated proteins are functional transcriptional activators. These experiments further showed that Sf9 cell–derived and HepG2 cell–derived 6H–MHBst do not differ in their functionality. This system allowed production and purification of functional 6H–MHBst in amounts sufficient enough to allow a further detailed analysis of MHBst activators on the protein level.
Autoantibody Prevalence in Chronic Hepatitis B Virus Infection: Effect in Interferon Alfa
Authors: Gregorio G V , Jones H , Choudhuri K , Vegnente A , Bortolotti F , Mieli-Vergani G , Vergani D
Published: November 2004

The most effective treatment of chronic hepatitis B virus (HBV) infection is interferon alfa (IFN–α), a potentially severe side effect of which is the induction of autoimmunity. To assess whether IFN– α causes clinical or serological autoimmune manifestations, we studied 61 children randomized to receive 5 MU/m2 of IFN–α three times per week for 12 weeks, with or without steroid priming or no treatment. Autoantibodies to antinuclei (ANA), smooth muscle (SMA), gastric parietal cell (GPC), liver kidney microsomal type 1, mitochondrial, liver cytosolic antigen, thyroid microsomal, and thyroid globulin were detected by standard techniques. Over a median of 4 years (range, 1–5 years) from randomization, no clinical signs of autoimmunity were observed. Autoantibody positivity for nuclei, smooth muscle, and/or gastric parietal cells was observed on at least one occasion in 42 of 61 children (69%), with no overall difference in the prevalence between patients treated with interferon alone (19 of 24 [79%]), steroids plus interferon (13 of 20 [65%]), or untreated controls (10 of 17 [59%]). There was also no difference in the autoantibody prevalence before, during, and at follow–up after cessation of treatment in both interferon–treated and interferon– untreated patients. Autoantibodies are common in chronic HBV infection, and their prevalence is uninfluenced by IFN–α.