bluuebunny/crossref_metadata_2025 huggingface dataset:
Antifibrogenic Effect of A Deletion Variant of Hepatocyte Growth Factor on Liver Fibrosis in Rats
Authors: Yasuda H , Imai E , Shiota A , Fujise N , Morinaga T , Higashio K
Published: November 2004

Hepatic fibrosis, which may lead to cirrhosis, is associated with most chronic liver diseases. Current therapies for hepatic fibrosis are, however, generally ineffective. In this report we assessed the efficacy of the treatment of hepatic fibrosis with a naturally occurring deletion variant of hepatocyte growth factor (dHGF). The administration of dHGF increased liver weight and suppressed the increase of hepatic collagen content in rats treated with dimethylnitrosamine (DMN) to induce hepatic fibrosis. Furthermore, dHGF exerted its mitogenic and antifibrogenic activities even after the liver fibrosis had been established with DMN. Northern blot analysis showed that dHGF suppressed the increase of messenger RNA (mRNA) levels of procollagen α2(I), αl(III), α1(IV), transforming growth factor β1 (TGF–β1), desmin (a marker of hepatic lipocytes), and α–smooth muscle (sm)–actin (a marker of activated hepatic lipocytes). In addition to suppressing the elevated TGF–β1, mRNA level in hepatic fibrosis, dHGF had a potent ability to decrease TGF–β1 mRNA level even in a normal liver. Immunohistochemical analysis revealed that desmin–positive cells and α–sm–actin–positive cells were increased in the hepatic fibrosis, whereas neither cells were seen in livers of DMN–treated rats given dHGF. We conclude that dHGF prevents and improves the DMN–induced hepatic fibrosis in rats by reducing mRNA levels of procollagens and TGF–β1, by inhibiting an activation of hepatic lipocytes, and by stimulating liver regeneration. dHGF may be useful for and applicable to the treatment of fibrosis in chronic liver diseases.
The Course of Galactose Elimination Capacity in Patients With Alcoholic Cirrhosis: Possible Use As A Surrogate Marker for Death
Authors: Merkel C , Marchesini G , Fabbri A , Bianco S , Bianchi G , Enzo E , Sacerdoti D , Zoli M , Gatta A
Published: November 2004

There is increasing interest for the use of surrogate end points in the evaluation of treatments in patients with liver disease, but adequate validation is seldom available. This study aimed to describe the different course of galactose elimination capacity in patients with alcoholic cirrhosis who continued to drink or abstained from alcohol consumption during follow–up, and to validate changes in galactose elimination as a surrogate end point for death from liver–related causes. Forty–five patients with alcoholic cirrhosis (22 who continued drinking throughout the study period, and 23 who stopped drinking and were abstinent throughout the study period) were retrospectively selected among patients who had galactose elimination capacity measured at 6–month intervals. During follow–up 10 drinkers and 3 abstainers died of liver–related causes (

P

= .025). Abstainers showed a transient improvement in galactose elimination capacity, followed by a decrease. Continuous drinkers showed a reduction from the beginning. According to Cox's regression analyses, persistent alcohol abuse and galactose elimination capacity were separately related to the risk of death, but, when a time–dependent model was fitted containing galactose elimination capacity and persistent alcohol abuse, only the former remained significant. This implies that variations in the risk of death occurring as a consequence of abstinence from alcohol consumption may be predicted from changes in galactose elimination capacity, and that the mechanisms through which abstinence influences survival are strictly linked to the mechanisms responsible for the changes in the test. Because of the strict association of decrease in galactose elimination capacity and short survival, as proved in several series, this observation represents adherence to the criteria requested for adequacy of a surrogate end point. In conclusion, in alcoholic cirrhosis the decrease in galactose elimination capacity is an adequate surrogate end point for death from liver–related causes, which is worth testing in other conditions and in response to other treatments.
Progressive Decrease in Tissue Glycogen Content in Rats With Long–Term Cholestasis
Authors: Krahenbuhl L , Talos C , Reichen J , Krahenbuhl S
Published: November 2004

Liver and skeletal muscle glycogen metabolism were investigated in rats 1 and 4 weeks after bile duct ligation (BDL) and in pair–fed, sham– operated control rats. Livers were subjected to morphometric analysis to express glycogen content and enzyme activities per mL hepatocytes. One week after BDL, the hepatic glycogen content was 28.8 ± 13.8 versus 38.6 ± 16.4 mg/mL hepatocyte in BDL and control rats, respectively. Total activity of glycogen synthase (50.2 ± 7.0 vs. 63.5 ± 9.4 mU/mL hepatocytes) and glycogen phosphorylase (59.4 ± 12.9 vs. 90.8 ± 18.9 U/mL) were significantly reduced in BDL whereas the active fraction of glycogen synthase (27 ± 6 vs. 38 ± 5%) but not of glycogen phosphorylase was reduced. The skeletal muscle glycogen content was not different between BDL and control rats. Four weeks after BDL, hepatic glycogen content was further reduced (20.5 ± 14.2 vs. 52.9 ± 6.4 mg/mL). Total activity of glycogen synthase (38.8 ± 12.1 vs. 60.1 ± 4.6 mU/mL hepatocytes) and glycogen phosphorylase (127 ± 19 vs. 178 ± 33 U/mL hepatocytes) were both reduced in BDL rats as were their corresponding active fractions (30 ± 18 vs. 66 ± 8% and 58 ± 10 vs. 76 ± 10). At this time point, the glycogen content in soleus muscle was decreased by 64% in BDL. The glucagon plasma concentration was increased in BDL rats at both time points. There were positive correlations between the volume fraction and both hepatic glycogen content and total activity of hepatic glycogen synthase. Plasma glucagon and the active fraction of hepatic glycogen synthase were negatively correlated. The current studies show a progressive decrease in the hepatic and skeletal muscle glycogen content in BDL rats. The observed decrease in the activities of glycogen synthase and phosphorylase suggest that reduced glycogen synthesis is the major mechanism leading to the reduction in the hepatic glycogen content in BDL rats.
Spermidine Acetyltransferase in Rat Hepatocytes Cultured At Different Oxygen Tensions
Authors: Vargiu C , Colombatto S , Giribaldi G , Grillo M A
Published: November 2004

To understand the mechanism involved in the liver zonation of polyamines, we have studied the possible role of oxygen tension. When hepatocytes were cultured at 21% and at 5% oxygen in atmosphere to mimic periportal and perivenous conditions, polyamine content was modified. The observed modifications suggested an effect on the interconversion pathway. Spermidine acetyltransferase (SAT) activity and

N

1–acetylspermidine were therefore measured in the same conditions. SAT activity was markedly increased after 6 hours and

N

1–acetylspermidine was accumulated in the cells. This was caused by new enzyme synthesis. The higher expression of SAT was accompanied by an increase in the content of the specific messenger RNA (mRNA). When liver cells were depleted of polyamines, SAT activity and the specific mRNA content were not enhanced by oxygen deprivation, but they increased when polyamines were added again. Polyamines therefore appear to be necessary to promote the increase in SAT mRNA.
Comparison of Vascular Nitric Oxide Production and Systemic Hemodynamics in Cirrhosis Versus Prehepatic Portal Hypertension in Rats
Authors: Niederberger M , Gines P , Martin P , Tsai P , Morris K , McMurtry I , Schrier R W
Published: November 2004

Nitric oxide (NO) is postulated to play a role in the pathogenesis of arterial vasodilation in chronic portal hypertension. This present study investigates the relationship between systemic hemodynamics and the vascular production of NO, as estimated by measuring cyclic guanosine monophosphate (cGMP) in aortic tissue in two models of chronic portal hypertension in the rat: the partial portal vein ligation (PVL) model and CCl4–induced cirrhosis. NOS was also examined by Western blotting in aortic and mesenteric vessels. Sham–operated rats and rats given phenobarbital were used as controls. PVL rats and rats with cirrhosis and ascites showed a typical pattern of a hyperdynamic circulatory state, when compared with their respective controls: mean arterial pressure; PVL: 113 ± 2 versus 124 ± 2,

P

< .01 and cirrhotics: 103 ± 5 versus 130 ± 4 mm Hg,

P

< .01. Cardiac index; PVL: 32 ± 2 versus 26 ± 1,

P

< .01 and cirrhotics: 51 ± 3 versus 30 ± 1 mL · min–1 · 100 gm–1,

P

< .0001. Systemic vascular resistance; PVL: 3.7 ± 0.1 versus 4.9 ± 0.2,

P

< .01 and cirrhotics: 2.1 ± 0.2 versus 4.4 ± 0.2 mm Hg · min–1 100 g–1,

P

< .0001. Aortic cGMP was markedly increased in cirrhotic rats with ascites (728 ± 83 fmol/ mg protein) as compared with phenobarbital– treated controls (244 ± 31 fmol/mg,

P

< .001). This increase was abolished by chronic administration of N(omega)–nitro–L–arginine methyl ester. By contrast, PVL rats had an aortic cGMP concentration similar to sham–operated controls (282 ± 16 fmol/mg vs. 274 ± 33 fmol/mg,

P

= not significant) and significantly lower than that found in cirrhotic rats with ascites. Expression of cirrhotic aortic endothelial nitric oxide synthase (eNOS) was increased but PVL aortic eNOS did not differ from that of controls, whereas the mesenteric eNOS was increased in both PVL and cirrhotic rats as compared with the controls. These results suggest that vascular NO production is higher in cirrhotic rats than in PVL rats. This increased production may contribute to the more marked abnormalities in systemic hemodynamics seen in experimental cirrhosis as compared with PVL.
Impact of Complete Inhibition of Viral Replication on the Cellular Immune Response in Chronic Hepatitis B Virus Infection
Authors: Marinos G , Naoumov N V , Williams R
Published: November 2004

Interferon alfa (IFN–α) treatment is effective in only a proportion of patients with chronic hepatitis B virus (HBV) infection. The mechanisms for therapeutic failure remain unknown but high levels of HBV replication are known to inhibit the immunopotentiating effects of IFN–α. In nine patients with chronic hepatitis B not responding to IFN–α monotherapy, we determined the virus–specific T–helper–cell responses during two consecutive therapeutic regimens: IFN–α alone and IFN–α in combination with a new potent inhibitor of HBV replication, lamivudine. By comparing the results obtained during the initial IFN–α monotherapy to those during the combination treatment, it was investigated whether complete inhibition of virus replication will enhance the interferon–induced immunoreactivity to HBV. Despite the rapid reduction to undetectable serum HBV DNA in all nine patients during the combination treatment, none sustained permanent hepatitis B e antigen (HBeAg) clearance during subsequent 12–month follow–up. HLA class II–restricted T–helper–cell responses to hepatitis B core antigen (HBcAg) showed no difference during IFN–α monotherapy and during the combination of lamivudine plus IFN–α. In contrast, a delayed T–cell activation occurred after a rebound in serum HBV DNA postcombination treatment, which lead to increased hepatocytolysis. These findings suggest that the profound inhibition of HBV replication by a nucleoside analogue does not restore the impaired virus–specific T–cell response in chronic HBV infection.
Response to Higher Doses of Interferon Alfa–2B in Patients With Chronic Hepatitis C: A Randomized Multicenter Trial
Authors: Lindsay K L , Davis G L , Schiff E R , Bodenheimer H C , Balart L A , Dienstag J L , Perrillo R P , Tamburro C H , Goff J S , Everson G T , Silva M , Katkov W N , Goodman Z , Lau J Y , Maertens G , Gogate J , Sanghvi B , Albrecht J
Published: November 2004

To evaluate response rates to 3, 5, or 10 million units (MU) of interferon alfa–2b, given thrice weekly, and to determine whether higher doses of interferon increase the likelihood or durability of the response, a multicenter, randomized trial was performed at nine academic medical centers in the United States. Two hundred forty eight patients with chronic hepatitis C were randomized to receive 3, 5, or 10 MU of interferon alfa–2b thrice weekly for 12 weeks. Based on the alanine aminotransferase (ALT) response at treatment–week 12, the patients were rerandomized to additional therapy at the same or at increased doses for an additional 12 to 36 weeks; in the case of no response to the highest dose, the patients were discontinued from the study. Serum ALT concentrations and liver histology were measured. The overall complete response rates to 3, 5, or 10 MU were not different at treatment–week 12 (31% vs. 42% vs. 40%, not significant). The majority of week–12 responders continued to respond during additional treatment. When the treatment was discontinued, 15.4% to 19.0% of patients maintained their response. Of the nonresponders to 3 MU at week 12, who were continued on 3 MU for an additional 12 weeks, none responded. However, response to additional therapy occurred in 12% of week–12 nonresponders, whose dose was escalated from 3 or 5 MU to 10 MU. The only baseline features associated with the treatment response were the absence of fibrosis or cirrhosis on the pretreatment liver biopsy and viral genotype. We conclude that the initial response to interferon in patients with chronic hepatitis C is not increased by treatment with higher doses of the drug. Patients who do not respond to 3 MU by treatment–week 12 will not respond with continued therapy at that dose; however, a proportion of patients who do not respond to 12 weeks of treatment with 3 or 5 MU may respond to higher doses. Although the long–term sustained response rates are marginally increased with interferon doses above 3 MU three times per week, the side effects are difficult to tolerate. The analysis of baseline factors in relation to response identified no single baseline factor associated with a low–enough response rate to warrant withholding interferon therapy from patients with chronic hepatitis C.
Severity of Liver Disease in Liver Transplantation Recipients With Hepatitis C Virus Infection: Relationship to Genotype and Level of Viremia
Authors: Zhou S , Terrault N A , Ferrell L , Hahn J A , Lau J Y , Simmonds P , Roberts J P , Lake J R , Ascher N L , Wright T L
Published: November 2004

Infection with hepatitis C virus (HCV) genotype 1b has been reported to be associated with more severe posttransplantation liver disease than infection with non–1b genotypes. To address this issue, we evaluated the outcome in 124 patients who underwent liver transplantation for chronic HCV infection. The HCV genotype and/or serotype responsible for infection was determined by four different methods. HCV RNA was detected in serum samples by polymerase chain reaction (PCR) amplification, and quantified by branched DNA assay. Disease severity was expressed as a histological score (which included grading of portal inflammation, lobular activity, fibrosis, and cytopathic changes). Median duration of histological follow–up was 25 months (range 1–75 months). Genotype was assignable in 112 (92.5%) patients. Genotypes responsible for infection were as follows: 1a = 32.2%, 1b = 27.3%, 2a = 7.4%, 2b = 8.3%, 3a = 14%, and mixed infection (more than one subtype) = 3.3%. Level of viremia, alanine aminotransferase (ALT), aspartate aminotransferase (AST), bilirubin, and total histological score were not significantly different in patients infected with type 1b compared with patients infected with other genotypes. While duration of histological follow–up was greater in patients infected with type lb versus other types (

P

= .02), by univariate and multivariate analysis neither HCV genotype (lb versus others), level of viremia nor duration of histological follow–up were associated with disease severity. Moreover, there was no significant difference in the actuarial graft survival in patients infected with type lb compared with that of patients infected with non–lb types (82% and 87% at 3 years, respectively). Reanalysis using HCV genotype 1 showed no association with disease severity, graft survival, and patient survival. We conclude that HCV genotype 1 and subtype 1b are not associated with disease severity or graft survival in liver transplantation recipients.
Evaluation of A New Endoscopic Index to Predict First Bleeding From the Upper Gastrointestinal Tract in Patients With Cirrhosis
Authors: Zoli M , Merkel C , Magalotti D , Marchesini G , Gatta A , Pisi E
Published: November 2004

The aim of this study was to prospectively evaluate the usefulness of the Italian Liver Cirrhosis Project (ILCP) classification of esophageal varices, together with the gastric features of portal hypertension, in predicting the first upper gastrointestinal bleeding in cirrhosis. The efficiency of these endoscopic parameters in predicting bleeding was also compared with the efficiency of the North Italian Endoscopic Club (NIEC) index. Three hundred forty–four cirrhotic patients with esophageal varices but without any previous bleeding were enrolled in the study. The following endoscopic parameters of esophageal varices were recorded: location, size, occupancy, blue tone, and red color signs. Gastric varices were graded as absent or present, while congestive gastropathy was considered as absent, mild to moderate, or severe. All patients were followed until the first upper gastrointestinal bleeding and/or death, or for at least 24 months. No patient received any treatment to prevent bleeding. Sixty–five patients bled from the upper gastrointestinal tract during the study. Univariate analysis showed that all endoscopic parameters were predictors of bleeding. According to multivariate analysis (Cox's model), size, gastric varices and congestive gastropathy were the only independent predictors of bleeding, and the following prognostic index (PI) was developed: PI = (size × 0.0395) + (congestive gastropathy × 0.878) + (gastric varices × 0.705). This index, which was validated using a split–sample technique, and which appears to be superior to the NIEC index in predicting bleeding, may be useful in decision making for primary prophylaxis.
Reperfusion Damage to the Bile Canaliculi in Transplanted Human Liver
Authors: Cutrin J C , Cantino D , Biasi F , Chiarpotto E , Salizzoni M , Andorno E , Massano G , Lanfranco G , Rizzetto M , Boveris A , Poli G
Published: November 2004

In 19 patients who have undergone orthotopic liver transplantation (OLT), the trend and degree of cholestasis was statistically monitored in terms of plasma levels of l–gamma–glutamyl transferase (GGT) and total bilirubin. In addition, the ultrastructure of the bile canaliculus was examined during the entire OLT procedure, i.e., during explantation, cold ischemia, and after 60 to 90 minutes of organ reperfusion. Cholestasis was evident from the second day after surgery, with a peak after approximately 10 to 16 days. Defined, small changes in the functional state of actin filaments were noted in the bile canalicular area after prolonged ischemia. But the morphological status of the bile canaliculi changed dramatically after reperfusion. In fact, the mean area and perimeter of the canaliculi had increased significantly, and there was a marked loss in the number of bile microvilli per unit of canalicular area. The bile canaliculus appears to be one of the liver structures most susceptible to ischemia– reperfusion damage. A series of biochemical changes occurring during ischemia and after reoxygenation of the transplanted liver, especially, would provide a reason for the observed early morphological damage of the bile canaliculus, which, in turn, would explain the cholestasis of these patients in the first posttransplantation period.
Impaired Cardiovascular Autonomic Response to Passive Tilting in Cirrhosis With Ascites
Authors: Laffi G , Lagi A , Cipriani M , Barletta G , Bernardi L , Fattorini L , Melani L , Riccardi D , Bandinelli G , Mannelli M , La Villa G , Gentilini P
Published: November 2004

The autonomic regulation of cardiovascular function was evaluated in 15 cirrhotic patients with ascites and in 13 healthy subjects by the autoregressive power spectral analysis (PSA) of the intervals between adjacent R waves of the electrocardiogram (RR) interval and arterial pressure variability. Total power, low frequency (LF; index of the sympathetic activity of the heart and circulation), and high frequency (HF; index of vagal tone to the heart) components of the RR interval, systolic, and diastolic arterial pressure were evaluated in the supine position and during passive tilting, together with plasma norepinephrine levels. In the supine position, no significant differences in the PSA data were observed between the control subjects and cirrhotic patients, who had higher plasma norepinephrine levels. In healthy subjects, tilting was associated with an increase in the LF of the RR interval and arterial pressure and a decrease in the HF of the RR interval. In contrast, patients with cirrhosis showed a decrease of both LF and HF. Consequently, the LF/HF ratio significantly increased in healthy subjects, whereas it was unchanged in cirrhotic patients. The LF component of the diastolic pressure also decreased during tilting in cirrhotic patients. Plasma norepinephrine increased after tilting in both groups. These results indicate that the autonomic response to passive tilting is impaired in cirrhotic patients with ascites at both the cardiac and vascular levels, as a result of an altered sympatho–vagal balance, with reduced sympathetic predominance. These alterations occurred despite an appropriate response to the tilting of plasma norepinephrine, pointing to a receptorial or postreceptorial site of the autonomic impairment.
Characterization of Apomucin Expression in Intrahepatic Cholangiocarcinomas and Their Precursor Lesions: An Immunohistochemical Study
Authors: Sasaki M , Nakanuma Y , Kim Y S
Published: November 2004

To date, seven apomucins have been characterized and their expression in malignant and premalignant lesions is under evaluation. In this study, we examined the expression of MUC1, MUC2, MUC3, and MUC5/6 apomucins in cholangiocarcinoma (CC) and biliary epithelial dysplasia. We used 14 liver samples from patients with hepatolithiasis and CC, 11 with hepatolithiasis showing biliary epithelial dysplasia, 31 with CC alone (19 hilar, 10 peripheral, and 2 unclassifiable), and 14 with combined hepatocellular–cholangiocellular carcinoma (HC–CC) and immunohistochemically characterized the expression profiles of apomucins. Nondysplastic biliary epithelial cells in the intrahepatic large bile ducts constantly expressed MUC3 apomucin. MUC5/6 and MUC3 apomucin expression was widespread in dysplastic biliary epithelial cells in hepatolithiasis, although the latter was reduced or absent in dysplastic foci. CC extensively expressed MUC1 apomucin and focally expressed MUC2 apomucin. In addition, CC of the hilar type, including those with hepatolithiasis, frequently expressed MUC3 apomucin (68% and 57%, respectively), whereas those of the peripheral type infrequently expressed MUC3 apomucin (10%) (

P

< .01). MUC5/6 apomucin was more frequently expressed in well–differentiated (89%), compared with poorly differentiated CC (42%) (

P

< .01). Cholangiocellular elements of combined HC–CC frequently expressed MUC1 apomucin, although they rarely expressed MUC2 and MUC3 apomucin and infrequently expressed MUC5/6 apomucin. The frequent and aberrant expression of “gastric type” MUC5/6 apomucin in biliary epithelial dysplasia, as well as in CC, suggests that biliary epithelial cells gain a gastric apomucin phenotype during carcinogenesis. MUC3 apomucin expression in CC is a marker that suggests that CC arises in the intrahepatic large bile ducts.
Changes in S–Adenosylmethionine Synthetase in Human Liver Cancer: Molecular Characterization and Significance
Authors: Cai J , Sun W , Hwang J , Stain S C , Lu S C
Published: November 2004

S

–adenosylmethionine synthetase (SAMS) catalyzes the formation of

S

–adenosylmethionine (SAM) and is essential to normal cell function. There are two forms of SAMS, liver–specific and nonliver–specific (often referred to as “kidney”), which are products of two different genes. SAMS isoenzymes differ greatly in kinetic parameters and sensitivity to inhibition by methionine analogs. The current work studied changes in SAMS and their significance in liver cancer. Northern blot analysis showed that while normal liver expresses only liver–specific SAMS, both HepG2 and HuH–7 cells express only nonliver–specific SAMS. Absence of liver–specific SAMS messenger RNA (mRNA) was not because of gene deletion or rearrangement but complete lack of gene transcription. Reverse–transcription polymerase chain reaction (RT–PCR) with liver–and kidney–specific SAMS primers showed that liver–specific SAMS mRNA was absent with only kidney SAMS mRNA present in HepG2, HuH–7, Hep3B, and HuH–1 cells, and four consecutive hepatocellular carcinoma (HCC) specimens. Normal liver tissues from the same patients express both forms of SAMS mRNA. As a result of the change in SAMS expression, SAMS activity was higher in HepG2 and HuH–7 cells at physiologically relevant methionine concentrations but lower at high (mmol/l) methionine concentrations than rat hepatocytes. Treatment with ethionine and seleno–D,l–ethionine, two inhibitors known to have I50 values 50 to 60 times lower against SAMS purified from Novikoff hepatoma cells as compared with SAMS purified from normal rat liver, resulted in increased cell lysis in HepG2 and HuH–7 cells but not cultured rat hepatocytes. These agents did not affect cellular adenosine triphosphate (ATP) levels but inhibited SAMS activity in HepG2 and HuH–7 cells when added to their protein extracts. In summary, expression of SAMS is altered in human liver cancer. This occurrence may provide a potentially exploitable target for cancer chemotherapy.
The Plasminogen–Activating System in Hepatic Stellate Cells
Authors: Leyland H , Gentry J , Arthur M J , Benyon R C
Published: November 2004

Urokinase plasminogen activator (uPA) generates plasmin, a process inhibited by plasminogen–activator inhibitor (PAI)–1 and localized to the cell surface by binding of uPA to a specific receptor. Plasmin degrades extracellular matrix (ECM) both directly and by activation of matrix metalloproteinases (MMPs). Because stellate cells play a central role in the pathogenesis of liver fibrosis both via production of ECM proteins and through secretion of MMPs, their contribution to plasmin generation was assessed. Stellate cells were prepared from rat liver and cultured on plastic. Northern analysis showed cellular expression of messenger RNA (mRNA) for PAI–1, uPA, and uPA receptor. Zymography/reverse zymography identified cell–surface–associated uPA activity and uPA and PAI–1 in culture media. Net uPA activity in culture media was maximal after 7 days in culture and then declined, whereas PAI–1 antigen levels remained consistently elevated between 7 and 21 days in culture. Stellate cell–mediated plasmin generation was also seen in

in vitro

cultures supplemented with plasminogen. Because hepatic stellate cells (HSCs) contain retinoids and release them on activation, the effect of retinoic acid on the plasminogen–activating system was also assessed. Treatment of cultured HSCs with retinoic acid (1 μmol/L) increased uPA secretion 2.6–fold but did not alter PAI–1. We conclude that stellate cells synthesize key components of the plasminogen–activating system and generate plasmin and therefore have the ability to regulate MMP activation. Upregulation of uPA synthesis by retinoic acid may have implications in matrix remodeling in sites of stellate cell activation in which high concentrations of retinoids may be achieved.
Aging of the Liver: Age–Associated Mitochondrial Damage in Intact Hepatocytes
Authors: Sastre J , Pallardó F V , Plá R , Pellín A , Juan G , O'Connor J E , Estrela J M , Miquel J , Viña J
Published: November 2004

Mitochondrial damage may be a major cause of cellular aging. So far, this hypothesis had only been tested using isolated mitochondria. The aim of this study was to investigate the involvement of mitochondria in aging using whole liver cells and not isolated mitochondria only. Using flow cytometry, we found that age is associated with a decrease in mitochondrial membrane potential (30%), an increase in mitochondrial size, and an increase in mitochondrial peroxide generation (23%). Intracellular peroxide levels were also increased. The number of mitochondria per cell and inner mitochondrial membrane mass did not change. Gluconeogenesis from glycerol or fructose (mitochondrial– independent) did not change with age, whereas it did from lactate (mitochondrial–dependent). The change in the rate of gluconeogenesis was not accompanied by changes in any of the following parameters: phosphoenolpyruvate carboxykinase or pyruvate carboxylase activities or mitochondrial ATP/ADP or cytosolic NADH/NAD+ ratios. This was caused by a decreased rate of malate export (to 20% of the controls) from mitochondria. The impairment of the mitochondrial malate transporter is posttranscriptional because its expression in

Xenopus

oocytes using polyadenylated RNA from livers of young or old animals did not change. Ketogenesis from oleate also fell in hepatocytes from old rats. Our results show, for the first time in intact cells, a correlation between age–associated impairment of cell metabolism and specific changes in mitochondrial function and morphology, supporting the hypothesis that mitochondrial damage plays a key role in aging.
Effect of Alanine on D–Galactosamine–Induced Acute Liver Failure in Rats
Authors: Maezono K , Mawatari K , Kajiwara K , Shinkai A , Maki T
Published: November 2004

The effect of high–dose alanine on survival and liver function in rats with acute liver failure caused by a lethal dose of d–galactosamine (d–gal) was studied. Greater than 90% of control animals died within 5 days after d–gal injection, but alanine significantly decreased mortality, even when treatment was started at 12 hours after d–gal injection. Alanyl–glutamine had a slight effect, but glucose produced no improvement. There was marked elevation of the plasma aspartate transaminase (AST) level, prolongation of the prothrombin time, and a decrease of the arterial ketone body ratio (AKBR) and hepatic adenosine triphosphate (ATP) content within 12 hours after d–gal injection. The AKBR decreased in parallel with the decrease of the hepatic ATP content. These parameters were significantly improved in alanine– treated rats at 48 hours after the induction of liver damage, which was just before control rats began to die. The hepatic ATP content was significantly greater in alanine–treated rats than in the other rats (including normal controls), but glucose pretreatment had no effect. It was also found that the liver labeling index of partially hepatectomized rats was significantly elevated by alanine administration at 3 hours before measurement. In conclusion, alanine is effective for the treatment of experimental acute liver failure, probably caused by promotion of ATP synthesis. Ala may be a good candidate for clinical application because of its preventive effect on hepatocyte necrosis and its promotive effect on liver regeneration.
A Novel Two–Nucleotide Deletion in the Ornithine Transcarbamylase Gene Causing Fatal Hyperammonia in Early Pregnancy
Authors: Schimanski U , Krieger D , Horn M , Stremmel W , Wermuth B , Theilmann L
Published: November 2004

Ornithine transcarbamylase (OTC) deficiency shows X–linked inheritance. Typically, symptomatic females (who constitute 15%–20% of all carriers) have markedly reduced enzyme activity and show first symptoms in late infancy or early childhood. Here we present the case of a previously asymptomatic 24–year–old woman who died of severe hyperammonemia associated with orotic aciduria but normal OTC activity in the fourth month of pregnancy. DNA analysis revealed a novel mutation in form of the deletion of two nucleotides (T892, G893) in exon 9 of the OTC gene, leading to a frame shift and an aberrant gene product. We suggest that OTC deficiency should be suspected in any patient who presents with hyperammonia in the presence of otherwise normal liver function.
Phenprocoumon for Prevention of Shunt Occlusion After Transjugular Intrahepatic Portosystemic Stent Shunt: A Randomized Trial
Authors: Sauer P , Theilmann L , Herrmann S , Bruckner T , Roeren T , Richter G , Stremmel W , Stiehl A
Published: November 2004

Development of stenosis or occlusion of the transjugular intrahepatic portosystemic stent shunt (TIPSS) is one of the major limiting factors in the long–term viability of this procedure. The efficacy of anticoagulation with heparin which is used in different centers is still unclear. In the present study, we evaluated the effect of phenprocoumon on shunt patency after TIPSS placement using Palmaz stents; 49 patients with Child's A and B cirrhosis, who underwent successful TIPSS placement were randomized into the treatment group (n = 24) who received phenprocoumon and a control group (n = 25). After 11 to 13 weeks, all patients were admitted and had a reevaluation that included control angiography by transjugular approach. Phenprocoumon treatment was stopped after the first reevaluation and both groups were followed for 1 year after randomization. During the 3–month treatment period 11 of 22 patients of the treatment group and 12 of 23 patients of the control group required reintervention because of an increased portosystemic gradient. Five of the 12 patients in the control group showed complete occlusion of the shunt, whereas no occlusion in the treatment group was observed (

P

< .05). During the mean follow–up of 8 months after the treatment was stopped, in both groups stenosis occurred in 50% of patients, but no further occlusion of the stent was observed. These data indicate that occlusion of the stent is related to thrombosis, whereas stenosis does not appear to be dependent on blood coagulation. In patients with preserved liver function occlusion of the shunt may be prevented by phenprocoumon treatment in the first 3 months after TIPSS placement. Thereafter shunt occlusion was not observed and further phenprocoumon treatment seemed unnecessary.
E–Cadherin Is Involved in the Intrahepatic Metastasis of Hepatocellular Carcinoma
Authors: Osada T , Sakamoto M , Ino Y , Iwamatsu A , Matsuno Y , Muto T , Hirohashi S
Published: November 2004

In human hepatocellular carcinoma (HCC), the liver is the major target organ of metastasis, which is known as intrahepatic metastasis. To analyze the mechanism of this metastasis, we established two sublines from the human HCC cell line Li7. Subline Li7HM produced multiple liver metastasis, whereas subline Li7NM never did so after intrasplenic injection into nude mice. Two–dimensional gel electrophoresis and immunoblot analysis showed that only Li7NM expressed vimentin and lacked E–cadherin expression, indicating that this clone had undergone epithelial–mesenchymal transition. We then transfected mouse E–cadherin complementary DNA into Li7NM cells and found that the transfectant cells (EM16.21B.3) formed liver metastasis (8/16 mice) after intrasplenic injection and liver tumors (11/13 mice) after intrahepatic injection, whereas the control cell line formed no tumors. These results suggest that E–cadherin plays an important role in the process of intrahepatic metastasis of HCC.
The Role of Adhesion Molecules in the Recruitment of Hepatic Natural Killer Cells (Pit Cells) in Rat Liver
Authors: Luo D , Vanderkerken K , Bouwens L , Kuppen P J , Baekeland M , Seynaeve C , Wisse E
Published: November 2004

Previous studies showed that blood large granular lymphocytes (LGL), which possess natural killer (NK) activity, develop within rat liver sinusoids into high–density (HD) and subsequently into low–density (LD) pit cells which show an increasing level and spectrum of tumor cytotoxicity. In this study, we investigated the role of adhesion molecules, such as CD2, CD11a, CD18, and CD54 in the recruitment of pit cells to the liver. Immunostaining for electron microscopy, and two color flow cytometry showed that most pit cells expressed CD2, CD11a, CD18, and CD54. After intravenous injections into rats with anti–CD2, anti–CD11a, and anti–CD18 antibodies, the number of pit cells per square millimeter in frozen sections of liver tissue decreased. Treatment of rats with zymosan increased the number of pit cells fivefold, whereas subsequent treatment with anti–adhesion–molecule antibodies resulted in approximately 60% lower number of pit cells. Anti–CD54, supposed to block CD54 expression on sinusoidal endothelial cells, also decreased the number of pit cells. The number of blood LGL was, however, not affected by these antibodies. These results indicate that blocking of CD2, CD11a, CD18, and CD54 antigens on blood LGL and/or liver endothelium decreased the number of pit cells in the liver. These adhesion molecules therefore play an important role in the recruitment of pit cells in the liver.
Effects of Varying Doses of Spironolactone Without and With Nitrates on Portal Vein Pressure and Kidney Function in Partial Portal Vein Ligated Rats
Authors: Van de Castelle M , Van Roey G , Nevens F , Fevery J
Published: November 2004

The optimal dose of spironolactone to reduce portal vein pressure, alone or in combination with isosorbide–5–mononitrate (IsMn), has not been identified. We studied the effects of 8 days oral treatment with spironolactone, IsMn or both on portal pressure, plasma volume and renal sodium handling in rats with partial portal vein ligation. At daily doses of 0.33; 0.50; 1, and 1.50 mg/kg, spironolactone reduced portal pressure (all

P

< .05) as compared with placebo. Only the highest dose significantly lowered plasma volume (10.1 ± 0.7 vs. 13.0 ± 0.3 mL;

P

< .03) and enhanced urinary fractional sodium excretion (0.73 ± 0.04 vs. 0.58 ± 0.03%;

P

< .03) and the (Na+)/(K+) ratio in urine (0.43 ± 0.04 vs. 0.30 ± 0.03;

P

< .02). IsMn at doses of 0.25; 0.50, and 1 mg/kg decreased portal pressure (all

P

< .05) without a change in plasma volume but with a tendency (not significant) to lower fractional sodium excretion. IsMn impaired free water clearance at doses of 0.5 and 1 mg/kg (

P

< .05). Combinations of spironolactone 1 mg/kg with IsMn 0.5 or 1 mg/kg have no additive effect on portal pressure compared with spironolactone or IsMn alone. The higher the dose of IsMn in the combination, the more the natriuretic effect of spironolactone is opposed. Low doses of spironolactone are as effective as a higher dose to reduce portal pressure. This reduced portal pressure was independent of changes in plasma volume and diuretic effect, which suggests that spironolactone might have a direct vasoactive effect on the splanchnic circulation. To counteract sodium retention of nitrovasodilators, combination with high doses of spironolactone seems advantageous.
Development of thermal flow simulation method for switch mode power supplies and its integration with electric circuit analysis (in the case of natural convection air cooled switch mode power supply)
Authors: Koizumi Katsuhiro , Joboji Akito , Nagahara Kuniaki , Ishizuka Masaru
Published: November 2004

This paper describes the development of a thermal flow simulation method for the design of electronic equipment. In the proposed sequence of analyses, the electric circuit analysis for estimating power loss in power semiconductor devices was integrated with the thermal analysis. The method was applied to the thermal design of a new switch mode power supply (SMPS). The analysis was carried out using a commercially available computational fluid dynamics (CFD) code. In reducing actual structural organizations of the printed circuit board (PCB) and the power semiconductor devices to simpler models, the method of experimental design (MED) was employed. Following the prescription of MED, the PCB was treated as a simple plate having the same thermal conductivity as epoxy resin, and the power semiconductor devices were modeled by hexahedral resistance network. The heat sources were a field effect transistor (FET) and a diode. Computation of power loss from the FET and the diodes is described and the accuracy of different estimation methods is discussed. The difference between measured and calculated temperature rises of the power semiconductor devices was found to be within approximately 10 K. © 2004 Wiley Periodicals, Inc. Heat Trans Asian Res, 33(8): 473–489, 2004; Published online in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/htj.20037
Differential expression of 37 selected genes in hormone‐refractory prostate cancer using quantitative taqman real‐time RT‐PCR
Authors: Fromont Gaëlle , Chene Laurent , Vidaud Michel , Vallancien Guy , Mangin Philippe , Fournier Georges , Validire Pierre , Latil Alain , Cussenot Olivier
Published: November 2004

Progression of prostate cancer to androgen independence remains the primary obstacle to improved survival. The development of more effective treatments depends on our understanding of the molecular events associated with the hormone‐refractory stage. We quantified, among 90 screened genes, the expression of 37 target genes, using real‐time quantitative RT‐PCR. Gene expression was studied in 13 samples of HPRC compared to 33 clinically localised cancers and normal prostate tissue. We identify 19 genes with significant differential expression in HRPC compared to localised prostate cancer. Genes with decreased expression included receptors for growth factors, MMR genes and the serine protease hepsin. Analysis of increased gene expression confirmed the importance of AR upregulation and highlighted genes not previously linked to HRPC, including enzymes involved in steroid synthesis and the antiapoptotic factor survivin. Progression of prostate cancer to the hormone‐refractory state is associated with differential gene expression, which may prove useful for both understanding disease progression and the development of new therapeutic approaches. © 2004 Wiley‐Liss, Inc.
Constraint satisfaction using soft quantifiers
Authors: Yager Ronald R.
Published: November 2004

Fuzzy sets and other methods have been used to model a softening of constraints in constraint propagation (CP) problems. Here, we suggest an approach to the softening of the CP problem at the meta level, in the process used to aggregate the satisfactions to the individual constraints. We discuss the possibility of using soft quantiﬁers such as ‘most’ to guide the process of aggregating the satisfactions to the individual constraints. Use is made of the ability to represent these soft quantiﬁers by fuzzy sets and the ability to implement their authorized aggregation by the ordered weighted averaging operator. Copyright © 2004 John Wiley & Sons, Ltd.
Differentiation of human colon adenocarcinoma cells alters the expression and intracellular localization of annexins A1, A2, and A5
Authors: Guzmán‐Aránguez Ana , Olmo Nieves , Turnay Javier , Lecona Emilio , Pérez‐Ramos Pablo , de Silanes Isabel López , Lizarbe M. Antonia
Published: November 2004

Butyrate induces differentiation and alters cell proliferation in intestinal–epithelial cells by modulation of the expression of several genes. Annexins are a superfamily of ubiquitous proteins characterized by their calcium‐dependent ability to bind to biological membranes; their involvement in several physiological processes, such as membrane trafficking, calcium signaling, cell motility, proliferation, and differentiation has been proposed. Thus, we have analyzed changes in annexin A1 (AnxA1), annexin A2 (AnxA2), and annexin A5 (AnxA5) levels and localization in human colon adenocarcinoma cells differentiated by butyrate treatment or by culture in glucose‐free inosine‐containing medium. The acquired differentiated phenotype increased dipeptidyl peptidase‐IV (DPP‐IV) expression and alkaline phosphatase (ALP) activity, two well known brush border markers. Butyrate induces cell differentiation and growth arrest in BCS‐TC2, BCS‐TC2.2, HT‐29, and Caco‐2 cells, increasing the levels of AnxA1 and AnxA5, whereas AnxA2 decreases except in Caco‐2 cells. Inosine‐differentiated cells present increased amounts of the three studied annexins, as occurs in spontaneously differentiated Caco‐2 cells. AnxA2 down‐regulation is not due to proteasome activation and seems to be related to the butyrate‐induced cell proliferation arrest; AnxA1 and AnxA5 expression is growth‐state independent. AnxA1 and AnxA5 are mainly found in the cytoplasm while AnxA2 is localized underneath the plasma membrane in cell‐to‐cell contacts. Butyrate induces changes in subcellular localization towards a vesicle‐associated pattern. Human colon adenocarcinoma cell differentiation is associated with an up‐regulation of AnxA1, AnxA2, and AnxA5 and with a subcellular relocation of these proteins. No correlation between annexin levels and tumorigenicity was found. Up‐regulation of AnxA1 could contribute to the reported anti‐inflammatory effects of butyrate in colon inflammatory diseases. © 2004 Wiley‐Liss, Inc.
Spatio‐temporal dynamics at rDNA foci: Global switching between DNA replication and transcription
Authors: Pliss Artem , Koberna Karel , Večeřová Jaromíra , Malínský Jan , Mašata Martin , Fialová Markéta , Raška Ivan , Berezney Ronald
Published: November 2004

We have investigated the in situ organization of ribosomal gene (rDNA) transcription and replication in HeLa cells. Fluorescence in situ hybridization (FISH) revealed numerous rDNA foci in the nucleolus. Each rDNA focus corresponds to a higher order chromatin domain containing multiple ribosomal genes. Multi‐channel labeling experiments indicated that, in the majority of cells, all the rDNA foci were active in transcription as demonstrated by co‐localization with signals to transcription and fibrillarin, a protein involved in ribosomal RNA processing. In some cells, however, a small portion of the rDNA foci did not overlap with signals to transcription and fibrillarin. Labeling for DNA replication revealed that those rDNA foci inactive in transcription were restricted to the S‐phase of the cell cycle and were replicated predominantly from mid to late S‐phase. Electron microscopic analysis localized the nucleolar transcription, replication, and fibrillarin signals to the dense fibrillar components of the nucleolus and at the borders of the fibrillar centers. We propose that the rDNA foci are the functional units for coordinating replication and transcription of the rRNA genes in space and time. This involves a global switching mechanism, active from mid to late S‐phase, for turning off transcription and turning on replication at individual rDNA foci. Once all the rRNA genes at individual foci are replicated, these higher order chromatin domains are reprogrammed for transcription. © 2004 Wiley‐Liss, Inc.