bluuebunny/crossref_metadata_2025 huggingface dataset:
Hepatic tocopherol content in primary hepatocellular carcinoma and liver metastases
Authors: Rocchi E , Seium Y , Camellini L , Casalgrandi G , Borghi A , D'Alimonte P , Cioni G
Published: July 2004

The high incidence of hepatocellular carcinoma (HCC) in cirrhosis, where previous studies have indicated a severe reduction in several antioxidant vitamin factors, prompted us to compare plasma liposoluble vitamins with tocopherol content in healthy and neoplastic liver tissue in humans. This, with a view to a more positive preventive dietary approach, given the conflicting results obtained by liposoluble vitamin dietary supplementation in different malignancies. Eleven patients with cirrhosis, 18 patients affected by cirrhosis with HCC, and 10 patients with liver metastases (LM) from digestive tract adenocarcinomas were compared with controls who had undergone perlaparoscopic cholecistectomy. Plasma α- and β-carotene, retinol and tocopherol, together with liver tocopherol, from both nonmalignant portions and malignant nodules of the same organ, were determined by high-performance liquid chromatography following a well-assessed technique. The results confirm a trend towards a reduction in circulating carotenoids and tocopherol in cirrhosis and in patients affected by cirrhosis with HCC. Tocopherol content in liver tissue is significantly decreased in cirrhosis (0.26 + 0.03 μmol/g prot., mean + SEM, P < .001) and in cirrhotic areas of the HCC group (0.31 + 0.02, P < .002), with respect to its content in liver specimens of healthy controls (0.46 + 0.03) and in healthy areas of the same organ in patients with LM (0.41 + 0.03). Tocopherol concentration is further reduced by 50% in malignant liver nodules of HCC, with respect to surrounding cirrhotic tissue, whereas in metastatic liver nodules from digestive neoplasms the tocopherol content is almost twice that of healthy surrounding areas. This unpredictable tocopherol behavior in liver specimens, of secondary as opposed to primary malignancies of the liver, affords further insight into the conflicting effects of liposoluble vitamins employed in the chemopreventive treatment of different malignant diseases, where hepatic tocopherol concentration show opposite trends: halved in primary HCC and doubled in LM of digestive adenocarcinomas, with respect to healthy controls.
Preventive and therapeutic effects in rats of hepatocyte growth factor infusion on liver fibrosis/cirrhosis
Authors: Matsuda Y , Matsumoto K , Yamada A , Ichida T , Asakura H , Komoriya Y , Nishiyama E , Nakamura T
Published: July 2004

Liver fibrosis/cirrhosis is characterized by hyper-accumulation of fibrous tissue components and is commonly observed in later or terminal states of chronic hepatic diseases. In ongoing work, we found that the administration of human recombinant hepatocyte growth factor (hrHGF) suppressed the onset of liver fibrosis/cirrhosis in several distinct models and accelerated the recovery from liver fibrosis/cirrhosis in rats. Repeated administration of porcine serum for 10 weeks to rats induced liver fibrosis without any accompanying hepatocellular injuries; in addition, the intravenous (i.v.) administration of hepatocyte growth factor (HGF) to these rats suppressed increases in fibrous components and hydroxyproline contents in the liver, thus preventing the onset of liver fibrosis. Repeated administration of dimethylnitrosamine (DMN) for four weeks induced liver cirrhosis, as characterized by the hyper-accumulation of fibrous components, infiltration of mononuclear leukocytes, and hepatic dysfunction. When HGF was injected daily for four weeks along with DMN-treatment, the onset of DMN-induced hepatic fibrosis/cirrhosis was suppressed; the numbers of infiltrating mononuclear cells, fibrous tissue components, and hydroxyproline content in the liver were decreased. When HGF was injected for two weeks following four weeks of DMN-treatment, HGF accelerated the recovery from liver cirrhosis and prevented death due to hepatic dysfunction. Likewise, HGF-injection suppressed the onset of liver fibrosis, when liver fibrosis had been induced by long-term treatment with carbon tetrachloride (CCl4). Thus, the administration of HGF holds great promise for treating subjects with liver fibrosis/cirrhosis as a result of chronic hepatic injury.
Cholesterol supplementation prevents necrosis and inflammation but enhances fibrosis in alcoholic liver disease in the rat
Authors: Nanji A A , Rahemtulla A , Daly T , Khwaja S , Miao L , Zhao S , Tahan S R
Published: July 2004

Based on studies that show a role for the low-density lipoprotein (LDL)-receptor in arachidonic acid delivery and eicosanoid synthesis in macrophages, the present study investigated the effect of cholesterol supplementation on pathological changes and thromboxane (TX) synthesis in alcoholic liver injury. Male Wistar rats were intragastrically fed ethanol with either corn oil or fish oil for 1 month. Control rats received isocaloric amounts of dextrose instead of ethanol. An additional group of rats fed either ethanol or dextrose with fish oil or corn oil were supplemented with 1% cholesterol. At the time of killing, all rats had the following evaluated: liver histopathology, lipid peroxidation, liver and plasma thromboxane levels, plasma endotoxin and messenger RNA (mRNA) levels of LDL-receptor, tumor necrosis factor α (TNF-α), cyclooxygenase (Cox)-1 and -2, and transforming growth factor β (TGF-β). Rats fed ethanol with either fish oil or corn oil developed fatty liver, necrosis, inflammation, and central vein collagen deposition. Cholesterol supplementation enhanced the degree of fibrosis but prevented necrosis and inflammation. These alterations in pathological changes by cholesterol were accompanied by absent TNF-α and Cox-2 mRNAs, decreased thromboxane levels, decreased lipid peroxidation, and increased TGF-β mRNA. Cholesterol enrichment of the diet thus decreases proinflammatory components, but enhances fibrosis in ethanol-fed rats.
ClQ synthesis by tissue mononuclear phagocytes from normal and from damaged rat liver: Up-regulation by dexamethasone, down-regulation by interferon gamma, and lipopolysaccharide
Authors: Armbrust T , Nordmann B , Kreiβig M , Ramadori G
Published: July 2004

The subcomponent of complement C1, C1q, mediates complement activation via the classical pathway, and therefore may play an important role in the inflammatory processes in which complement activation is involved. The aim of our study was to investigate C1q synthesis by macrophages of normal and of acutely damaged livers. The localization of C1q in liver tissue was studied by immunohistochemistry. Rat liver tissue macrophages were isolated from normal as well as from acutely damaged (carbon tetrachloride model) liver, and were separated into small, monocyte-like phagocytes and large, mature tissue macrophages, as revealed by immunocytochemistry. C1q gene expression was studied by endogeneous labeling of newly synthesized proteins, immunoprecipitation, and sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), and by reverse-transcription polymerase chain reaction (RT-PCR) of C1qB messenger RNA (mRNA). Semiquantitative analysis was performed by Northern blotting of total RNA and hybridization with the radioactively labeled RT-PCR product. C1 esterase inhibitor synthesis was studied in parallel. For comparison, C1q and C1-inhibitor synthesis were also investigated in blood monocytes and peritoneal macrophages. C1q was weakly detectable in sinusoidal cells of the normal liver. C1qB mRNA, as well as constitutive synthesis and secretion of C1q, was clearly detected in freshly isolated and cultured Kupffer cells from normal rat liver. In comparison, newly recruited “inflammatory” macrophages from damaged rat liver synthesized considerably lower amounts of the protein, similar to what was found in the monocyte-like macrophages of normal liver and in peritoneal macrophages. Monocyte C1qB mRNA was not detected even by RT- PCR, and remained undetectable during the time in culture. Similar behavior was observed for C1-inhibitor synthesis. Treatment of the cultures with interferon gamma (IFN-γ) or lipopolysaccharide (LPS) strongly decreased, whereas treatment with dexamethasone strongly increased C1q gene expression in the macrophage populations, and induced C1qB mRNA in cultured monocytes, as revealed by RT-PCR. Kupffer cells of normal liver may produce considerable amounts of C1q, whereas the inflammatory macrophages of the acutely damaged liver may not be so important for the synthesis of C1q.
Glycine protects against hepatocyte killing by KCN or hypoxia by preventing intracellular Na+ overload in the rat
Authors: Carini R , Bellomo G , De Cesaris M , Albano E
Published: July 2004

Glycine has been shown to prevent hepatocyte death induced by anoxia and by several toxic agents. However, the mechanisms responsible for such a cytoprotective effect have not yet been entirely clarified. We have previously shown that an uncontrolled increase in intracellular Na+ is critical for hepatocyte killing induced by adenosine triphosphate (ATP) depletion. We herein report that protection by glycine (2 mmol/L) against cytotoxicity induced in isolated rat hepatocyte by potassium cyanide (KCN) or hypoxia was associated with the prevention of cytosolic Na+ accumulation. The addition of the Na+ ionophore, monensin, abolished the effects of glycine on both Na+ increase and cytotoxicity. Pretreating hepatocytes with the glycine- receptor antagonist, strychnine (1 mmol/L), similarly prevented Na+ overload and cell killing. Glycine at high concentrations and strychnine are known to block Cl− channels in many cell types. Consistently, we have observed that glycine and strychnine prevented the increase of intracellular Cl−levels caused by hypoxia or KCN. Incubation of hepatocytes in a Cl−-free medium, obtained by substituting chloride with membrane-impermeable gluconate, significantly reduced Na+ accumulation and cell killing triggered by hypoxia or KCN. Both these effects were abolished by the addition of monensin. The cytoprotective action exerted by hepatocyte incubation in the Cl(−)-free medium was, however, lost when membrane-permeable nitrate, which allowed Na+ accumulation, was used instead to replace chloride. Altogether, these results indicate that glycine inhibition of Cl− conductance protects against hepatocyte killing induced by KCN and hypoxia by interfering with intracellular Na+ accumulation triggered by ATP depletion.
Tumor necrosis factor-induced, superoxide-mediated neutrophil accumulation in cold ischemic/reperfused rat liver
Authors: Shibuya H , Ohkohchi N , Tsukamoto S , Satomi S
Published: July 2004

The mechanisms of hepatic ischemia/reperfusion injury are complicated and multifactorial. This study was designed to examine superoxide generation and neutrophil accumulation in cold ischemic-reperfused rat livers after elimination of Kupffer cells and to determine the role of superoxide/tumor necrosis factor (TNF) interactions. Rat Kupffer cells were eliminated by liposome-encapsulated dichloromethylene diphosphonate injected intravenously. Livers from control and treated rats were isolated and preserved in University of Wisconsin solution (4°C) for 0, 12, and 24 hours and then perfused for 60 minutes with oxygenated Krebs-Henseleit bicarbonate buffer (37°C) by adding neutrophils into the perfusate. Superoxide generation was measured by using real-time chemiluminescence (CL) during perfusion, and neutrophil accumulation was assessed by measuring myeloperoxidase activity in the liver tissue. In the control livers, CL intensity markedly increased on reoxygenation, and after neutrophil infusion it increased again with a lag period of 10 minutes. Total CL intensity and myeloperoxidase activity increased with the duration of cold preservation. TNF release into the effluent perfusate was detectable only after 24 hours of preservation, and lactate dehydrogenase release was high. Elimination of Kupffer cells attenuated CL intensity and TNF and lactate dehydrogenase release and resulted in reduced myeloperoxidase activity. Electron microscopy revealed amelioration of hepatocyte swelling and endothelial cell disruption when Kupffer cells were eliminated. After 24 hours of preservation, superoxide generation was inhibited in the control livers by anti-TNF antiserum, whereas TNF release was not inhibited by superoxide dismutase. These results suggest that TNF induces superoxide generation by Kupffer cells, which mediates neutrophil accumulation and causes cellular injury in the initial phase of reperfusion.
Effects of S -adenosyl-L-methionine on hepatic and renal oxidative stress in an experimental model of acute biliary obstruction in rats
Authors: Gonzalez-Correa J A , De La Cruz J P , Martin-Aurioles E , Lopez-Egea M A , Ortiz P , de la Cuesta Sanchez F
Published: July 2004

We used an animal model of extrahepatic biliary obstruction of 7 days' duration to study the production of thiobarbituric acid reactive substances (TBARS), total glutathione (TG), reduced glutathione (GSH), and oxidized glutathione (GSSG), and the enzymatic activities of GSH- peroxidase, GSSG-reductase, and GSH-transferase. Four groups of six rats each were treated with saline, drug solvent, S-adenosyl-L-methionine (SAM) 5 mg/kg/d, subcutaneously, or SAM 10 mg/kg/d, subcutaneously. Extrahepatic biliary obstruction increased TBARS. SAM had the dose-dependent effects of inhibiting TBARS production and increasing TG content, mainly as a result of the increase in GSH. The activity of GSH-peroxidase and GSH-transferase was also significantly increased. In renal tissue these effects were statistically significant only in animals given the higher dose of SAM. In liver we found a reduction in biochemical values indicative of liver damage. We conclude that effect of SAM on hepatorenal function is strongly influenced by the drug's ability to reestablish equilibrium after oxidative tissue stress.
Severe radiation-induced liver disease following localized radiation therapy for biliopancreatic carcinoma: Activation of hepatic stellate cells as an early event
Authors: Sempoux C , Horsmans Y , Geubel A , Fraikin J , Van Beers B E , Gigot J , Lerut J , Rahier J
Published: July 2004

Radiation-induced liver disease is recorded as a form of veno-occlusive disease. Its pathogenesis remains unclear even if the initial injury likely occurs in the endothelial cells of central veins. The aim of our study was to investigate liver morphological features in relation to α-isoform of smooth muscle actin expression in hepatic stellate cells in six patients treated by localized radiotherapy on the biliopancreatic area. Within the month after completion of treatment, an activation of hepatic stellate cells strictly confined to irradiated areas and coinciding with congestive changes was observed. At a later stage, collagen deposition gradually increased, replacing the congestive and destroyed areas. This new fibrotic tissue also contained numerous α-smooth muscle positive cells. Our data suggest that early hepatic stellate cells activation coinciding with congestive changes plays an important role in radiation liver injury and ensuing fibrosis.
Association of a tumor necrosis factor promoter polymorphism with susceptibility to alcoholic steatohepatitis
Authors: Grove J , Daly A K , Bassendine M F , Day C P
Published: July 2004

Twin concordance studies suggest that genetic factors play a role in determining why only a minority of heavy drinkers develop hepatitis and cirrhosis. Tumor necrosis factor α (TNF-α) has emerged as the “final common pathway” in the pathogenesis of alcohol-related hepatic necro-inflammation. We have examined the frequency of the two recently described polymorphisms of the TNF-α promoter in 150 patients with biopsy-proven alcoholic liver disease and 145 healthy volunteers. There was a significant excess of the rare allele (TNFA-A; G(−238) → A) at position −238 in patients with steatohepatitis compared with controls or patients without this lesion. This is consistent with previous suggestions that the TNFA-A allele, which falls within a putative Y regulation box of the TNF-α promoter, is associated with increased TNF-α expression. No differences were observed for the polymorphism at position −308.
Nitric oxide formation lowers norepinephrine-induced intrahepatic resistance without major effects on the metabolism in the perfused rat liver
Authors: Weidenbach H , Nussler A K , Shu Z , Adler G , Beckh K
Published: July 2004

Nitric oxide (NO) and norepinephrine are potent vasoactive agents that are involved in the control of portal blood flow. We have studied NO and norepinephrine in a non-recirculated rat liver perfusion to analyze their influence on portal flow and hepatic metabolism. Animals were either pretreated with endotoxin (4 hours; 10 mg/kg intraperitoneally) to activate the inducible NO synthase (NOS2), or used without pretreatment for the constitutive NO synthase (NOS3). In both groups, portal flow, bile flow, bile secretion, and the sinusoidal bile acid uptake were reduced by norepinephrine with a simultaneous increase of glucose and lactate output. The addition of the substrate for NO synthesis, L-arginine (0.5 mmol/L), to the perfusate markedly inhibited the effect of 0.1 μmol/L norepinephrine on portal flow from −2.6 ± 0.32 to 0.3 ± 0.1 mL/g/10 min in endotoxin-treated animals, and from −2.9 ± 0.45 to 0.77 ± 0.29 mL/g/10 min in the untreated ones. In contrast, neither NO formation after L-arginine supplementation nor inhibition of NO synthesis via the structural analogue (N
G-monomethyl-L-arginine [L-NMMA]) changed cholestatic and glycogenolytic effects caused by norepinephrine. Only the sinusoidal bile acid uptake was reduced following increased NO formation. Thus, we conclude that endogenous NO formation prevents α-catecholaminergic-increased intrahepatic resistance without a major influence on the metabolic effects.
Genetic and morphological findings in progressive familial intrahepatic cholestasis (Byler disease [PFIC-1] and Byler syndrome): Evidence for heterogeneity
Authors: Bull L N , Carlton V E , Stricker N L , Baharloo S , DeYoung J A , Freimer N B , Magid M S , Kahn E , Markowitz J , DiCarlo F J , McLoughlin L , Boyle J T , Dahms B B , Faught P R , Fitzgerald J F , Piccoli D A , Witzleben C L , O'Connell N C , Setchell K D , Agostini R M , Kocoshis S A , Reyes J , Knisely A S
Published: July 2004

Byler disease (ByD) is an autosomal recessive disorder in which cholestasis of onset in infancy leads to hepatic fibrosis and death. Children who have a clinically similar disorder, but are not members of the Amish kindred in which ByD was described, are said to have Byler syndrome (ByS). Controversy exists as to whether ByD and ByS (subtypes of progressive familial intrahepatic cholestasis [PFIC]) represent one clinicopathological entity. The gene for ByD has been mapped to a 19-cM region of 18q21-q22. PFIC caused by a lesion in this region, including ByD, can be designated PFIC-1. Examination of haplotypes in siblings with ByS in two unrelated non-Amish families showed that the gene(s) responsible for their disorder(s) did not lie in the PFIC-1 candidate region. On light microscopy and transmission electron microscopy (TEM), liver tissue differed between Amish children with PFIC-1, who had coarsely granular bile and at presentation had bland intracanalicular cholestasis, and the children with ByS in the two non-Amish families, who had amorphous or finely filamentous bile and at presentation had neonatal hepatitis. Bile acid composition of bile also differed: In the Amish children with PFIC-1 and in one ByS family, the proportional concentration of chenodeoxycholic acid (CDCA) in bile was low compared with normal bile; in the other ByS family, it was only slightly reduced. Genetic analysis and light microscopy and TEM of liver may help distinguish PFIC-1 from other forms of ByS.
Extramedullary hematopoiesis in the adult mouse liver is associated with specific hepatic sinusoidal endothelial cells
Authors: Cardier J E , Barberá-Guillem E
Published: July 2004

In previous work, two anatomically distinct-liver sinusoid endothelial cells (LEC): LEC-1 and LEC-2, have been described. We also reported that extramedullary hepatic hematopoiesis occurs only in close contact with LEC-1, suggesting that these cells may provide the microenvironment necessary for the maintenance and growth of hematopoietic cells. In the present work, we studied the capacity of LEC-1 and LEC-2 to maintain in vitro hematopoiesis. LEC-1 and LEC-2 were isolated and cloned from livers of adult mice. Bone marrow cells (BM) enriched with primitive hematopoietic progenitors were isolated from day-2, post-5-FU-treated mice (5-FUBMC). LEC-1 supported the maintenance and differentiation of hematopoietic progenitors for more than 6 weeks in vitro. In contrast, LEC-2 cells poorly supported the proliferation of hematopoietic cells for only two weeks of the co- culture. LEC-1 and 5-FUBMC cocultures showed cobblestone-area formation and the presence of hematopoietic progenitors that are able to form colonies (CFC) in the adhering fraction after six weeks of coculture. LEC-1 co-cultures treated with a cocktail of cytokines (stem cell factor, interleukin [IL]1α, IL-3, and Epo) showed that megakaryocyte (CFU-Mk) and erythrocyte progenitors (BFU-e) were present during the entire period of the culture. Granulocyte-macrophage progenitors (CFU-GM) were present only during the first three weeks of the culture. These results suggest that LEC-1, but not LEC-2, provide an appropriate hematopoietic microenvironment for supporting the proliferation and differentiation of primitive hematopoietic cells. This could explain the anatomical restriction of hematopoietic cells for growing in LEC-1 domains during liver extramedullary hematopoiesis.
Transferrin receptor recycling in rat hepatocytes is regulated by protein phosphatase 2A, possibly through effects on microtubule-dependent transport
Authors: Runnegar M , Wei X , Berndt N , Hamm-Alvarez S F
Published: July 2004

To understand the regulation of receptor-mediated endocytosis in hepatocytes, we have used two specific inhibitors of serine-threonine protein phosphatases (PP), microcystin (MCYST) and okadaic acid (OKA) as probes to alter protein phosphorylation in hepatocytes. We have then examined the impact of these changes on the specific binding and uptake of transferrin (Tf) in hepatocytes. The measurement of PP activity in hepatocyte lysates showed that OKA and MCYST shared a common inhibition of protein phosphatase 2A (PP2A). Our results showed that both OKA (250 nmol/L) and MCYST (500 nmol/L) significantly reduced Tf uptake at steady state (P ≤ .05). The measurement of Tf internalization after 15 minutes in protein phosphatase inhibitor-pretreated cells revealed that the initial uptake was also significantly reduced. Binding studies showed that pretreatment with either of the phosphatase inhibitors did not result in significant changes in the Kd for Tf binding to transferrin receptor (TfR). Additionally, no significant changes in the number of TfR in the plasma membrane were observed in phosphatase inhibitor-pretreated cells. The treatment of hepatocytes with nocodazole (NOC), which results in microtubule disassembly and inhibition of microtubule-based vesicle transport, caused comparable reductions in initial and steady state levels of transferrin accumulation. The changes in transferrin accumulation by both phosphatase inhibitors and nocodazole were accompanied by redistribution of the microtubule-anchored Golgi apparatus and lysosomal network from the perinuclear region to the cell periphery. Our data show that the regulation of Tf uptake by receptor-mediated endocytosis is mediated by PP2A and additionally may occur through regulation of microtubule-based vesicle transport.
Mini-microabscess syndrome in liver transplant recipients
Authors: MacDonald G A , Greenson J K , DelBuono E A , Grady W M , Merion R M , Frank T S , Lucey M R , Appelman H D
Published: July 2004

Cytomegalovirus (CMV) is a significant cause of morbidity in immunosuppressed patients. It is characterized in the liver by parenchymal microabscesses, usually containing CMV-infected cells. However, not all hepatic microabscesses are due to CMV infection. In 1992, we described “mini” microabscess (MMA) syndrome, a distinct clinical syndrome that occurs in transplanted livers. This report analyzes the clinical and laboratory features of 57 cases of MMA syndrome occurring in 52 patients and compares these with 19 biopsy-proven cases of CMV infection. The diagnosis of MMA syndrome can only be made histologically. The microabscesses are smaller and more numerous than in CMV infection, and there are no viral inclusions present. CMV DNA could not be detected in liver biopsy specimens with MMAs by using “nested” polymerase chain reaction (PCR), indicating that MMA syndrome is not caused by CMV infection. The pattern of liver enzyme and bilirubin elevation is predominantly hepatocellular, with transaminase levels elevated, on average, six to eight times the upper limit of normal. The clinical features of MMA syndrome are that it predominantly affects female (40 of 52 patients) orthotopic liver transplant (OLT) recipients of all ages (range, 11 months to 66.9 years). MMA syndrome is unrelated to the indication for initial OLT and tends to occur later after transplantation than CMV infection (median, 91 days post-OLT vs. 32 days for CMV hepatitis). Although the etiology of MMA syndrome is not clear, it does not appear to adversely affect graft or patient survival.
Long-term consequence of rat orthotopic liver transplantation with and without hepatic arterial reconstruction: A clinical, pathological, and hemodynamic study
Authors: Imamura H , Rocheleau B , Côté J , Huet P M
Published: July 2004

Our aim was to investigate the time-related changes in various parameters following orthotopic rat liver transplantation with (AOLT) and without (NOLT) arterial reconstruction in male Lewis rats. Body weight and biochemical parameters were measured weekly, and a liver biopsy was obtained at 4, 8, and 12 weeks. Hemodynamics were evaluated at 12 weeks using the microsphere technique and compared with matched controls. Following AOLT, rats gained weight normally without any noticeable complication. In NOLT, two subgroups (NOLT-1 and NOLT-2) could clearly be identified retrospectively. In the NOLT-1 group, the body weight increased normally, although animals presented transient cholestasis. In these rats, the ductular proliferation found at 4 weeks had regressed by the 12th week with near-normal biopsies. By contrast, in the NOLT-2 group, rats did not gain body weight and had persistent cholestasis. Marked ductular proliferation with increasing fibrosis was observed, resulting in a secondary biliary cirrhosis by the 12th week. Surprisingly, rearterialization of the grafted liver occurred in both NOLT-1 and NOLT-2 irrespective of their clinical course. All transplanted rats showed portal hypertension with marked portosystemic shunts, probably caused by the portal cuff. However, a hyperdynamic circulatory state was only observed in the NOLT-2 group with cirrhotic changes. These findings further show the combined role of an intact hepatic innervation and of hepatocellular insufficiency in the genesis of the hyperdynamic circulatory state associated with portal hypertension.
Thyroid abnormalities in chronic viral hepatitis and their relationship to interferon alfa therapy
Authors: Deutsch M , Dourakis S , Manesis E K , Gioustozi A , Hess G , Horsch A , Hadziyannis S
Published: July 2004

The prevalence of antithyroid peroxidase antibodies (ATPO) and/or of thyroid dysfunction was studied in 422 patients with chronic viral hepatitis C, B, and D. Baseline results were compared with those during and 6 months after interferon alfa (IFN-α) therapy. The overall prevalence of ATPO among untreated patients was 14.1%, with no significant differences between chronic hepatitis C, B, or D, as well as between males and females. However, high ATPO titers (≥ 18 IU/mL) clustered significantly among females (8.7% vs. 3.4%; P = .022), especially those with chronic hepatitis C (11.2% vs. 3.6%; P = .036). Before treatment, 3.7% of the patients had thyroid dysfunction, mostly hypothyroidism (3.5%), the latter increasing to 14.3% among patients with ATPO titers ≥ 18 IU/mL. IFN-α treatment significantly increased overall thyroid dysfunction (9.7%; P = .001) and hypothyroidism (7.8%; P = .01), particularly among patients with high baseline ATPO (38.5%; P = .0002). Six months after stopping IFN-α treatment, the prevalence of thyroid dysfunction was 8.0%, still significantly higher than at baseline. By multivariate analysis, the only predictor positively associated with pre- or on-treatment hypothyroidism was the baseline titer of the ATPO antibodies (relative risk [RR], 3.0 and 3.8 per each log titer increase, respectively). In conclusion, patients with chronic viral hepatitis on IFN-α treatment exhibit an almost threefold increase of baseline thyroid dysfunction, persisting long after the end of therapy. High ATPO titers, clustering among females, particularly those with hepatitis C, represent the only predictor of pre- and on-treatment hypothyroidism by multivariate analysis. Patients with chronic viral hepatitis, especially females, should be tested for ATPO and thyroid function and monitored during and posttreatment for free thyroxin (FT4) and thyroid- stimulating hormone (TSH) levels.
Effect of lamivudine on morphology and function of mitochondria in patients with chronic hepatitis B
Authors: Honkoop P , De Man R A , Scholte H R , Zondervan P E , Van Den Berg J W , Rademakers L H , Schalm S W
Published: July 2004

Nucleoside analogues can induce mitochondrial dysfunction leading to severe clinical syndromes. Lamivudine, a new nucleoside analogue, is an active inhibitor of hepatitis B viral replication without apparent clinical toxicity. To assess subclinical mitochondrial toxicity, we studied the morphology and function of the mitochondrial system in 15 patients treated with lamivudine. Morphology was investigated by routine histological evaluation and electron-microscopic studies of mitochondria in liver biopsy specimens. Mitochondrial function was assessed by 2-keto[1-14C] isocaproic acid decarboxylation (KICA breath test) and by measuring the activity in liver biopsy specimens of the mitochondrial enzymes encoded by nuclear and mitochondrial DNA (mt-DNA) (complex I and IV) as well as a mitochondrial and a cytosolic enzyme both encoded by nuclear DNA only (complex II and lactic dehydrogenase [LDH]). All 15 patients underwent a liver biopsy before treatment and a KICA breath test before and during treatment; 13 agreed to undergo a repeat liver biopsy during lamivudine treatment. Liver tissue with no or minimal fibrotic changes from 7 patients treated for 6 months with lamivudine was suitable for assessment of the mitochondrial enzyme activity. We observed no signs of toxicity by routine histological or electron-microscopic evaluation. KICA breath tests revealed no differences in either peak exhalation or the area under the curve from 0 to 60 minutes between healthy controls (3.0% and 19.3%), untreated patients with chronic hepatitis B (3.4% and 19.3%), and patients treated with lamivudine (3.1% and 20.6%). The activities of the mt-DNA-encoded enzymes remained normal after lamivudine therapy. Unexpectedly a significant decrease in the activity of nuclear-DNA-encoded enzymes in patients with chronic hepatitis B in comparison with normal controls was found. The mean activity of complex II dropped from 45.3 to 20.0 μmol · min−1, that of lactic dehydrogenase from 106 to 44 μmol · min−1 (Wilcoxon rank sum; P < .05). In conclusion, no subclinical signs of mitochondrial toxicity resulting from lamivudine therapy for 6 months were observed.
Synergistic inhibition of hepadnaviral replication by lamivudine in combination with penciclovir in vitro
Authors: Colledge D , Locarnini S , Shaw T
Published: July 2004

Lamivudine ([-]-β-L-2′,3′-dideoxy-3′-thiacytidine [3TC]) and penciclovir (9-[2-hydroxy-1-(hydroxymethyl)ethoxymethyl]guanine [PCV]) are potent inhibitors of hepatitis B virus (HBV) replication. Both drugs have entered phase III clinical trials for treatment of chronic HBV infection. 3TC and PCV are deoxycytidine and deoxyguanosine analogs, respectively, and their modes of action and how they interact are matters of both theoretical and practical interest. We compared the antiviral activities of 3TC and PCV alone and in combination in primary duck hepatocyte (PDH) cultures derived from ducklings congenitally infected with the duck hepatitis B virus (DHBV). 3TC and PCV inhibited DHBV replication to a comparable extent when used alone (50% inhibitory concentrations with 95% confidence intervals were 0.55 [0.50-0.59] μmol/L for 3TC and 0.35 [0.27-0.43] μmol/L for PCV), and in combination, the two nucleoside analogs acted synergistically over a wide range of clinically relevant concentrations. Synergy between PCV and 3TC was also observed in acutely infected cells and in “washout” experiments designed to assess the persistence of antiviral activity after drug removal. Furthermore, the combination was more effective in reducing the normally recalcitrant viral covalently closed circular (CCC) DNA form of DHBV than either drug alone. These results suggest that combinations of 3TC and PCV may act synergistically against HBV in vivo.
Inhibition of pancreaticobiliary secretion by loperamide in humans
Authors: Thimister P W L , Hopman W P , van Roermund R F C , Willems H L , Rosenbusch G , Woestenborghs R , Jansen J B
Published: July 2004

Loperamide, a peripherally acting opiate receptor agonist with antidiarrheal action, inhibits ileal and colonic motor function. It was determined whether loperamide also affects gallbladder emptying and pancreatic enzyme secretion in humans. Plasma cholecystokinin (radioimmunoassay), gallbladder volume (ultrasonography), and intraduodenal bilirubin and amylase output (spot sampling) were measured at regular intervals before and during intraduodenal perfusion of an amino acid meal in 8 healthy subjects: once without and once with pretreatment of 8 mg loperamide, ingested 13 and 4 hours before the start of the meal. Loperamide decreased basal amylase output from 3.2 ± 0.5 to 1.0 ± 0.5 kU/h (P < .005) and abolished basal bilirubin output (21 ± 5 vs. 0 ± 0 micromol/h; P < .005) into the duodenum. Loperamide increased basal gallbladder volume from 28 ± 4 to 39 ± 4 mL (P < .0001) but was without effect on basal plasma cholecystokinin (2.7 ± 0.3 vs. 3.0 ± 0.3 pmol/L). During the amino acid meal, pretreatment with loperamide inhibited amylase output from 5.1 ± 0.8 to 1.6 ± 0.4 kU/h (P < .001), bilirubin output from 39 ± 6 to 18 ± 6 micromol/h (P < .0005) and gallbladder contraction from 47% ± 3% to 26% ± 6% (P < .05), whereas loperamide enhanced amino acid-stimulated plasma cholecystokinin from 4.5 ± 1.6 to 7.6 ± 1.0 pmol/L (P < .05). It is concluded that loperamide inhibits basal and amino acid-stimulated gallbladder motility and intraduodenal output of bilirubin and amylase, despite an enhanced postprandial cholecystokinin release.
Elevated glomerular and blood mononuclear lymphocyte nitric oxide production in rats with chronic bile duct ligation: Role of inducible nitric oxide synthase activation
Authors: Criado M , Flores O , Ortiz M C , Hidalgo F , Rodriguez-Lopez A M , Eleno N , Atucha N M , Sanchez-Rodriguez A , Arevalo M , Garcia-Estan J , Lopez-Novoa J M
Published: July 2004

Recent work indicates that nitric oxide (NO) plays an important role in the systemic and renal alterations of cirrhosis. In the present study, we have evaluated whether the inducible NO synthase (iNOS) isoform participates in the enhanced renal and systemic NO production of a rat model of cirrhosis. In vitro and in vivo experiments were performed in rats subjected to chronic bile duct ligation (BDL) and in sham-operated (SO) animals. Plasma nitrite (3.1 ± 0.1 micromol/L in SO and 6.6 ± 0.2 micromol/L in BDL), glomerular nitrite production (6.4 ± 0.1 vs. 9.8 ± 0.1 nmol/24h/7,000 glomeruli, respectively), and mononuclear lymphocyte cells nitrite production (0.3 ± 0.04 vs. 0.6 ± 0.12 nmol/10(6) cells, respectively) were all significantly higher in BDL than in SO. Moreover, mononuclear lymphocytes and glomeruli from BDL rats showed an increased expression of macrophage-type iNOS, detected by Western blot. Kidneys from BDL animals also showed an increased calcium-independent NO synthase activity, compared with those from SO rats. Constitutive endothelial-type NO synthase expression in glomeruli or the activity of calcium-dependent NO synthase in whole kidney did not show differences between BDL and SO rats. In cultured mesangial cells from normal rats, the addition of plasma from BDL but not of plasma from SO significantly stimulated (35%) nitrite production and increased the expression of macrophage-type iNOS. In addition, administration of aminoguanidine (AG), a preferential iNOS inhibitor, elevated dose-dependently mean arterial pressure in both groups, but this increase was greater in BDL (26.5 ± 4.4 mm hg) than in SO (13.6 ± 2.6). In BDL, AG also increased sodium and water excretion and glomerular filtration rate. In contrast, there were only small nonsignificant changes in SO animals. Therefore, these results indicate that the expression, activity and production of NO in kidneys, glomeruli, and mononuclear lymphocyte cells is elevated in BDL rats, and this is partly because of a plasma-derived substance(s), which stimulates iNOS formation. The amelioration of the arterial hypotension and the associated reduced excretory levels of these cirrhotic animals by aminoguanidine further support the involvement of the inducible NO synthase isoform in the renal alterations observed in BDL animals.
Factors regulating tumor pressure in hepatocellular carcinoma and implications for tumor spread
Authors: Tanaka T , Yamanaka N , Oriyama T , Furukawa K , Okamoto E
Published: July 2004

This study was performed to investigate factors determining tumor pressure (TP) in hepatocellular carcinoma (HCC) and a correlation of pressure gradient between TP and portal vein pressure (PVP) with tumor spread. TP and hepatic tissue pressure (HTP) were measured in 70 patients who underwent hepatic resection for HCC. Changes in TP after hepatic artery occlusion (HAO) or portal vein occlusion (PVO) were observed. HTP was elevated with worsening chronic liver disease, and exhibited a significant positive correlation with PVP. The TP was significantly higher in the encapsulated and the nonnecrotic HCCs. The pressure gradient (TP-PVP) was significantly greater in the encapsulated HCCs, especially in those with microscopic portal vein invasion (vp) or intrahepatic metastasis (im). HAO caused a greater decrease in TP than did PVO, with the decrease being particularly marked in the encapsulated HCCs. The TP of HCC was found to be regulated mainly by the presence or absence of a tumor capsule or necrosis, and partly by the hepatic arterial flow and PVP. Our study suggests that TP increases with capsule formation and that the pressure gradient between TP and PVP may be a causal factor in the dispersement of tumor cells into the portal vein.
Hepatocytic phenotypes induced in sarcomatous cholangiocarcinoma cells treated with 5-azacytidine
Authors: Enjoji M , Nakashima M , Honda M , Sakai H , Nawata H
Published: July 2004

The sarcomatoid cells found in cholangiocarcinoma (CC) or hepatocellular carcinoma (HCC) are not well characterized. In this study, a human sarcomatoid CC cell line, ETK-1, was established from a patient, and then morphological and phenotypical characteristics of the ETK-1 cells were evaluated before and after treatment with differentiation-inducing 5-azacytidine (5-azaCR). Phenotypically, the ETK-1 cells appeared immature. Exposure to 5-azaCR induced morphological transformation; a converted cell line, MEK, was successfully established. The MEK cells expressed such hepatocyte-specific proteins as α-fetoprotein, albumin, integrin α1, and thrombopoietin, but lost such bile duct-specific proteins as integrin α3 and integrin β4. The histopathology of MEK xenografts resembled that of HCC. The ETK-1 cells appeared to be converted into hepatocytes by exposure to 5-azaCR. On the other hand, ETK-1 xenografts were diagnosed as tubular adenocarcinoma, and the tumor cells had a ductal phenotype. This suggests the possibility that ETK-1 cells can differentiate along a biliary epithelial cell lineage. ETK-1 and MEK will be useful in studying hepatocytic differentiation and the transformation from a biliary epithelial cell to a hepatocytic lineage.
Possible involvement of pertussis toxin-sensitive G protein in hepatocyte growth factor-induced signal transduction in cultured rat hepatocytes: Pertussis toxin treatment inhibits activation of phospholipid signaling, calcium oscillation, and mitogen-activated protein kinase
Authors: Adachi T , Nakashima S , Saji S , Nakamura T , Nozawa Y
Published: July 2004

Treatment of primary cultured rat hepatocytes with hepatocyte growth factor (HGF) gives rise to inositol phosphate formation, cytosolic calcium oscillation, activation of mitogen-activated protein (MAP) kinase and phospholipase D (PLD), and arachidonic acid release, leading to DNA synthesis. Pretreatment of cultured hepatocytes with pertussis toxin (PT), which is known to adenosine diphosphate-ribosylate Gi and Go guanine nucleotide -binding proteins and to inhibit their functions, partially inhibited HGF-induced [3H]thymidine incorporation in a concentration-dependent manner. These results suggest that HGF-mediated DNA synthesis of hepatocytes is partly regulated via PT-sensitive guanine nucleotide-binding protein. Therefore, the effects of PT treatment on HGF-induced signal-transduction pathways were investigated. HGF-induced MAP kinase activation and arachidonic acid release were decreased by PT treatment, whereas PLD activation was diminished by PT to the level of unstimulated control. PT also interfered with HGF-induced inositol phosphate formation and cytosolic calcium oscillation. These results suggest that both PT-sensitive and PT-insensitive pathways are involved in HGF-induced signaling.
Mouse liver T cells: Their change with aging and in comparison with peripheral T cells
Authors: Tsukahara A , Seki S , Iiai T , Moroda T , Watanabe H , Suzuki S , Tada T , Hiraide H , Hatakeyama K , Abo T
Published: July 2004

Mouse liver contains both IL-2Rβ- (or low positive) high T-cell receptor (TCR(hi)) cells and IL-2Rβ+ intermediate TCR (TCR(int)) cells. TCR(int) cells consist of natural killer 1.1 (NK1)+ and NK1- subsets. NK1- TCR(int) cells increase constantly with age whereas TCR(hi) cells decrease. NK1+ TCR(int) cell proportions in the liver increase until middle age and decrease thereafter. Although NK1+ TCR(int) cells in other organs are few regardless of age, NK1- TCR(int) cells gradually appear in other lymphoid organs with aging. Skewed usage of Vβ7 and Vβ8 TCR was observed in NK1+ TCR(int) cells in the liver but the predominance was less obvious in NK1- TCR(int) and TCR(hi) cells in the liver and other organs. TCR V α14 messenger RNA (mRNA) was detected in NK1+ TCR(int) cells but not in the other two populations. In contrast, although NK1+ TCR(int) cells contain virtually no V α11+ T cells, NK1- TCR(int) cells contain a much higher proportion (approximately 12%) of V α11+ T cells, whereas approximately 4% of TCR(hi) cells are V α11+. NK activities of liver mononuclear cells (MNC) and splenocytes decrease with aging, although the former is always greater than the latter. NK activity of liver MNC is a function of NK cells, partly NK1+ TCR(int) cells but not NK1- TCR(int) cells or TCR(hi) cells. These results suggest that lymphocytes of liver and other organs at old age are no longer occupied solely by conventional thymus-derived T cells, and the increase of extrathymic IL-2Rβ+ NK1- TCR(int) cells in liver and periphery could be closely related to immunological changes with aging.
Hepatic stellate cell immunodetection and cirrhotic evolution of viral hepatitis in liver allografts
Authors: Guido M , Rugge M , Leandro G , Fiel I M , Thung S N
Published: July 2004

Patients with chronic viral hepatitis are at high risk of developing cirrhosis, but the outcome of the disease in a given patient is unpredictable. Hepatic stellate cells have been demonstrated to be the most important cell type involved in hepatic fibrogenesis, regardless of the cause of the liver injury. The α isotype of actin (a phenotypic marker of smooth muscle cells) may be expressed by hepatic stellate cells, reflecting their “activation” to myofibroblast-like cells. The present study aimed to analyze the expression of α-smooth muscle actin-positive hepatic stellate cells in liver allografts with recurrent viral hepatitis, and to evaluate whether and how such expression may be related to the outcome of the disease. Using immunohistochemistry and a semi-quantitative scoring system, the expression of α-smooth muscle actin in hepatic stellate cells was analyzed in liver allografts of 17 patients with recurrent viral hepatitis. They included nine patients who developed cirrhosis at the end of follow-up (mean time 23.6 months), and eight patients with no cirrhosis at the end of a comparable follow-up time (mean 30.1 months). In all patients, liver biopsy specimens were obtained between 3 and 6 months (t1) and between 10 and 15 months (t2) after transplantation. Preperfusion biopsy specimens of donor livers served as a baseline (t0). By comparison with the baseline biopsy, an increased number of α-smooth muscle actin-expressing hepatic stellate cells was observed in all cases in t1 biopsies. An increase in the amount of α-smooth muscle actin-positive hepatic stellate cells in zone 1 at t1 was significantly (P < .006) related to subsequent cirrhotic evolution. In conclusion, in liver allografts with recurrent viral hepatitis, the activation of hepatic stellate cells is an early event. An increased number of α-smooth muscle actin-positive hepatic stellate cells in zone 1 may represent an unfavorable event related to cirrhotic evolution.
Pentoxifylline inhibits growth and collagen synthesis of cultured human hepatic myofibroblast-like cells
Authors: Preaux A , Mallat A , Rosenbaum J , Zafrani E , Mavier P
Published: July 2004

During the course of liver fibrogenesis, myofibroblast-like cells (MFLC), mostly derived from hepatic stellate cells, proliferate and synthesize excessive amounts of extracellular matrix components. Pentoxifylline (PTX) elicits antiproliferative and antifibrogenic effects in human dermal fibroblasts. The aim of this study was to test the effects of PTX on the proliferation and the synthesis of collagen and gelatinase A in cultured human hepatic MFLC. MFLC were obtained by outgrowth from human liver explants. PTX markedly reduced serum-driven cell proliferation, as assessed by nuclear autoradiography experiments and measurement of actual cell growth. Growth inhibition was totally reversed after removal of the drug. PTX also affected collagen synthesis, as measured by [3H]hydroxyproline incorporation into proteins. Synthesis of secreted collagen was reduced by 24% and 67% at concentrations of 100 microg/mL and 500 microg/mL, respectively. This was associated with a decrease in type I and III procollagen messenger RNA (mRNA), indicating an effect at a pretranslational level. In contrast, PTX did not affect either gelatinase A activity released in culture medium or the expression of its specific mRNA. In conclusion, PTX exhibits potent antiproliferative and antifibrogenic effects toward hepatic MFLC. These results suggest that PTX might have therapeutic implications in chronic liver disease.