bluuebunny/crossref_metadata_2025 huggingface dataset:
Enhanced Gi–Protein-Mediated Mitogenesis Following Chronic Ethanol Exposure in A Rat Model of Experimental Hepatocellular Carcinoma
Authors: McKillop Iain H. , Vyas Nilesh , Schmidt Max C. , Cahill Paul A. , Sitzmann James V.
Published: July 2004

Hepatocellular carcinoma (HCC) is associated with increased expression and function of inhibitory guanine nucleotide regulatory proteins (Gi–proteins). This study addresses the effects of chronic ethanol exposure on the expression and function of adenylyl cyclase (AC)–linked G–proteins (Gs and Gi) and growth in experimental HCC. G–protein expression and function was determined by immunoblot in the hepatic tumorigenic H4IIE cell line and isolated cultured hepatocytes in the absence or presence of ethanol (5–100 mmol/L). Chronic exposure (24 hours) to ethanol dose–dependently increased Giα1/2 expression in the H4IIE cell line, but not in cultured hepatocytes. Gsα–protein expression remained unchanged in both H4IIE cells and cultured hepatocytes following ethanol treatment. In addition, ethanol directly activated a Gi–protein, because pertussis toxin (PTx)–catalyzed, adenosine diphosphate (ADP)–dependent ribosylation of Giα substrates decreased following ethanol treatment. The increased functional activity of Giα1/2 –protein expression was confirmed by demonstrating that ethanol dose–dependently inhibited basal and stimulated AC activity in H4IIE cells, while not significantly altering basal AC activity in isolated cultured hepatocytes. Furthermore, while ethanol had no significant effect on basal mitogenesis in H4IIE cells or hepatocytes, increased mitogenesis caused by direct Giα–protein stimulation (mastoparan M7; 10–5,000 nmol/L) was further enhanced in the presence of ethanol, an effect that was completely blocked following Gi–protein inhibition (PTx; 100 ng/mL). In contrast, activation of Gi–proteins using M7 failed to alter cellular mitogenesis in isolated cultured hepatocytes, whether in the absence or presence of ethanol. Finally, analysis of mitogen–activated protein kinase (MAPK) activity demonstrated that chronic ethanol treatment further enhanced Gi–protein-stimulated MAPK activity in hepatic tumorigenic cells. In conclusion, these data demonstrate that ethanol enhances cellular mitogenesis in experimental HCC as a result of, at least in part, a Gi–MAPK-dependent pathway. Furthermore, this effect may be caused by ethanol's direct up–regulation of the expression and activity of Gi–proteins in HCC
A Prospective, Community–Based Evaluation of Liver Enzymes in Individuals With Hepatitis C After Drug Use
Authors: Inglesby Thomas V. , Rai Rudra , Astemborski Jacquie , Gruskin Leslie , Nelson Kenrad E. , Vlahov David , Thomas David L.
Published: July 2004

Serum alanine transaminase (ALT) levels are used to select hepatitis C virus (HCV)–infected patients for treatment and liver biopsy. However, the natural history of these measurements is poorly understood. To examine the hypothesis that ALT levels vary over time in HCV–infected patients, serial serum ALT levels were prospectively measured in a cohort of 1,235 persons with a history of prior illicit drug use. Over 25 months of follow–up, there was a median of four evaluations per patient. ALT values were higher in 1,164 (94%) HCV–infected individuals than in 71 (6%) HCV–uninfected individuals. The remainder of the analysis focused on these HCV–infected individuals, 647 (62%) of whom had normal ALT values at their initial visit. However, 323 (49%) of these had at least one elevated ALT over the next 25 months. Of the 395 patients whose ALT was initially abnormal, 332 (84%) had at least one normal value over the next 25 months. Overall, among individuals with four or more visits, ALT values were persistently normal in 42%, persistently elevated in 15%, and intermittently elevated in 43%. Because serum ALT levels have high visit–to–visit variability, single assessments should not be used to manage HCV–infected individuals. Further investigation is needed to ascertain the correlation of serial ALT trends with important disease outcomes.
Glutathione Peroxidase-Deficient Mice Are More Susceptible to Neutrophil–Mediated Hepatic Parenchymal Cell Injury During Endotoxemia: Importance of An Intracellular Oxidant Stress
Authors: Jaeschke Hartmut , Ho Ye-Shih , Fisher Michael A. , Lawson Judy A. , Farhood Anwar
Published: July 2004

Neutrophils contribute to hepatocellular injury in a number of acuteinflammatory reactions. However, the molecular mechanism of parenchymal cell injury remains controversial. To address the issue of whether or not reactive oxygen species (ROS) are important in the injury process, we used the galactosamine/endotoxin (Gal/ET) model of acute liver failure, which involves a neutrophil–mediated parenchymal cell injury. In C3Heb/FeJ mice, Gal/ET induced a significant increase of hepatic and plasma levels of glutathione disulfide (GSSG), an indicator of oxidant stress, selectively during the neutrophil–mediated injury phase. In glutathione peroxidase-deficient mice (Gpx1-/-
), Gal/ET or Gal/tumor necrosis factor α (TNF–α) caused more severe neutrophil–mediated liver injury compared with wild–type animals. However, there was no significant difference in other critical parameters, e.g., activation of the transcription factor, nuclear factor–κB (NF–κB), and soluble intercellular adhesion molecule–1 (sICAM–1), parenchymal cell apoptosis, and neutrophil sequestration in the liver. Our results suggest that neutrophil–derived ROS are responsible for an intracellular oxidant stress in hepatocytes after Gal/ET treatment. Because of the higher susceptibility of Gpx1-/-
mice to a neutrophil–mediated injury, we conclude that peroxides generated by neutrophils diffused into hepatocytes and contributed to parenchymal cell death in vivo. Thus, strengthening defense mechanisms against ROS in target cells can attenuate excessive inflammatory injury without affecting host defense reactions
Detection of Genomic– and Minus–Strand of Hepatitis C Virus Rna in the Liver of Chronic Hepatitis C Patients by Strand–Specific Semiquantitative Reversetranscriptase Polymerase Chain Reaction
Authors: Negro Francesco , Krawczynski Krzysztof , Quadri Rafael , Rubbia-Brandt Laura , Mondelli Mario , Zarski Jean-Pierre , Hadengue Antoine
Published: July 2004

Studies aimed at correlating the intrahepatic hepatitis C virus (HCV)–RNA level and anatomo–clinical features have been difficult because of sensitivity and specificity shortcomings of available techniques. We titered the genomic– and minus–strand HCV RNAs by a strand–specific, semiquantitative, genotype–independent reverse–transcriptase polymerase chain reaction (RT–PCR) in the liver tissue of 61 patients with chronic hepatitis C. Findings were correlated with the levels of HCV RNA in the serum, the HCV genotype, the expression of intrahepatic HCV antigens, the histological activity (using separate scores for the lobular and the portal/periportal necroinflammatory activity and for the fibrosis), and the response to interferon alfa (IFN–α) treatment. Genomic– and minus–strand HCV RNA were detected in 59 and 57 liver specimens, respectively. The HCV–RNA level in the serum correlated with the genomic–strand, but not with the minus–strand, HCV–RNA titer in the liver. No correlations were found between either strand of the intrahepatic HCV RNA and the level of expression of HCV antigens in the liver, or with the grading/staging of the underlying liver disease. The response to IFN–α treatment could be predicted by the serum HCV–RNA level and genotype, but not by the intrahepatic level of genomic– or minus–strand HCV RNA. These results suggest that, although the detection of the minus–strand HCV RNA reliably identifies the presence of replicating HCV in its target organ, the quantitative measurement of viremia remains the clinically meaningful “golden standard” for assessing the level of HCV replication
Detection of Occult Low–Grade B–Cell Non–Hodgkin's Lymphoma in Patients With Chronic Hepatitis C Infection and Mixed Cryoglobulinemia
Authors: Rasul Imran , Shepherd Frances A. , Kamel-Reid Suzanne , Krajden Mell , Pantalony Dominic , Heathcote Jenny E.
Published: July 2004

Hepatitis C virus (HCV) is both hepatotropic and lymphotropic and has been shown to be associated with the benign lymphoproliferative disorder, mixed cryoglobulinemia (MC). Preliminary studies suggest that there may be an association between chronic hepatitis C, MC, and non–Hodgkin's lymphoma (NHL). The aim of this study was to determine whether patients with chronic HCV and MC have occult bone marrow NHL. Sixteen patients with chronic HCV and clinically active MC underwent bone marrow biopsy and aspiration. Flow cytometry was performed looking for abnormal B–cell lineage. Molecular genetic studies were performed to identify B–cell monoclonality. Nine of 16 patients (56%) had abnormal marrow morphology, 7 (44%) were interpreted as suspicious for lymphoma, and 2 (13%) as consistent with lymphoma. Flow cytometry on 13 patients identified5 (39%) with increased B–cell populations. Molecular analysis on 13 patients identified 3 (23%) with monoclonal proliferation ofthe B–cell lineage. All 13 patients tested for Epstein–Barr virus were negative by polymerase chain reaction (PCR). Four of 16 patients(25%) had marrow morphology and evidence of a monoclonal B–cell population by flow cytometry and/or molecular studies, consistent with B–cell NHL. These findings confirm the presence of lymphoproliferative disorders in patients with chronic HCV and MC. Some of these disorders meet the morphological and molecular criteria for the diagnosis of malignantlymphoma
Modulation of Protein Kinase C by Taurolithocholic Acid in Isolated Rat Hepatocytes
Authors: Beuers Ulrich , Probst Irmelin , Soroka Carol , Boyer James L. , Kullak-Ublick Gerd A. , Paumgartner Gustav
Published: July 2004

The protein kinase C (PKC) family of isoenzymes plays a key role in the regulation of hepatocellular secretion. The hydrophobic and cholestatic bile acid, taurolithocholic acid (TLCA), acts as a potent Ca++ agonist in isolated hepatocytes. However, its effect on PKC isoforms has not been elucidated. Here we investigate the effects of TLCA at low micromolar concentrations on the distribution of PKC isoforms and on membrane–associated PKC activity. The distribution of PKC isoforms was determined in isolated rat hepatocytes in short–term culture using Western blotting and immunofluorescence techniques. PKC activity was measured radiochemically. TLCA (10 μmol/L) induced selective translocation of ε–PKC by 47.9% ± 20.5% (P < .02 vs. controls; n = 7), but not of α–, δ–, and ζ–PKC to the hepatocellular membranes, whereas the phorbol ester, phorbol 12–myristate 13–acetate (PMA) (1 μmol/L) caused translocation of all mobile isoforms, α–, δ–, and ε–PKC, as shown by immunoblotting. Immunofluorescence studies demonstrated selective translocation of ε–PKC to the canalicular membranes of isolated rat hepatocyte couplets by TLCA (10 μmol/L), but predominant translocation to intracellular and basolateral membranes by PMA (1 μmol/L). Both TLCA (10 μmol/L) and PMA (1 μmol/L) stimulated membrane–bound PKC activity by 60.5% ± 45.8% (P < .05 vs. controls; n = 5) and 72.4% ± 37.2% (P < .05; n = 5), respectively. TLCA at lower concentrations (5 μmol/L) was less effective. Because activation of ε–PKC has been associated with impairment of vesicle–mediated targeting and insertion of membrane proteins in secretory cells, it is attractive to speculate that TLCA reduces bile secretory capacity of the liver cell by activation of ε–PKC at the canalicular membrane
Expression of Inducible Nitric Oxide Synthase in Endotoxemic Rat Hepatocytes Is Dependent on the Cellular Glutathione Status
Authors: Vos Thera A. , van Goor Harry , Tuyt Leonore , de Jager-Krikken Alie , Leuvenink Ron , Kuipers Folkert , Jansen Peter L. , Moshage Han
Published: July 2004

The inducible nitric oxide synthase (iNOS) promoter contains nuclear factor κB (NF–κB) binding sites. NF–κB activation is determined, in part, by the intracellular redox status. The aim of this study was to determine the importance of the cellular glutathione status in relation to NF–κB activation and iNOS expression in hepatocytes in vivo and in vitro. For in vivo experiments, rats were injected with endotoxin and sacrificed 6 hours later. Glutathione was depleted by diethylmaleate. For in vitro experiments, cultured hepatocytes from untreated rats were exposed to a cytokine mixture. Glutathione levels were depleted by diethylmaleate and restored by N–acetylcysteine. iNOS expression was assessed by Western blot, reverse transcription polymerase chain reaction, nitric oxide (NO) metabolites, and immunohistochemistry. NF–κB binding was assessed by electrophoretic mobility shift assay. Endotoxin–induced iNOS expression in rat liver was prominent in hepatocytes, Kupffer cells, and inflammatory cells, in particular neutrophils. Glutathione depletion prevented iNOS induction in hepatocytes, but not in inflammatory cells. iNOS protein levels were in accordance with iNOS messenger RNA and NO metabolites in plasma. Glutathione depletion did not affect neutrophil infiltration. Cytokines strongly induced iNOS in cultured hepatocytes. Induction was prevented by glutathione depletion and could be restored by addition of N–acetylcysteine. NF–κB binding correlated with iNOS induction. In conclusion, in this study we show that iNOS induction in hepatocytes in vivo and in vitro is dependent on the intracellular glutathione status and correlates with NF–κB binding. Glutathione–depletion has no effect on the expression of iNOS in inflammatory cells, nor on neutrophil infiltration
Class Vi Intermediate Filament Protein Nestin Is Induced During Activation of Rat Hepatic Stellate Cells
Authors: Niki Toshiro , Pekny Milos , Hellemans Karine , De Bleser Pieter , Van Den Berg Kit , Vaeyens Freya , Quartier Erik , Schuit Frans , Geerts Albert
Published: July 2004

Hepatic stellate cells are considered to be liver–specific pericytes that play a key role in liver fibrosis. Because these cells express desmin and smooth muscle α–actin, they were assumed to be of myogenic origin. This hypothesis became doubtful when it was reported that stellate cells also express glial fibrillary acidic protein and neural cell adhesion molecule. In the present study, we show that activated stellate cells express nestin, a class VI intermediate filament protein originally identified as a marker for neural stem cells. Expression of nestin was first studied during spontaneous activation of stellate cells in culture. Immunohistochemistry showed that nestin–positive stellate cells already appeared at day 3, and nearly all the cells became positive for nestin at day 6 and 15. The immunoreaction was present in filaments except in dividing cells. The presence of messenger RNA transcript for nestin was shown by reverse transcription polymerase chain reaction and sequencing of amplified complementary DNA. We then compared the presence of nestin with that of other intermediate filament proteins and smooth muscle α–actin. Immunoblotting showed that the relative concentrations of nestin, desmin, and vimentin increased between day 2 and 6 in primary culture. After the initial increase vimentin leveled off, while nestin and desmin showed a tendency to decrease. This pattern was quite different from that of glial fibrillary acidic protein, which kept declining, and smooth muscle α–actin, which increased continuously up to day 13 in culture. We then studied the presence of nestin in normal and CCl4–injured rat liver. In normal liver, minimal immunoreaction for nestin was observed within the liver parenchyma. During induction of fibrosis by carbon tetrachloride, nestin–positive stellate cells appeared at 6 weeks, which was late in comparison with the induction of desmin and smooth muscle α–actin. We conclude that nestin is induced in stellate cells during transition from the quiescent to the activated phenotype; culture activation is a stronger stimulus thanin vivoactivation by injection of CCl4. Taken together with reports on expression of glial fibrillary acidic protein and neural cell adhesion molecule by stellate cells, new experimental studies on the embryonic origin of these cells are required
Expression of Vascular Endothelial Growth Factor Promotes Colonization, Vascularization, and Growth of Transplanted Hepatic Tissues in the Mouse
Authors: Ajioka Itsuki , Akaike Toshihiro , Watanabe Yoshifumi
Published: July 2004

A complex vascular network forms an important component of the liver architecture. This network is essential for the supply of oxygen and nutrients to cells and delivery of molecules for metabolic exchange. In this study, we attempted to construct a vascular network in transplanted hepatic tissues and examined the effect of such network on tissue formation. Primary hepatocytes of adult mice were transfected with vascular endothelial growth factor (VEGF) gene in vitro then transplanted with collagen beads intraperitoneally in mice. VEGF–transfected hepatocytes secreted sufficient protein of the transgene in vitro to induce proliferation of endothelial cells. In vivo, VEGF–transfected hepatocytes formed a large number of colonies and developed a significant vascular network in established tissues compared with control tissues. In addition, hepatocytes of VEGF–transfected, established tissues proliferated and formed a substantial parenchymal region. These hepatocytes were also functional as confirmed by the production of albumin. Our results suggested that VEGF expression conferred not only the formation of a vascular network but also promoted tissue formation. Our study showed that ex vivo gene transfection into hepatocytes is a useful method for the induction of liver reconstitution in vivo
Association of Diabetes Mellitus and Chronic Hepatitis C Virus Infection
Authors: Mason Andrew L. , Lau Johnson Y. , Hoang Nicole , Qian KePing , Alexander Graeme J. , Xu Lizhe , Guo Linsheng , Jacob Sheraj , Regenstein Fredric G. , Zimmerman Robert , Everhart James E. , Wasserfall Clive , Maclaren Noel K. , Perrillo Robert P.
Published: July 2004

While patients with liver disease are known to have a higher prevalence of glucose intolerance, preliminary studies suggest that hepatitis C virus (HCV) infection may be an additional risk factor for the development of diabetes mellitus. To further study the correlation of HCV infection and diabetes, we performed a retrospective analysis of 1,117 patients with chronic viral hepatitis and analyzed whether age, sex, race, hepatitis B virus (HBV) infection, HCV infection, and cirrhosis were independently associated with diabetes. In addition, a case–control study was conducted to determine the seroprevalence of HCV infection in a cohort of 594 diabetics and 377 clinic patients assessed for thyroid disease. In the former study after the exclusion of patients with conditions predisposing to hyperglycemia, diabetes was observed in 21% of HCV–infected patients compared with 12% of HBV–infected subjects (P = .0004). Multivariate analysis revealed that HCV infection (P= .02) and age (P = .01) were independent predictors of diabetes. In the diabetes cohort, 4.2% of patients were found to be infected with HCV compared with 1.6% of control patients (P= .02). HCV genotype 2a was observed in 29% of HCV–RNA-positive diabetic patients versus 3% of local HCV–infected controls (P< .005). In conclusion, the data suggest a relatively strong association between HCV infection and diabetes, because diabetics have an increased frequency of HCV infection, particularly with genotype 2a. Furthermore, it is possible that HCV infection may serve as an additional risk factor for the development of diabetes, beyond that attributable to chronic liver disease alone.
Acute Liver Injury Following Infection With A Cytopathic Strain of Duck Hepatitis B Virus
Authors: Lenhoff Raymond J. , Luscombe Carolyn A. , Summers Jesse
Published: July 2004

A variant avian hepadnavirus that has been shown to destroy hepatocytes in vitro was found to be cytopathic in vivo. A single amino acid change of glycine to glutamic acid at position 133 (G133E) in the preS protein of duck hepatitis B virus (DHBV) caused an increase in the intranuclear pool of viral covalently closed circular DNA (cccDNA), resulting in a transient elevation of viral replication and eventual hepatocyte destruction. In vivo viral infection with the G133E virus was compared with infection with wild–type virus over a 72–day period. Birds were inoculated with virus at day 2 post–hatch to ensure a high percentage of infected hepatocytes and potential persistence of virus. Birds infected with the G133E virus had increased periportal cellular proliferation and numerous lysed apoptotic hepatocytes following 100% infection of hepatocytes. The liver damage within G133E virus-infected birds subsided over time, resulting in mild chronic hepatitis that was similar to that observed within wild–type virus–infected birds. The subsidence of liver damage in G133E virus-infected birds coincided with a reduction of viral cccDNA to wild–type virus levels in the liver. Our study indicates that maintenance of wild–type levels of viral cccDNA promotes persistence of virus infection by establishing a noncytopathic infection.
Comparative Evaluation of Hepatitis C Virus Rna Quantitation by Branched Dna, Nasba, and Monitor Assays
Authors: Lunel Françoise , Cresta Pascale , Vitour Damien , Payan Christopher , Dumont Bruno , Frangeul Lionel , Reboul Dorothée , Brault Christine , Piette Jean-Charles , Huraux Jean-Marie
Published: July 2004

Several studies have shown a relationship between pretreatment hepatitis C virus (HCV) viral load and the response to interferon (IFN) therapy, creating a need for quantitative HCV–RNA assays. Here, we compared three commercial methods: nucleic acid sequence-based amplification NASBA (Organon), branched DNA 2.0 (bDNA) (Chiron), and Monitor (Roche), with reverse–transcription polymerase chain reaction (RT–PCR) as the reference. We assessed sensitivity and reproducibility on a well–characterized panel of sera (EUROHEP), a Chimp Rodney plasma pool, and samples from IFN–treated and –untreated patients with chronic hepatitis C caused by different HCV genotypes. The reproducibility of the NASBA and bDNA methods was slightly better than that of Monitor, especially for genotypes 2 and 4. NASBA had the highest sensitivity (99% vs. 94% and 88% with Monitor and bDNA, respectively), especially for the follow–up of patients on IFN. NASBA gave the highest HCV–RNA concentrations, which were approximately 10–fold more than with the bDNA assay and 100–fold more than with the Monitor kit. The linearity, tested on the chimp Rodney plasma pool, was better with bDNA for high viral load than with NASBA and Monitor, although for low concentration of HCV RNA, bDNA was negative. Pretreatment viral load was lower in patients who had a sustained virological response to IFN, although the bDNA method was not sensitive enough to quantify all pretreatment samples. This study indicates that gene amplification methods (NASBA or Monitor) have better sensitivity than bDNA assays for quantification of HCV RNA in patients with chronic HCV infection, although the bDNA and NASBA methods are more likely to quantify all genotypes. Prospective studies are needed to demonstrate the usefulness of quantitative assays for the follow–up of patients with chronic hepatitis C
Transfection of A Rat Hepatoma Cell Line With A Construct Expressing Human Liver Annexin V Confers Susceptibility to Hepatitis B Virus Infection
Authors: Gong Zuo Jiong , De Meyer Sandra , van Pelt Jos , Hertogs Kurt , Depla Erik , Soumillion Ann , Fevery Johan , Yap Sing-Hiem
Published: July 2004

Previously, we have found that human liver annexin V (hA–V; in earlier reports referred as Endonexin II) is a specific hepatitis B surface antigen (HBsAg) binding protein. In this study, we demonstrate that transfection of rat hepatoma FTO 2B cells, a cell line that is not infectable by hepatitis B virus (HBV) and does not express hA–V, with a construct containing the hA–V gene, resulted in hA–V expressing cells susceptible to HBV infection. Afterin vitroinfection, transfected FTO cells (assigned as FTO 9.1 cells) expressing hA–V in cultures were shown to contain HBV–precore/core, X mRNAs, and covalently closed circular (ccc) DNA as detected by polymerase chain reaction (PCR). The presence of HBV ccc and replicative intermediate DNA was also demonstrated by Southern blot hybridization assay. HBV DNA secreted in the culture medium was also evident as determined by quantitative branched DNA (bDNA) assay. HBsAg and hepatitis B core antigen (HBcAg) could also be detected by an immunocytochemical method in 10% to 15% of the cells at day 3 and day 5 after infection. Infectivity ofin vitro-propagated HBV was demonstrated by infection of the naive FTO 9.1 cells with the culture supernatant from HBV–carrier cultures. In contrast to primary cultures of human hepatocytes and FTO 9.1 cells, primary rat and mouse hepatocytes, as well as rat hepatoma cell lines that do not express hA–V, are not susceptible to HBV infection. These findings suggest that hA–V plays a key role in the initial step of HBV infection and that the species–specific susceptibility to HBV infection and replication in hepatocytes is associated with the expression of hA–V.
Reduced C–Terminal Src Kinase (Csk) Activities in Hepatocellular Carcinoma
Authors: Masaki Tsutomu , Okada Masato , Tokuda Masaki , Shiratori Yasushi , Hatase Osamu , Shirai Mutsunori , Nishioka Mikio , Omata Masao
Published: July 2004

The proto–oncogene product pp60c–src is the cellular homologue of the Rous sarcoma transforming gene, and it is a non-receptor–linked and membrane–associated tyrosine kinase. There is a close correlation between elevated pp60c–src activity and cell transformation. We have recently reported that pp60c–src was activated in hepatocellular carcinoma (HCC) of human and Long–Evans cinnamon (LEC) rats. However, the mechanisms involved in this process remain unknown. C–terminal Src kinase (Csk) is a novel cytoplasmic protein tyrosine kinase that inactivates the members of the Src family protein tyrosine kinase in vitro. We investigated the role of Csk in hepatocarcinogenesis by analyzing the location, amount of Csk, and its kinase activity levels in nontumorous cirrhotic and tumorous sections of HCC of patients and an animal model of LEC rats. Csk tyrosine kinase activity was significantly reduced in tumorous tissues compared with nontumorous sections of patients as well as LEC rats. A single immunoreactive band at 50 kd was detected with Csk antibody in normal liver (NL), chronic hepatitis (CH), and nontumorous cirrhotic (NTC) segments of HCC of patients and LEC rats. In human tumorous tissues, Western blot revealed a 53–kd immunoreactive band, which was slightly larger than the usual 50–kd band of Csk. These results suggest that the reduced activity of tyrosine kinase of Csk may play an important role in the malignant transformation of hepatocytes in human and LEC rat, and the appearance of 53–kd Csk–related protein may be closely involved in the progression of cirrhosis to HCC in humans, and that 50–kd Csk may act as an antioncogene through the negative regulation of pp60c–src in the development of human HCC
Acute Carbon Tetrachloride Feeding Selectively Damages Large, But Not Small, Cholangiocytes From Normal Rat Liver
Authors: LeSage Gene D. , Benedetti Antonio , Glaser Shannon , Marucci Luca , Tretjak Ziga , Caligiuri Alessandra , Rodgers Rebecca , Phinizy Jo Lynne , Baiocchi Leonardo , Francis Heather , Lasater John , Ugili Laura , Alpini Gianfranco
Published: July 2004

The aim of this study was to develop a model of selective duct damage restricted to hormone–responsive segments corresponding to the ducts damaged in primary biliary cirrhosis (PBC). Carbon tetrachloride (CCl4) was fed by gavage to rats, and 2, 7, 14, and 28 days later, small and large cholangiocytes were isolated. Apoptosis was determined in situ by morphology and in purified cholangiocytes by assessment of nuclear fragmentation by 4,6–diamidino–2–phenylindole (DAPI) staining. Cholangiocyte proliferation was evaluated in situ by morphometry of liver sections stained for cytokeratin–19 (CK–19) and by proliferating cellular nuclear antigen (PCNA) staining in liver sections and in purified cholangiocytes by PCNA gene expression. Ductal secretion was assessed by measurement of secretin receptor (SR) gene expression and secretin–induced cyclic adenosine 3′,5′–monophosphate (cAMP) synthesis and secretin–induced choleresis. Two days after CCl4 administration, there was an increased number of small ducts, but a reduction of large ducts. Apoptosis, observed only in large ducts, was associated with decreased DNA synthesis and ductal secretion. Conversely, small cholangiocytes expressed de novo the SR gene and secretin–stimulated cAMP synthesis 2 days after CCl4 treatment. Proliferation of large cholangiocytes was delayed until 7 days, which was associated with a transient increase in ductal secretion in vivo.CCl4 effects on cholangiocytes were reversed by day 28. CCl4treatment causes a decrease in large duct mass as a result of a higher rate of apoptosis and absence of initial proliferation in large cholangiocytes. These processes were concomitant with a decrease of ductal secretion in large cholangiocytes. Small cholangiocytes appear resistant to CCl4–induced apoptosis, and proliferate and transiently compensate for loss of proliferative and secretory activity of large cholangiocytes.
Three–Dimensional Reconstruction of Parenchymal Units in the Liver of the Rat
Authors: Teutsch Harald F. , Schuerfeld Dirk , Groezinger Elke
Published: July 2004

To investigate the parenchymal units in the liver of the rat three–dimensionally, 15 μm cryosections were used for the demonstration of glucose–6–phosphatase (G6Pase) activity to visualize the borders of the individual units. Together with the supplying and draining vessels, they were traced through a sequence of 146 sections and reconstructed. A cone–shaped secondary unit with a height of 2.1 mm and a volume of 3.3 mm3 was reconstructed. It was “covered” by a continuous vascular surface, consisting of portal tracts and vascular septa, connecting the portal venular branches. The secondary unit was subdivided by portal tracts and vascular septa, and by branches of a draining central venular tree into 14 primary units. Most of them were tri– to heptahedral in shape. The height varied between 330 and 840 μm, and the volume varied between 0.094 and 0.621 mm3. The branches of the portal venular tree, with diameters from 28 ± 5 μm to 61 ± 14 μm, were oriented preferentially along the vertical axis of the units. Most of the primary units were drained by single branches of the central venular tree, located in the center and oriented along the vertical axis of the units. Vessel diameters ranged from 62 ± 14 μm to 216 ± 9 μm. The average length of the sinusoids was 355 ± 3 μm. From the results of this reconstruction study, it was concluded that the concept of the liver acinus cannot be applied to the liver of the rat
Methyl Homologues of Methyl Jasmonate and Methyl Dihydrojasmonate (Hedione®) from Sorbyl Alcohol
Authors: Giersch Wolfgang , Farris Iris
Published: July 2004

Treatment of cycloalkanone dimethyl acetals 3–6 with sorbyl alcohol (=(2E,4E)‐hexa‐2,4‐dien‐1‐ol; 1) in the presence of acids afforded the novel cycloalkenones 8, 9, 11, and 13 via a domino reaction (Claisen rearrangement with intramolecular ene reaction and retro‐ene reaction). Cyclopentenone 8 was readily transformed into 14 and 15, methyl homologues of racemic methyl jasmonate (16) and methyl dihydrojasmonate (=Hedione®; 17), respectively. The organoleptic properties of 14 and 15 are also discussed.
Spirocyclic Ethers Related to Ambrox®: Synthesis and Structure‐Odor Relationships
Authors: Winter Beat
Published: July 2004

The seven spirocyclic ethers 4–10, related to the tricyclic odorant Ambrox ® (1) and its diastereoisomers 2 and 3, were synthesized. Their odoriferous activity/inactivity was correlated with the steric accessibility of the ether O‐atom, calculated by computer‐aided molecular modeling. The olfactory properties of the active compounds are discussed.
Diradical‐Promoted Two‐Carbon Ring‐Expansion Reactions by Thermal Isomerization: Synthesis of Functionalized Macrocyclic Ketones
Authors: Rüedi Georg , Hansen Hans‐Jürgen
Published: July 2004

A new method for the smooth and highly efficient preparation of functionalized macrocyclic ketones has been developed. Pyrolysis of medium‐ and large‐ring 3‐vinylcycloalkanones by dynamic gas‐phase thermo‐isomerization (DGPTI) at 600–630° yielded, under insertion of a previously attached vinyl side chain by means of a 1,3‐C shift, the corresponding γ,δ‐unsaturated cycloalkanones. The yield of the two‐carbon ring‐expanded ketones greatly depended on the relative ring strains of substrate and product (5–87%, cf. Table 5). The formation of minor amounts of one‐carbon ring‐expanded cycloalkenes (<10%) can be ascribed to a subsequent decarbonylation step. A reaction mechanism involving initial cleavage of the weakest single bond in the molecule has been established (cf. Scheme 6). Recombination within the generated diradical intermediate in terminal vinylogous position led to the observed products, while reclosure gave recovered starting material. Substituents on the vinyl moiety were transferred locospecifically into the ring‐expanded products. An isopropenyl group did not significantly affect the isomerization process, whereas substrates bearing a prop‐1‐enyl group in β‐position enabled competing intramolecular H‐abstraction reactions, leading to acyclic dienones (cf. Schemes 9–11). DGPTI of the 13‐membered analogue directly yielded 4‐muscenone, which, upon hydrogenation, led to the valuable musk odorant (±)‐muscone. Increasing the steric hindrance on the vinyl moiety gave rise to diminishing amounts of the desired γ,δ‐unsaturated cycloalkanones. This novel two‐carbon ring‐expansion protocol was also successfully applied to 3‐ethynylcycloalkanones, giving rise to the corresponding ring‐expanded cyclic allenes (cf. Scheme 13).
Enantioselectivity in Odor Perception
Authors: Hölscher Bernd , Braun Norbert A. , Weber Berthold , Kappey Claus‐Hermann , Meier Manfred , Pickenhagen Wilhelm
Published: July 2004

The 3‐methyl‐4‐(tricyclo[5.2.1.02,6]dec‐4‐en‐8‐ylidene)butan‐2‐ols (=Fleursandol®; rac‐10), a new class of sandalwood odorants, were synthesized in their enantiomerically pure forms by use of tricyclo[5.2.1.02,6]dec‐4‐en‐8‐ones 17 and ent‐17 and (tetrahydro‐2H‐pyran‐2‐yl)‐protected 4‐bromo‐3‐methylbutan‐2‐ols 22 and ent‐22 as starting materials (Schemes 2–4). Only four of 16 possible stereoisomers of rac‐10 possess the typical, very pleasant, long‐lasting sandalwood odor (Table 1). The (2S,3R,4E,1′R,2′R,6′R,7′R)‐isomer ent‐10a is by far the most important representative, with an odor threshold of 5 μg/l in H2O.
Towards Functionalized Silicon‐Containing α‐Amino Acids: Asymmetric Syntheses of Sila Analogs of Homoserine and Homomethionine
Authors: Smith Richard J. , Bienz Stefan
Published: July 2004

The homoserine and homomethionine sila analogs, (R)‐HOSi(Me2)CH2CH(NH2)CO2H ((R)‐21) and (R)‐MeSCH2Si(Me2)CH2CH(NH2)CO2H ((R)‐24), respectively, were each synthesized in nine steps and in 30 and 23% overall yield, respectively, from commercially available ClSiMe2CH2Cl. The key step of both syntheses was the asymmetric α‐bromination of an Evans amide to introduce the stereogenic center of the amino acids with defined absolute configuration. While the preparation of the homomethionine analog (R)‐24 followed the expected pathway, the sila analog of homoserine, (R)‐21, was unexpectedly formed during the catalytic hydrogenation of an N3CH2‐substituted silane derivative.
γ,δ‐ and δ,ε‐Unsaturated Aldehydes from γ‐ and δ‐Lactones in One Step. Preliminary Communication
Authors: Giersch Wolfgang , Naef Ferdinand
Published: July 2004

A one‐step transformation of γ‐ and δ‐(spiro)lactones into γ,δ‐ and δ,ε‐unsaturated aldehydes with an excess of formic acid in the vapor phase over a supported manganese catalyst is described for the first time. The scope and limitations of this new reaction are shown with different lactones as substrate, and a mechanistic rationale is proposed.
Efficient Synthesis of (−)‐(R)‐Muscone by Enantioselective Protonation
Authors: Fehr Charles , Galindo José , Farris Iris , Cuenca Ana
Published: July 2004

A new synthesis of (−)‐(R)‐muscone ((R)‐1) by means of enantioselective protonation of a bicyclic ketone enolate as the key step (see 6→(S)‐4 in Scheme 2) is presented. The C15 macrocyclic system is obtained by ozonolysis (Scheme 7).
Comparative Infrared, Raman, and Natural‐Bond‐Orbital Analyses of King's Sultam
Authors: Hagemann Hans , Dulak Marcin , Wesolowski Tomasz A. , Chapuis Christian , Jurczak Janusz
Published: July 2004

By means of 1H‐NOESY‐ and Raman‐spectroscopic analyses, we experimentally demonstrated the presence of the equatorial NMe conformer of King's sultam 4b in solution, resulting from a rapid equilibrium. As a consequence, the value of the N lone‐pair anomeric stabilization should be revised to 1.5–1.6 kcal/mol. Independently from the N tilting, natural bond orbital (NBO)‐comparative analyses suggest that the S d* orbitals do not appear as primordial and stereospecific acceptors for the N lone pair. Second, the five‐membered‐ring sultams do not seem to be particularly well‐stabilized by the SC σ* orbital in the N‐substituted pseudo‐axial conformation, as opposed to an idealized anti‐periplanar situation for the six‐membered‐ring analogues. In this latter case, the other anti‐periplanar CC σ* and C(1′)H/C(2′) σ*orbitals are as important, if not more, when compared to the SC σ* participation. In the pseudo‐equatorial conformation, γ‐sultams particularly benefit from the N lone‐pair hyperconjugation with the anti‐periplanar SO1 σ* and C(2)H/C or C(1′)H/C σ* orbitals. This is also the case for δ‐sultams when the steric requirement of the N‐substituent exceeds 1.6 kcal/mol. When both axial and equatorial conformations are sterically too exacting, the N‐atom is prone to sp2 hybridization or/and conformational changes (i.e., 12c). In that case also, the mode of stereoelectronic stabilization differs from γ‐ to δ‐sultams.
Studies on the Synthesis of Trisaccharide Analogues of the Antibiotic Moenomycin A
Authors: Yang Guangbin , Hennig Lothar , Findeisen Matthias , Oehme Ramona , Giesa Sabine , Welzel Peter
Published: July 2004

A new approach for the synthesis of moenomycin trisaccharide analogues is reported in which three monosaccharide building blocks are used and allows the introduction of different substituents at the 6‐position of unit E (Scheme 2). Furthermore, a new procedure for the introduction and manipulation of unit F is described (Scheme 3).
Synthesis of 2‐Aryl‐ and 2‐Heteroaryl‐3,5‐dimethoxy‐1,4‐benzoquinones Involving Pd‐Catalyzed Cross‐Coupling of (2,3,4,6‐Tetramethoxyphenyl)boronic Acid
Authors: Leão Lana Enio José , Carazza Fernando , Aparacida de Oliveira Rosilene
Published: July 2004

A series of 2‐aryl‐ and 2‐heteroaryl‐substituted 3,5‐dimethoxy‐1,4‐benzoquinones (compounds 27–36) have been synthesized by cross‐coupling of (2,3,4,6‐tetramethoxyphenyl)boronic acid (2) with aromatic bromides or iodides in the presence of [Pd0(Ph3)4] and Na2CO3, followed by AgO‐promoted oxidation of the resulting biaryl compounds 17–26.
Synthesis of A/B Ring Analogs of Territrem B and Evaluation of Their Biological Activities
Authors: Zhao Jinhao , Zhao Feng , Wang Yanguang , Li Haibo , Zhang Qijun , Guénard Daniel , Ge Qiufu , Wei Erqing , Jiang Hao , Wu Yihang , Wang Lin , Jiang Hualiang , Guéritte Françoise , Wu Xiumei , Cheng Christopher H. K. , Lee Shoei‐Sheng , Zhao Yu
Published: July 2004

Two series of territrem B analogs, i.e., 5–10, containing both the 2‐en‐1‐one‐A‐ring and the aromatic‐E‐ring pharmacophores were designed and synthesized from jujubogenin (4a). The anti‐acetylcholinesterase (anti‐AChE), anti‐caspase‐3, and other biological activities of these territrem‐B analogs and their intermediates were assessed. Compound 9b, 22a, and 24f were shown to be weak inhibitors of AChE. None of the synthesized compounds exhibited significant inhibitory activity on caspase‐3. On the other hand, compounds 22e, 24a, 7b, and 8a showed mild cytotoxicity on cultured KB cells, with IC50 values of 2.0, 3.5, 6.5, and 14 μM, respectively. In addition, compounds 23b and 5f were active against injury arising from oxygen‐glucose deprivation.
Regioselective Acylation of Ginsenosides by Novozyme 435 to Generate Molecular Diversity
Authors: Teng Rongwei , Ang Chingseng , McManus David , Armstrong David , Mau Shaiolim , Bacic Antony
Published: July 2004

Ginsenosides are major bioactive constituents of ginseng (Panax spp.; Araliaceae), a traditional Chinese medicinal herb. In order to increase the molecular diversity and broaden the potential usage of ginsenosides, ginsenosides Rd (1), Rg3 (2), (20R)‐Rg3 (3), Rh2 (4), Re (5), Rh1 (8), Rg2 (9), gypenoside XVII (6), and pseudoginsenoside F11 (7) were regioselectively acylated with vinyl acetate, catalyzed by Novozyme 435 (lipase B from Candida antarctica), in organic solvents to afford different mono‐acetyl ginsenosides. Ginsenoside Rd (1) was also acylated with vinyl decanoate or vinyl cinnamate to generate 1b and 1c, respectively. Acylation of glucosylated ginsenosides (1–4, 6, 8) occurred at the primary 6‐OH function of the terminal glucose (Glc) moiety of the sugar at C(3) or C(20) of the dammarane‐type aglycone. In contrast, ginsenosides 5, 7, and 9, containing mixed sugar moieties, resulted in acylation of both the rhamnose (Rha) and the glucose (Glc) moieties. In the case of ginsenoside Re (5) and pseudoginsenoside F11 (7), acylation at the secondary 4‐OH function of the terminal Rha moiety, attached at C(3) of the aglycone, is preferred. The structures of all acylated products were determined by extensive MALDI‐TOF‐MS and NMR analyses.
Molecular‐Sieve Honeycomb for Air Separation from Picea abies
Authors: Onyestyák György , Rees Lovat V. C. , László Krisztina
Published: July 2004

The cellular structure of Norway spruce (Picea abies) was transformed via a simple, single‐step carbonization process into a carbon monolith with molecular‐sieve properties. The monolith exhibited a genuine honeycomb structure derived from the original intrinsic H2O channels of the wood. The micropores formed during carbonization from the walls of the channels were shown to have a high adsorption capacity. The honeycomb monolith was tested for air separation. Micropore diffusion of N2 and O2 was found by the frequency‐response (FR) technique to be the rate‐controlling process of mass transport.
Crystal Structure of Melaminium Orthophosphate from High‐Resolution Synchrotron Powder‐Diffraction Data
Authors: De Ridder Dirk J. A. , Goubitz Kees , Brodski Vladimir , Peschar René , Schenk Henk
Published: July 2004

The crystal structure of melaminium orthophosphate (MP) has been determined from high‐resolution synchrotron powder‐diffraction data. The crystal packing consists of melaminium layers and pairs of orthophosphate chains connected by H‐bonds almost perpendicular to the layers. The distance between melaminium layers is 3.62 Å. Neighboring melaminium molecules do not lie in the same plane, but in two parallel planes at close distance (0.79 Å), and are shifted with respect to each other. The orthophosphate chains are connected by both intra‐chain and inter‐chain H‐bonds. The melamine is singly protonated at an endocyclic N‐atom. The powder‐diffraction data were corroborated by solid‐state NMR experiments.
Vibratilicin: a Novel Compound from the Basidiomycete Cortinarius vibratilis
Authors: Wang Fei , Tan Jian‐Wen , Liu Ji‐Kai
Published: July 2004

A novel N‐containing compound, vibratilicin (=3‐[3‐(dimethylamino)‐4‐(hydroxyamino)‐4‐oxobutoxy]‐2‐(palmitoyloxy)propyl (9E,12E)‐octadeca‐9,12‐dienoate; 1), was isolated from the fruiting bodies of the basidiomycete Cortinarius vibratilis Fr. Compound 1 is a representative of the rare natural products containing hydroxamic acid moieties, and can be viewed as a derivative of neoengleromycin (2). Also isolated from the same fungus were five known compounds: ergosta‐5,7,22‐trien‐3β‐ol, 5α,8α‐epidioxyergosta‐6,22‐dien‐3β‐ol, p‐anisic acid, N‐(2‐hydroxyhexadecanoyl)‐4‐sphingenine, and (4E,8E)‐2‐N‐(2′‐hydroxypalmitoyl)‐1‐O‐(β‐D‐glucopyranosyl)‐9‐methyl‐4,8‐sphingadienine. Their structures were determined mainly spectroscopically, including 2D‐NMR techniques (HMBC, HMQC, 1H,1H‐COSY).
Alumina hip joints characterized by run‐in wear and steady‐state wear to 14 million cycles in hip‐simulator model
Authors: Oonishi Hironobu , Clarke Ian C. , Good Victoria , Amino Hirokazu , Ueno Masuru
Published: July 2004

The biphasic wear performance (run‐in; steady‐state phase) of 28‐mm alumina‐alumina hip implants was studied by hip simulator methods using bovine serum as the lubricant. The Biolox™ implants were run to 5.7 million cycles and Bioceram™ implants to 14.4 million cycles (Mc). Wear with all‐alumina total hip replacements (THR) first showed a high wear rate of the order 1.2 mm3/Mc, lasting approximately 0.17 Mc. Overall to 0.7 Mc, the run‐in phase appeared curvilinear but could be described by a linear phase averaging 0.3 mm3/Mc. From 0.7 to 1 Mc duration, the wear trend transitioned into a steady‐state phase. Wear rates from 1 to 14 Mc were of the order 0.02 mm3/Mc. Surface contamination from the serum lubricant resulted in cyclic weight fluctuations of the order 0.2 mg. The transition from average run‐in to steady‐state phase represented a wear reduction of 13‐fold. Comparing the steady‐state wear value to that in standard 28‐mm UHMWPE CUPS approaching 75 mm3/year, there was clearly a three‐orders‐of‐magnitude wear superiority in favor of ceramic cups. © 2004 Wiley Periodicals, Inc. J Biomed Mater Res 70A: 523–532, 2004
Effects of adsorbed proteins and surface chemistry on foreign body giant cell formation, tumor necrosis factor alpha release and procoagulant activity of monocytes
Authors: Shen Mingchao , Garcia Iris , Maier Ronald V. , Horbett Thomas A.
Published: July 2004

The adhesion and activation of monocytes and macrophages are thought to affect the foreign body response to implanted medical devices. However, these cells interact with devices indirectly, because of the prior adsorption of proteins. Therefore, we preadsorbed several “model” biomaterial surfaces with proteins and then measured foreign body giant cell (FBGC) formation, tumor necrosis factor alpha (TNFα) release, and procoagulant activity. The model surfaces were tissue culture polystyrene (TCPS), untreated polystyrene (PS), and Primaria, whereas the proteins used were albumin, fibronectin, fibrinogen, and immunoglobulin. FBGC formation, TNFα release, and procoagulant activity of monocytes were the highest for surfaces preadsorbed with IgG. FBGC formation was lower on surfaces with adsorbed fibrinogen and fibronectin than on uncoated surfaces. TNFα release and procoagulant activity of monocytes were similar on surface adsorbed with fibrinogen, fibronectin, or albumin. Monocyte activation was also affected by the surface chemistry of the substrates, because FBGC formation was the highest on PS and the lowest on TCPS. Monocyte procoagulant activity was the highest on Primaria. Adsorbed proteins and surface chemistry were found to have strong effects on FBGC formation, monocyte TNFα release, and procoagulant activityin vitro, providing support for the idea that these same variables could affect macrophage‐mediated foreign body response to biomaterialsin vivo. © 2004 Wiley Periodicals, Inc. J Biomed Mater Res 70A: 533–541, 2004