Possible involvement of arachidonic acid metabolism in phenobarbital promotion of hepatocarcinogenesis

The effects of inhibitors of arachidonic acid metabolism andantioxidants on the rat liver tumor promotion activity of phenobarbital(PB) were assessed using the enzyme-altered focus as the end-pointlesion. Fischer 344 male rats were initiated with N-nitrosodiethylamine(200 mg/kg) and then divided into five groups placed on basaldiet, diet containing 0.05% PB, diet containing 0.05% PB plus0.75%, 1% or 1.5% levels of various inhibitors of arachidonicacid metabolism or antioxidants, or diet containing 1% or 1.5%inhibitors or antioxidants alone for 10 weeks, and then killed.-Bromo phenacyl bromide, an inhibitor of phospholipase A₂ significantly inhibited the promotion activity of PB at dose levelsof 0.75% and 1.5%, reaching plateau at 0.75%. Both quercetin,an inhibitor of lipoxygenase, and morin, a dual inhibitor oflipoxygenase-cyclooxygenase, significantly reduced the promotionactivity of PB at the 1.5% but not 0.75% dose levels. Moreover,acetylsalicylic acid, an inhibitor of cyclooxygenase dose-dependentlyinhibited the promotion activity of PB. Among the antioxidantsinvestigated, vitamin E did not affect, but n-propyl gallateand ethoxyquin exerted a dose-dependent inhibition of PB promotion.These results are strongly suggestive of an involvement of phospholipaseA₂ lipoxygenase and cyclooxygenase arachidonic acid metabolicpathways in the mechanisms underlying PB enhancement of hepatocarcinogenesis.     

Anti-cholinergic effect of verapamil on the muscarinic acetylcholine receptor-gated K+ channel in isolated guinea-pig atrial myocytes

Summary Effects of verapamil on the acetylcholine (ACh)-induced K⁺ current were examined in single atrial cells, using the tight-seal whole-cell clamp technique. The pipette solution contained guanosine-5-triphosphate (GTP) or guanosine-5-O-(3-thiotriphosphate) (GTP-S, a non-hydrolysable GTP analogue). In GTP-loaded cells, ACh induced a specific K⁺ current, which is known to be mediated by pertussis toxin-sensitive GTP-binding (G) proteins. Verapamil (0.1–100 M) depressed the ACh-induced K⁺ current in a concentration-dependent fashion. In GTP-S-loaded cells, the K⁺ current remained persistently after wash-out of ACh, probably due to irreversible activation of G proteins by GTP-S. Verapamil (0.1–100 M) also depressed the intracellular GTP-S-induced K⁺ current. However, the magnitude of verapamil-depression of the K⁺ current in GTP-S-loaded cells was significantly smaller than that in GTP-loaded cells at concentrations between 1 and 10 M of the drug. From these results, it is suggested that verapamil may block not only the function of muscarinic ACh receptors but also of G proteins and/or the K⁺ channel itself and thereby depress the ACh-induced K⁺ current in isolated atrial myocytes.Supported by grants from the Ministry of Education, Science and Culture of Japan and the Research Program on Ca Signal Control Send offprint requests to Y. Kurachi at the above address     

Microarray analysis on Phase II drug metabolizing enzymes expression in pregnant rats after treatment with pregnenolone-16alpha-carbonitrile or phenobarbital

We previously reported the expression profiles of 9 cytochrome P450 isozymes (CYPs) proteins and those of 40 CYPs genes in pregnant rat's liver, placenta and fetal liver after treatment with pregnenolone-16alpha-carbonitrile (PCN) or phenobarbital (PB). This study was carried out focusing on the gene expression profiles of Phase II drug metabolizing enzymes, Glutathione S-transferase isozymes (GSTs) and UDP-glycosyltransferase isozymes (UDPGTs). Fischer 344 (F344) pregnant rats were daily treated intraperitoneally with 50 mg/kg of PCN or 80 mg/kg of PB from 13 to 16 days of gestation (DG). They were sacrificed on 17 DG, and microarray analysis using Affymetrix Rat Expression Array 230 A was performed. Among 16 GSTs genes examined in this study, 7 genes were significantly induced in dam's liver and 3 genes in fetal liver, respectively, in the PCN-group, while 8 genes were significantly induced in dam's liver and 1 gene in fetal liver, respectively, in the PB-group. On the other hand, among 11 UDPGTs genes examined, 5 genes were significantly induced in dam's liver and 3 genes in fetal liver, respectively, in the PCN-group, while 5 genes were significantly induced in dam's liver and 1 gene in fetal liver, respectively, in the PB-group. There were no significant changes in the placenta of all groups. This is the first report of the gene expression profiles of Phase II drug metabolizing enzymes in pregnant rat and fetal livers and placenta after treatment with typical inducers of drug metabolizing enzymes.     

The TRPV4 channel: structure-function relationship and promiscuous gating behaviour

Transient receptor potential (TRP) channels provide an enormous variability of Ca(2+) influx mechanisms triggered by a wide range of stimuli. In this review, we discuss the activation properties of the Ca(2+)- and Mg(2+)-permeable TRP channel of the vanilloid subfamily TRPV4. This channel is activated by various physical and chemical stimuli, such as cell swelling, heat, phorbols and, probably, by endogenous ligands, which are able to induce Ca(2+) entry. Not much is known about the regulation of this channel. We will refer only to a mechanism of Ca(2+)-dependent inhibition of TRPV4. Possible functional roles of this channel will be correlated with its observed expression pattern. Finally, we discuss the structural determinants of TRPV4 channel function.     

Increased susceptibility to HIV-1 of peripheral blood lymphocytes in acute infection with Epstein-Barr virus

Epstein-Barr virus (EBV) is an important pathogen in human immunodeficiency virus (HIV)-infected individuals that causes lymphoma and other lymphoproliferative disorders upon disease progression; however, interaction between the two viruses during acute infection is not well known. Expression of CCR5, a major coreceptor for HIV, was enhanced on CD4+ T cells from patients with acute EBV infection. Furthermore, susceptibility of those cells to R5-HIV-1, but not X4-HIV-1, was increased. EBV effects on CCR5 expression on or susceptibility to R5-HIV-1 of CD4+ T cells did not require coinfection of the same cell with the two viruses, because CD4+ T cells from patients with acute EBV infection were not infected with EBV. Considering that both HIV and EBV are transmitted by intimate contact, such possible interaction between the two viruses may have implications for viral transmission and the pathogenesis of HIV disease.     

Immunogenetic analysis of gastric MALT lymphoma-like lesions induced by Helicobacter pylori infection in neonatally thymectomized mice

Most gastric mucosa-associated lymphoid tissue (MALT) lymphomas are caused by Helicobacter pylori (H. pylori) infection. We previously reported that acquired lymphoid follicles with germinal centers were induced by H. pylori infection in neonatally thymectomized (nTx) mice. In the present study, we developed gastric MALT lymphoma-like lesions in nTx mice by long-term H. pylori infection, and performed immunogenetic analyses. BALB/c mice were thymectomized on the 3rd day after birth. At 6 weeks of age, mice were orally infected with 10(8) H. pylori and serially killed 2, 4, 6, and 12 months later. Normal BALB/c and noninfected nTx mice served as controls. Follicle formation occurred after 2 months of H. pylori infection in the nTx mice. Follicle formation and infiltration of intraepithelial lymphocytes progressed in a time-dependent manner. Lymphoepithelial lesions, a characteristic feature of MALT lymphoma, also occurred in a time-dependent manner (100% at 12 months). Serum immunoelectrophoresis revealed a monoclonal band (M-protein) in 30% (3/10) of mice 6 months after infection. M-protein-positive mice had amplification of one or two IgM and/or IgG heavy-chain genes in the gastric B lymphocytes, as determined with polymerase chain reaction, suggesting mono- or oligoclonality. Overexpression of Bcl-X(L) protein was immunohistologically observed in the infiltrating B lymphocytes and in some follicular B lymphocytes in 80% (8/10) of the cases at 12 months. Thus, H. pylori infection is involved in the development of gastric MALT lymphoma-like lesions in nTx mice. Our mouse model is useful for clarifying the pathogenetic mechanism of gastric MALT lymphoma by H. pylori infection.     

Simulation studies of data classification by artificial neural networks: Potential applications in medical imaging and decision making

Artificial neural networks are being investigated in the field of medical imaging as a means to facilitate pattern recognition and patient classification. In the work reported here, the effects of internal structure and the nature of input data on the performance of neural networks were investigated systematically using computer-simulated data. Network performance was evaluated quantitatively by means of receiver operating characteristic analysis and compared with the performance of an ideal statistical decision maker. We found that the relatively simple neural networks investigated in this study can perform at the level of an ideal decision maker. These simple networks were also found to learn accurately even when the training data are extremely unbalanced with respect to the prevalence of actually positive cases and to differentiate input data patterns by recognizing their unique characteristics.     

Statistical and integrative approach for constructing biological network maps

A goal of systems biology is to build a concrete biochemical network map, which provides an important instruction to trace the pathways of interest or to understand the mechanism of a biological system. In the postgenomic era, not only the concrete biochemical maps, but also postgenomic maps (mRNA coexpression and protein-protein interaction networks) have been extensively produced. In the biochemical map, the individual reactions are reliable, but the number of the reactions is limited, because molecular biology requires extensive experiments to verify them. By contrast, postgenomic data provide much information regarding interactions, but are coarse-grained. To expand the biochemical network, an intuitional approach, which superposes postgenomic data on the map one by one, has been carried out, but it is not effective when a large amount of the coarse-grained data is handled. In order to effectively integrate such postgenomic interactions into a biochemical map, a statistical approach would be suitable rather than intuition. In this article, we proposed a novel statistical approach that integrates postgenomic interaction networks into the biochemical network, predicting novel pathways. A statistical correlation for such different types of networks identifies functional modules; subsequently the superposition of the different networks on the functional modules predicts inter-modular relations, which are the key pathways to construct a large-scale biochemical network.