Genistein effects on growth and cell cycle ofCandida albicans

Microbial virulence is generally considered to be multifactorial with infection resulting from the sum of several globally regulated virulence factors. Estrogen may serve as a signal for global virulence induction in Candida albicans. Nonsteroidal estrogens and estrogen receptor antagonists may therefore have interesting effects on yeast and their virulence factors. Growth of C. albicans was monitored by viable plate counts at timed intervals after inoculation into yeast nitrogen broth plus glucose. To determine if increased growth of yeast in the presence of estradiol was due to tyrosine kinase-mediated signaling, we measured growth in the presence of genistein, estradiol or genistein plus estradiol and compared these conditions to controls, which were not supplemented with either compound. Unexpectedly, genistein stimulated growth of C. albicans. In addition, genistein was found to increase the rate of germination (possibly reflecting release from G(0) into G(1) cell cycle phase) and also increased Hsp90 expression, demonstrated by a dot blot technique which employed a commercial primary antibody detected with chemiluminescence with horseradish peroxidase-labeled secondary antibody. These biological effects may be attributable to genistein's activity as a phytoestrogen. In contrast, nafoxidine suppressed growth of Candida and mildly diminished Hsp90 expression. This study raises the possibility of receptor cross-talk between estrogen and isoflavinoid compounds, and antiestrogens which may affect the same signaling system, though separate targets for each compound were not ruled out.     

Dopamine and alcohol relapse: D1 and D2 antagonists increase relapse rates in animal studies and in clinical trials

A considerable number of animal studies on the effects of dopaminergic agents on alcohol intake behavior have been performed. Acute alcohol administration in rats induces dopamine release in the caudate nucleus and in the nucleus accumbens, an effect related among others to reinforcement. It has been repeatedly suggested that D1 and D2 receptor activation mediates reward. As alcohol consumption and dopaminergic transmission seem to have a close relationship, all kinds of dopaminergic agents may be regarded as putative therapeutics for preventing relapse. In a prospective European double-blind multicenter clinical trial, comparing the D1, D2, D3 antagonist flupenthixol and placebo in 281 chronic alcohol-dependent patients (27.4% women), the application of the Lesch typology made an outcome differentiation possible. It could be shown in which patients flupenthixol administration was followed by a significantly higher relapse rate and in which patient groups no differences were found when compared to placebo.     

Microarray profiling of gene expression patterns in bladder tumor cells treated with genistein

Microarray technology was used to gain an insight into the molecular events of tumor cell growth inhibition mediated by the soy isoflavone genistein. For this, a susceptible bladder tumor line TCCSUP was treated with the inhibitory dose (50 microM) of genistein for various periods of time, followed by mRNA isolations, cDNA probe preparations, and hybridization individually to cDNA chips containing 884 sequence-verified known human genes. After analyzing the hybridization signals with a simple quantitative method developed by this study, we detected that egr-1, whose expression has been associated with proliferation and differentiation, was transiently induced and this expression pattern was later confirmed by RT-PCR. Thus, microarray technology is a reliable and powerful tool for profiling gene expression patterns in many biological systems related to cancer. We further detected many groups of genes with distinct expression profiles and most of them encode for proteins that regulate the signal transduction or the cell cycle pathways. These genes warrant further investigation as regards their roles in the susceptibility of the tumor cell line to the antitumor drug.     

Intragastric ethanol infusion model for cellular and molecular studies of alcoholic liver disease

The intragastric alcohol infusion rat model (IAIRM) of alcoholic liver disease (ALD) has been utilized in various laboratories to study various aspects of ALD pathogenesis including oxidative stress, cytokine upregulation, hypoxic damage, apoptosis, ubiquitin-proteasome pathway and CYP2E1 induction. The basic value of the model is that it produces pathologic changes which resemble ALD including microvesicular and macrovesicular fat, megamitochondria, apoptosis, central lobular and pericellular fibrosis, portal fibrosis, bridging fibrosis, central necrosis, and mixed inflammatory infiltrate including PMNs and lymphocytes. The model is valuable because the diet and ethanol intake are totally under the control of the investigator. A steady state can be maintained with high or low blood alcohol levels for long periods. The cycling of the blood alcohol levels, when a constant infusion rate of alcohol is maintained, simulates binge drinking. Using this model the importance of dietary fat, especially the degree of saturation of the fatty acids on the induction of liver pathology, has been documented. The role of endotoxin, the Kupffer cell, TNFalpha, and NADPH oxidase have been demonstrated. The importance of 2E1 in oxidative stress induction has been shown using inhibitors of the isozyme. The importance of dietary iron in the pathogenesis of cirrhosis has been documented. Acetaldehyde has been shown to play a role in preventing liver pathology by preventing NFkappaB activation. Using the model, to maintain high blood alcohol levels is found to be necessary to demonstrate proteasomal peptidase inhibition. Ubiquitin synthesis is also inhibited at high blood alcohol levels in the IAIRM model. Oxidized proteins accumulate in the liver at high blood alcohol levels. Neoantigens derived from protein adducts formed with products of oxidation induce autoimmune mechanisms of liver injury. Thus, in many ways the model has revolutionized our understanding of the pathogenesis of ALD.     

Oxygen and nitrate-dependent regulation of <Emphasis Type="Italic">dmsABC</Emphasis> operon expression in <Emphasis Type="Italic">Escherichia coli</Emphasis>: sites for Fnr and NarL protein interactions

Background  

Escherichia coli can respire anaerobically using dimethyl sulfoxide (DMSO) or trimethylamine-N-oxide (TMAO) as the terminal electron acceptor for anaerobic energy generation. Expression of the dmsABC genes that encode the membrane-associated DMSO/TMAO reductase is positively regulated during anaerobic conditions by the Fnr protein and negatively regulated by the NarL protein when nitrate is present.     

A comparison of cholinesterase activity after intravenous, oral or dermal administration of methyl parathion

Time-dependent changes in blood cholinesterase activity caused by single intravenous, oral or dermal administration of methyl parathion to adult female rats were defined. Intravenous and oral administration of 2.5 mg/kg methyl parathion resulted in rapid (<60 min) decreases in cholinesterase activity which recovered fully in vivo within 30-48 h. In contrast, spontaneous reactivation of cholinesterase in vitro was complete within 6 h at 37 degrees C. Dermal administration of methyl parathion caused dose-dependent inhibition of cholinesterase activity which developed slowly (> or =6 h) and was prolonged (> or =48 h). Time- and route-dependent effects of methyl parathion on cholinesterase activity in brain and other tissues generally paralleled its effects on activity in blood. In conclusion, pharmacodynamics of methyl parathion differ substantially with route of exposure. Recovery of cholinesterase in vivo after intravenous or oral exposure may partially reflect spontaneous reactivation and suggests a rapid clearance of methyl parathion or its active metabolite methyl paraoxon. The more gradual and prolonged inhibition of cholinesterase caused by dermal administration is consistent with disposition of methyl parathion at a site from which it or methyl paraoxon is only slowly distributed. Thus, dermal exposure to methyl parathion may pose the greatest risk for long-term adverse effects.     

The joys of HexNAc. The synthesis and function of N-andO-glycan branches

This review covers discoveries made over the past 30–35 years that were important to our understanding of the synthetic pathway required for initiation of the antennae or branches on complex N-glycans and O-glycans. The review deals primarily with the author's contributions but the relevant work of other laboratories is also discussed. The focus of the review is almost entirely on the glycosyltransferases involved in the process. The following topics are discussed. (1) The localization of the synthesis of complex N-glycan antennae to the Golgi apparatus. (2) The evolutionary boundary at the stage in N-glycan processing where there is a change from oligomannose to complex N-glycans; this switch correlates with the appearance of multicellular organisms. (3) The discovery of the three enzymes which play a key role in this switch, N-acetylglucosaminyltransferases I and II and mannosidase II. (4) The yellow brick road which leads from oligomannose to highly branched complex N-glycans with emphasis on the enzymes involved in the process and the factors which control the routes of synthesis. (5) A short discussion of the characteristics of the enzymes involved and of the genes that encode them. (6) The role of complex N-glycans in mammalian and Caenorhabditis elegans development. (7) The crystal structure of N-acetylglucosaminyltransferase I. (8) The discovery of the enzymes which synthesize O-glycan cores 1, 2, 3 and 4 and their elongation.     

Expression of a truncated retroviral envelope gene enhances expression of normal cellular phenotypes

The envelope gene of Moloney murine leukemia virus (Mo-MLV) and its various functional domains have been studied extensively but not as much in terms of their biological effects on cell growth. In this study, we report the biological characterization of a truncated Mo-MLV envelope gene, LN11, which is devoid of a signal peptide. Its expression in various cell types, as compared to the control, enabled the transduced cells to assume a more normal phenotype, which is defined by an increase in contact inhibition and factor dependence, as well as reduced tumorigenicity. LN11-transduced fibroblasts exhibited a higher degree of contact inhibition, assumed a more flattened morphology and were more adherent compared to the control. In v-abl transformed hematopoietic cells, expression of LN11 resulted in slower cell growth, which was due to an enhanced dependence on exogenous growth factors. Enforced expression of LN11 also resulted in a slower rate of tumor development and a reduced tumor load. Thus, modification of a retroviral genome could have a significant impact on cell growth and development. This is one example where we need to consider the safety issue carefully when constructing retrovirus vectors for gene therapy.     

p53 gene status modulates the chemosensitivity of non-small cell lung cancer cells

This study examined the effects of p53 gene status on DNA damage-induced cell death and chemosensitivity to various chemotherapeutic agents in non-small cell lung cancer (NSCLC) cells. A mutant p53 gene was introduced into cells carrying the wild-type p53 gene and also vice versa to introduce the wild-type p53 gene into cells carrying the mutant p53 gene. Chemosensitivity and DNA damage-induced apoptosis in these cells were then examined. This study included five cell lines, NCI-H1437, NCI-H727, NCI-H441 and NCI-H1299 which carry a mutant p53 gene and NCI-H460 which carries a wild-type p53 gene. Mutant p53-carrying cells were transfected with the wild-type p53 gene, while mutant p53 genes were introduced into NCI-H460 cells. These p53 genes were individually mutated at amino acid residues 143, 175, 248 and 273. The representative cell line NCI-H1437 cells transfected with wild-type p53 gene (H1437/wtp53) showed a dramatic increase in susceptibility to three anticancer agents (7-fold to cisplatin, 21-fold to etoposide, and 20-fold to camptothecin) compared to untransfected or neotransfected H1437 cells. An increase in chemosensitivity was also observed in wild-type p53 transfectants of H727, H441, H1299 cells. The results of chemosensitivity were consistent with the observations on apoptotic cell death. H1437/wtp53 cells, but not H1437 parental cells, exhibited a characteristic feature of apoptotic cell death that generated oligonucleosomal-sized DNA fragments. In contrast, loss of chemosensitivity and lack of p53-mediated DNA degradation in response to anticancer agents were observed in H460 cells transfected with mutant p53. These observations suggest that the increase in chemosensitivity was attributable to wild-type p53 mediation of the process of apoptosis. In addition, our results also suggest that p53 gene status modulates the extent of chemosensitivity and the induction of apoptosis by different anticancer agents in NSCLC cells.