Cell wall component and mycotoxin moieties involved in the binding of fumonisin B1 and B2 by lactic acid bacteria.

AIMS: The ability of lactic acid bacteria (LAB) to bind fumonisins B1 and B2 (FB1, FB2) in fermented foods and feeds and in the gastrointestinal tract could contribute to decrease their bioavailability and toxic effects on farm animals and humans. The aim of this work was to identify the bacterial cell wall component(s) and the functional group(s) of FB involved in the LAB-FB interaction. METHODS AND RESULTS: The effect of physicochemical, enzymatic and genetic treatments of bacteria and the removal/inactivation of the functional groups of FB on toxin binding were evaluated. Treatments affecting the bacterial wall polysaccharides, lipids and proteins increased binding, while those degrading peptidoglycan (PG) partially decreased it. In addition, purified PG from Gram-positive bacteria bound FB in a manner analogue to that of intact LAB. For FB, tricarballylic acid (TCA) chains play a significant role in binding as hydrolysed FB had less affinity for LAB. CONCLUSIONS: Peptidoglycan and TCA are important components of LAB and FB, respectively, involved in the binding interaction. SIGNIFICANCE AND IMPACT OF THE STUDY: Lactic acid bacteria binding efficiency seems related to the peptide moiety structure of the PG. This information can be used to select probiotics with increased FB binding efficiency.

[Absolute risk fracture prediction by risk factors validation and survey of osteoporosis in a Brussels cohort followed during 10 years (FRISBEE study)]. / Prédiction du risque fracturaire absolu par validation de facteurs de risque et dépistage de l'ostéoporose dans une cohorte bruxelloise suivie pendant 10 ans (etude FRISBEE).

Osteoporosis is a major public health problem. For the time being, the diagnosis of osteoporosis relies on densitometry (T-score < -2.5 by DXA), although the risk of fracture depends also on other factors than the bone mass. Osteoporosis diagnosis (DXA) must be distinguished from the individual risk assessment of fracture. Different risk factors complementary to bone mass have been already validated in different populations. These include an old age, a history of fracture after the age of 50, a familial history of hip fracture (father or mother), a low BMI (< 20), corticoid treatment (> 3 months), tabagism and excessive alcohol consumption. A WHO taskforce has combined these different factors in order to integrate them in a 10-years predictive risk model of fracture (FRAX**). This model should still be validated in different populations, especially in populations not included in its development, which is the case for Belgium. We are evaluating these different risk factors for fracture in a Brussels population of 5000 women (60-80 years) who will be followed each year during 10 years. We also assess the predictive value of other risk factors for fracture not included in the WHO model (tendency to fall, use of sleeping pills, early non substituted menopause, sedentarity, ...). In an interim analysis of the first 452 women included and with data yet available at the time of this writing, we could find a significant (P < 0.05) relationship between diagnosis of osteoporosis at DXA and the number of risk factors, age > 70 years, a personal history of fracture after 50 years and a BMI < 20.

Effects of infant formula composition on long-term metabolic health.

Early nutrition may have long-lasting metabolic impacts in adulthood. Even though breast milk is the gold standard, most infants are at least partly formula-fed. Despite obvious improvements, infant formulas remain perfectible to reduce the gap between breastfed and formula-fed infants. Improvements such as reducing the protein content, modulating the lipid matrix and adding prebiotics, probiotics and synbiotics, are discussed regarding metabolic health. Numerous questions remain to be answered on how impacting the infant formula composition may modulate the host metabolism and exert long-term benefits. Interactions between early nutrition (composition of human milk and infant formula) and the gut microbiota profile, as well as mechanisms connecting gut microbiota to metabolic health, are highlighted. Gut microbiota stands as a key actor in the nutritional programming but additional well-designed longitudinal human studies are needed.

Splicing precedes polyadenylation during Drosophila E74A transcription.

The E74 gene is one of a small set of early genes induced by the steroid hormone ecdysone at the onset of metamorphosis in the fruit fly, Drosophila melanogaster. This complex gene directs the synthesis of a 60-kilobase (kb) primary transcript that is spliced to form the 6-kb E74A mRNA. In a previous study, we found that ecdysone directly activates the E74A promoter and determined that RNA polymerase II transcribes this gene at a rate of approximately 1.1 kb/min. This elongation rate accounts for most of the 1-hour delay seen between the addition of ecdysone and the appearance of cytoplasmic E74A mRNA (C. S. Thummel, K. C. Burtis, and D. S. Hogness, Cell 61:101-111, 1990). We show here that nascent E74A transcripts are spliced, and we propose a model for the order of that splicing. This study provides, for the first time, direct biochemical evidence for splicing of a low-abundance cellular RNA before transcription termination and polyadenylation.

Isolation and characterization of human orthologs of yeast CCR4-NOT complex subunits.

The yeast CCR4-NOT protein complex is a global regulator of RNA polymerase II transcription. It is comprised of yeast NOT1 to NOT5, yeast CCR4 and additional proteins like yeast CAF1. Here we report the isolation of cDNAs encoding human NOT2, NOT3, NOT4 and a CAF1-like factor, CALIF. Analysis of their mRNA levels in different human tissues reveals a common ubiquitous expression pattern. A multitude of two-hybrid interactions among the human cDNAs suggest that their encoded proteins also form a complex in mammalian cells. Functional conservation of these proteins throughout evolution is supported by the observation that the isolated human NOT3 and NOT4 cDNAs can partially com-plement corresponding not mutations in yeast. Interestingly, human CALIF is highly homologous to, although clearly different from, a recently described human CAF1 protein. Conserved interactions of this factor with both NOT and CCR4 proteins and co-immunoprecipitation experiments suggest that CALIF is a bona fide component of the human CCR4-NOT complex.

Functions of S-layers.

Although S-layers are being increasingly identified on Bacteria and Archaea, it is enigmatic that in most cases S-layer function continues to elude us. In a few instances, S-layers have been shown to be virulence factors on pathogens (e.g. Campylobacter fetus ssp. fetus and Aeromonas salmonicida), protective against Bdellovibrio, a depository for surface-exposed enzymes (e.g. Bacillus stearothermophilus), shape-determining agents (e.g. Thermoproteus tenax) and nucleation factors for fine-grain mineral development (e.g. Synechococcus GL 24). Yet, for the vast majority of S-layered bacteria, the natural function of these crystalline arrays continues to be evasive. The following review up-dates the functional basis of S-layers and describes such diverse topics as the effect of S-layers on the Gram stain, bacteriophage adsorption in lactobacilli, phagocytosis by human polymorphonuclear leukocytes, the adhesion of a high-molecular-mass amylase, outer membrane porosity, and the secretion of extracellular enzymes of Thermoanaerobacterium. In addition, the functional aspect of calcium on the Caulobacter S-layer is explained.

Xenon-mediated neuroprotection in response to sustained, low-level excitotoxic stress.

Noble gases such as xenon and argon have been reported to provide neuroprotection against acute brain ischemic/anoxic injuries. Herein, we wished to evaluate the protective potential of these two gases under conditions relevant to the pathogenesis of chronic neurodegenerative disorders. For that, we established cultures of neurons typically affected in Alzheimer's disease (AD) pathology, that is, cortical neurons and basal forebrain cholinergic neurons and exposed them to L-trans-pyrrolidine-2,4-dicarboxylic acid (PDC) to generate sustained, low-level excitotoxic stress. Over a period of 4 days, PDC caused a progressive loss of cortical neurons which was prevented substantially when xenon replaced nitrogen in the cell culture atmosphere. Unlike xenon, argon remained inactive. Xenon acted downstream of the inhibitory and stimulatory effects elicited by PDC on glutamate uptake and efflux, respectively. Neuroprotection by xenon was mimicked by two noncompetitive antagonists of NMDA glutamate receptors, memantine and ketamine. Each of them potentiated xenon-mediated neuroprotection when used at concentrations providing suboptimal rescue to cortical neurons but most surprisingly, no rescue at all. The survival-promoting effects of xenon persisted when NMDA was used instead of PDC to trigger neuronal death, indicating that NMDA receptor antagonism was probably accountable for xenon's effects. An excess of glycine failed to reverse xenon neuroprotection, thus excluding a competitive interaction of xenon with the glycine-binding site of NMDA receptors. Noticeably, antioxidants such as Trolox and N-acetylcysteine reduced PDC-induced neuronal death but xenon itself lacked free radical-scavenging activity. Cholinergic neurons were also rescued efficaciously by xenon in basal forebrain cultures. Unexpectedly, however, xenon stimulated cholinergic traits and promoted the morphological differentiation of cholinergic neurons in these cultures. Memantine reproduced some of these neurotrophic effects, albeit with less efficacy than xenon. In conclusion, we demonstrate for the first time that xenon may have a therapeutic potential in AD.

A stable open-shell redox active ditopic ligand.

Herein we describe the synthesis, structure and electronic properties of an unusual redox-active ditopic ligand with a stable open-shell configuration. This stable phenoxyl radical features intense and very low energy electronic transitions in the near infrared (NIR) part of the spectrum and is structurally set up to strongly spin couple coordinated transition metal ions in [2 × 2] grid-type structures.

Dose-response relationship of epirubicin-based first-line chemotherapy for advanced breast cancer: a prospective randomized trial.

PURPOSE: We compared prospectively the antitumor efficacy of two combination chemotherapy regimens with two different dose levels of epirubicin as first-line treatment for advanced breast cancer. PATIENTS AND METHODS: One hundred forty-one fully assessable patients were randomized to receive either our intensified schedule (group A, n = 71) of epirubicin 50 mg/m2 on days 1 and 8 (every 3 weeks), or a non-intensified program (group B, n = 70) in which epirubicin was only administered on day 1. Both groups also received fluorouracil (5 FU) and cyclophosphamide 500 mg/m2 on day 1 of each course. RESULTS: A statistically significant difference in response rate was observed (69% in group A v 41% in group B, P < .001) for both locally advanced (LA) and recurrent metastatic (RM) disease. Response duration (22 v 14 months, P < .01) and time to progression (TTP; 19 v 8 months, P < .02) were also significantly improved. Overall survival was similar in both groups. However, univariate and/or multivariate analyses showed a meaningful relationship between type of treatment allocated, dose-intensity (DI) of epirubicin, and response rate, as well as between TTP and survival. Ultimately, TTP and survival were also influenced by further treatment modalities, namely, hormonotherapy and chemotherapy. CONCLUSION: This study validates prospectively the concept of a dose-response relationship for an anthracycline-based chemotherapy in previously untreated advanced breast cancer.

Increased affective reactivity to neutral stimuli and decreased maintenance of affective responses in bipolar disorder.

BACKGROUND: Affective dysregulation is a core feature of bipolar disorder (BD) and a significant predictor of clinical and functional outcome. Affective dysregulation can arise from abnormalities in multiple processes. This study addresses the knowledge gap regarding the precise nature of the processes that may be dysregulated in BD and their relationship to the clinical expression of the disorder. METHODS: Patients with BD (n=45) who were either in remission or in a depressive or manic state and healthy individuals (n=101) were compared in terms of the intensity, duration and physiological response (measured using inter-beat intervals and skin conductance) to affective and neutral pictures during passive viewing and during experiential suppression. RESULTS: Compared to healthy individuals, patients with BD evidenced increased affective reactivity to neutral pictures and reduced maintenance of subjective affective responses to all pictures. This pattern was present irrespective of clinical state but was more pronounced in symptomatic patients, regardless of polarity. Patients, regardless of symptomatic status, were comparable to healthy individuals in terms of physiological arousal and voluntary control of affective responses. CONCLUSION: Our study demonstrates that increased affective reactivity to neutral stimuli and decreased maintenance of affective responses are key dimensions of affective dysregulation in BD.

Structure and magnetic properties of an unusual homoleptic iron(III) thiocyanate dimer.

We describe the structural and variable temperature magnetic susceptibility properties of an unusual homoleptic bimetallic iron(III) thiocyanate tetraanion. This work represents the first structurally characterized bis(µ-1,3-thiocyanato) dimer of iron(III). A weak antiferromagnetic exchange interaction is observed between the two iron(III) ions, which is supported by broken symmetry density functional theory (DFT) calculations.

Argon blocks the expression of locomotor sensitization to amphetamine through antagonism at the vesicular monoamine transporter-2 and mu-opioid receptor in the nucleus accumbens.

We investigated the effects of the noble gas argon on the expression of locomotor sensitization to amphetamine and amphetamine-induced changes in dopamine release and mu-opioid neurotransmission in the nucleus accumbens. We found (1) argon blocked the increase in carrier-mediated dopamine release induced by amphetamine in brain slices, but, in contrast, potentiated the decrease in KCl-evoked dopamine release induced by amphetamine, thereby suggesting that argon inhibited the vesicular monoamine transporter-2; (2) argon blocked the expression of locomotor and mu-opioid neurotransmission sensitization induced by repeated amphetamine administration in a short-term model of sensitization in rats; (3) argon decreased the maximal number of binding sites and increased the dissociation constant of mu-receptors in membrane preparations, thereby indicating that argon is a mu-receptor antagonist; (4) argon blocked the expression of locomotor sensitization and context-dependent locomotor activity induced by repeated administration of amphetamine in a long-term model of sensitization. Taken together, these data indicate that argon could be of potential interest for treating drug addiction and dependence.

Rising plasma levels of 19-nortestosterone throughout pregnancy: determination by radioimmunoassay and validation by gas chromatography-mass spectrometry.

A highly sensitive and specific radioimmunoassay was developed to measure 19-nortestosterone (NT), which allowed us to demonstrate this steroid in the plasma of pregnant women. Plasma NT was detectable throughout gestation, reaching values of 12 to 60 pg/ml in the 3rd trimester. These results were validated by gas chromatography-mass spectrometry. No NT was detectable in the plasma of 12 normal men and 12 nonpregnant women in the mid follicular phase of their cycle (detection limit 4 pg/ml). Our results are compatible with current concepts concerning the possible involvement of 19-norsteroids in an accessory biosynthetic pathway for estrogen in the placenta.

Protection against aspirin-induced gastric lesions by lansoprazole: simultaneous evaluation of functional and morphologic responses.

The protective effect of lansoprazole, a new proton pump inhibitor, against aspirin-induced gastric lesions was studied in a double-blind crossover trial with a simultaneous measure of the functional capacities of the mucosal barrier (by a recording of the gastric potential difference) and of the morphologic changes in the mucosa (by gastric endoscopy). After 1 week of treatment with lansoprazole (30 mg per day) or placebo, each healthy volunteer received 1 gm aspirin by mouth. Recording of the gastric potential difference lasted for 3 hours and was followed by gastric endoscopy. Morphologic lesions induced by aspirin were effectively prevented by lansoprazole: Lanza score was 0.67 +/- 0.98 (mean +/- SD) versus 2.25 +/- 1.1 with placebo (p < 0.005, ANOVA). Conversely, the decrease in the gastric potential difference was similar. The inhibition of acid secretion induced by lansoprazole was therefore sufficient to prevent aspirin-induced mucosal lesions without reinforcing the defense capacities of the mucosa. This simple pharmacologic model makes it possible to simultaneously evaluate the functional and morphologic effects of aspirin intake on the gastric mucosa.

Intravenous cyclosporine kinetics in renal failure.

Kinetics of the novel immunosuppressive cyclosporine were determined in four patients with terminal renal failure. After a short intravenous infusion (2.05 to 3.5 mg/kg in 4 hr), blood and plasma concentrations were measured (HPLC and radioimmunoassay [RIA] up to 36 hr. After infusion, concentration curves of the drug were characterized by a rapid initial fall (t 1/2 alpha 0.10 +/- 0.03 hr), followed by a biphasic elimination phase with corresponding t 1/2s of 1.08 +/- 0.25 hr (t 1/2 beta) and 15.8 +/- 8.4 hr (t 1/2 gamma). The volumes of distribution, calculated from whole blood concentrations (HPLC), were 0.140 +/- 0.48 l/kg (volume of the central compartment) and 3.49 +/- 2.65 l/kg (volume of distribution at steady state), whereas systemic clearances were 0.369 +/- 0.08 l/hr/kg. Blood levels measured by RIA exceeded the HPLC values after the fourth hour by up to 100%, indicating the production of cross-reacting cyclosporine metabolites. Plasma concentrations were considerably lower than in whole blood. Elimination of unchanged cyclosporine in patients with renal failure appears to be of the same order as in those with normal kidney function. Modification of the initial dosage regimens is therefore probably not required.

An active-site cysteine of sorghum leaf NADP-malate dehydrogenase studied by site-directed mutagenesis.

The chloroplast NADP-malate dehydrogenase is activated through the reduction of two different disulfides per subunit. The activated enzyme, as well as a permanently active mutant where all four regulatory cysteines were replaced are still sensitive to thiol reagents. This observation suggested the presence of an additional important cysteine at the active site. In an attempt to identify that cysteine, site-directed mutagenesis was performed on the cDNA encoding sorghum leaf NADP-malate dehydrogenase. The replacement of Cys-175 by an alanine yielded an enzyme whose sensitivity to thiol reagents was markedly decreased whereas its catalytic activity was enhanced. This finding suggests that Cys-175 has no catalytic function but is located close to the active site.

Direct evidence for the different roles of the N- and C-terminal regulatory disulfides of sorghum leaf NADP-malate dehydrogenase in its activation by reduced thioredoxin.

Plant NADP-dependent malate dehydrogenase is activated through thiol/disulfide interchange with reduced thioredoxin. Previous studies showed that this process involves the reduction of two different disulfides per subunit: one N-terminal, the other C-terminal. Substitution of regulatory cysteines at each end by site-directed mutagenesis and comparison of activation kinetics of the mutants led us to propose a model for the activation mechanism where the C-terminal end shielded the access to the catalytic residues, whereas the N- terminal end was involved in the slow conformational change of the active site. In the present study, we took advantage of the previous identification of the catalytic histidine residue which can be specifically derivatized by diethyl pyrocarbonate to test the accessibility of the active site. The results clearly show that in the mutants where the C-terminal bridge is open the active site histidine is freely accessible to the reagent, whereas in the mutants where the N-terminal bridge is open, the active site cannot be reached without activation, thus demonstrating the validity of the model.

Genomic exploration of the hemiascomycetous yeasts: 11. Kluyveromyces lactis.

Random sequencing of the Kluyveromyces lactis genome allowed the identification of 2235-2601 open reading frames (ORFs) homologous to S. cerevisiae ORFs, 51 ORFs which were homologous to genes from other species, 64 tRNAs, the complete rDNA repeat, and a few Ty1- and Ty2-like sequences. In addition, the complete sequence of plasmid pKD1 and a large coverage of the mitochondrial genome were obtained. The global distribution into general functional categories found in Saccharomyces cerevisiae and as defined by MIPS is well conserved in K. lactis. However, detailed examination of certain subcategories revealed a small excess of genes involved in amino acid metabolism in K. lactis. The sequences are deposited at EMBL under the accession numbers AL424881-AL430960.

Biphenyl-derivatives of 2-amino-7-phosphono-heptanoic acid, a novel class of potent competitive N-methyl-D-aspartate receptor antagonists--II. Pharmacological characterization in vivo.

A selection of biphenyl-analogues of 2-amino-7-phosphonoheptanoic acid (AP7), N-methyl-D-aspartate (NMDA) receptor antagonists with high affinity in vivo efficacy. The lead compound SDZ EAB 515 was found to inhibit L-phenylalanine uptake by the large neutral amino acid carrier in vitro and in vivo; active transport may thus confer a good bioavailability to this class of compounds. CNS effects were demonstrated by significant changes in 2-deoxyglucose-uptake in various brain regions at doses from 1 to 10 mg/kg i.p. With the most active agent, SDZ 220-581, full protection against maximal electroshock seizures (MES) was obtained at oral doses of 10 mg/kg in rats and in mice. The compound had a fast onset (< or = 1 hr) and a long duration (> or = 24 hr) of action. Motor-debilitating effects (impairment of rotarod performance) occurred at doses about 10 times higher than those required for protection against MES. Neuroprotective activity was demonstrated by the ability of the compounds to reduce the extent of quinolinic acid-induced striatal lesions in rats, in the dose range of 3-15 mg/kg (i.p.) or 10-50 mg/kg (p.o.). In the middle cerebral artery occlusion (MCAO) model of focal cerebral ischemia in rats, the test compounds reduced the infarct size by 40-50% when given i.v. before or by 20-30% when given i.v. 1 hr after MCAO. SDZ 220-581 provided 20-30% protection at > or = 2 x 10 mg/kg p.o. This compound also showed analgesic activity at low oral doses in a model of neuropathic pain, although higher doses were required in model of mechanical inflammatory hyperalgesia. Unexpectedly, SDZ 220-581 at low s.c. doses counteracted the antiparkinsonian effects of L-DOPA in MPTP-treated marmosets. (Sub)chronic administration of SDZ 220-581 did not reduce its ability to protect against quinolinic acid neurotoxicity, and no upregulation of NMDA receptors was detected using a [3H]CGP-39653 binding assay. In conclusion, from a series of biphenyl-AP7-derivatives, SDZ 220-581 is clearly the most active compound in vivo. Its pharmacological profile with a good, long-lasting oral activity might open up novel therapeutic applications for competitive NMDA receptor antagonists.

Pyrazole bioisosteres of leflunomide as B-cell immunosuppressants for xenotransplantation and chronic rejection: scope and limitations.

T-cell immunosuppressant-based therapies efficiently control early graft rejection in allotransplantation settings. They fail, however, to prevent those rejection events which are mediated by transplant-induced antibody (Ab) responses such as those involved in xenograft and chronic allograft rejection. This is mainly due to their inability to block T-cell-independent Ab production against the transplanted organs. The bioactive metabolite 2(Z) of leflunomide (1) inhibits the formation of such Ab, but the drug has pharmacokinetic properties and a therapeutic window incompatible with transplantation indications. Pyrazole 3, a constrained analogue of 2(Z), was designed and shown to be conformationally and biologically similar to 2(Z). Further investigations with derivatives of 3 demonstrated that the pyrazoles had very tight structure-activity relationships, the only equipotent compound being 3o. However, in contrast to 2(Z), both 3 and 3o were inactive in vivo due to short half-life and drug concentrations lower than the in vitro obtained IC50 values. Compound 3o inhibits T-cell-independent Ab production by a different biochemical mechanism from that of 2(Z) and 3 and may therefore represent a valuable tool for the identification of new targets for B-cell inhibition.