Detection of microcystin synthetase genes in health food supplements containing the freshwater cyanobacterium Aphanizomenon flos-aquae.

In this study we investigated the presence of toxin-producing cyanobacterial contaminants in food supplements manufactured from blooms of the non-toxic freshwater cyanobacterium Aphanizomenon flos-aquae. Previous reports investigating the contamination of health food supplements with toxin-producing cyanobacteria have used chemical and or biochemical methods such as HPLC, ELISA and protein phosphatase assays. Whilst these studies have drawn attention to the presence of hepatotoxic microcystins in some commercially available food supplements, the methods used do not provide any information on the source of the contaminant. Such information would be useful for the quality control of food supplements produced for human consumption. In this study we applied a molecular technique, involving the amplification of the 16s rRNA gene, the phycocyanin operon, and two genes of the microcystin synthetase gene cluster to show that all 12 food supplement samples, sourced from various internet distributors and containing non-toxic A. flos-aquae, also contained toxigenic cyanobacteria. Sequencing of the microcystin synthetase genes detected in all of the food supplements showed that M. aeruginosa was the organism responsible for the production of microcystins in the samples. The presence of microcystins in the food supplements was confirmed by ELISA, with concentrations within the range of 0.1--4.72 microgg(-1) (microcystin-LR equivalents). Given that the molecular methods applied here are highly sensitive, and show good agreement with the results obtained from ELISA, we believe that they could potentially be used as a quality control technique for food products that contain cyanobacteria.     

A double-blind, placebo-controlled trial to assess the efficacy of quetiapine fumarate XR in very heavy-drinking alcohol-dependent patients

Background: Despite advances in developing medications to treat alcohol dependence, few such medications have been approved by the Food and Drug Administration. Identified molecular targets are encouraging and can lead to the development and testing of new compounds. Atypical antipsychotic medications have been explored with varying results. Prior research suggests that the antipsychotic quetiapine may be beneficial in an alcohol‐dependent population of very heavy drinkers. Methods: In this double‐blind, placebo‐controlled trial, 224 alcohol‐dependent patients who reported very heavy drinking were recruited across 5 clinical sites. Patients received either quetiapine or placebo and Medical Management behavioral intervention. Patients were stratified on gender, clinical site, and reduction in drinking prior to randomization. Results: No differences between the quetiapine and placebo groups were detected in the primary outcome, percentage heavy‐drinking days, or other drinking outcomes. Quetiapine significantly reduced depressive symptoms and improved sleep but had no effect on other nondrinking outcomes. Results from a subgroup analysis suggest that patients who reduced their drinking prior to randomization had significantly better drinking outcomes during the maintenance phase (p < 0.0001). No significant interactions, however, were observed between reducer status and treatment group. Finally, quetiapine was generally well tolerated. Statistically significant adverse events that were more common with quetiapine versus placebo include dizziness (14 vs. 4%), dry mouth (32 vs. 9%), dyspepsia (13 vs. 2%), increased appetite (11 vs. 1%), sedation (15 vs. 3%), and somnolence (34 vs. 9%). Conclusions: This multisite clinical trial showed no efficacy for quetiapine compared with placebo at reducing alcohol consumption in heavy‐drinking alcohol‐dependent patients.     

Anti‐ACSA‐2 defines a novel monoclonal antibody for prospective isolation of living neonatal and adult astrocytes

Astrocytes are the most abundant cell type of the central nervous system and cover a broad range of functionalities. We report here the generation of a novel monoclonal antibody, anti‐astrocyte cell surface antigen‐2 (Anti‐ACSA‐2). Flow cytometry, immunohistochemistry and immunocytochemistry revealed that Anti‐ACSA‐2 reacted specifically with a not yet identified glycosylated surface molecule of murine astrocytes at all developmental stages. It did not show any labeling of non‐astroglial cells such as neurons, oligodendrocytes, NG2⁺ cells, microglia, endothelial cells, leukocytes, or erythrocytes. Co‐labeling studies of GLAST and ACSA‐2 showed largely overlapping expression. However, there were also notable differences in protein expression levels and frequencies of single‐positive subpopulations of cells in some regions of the CNS such as cerebellum, most prominently at early postnatal stages. In the neurogenic niches, the dentate gyrus of the hippocampus and the subventricular zone (SVZ), again a general overlap with slight differences in expression levels were observed. ACSA‐2 was unlike GLAST not sensitive to papain‐based tissue dissociation and allowed for a highly effective, acute, specific, and prospective purification of viable astrocytes based on a new rapid sorting procedure using Anti‐ACSA‐2 directly coupled to superparamagnetic MicroBeads. In conclusion, ACSA‐2 appears to be a new surface marker for astrocytes, radial glia, neural stem cells and bipotent glial progenitor cells which opens up the possibility of further dissecting the characteristics of astroglial subpopulations and lineages.     

Detection of TOAD-64 in adult rat brain as revealed by two-dimensional protein gel electrophoresis followed by MALDI mass spectrometry possible modulatory effect of chronic haloperidol treatment

The molecular mechanisms by which antipsychotic effects are achieved remain largely elusive. Possible mechanisms include the modulation of nerve cell gene expression. The antipsychotic drug haloperidol was administered orally (1.6 mg/kg) to adult rats for 3 weeks. Protein patterns in striata and forebrains were studied by two-dimensional gel electrophoresis (2-DE). One differentially regulated protein spot was identified by matrix-assisted laser desorption ionization mass spectrometry (MALDI-MS) after trypsin digest. Turned on after devision-64 kD (TOAD-64), the identified protein, was present in all gels and, in addition, was up-regulated in the striata but not in the forebrains of the haloperidol-treated animals by 43%. It is concluded that TOAD-64, typically regarded as a marker for commitment to neuronal differentiation during fetal development, also plays a role in adult rat forebrain and striatum and that its concentration is possibly modulated by haloperidol treatment.     

Identification of candidate antigens for serologic detection of Helicobacter pylori-infected patients with gastric carcinoma

Helicobacter pylori colonizes the stomach of almost half the world population and is a causative agent of gastric carcinomas and duodenal ulcers. Only a small fraction of infected people will develop these severe illnesses and a predictive test to identify people at high risk would greatly benefit disease management. Our study aimed to identify conserved bacterial antigens that may be useful for the development of such a diagnostic test. High-resolution immunoproteomics by 2-dimensional electrophoresis of H. pylori 26695 proteins was carried out with sera from infected patients with either duodenal ulcer (n=30) or gastric carcinoma (n=30), 2 clinically divergent conditions. According to their antigen recognition patterns clear groups of patients were identified. Although this classification did not correspond to the clinical status, it may be correlated to other bacterial or host factors that influence the outcome of infection. In general antigen recognition patterns were found to be highly variable, however by utilizing powerful image analysis and statistical tests the recognition of 14 antigenic protein species was found to differ significantly (p<0.01) between both diseases. Particular protein species of GroEL, HyuA, GroES and AtpA appear to be useful surrogate markers for gastric carcinoma detection and consequently should be considered for further prospective studies to assess their predictive value. For one protein species of AtpA, evidence was found that different post-translational modifications may confer different immunogenicities.     

Stimulation of plasminogen activation by recombinant cellular prion protein is conserved in the NH2-terminal fragment PrP23-110

The cellular prion protein (PrP(c)), tissue-type plasminogen activator (t-PA) and plasminogen are expressed in synaptic membranes in vivo. In the central nervous system the fibrinolytic system is associated with excitotoxin-mediated neurotoxicity and Alzheimer's disease. Recently binding of the disease associated isoform of the prion protein (PrP(Sc)) to plasminogen and stimulation of t-PA activity have been reported. In this study the interaction of PrP(c) and plasminogen was investigated using chromogenic assays in vitro. We found that plasmin is able to cleave recombinant PrP(c) at lysine residue 110 generating an NH(2)-terminal truncated molecule that has previously been described as a major product of PrP(c) metabolism. We further characterized the proteolytic fragments with respect to their ability to stimulate plasminogen activation in vitro. Our results show that the NH(2)-terminal part of PrP(c) spanning amino acids 23-110 (PrP23-110) together with low molecular weight heparin stimulates t-PA mediated plasminogen activation in vitro. The apparent rate constant was increased 57 fold in the presence of 800 nM PrP23-110. Furthermore, we compared the stimulation of t-PA activity by PrP(c) and beta-amyloid peptide (1-42). While the activity of the beta-amyloid was independent of low molecular weight heparin, PrP23-110 was approximately 4- and 37 fold more active than beta-amyloid in the absence or presence of low molecular weight heparin. In summary, plasmin cleaves PrP(c) in vitro and the liberated NH(2)-terminal fragment accelerates plasminogen activation. Cleavage of PrP c has previously been reported. Thus cleavage of PrP(c) enhancing plasminogen activation at the cell surface could constitute a regulatory mechanism of pericellular proteolysis.     

Novel orthobunyavirus in Cattle, Europe, 2011

In 2011, an unidentified disease in cattle was reported in Germany and the Netherlands. Clinical signs included fever, decreased milk production, and diarrhea. Metagenomic analysis identified a novel orthobunyavirus, which subsequently was isolated from blood of affected animals. Surveillance was initiated to test malformed newborn animals in the affected region.     

Disposition of desloratadine in healthy volunteers

The absorption, metabolism and excretion of desloratadine (DL, Clarinex) were characterized in six healthy male volunteers. Subjects received a single oral 10-mg dose of [(14)C]DL ( approximately 104 microCi). Blood, urine and feces were collected over 240 h. DL was well absorbed; drug-derived radioactivity was excreted in both urine (41%) and feces (47%). With the exception of a single subject, DL was extensively metabolized; the major biotransformation pathway consisted of hydroxylation at the 3 position of the pyridine ring and subsequent glucuronidation (3-OH-DL-glucuronide or M13). In five of the six subjects, DL was slowly eliminated (mean t((1/2)) = 19.5 h) and persisted in the plasma for 48-120 h post-dose. This is in contrast to a t((1/2)) of approximately 110 h and quantifiable plasma DL concentrations for the entire 240-h sampling period in one subject, who was identified phenotypically as a poor metabolizer of DL. This subject also exhibited correspondingly lower amounts of M13 in urine and 3-OH-DL (M40) in feces. Disposition of DL in this subject was characterized by slow absorption, slow metabolism and prolonged elimination. Further clinical studies confirmed the lack of safety issues associated with polymorphism of DL metabolism (Prenner et al. 2006, Expert Opinion on Drug Safety, 5: 211-223).