Effect of modulation of serotonergic, cholinergic, and nitrergic pathways on murine fundic size and compliance measured by ultrasonomicrometry

Reduced fasting or postprandial gastric volumes have been implicated in the pathophysiology of functional dyspepsia. The mechanisms that underlie the control of gastric fundic volume are incompletely understood, partly because of an inability to accurately measure fundic volume in vivo in small animals. Small animals are useful models to evaluate mechanisms, e.g., in knockout animals. The aim of this study was to determine whether an ultrasonometric technique accurately monitors fundic contraction and relaxation in mice in vivo and to determine the effect of modulation of cholinergic, nitrergic, and serotonergic pathways on fundic size and compliance in the intact mouse innervated stomach. Two to four piezoelectric crystals (diameter 1 mm, 24-microm resolution) were glued to the serosal side of fundus and used to measure distance. Validation studies showed excellent correlation between measured changes and actual changes in distances between crystals and excellent reproducibility. The expected responses to pharmacological modulation with bethanechol and nitroglycerin were demonstrated. Atropine increased the distance between the crystals, suggesting a baseline cholinergic regulation of fundic volume. Bethanechol, Nomega-nitro-L-arginine, and the 5-HT1B/D agonist sumatriptan decreased the distance between the crystals, suggesting fundic contraction. Atropine, nitroglycerin, and buspirone caused an increase in intercrystal distance consistent with fundic relaxation. Fundic compliance was investigated by changing intragastric pressure via an implanted catheter. Sumatriptan increased compliance, whereas buspirone increased the distance between crystals but did not change compliance. The data suggest that ultrasonomicrometry is a useful tool that can reproducibly and accurately measure changes in fundic size and the response to pharmacological agents.     

The Complete Genome and Phenome of a Community-Acquired Acinetobacter baumannii

Many sequenced strains of Acinetobacter baumannii are established nosocomial pathogens capable of resistance to multiple antimicrobials. Community-acquired A. baumannii in contrast, comprise a minor proportion of all A. baumannii infections and are highly susceptible to antimicrobial treatment. However, these infections also present acute clinical manifestations associated with high reported rates of mortality. We report the complete 3.70 Mbp genome of A. baumannii D1279779, previously isolated from the bacteraemic infection of an Indigenous Australian; this strain represents the first community-acquired A. baumannii to be sequenced. Comparative analysis of currently published A. baumannii genomes identified twenty-four accessory gene clusters present in D1279779. These accessory elements were predicted to encode a range of functions including polysaccharide biosynthesis, type I DNA restriction-modification, and the metabolism of novel carbonaceous and nitrogenous compounds. Conversely, twenty genomic regions present in previously sequenced A. baumannii strains were absent in D1279779, including gene clusters involved in the catabolism of 4-hydroxybenzoate and glucarate, and the A. baumannii antibiotic resistance island, known to bestow resistance to multiple antimicrobials in nosocomial strains. Phenomic analysis utilising the Biolog Phenotype Microarray system indicated that A. baumannii D1279779 can utilise a broader range of carbon and nitrogen sources than international clone I and clone II nosocomial isolates. However, D1279779 was more sensitive to antimicrobial compounds, particularly beta-lactams, tetracyclines and sulphonamides. The combined genomic and phenomic analyses have provided insight into the features distinguishing A. baumannii isolated from community-acquired and nosocomial infections.     

Neurons are MHC class I-dependent targets for CD8 T cells upon neurotropic viral infection

Following infection of the central nervous system (CNS), the immune system is faced with the challenge of eliminating the pathogen without causing significant damage to neurons, which have limited capacities of renewal. In particular, it was thought that neurons were protected from direct attack by cytotoxic T lymphocytes (CTL) because they do not express major histocompatibility class I (MHC I) molecules, at least at steady state. To date, most of our current knowledge on the specifics of neuron-CTL interaction is based on studies artificially inducing MHC I expression on neurons, loading them with exogenous peptide and applying CTL clones or lines often differentiated in culture. Thus, much remains to be uncovered regarding the modalities of the interaction between infected neurons and antiviral CD8 T cells in the course of a natural disease. Here, we used the model of neuroinflammation caused by neurotropic Borna disease virus (BDV), in which virus-specific CTL have been demonstrated as the main immune effectors triggering disease. We tested the pathogenic properties of brain-isolated CD8 T cells against pure neuronal cultures infected with BDV. We observed that BDV infection of cortical neurons triggered a significant up regulation of MHC I molecules, rendering them susceptible to recognition by antiviral CTL, freshly isolated from the brains of acutely infected rats. Using real-time imaging, we analyzed the spatio-temporal relationships between neurons and CTL. Brain-isolated CTL exhibited a reduced mobility and established stable contacts with BDV-infected neurons, in an antigen- and MHC-dependent manner. This interaction induced rapid morphological changes of the neurons, without immediate killing or impairment of electrical activity. Early signs of neuronal apoptosis were detected only hours after this initial contact. Thus, our results show that infected neurons can be recognized efficiently by brain-isolated antiviral CD8 T cells and uncover the unusual modalities of CTL-induced neuronal damage.     

Stem cell and cellular therapy developments

Recent discoveries on human and non-human stem cells have prompted the development of several studies aimed at the translation of laboratory evidences into novel clinical procedures collectively known as ‘cellular therapy’.In this regard, a number of features specifically related to the clinical setting require stringent evaluation, including, but not limited to: ethical appropriateness; donor and recipient informed consent; autologous versus allogeneic use; media and devices for cell collection, processing, characterization, storage and distribution; donor and recipient adverse events registration and management; risk-to-benefit and cost analysis; outcome analysis; production sites accreditation and management; regulatory oversight.This article describes recent national and international developments related to the distribution of cells and tissues for clinical use. Moreover, an example is reported of the implementation of a cellular therapy production site compliant with good manufacturing practices (GMPs) in a large European University hospital.     

Four tetranortriterpenoids from the stem bark of Khaya anthotheca

Eight limonoids, anthothecanolide (1), 3-O-acetylanthothecanolide (2), 2,3-di-O-acetylanthothecanolide (3), 6R,8alpha-dihydroxycarapin (4), 3beta-acetoxy-3-deoxo-6R-hydroxycarapin (5), methyl angolensate, methyl 6-hydroxyangolensate and khayalactone together with sitosterol glucoside, have been isolated from the extracts of the stem bark of Khaya anthotheca. Compounds 1-4 are described for the first time. Their structures were established by analysis of the high-field NMR and MS data. The structure of compound 4 was confirmed by a single crystal X-ray structure analysis.     

Syntrophin gamma 2 regulates SCN5A gating by a PDZ domain-mediated interaction

SCN5A encodes the alpha subunit of the cardiac muscle and intestinal smooth muscle mechanosensitive Na(+) channel. Mechanosensitivity in the intestine requires an intact cytoskeleton. We report, using laser capture microdissection, single cell PCR, and immunohistochemistry, that syntrophins, scaffolding proteins, were expressed in human intestinal smooth muscle cells. The distribution of syntrophin gamma 2 was similar to that of SCN5A. Yeast two-hybrid and glutathione S-transferase pull-down experiments show that SCN5A and syntrophin gamma 2 co-express and that the PDZ domain of syntrophin gamma 2 directly interacts with the C terminus of SCN5A. In native cells, disruption of the C terminus-syntrophin gamma 2 PDZ domain interaction using peptides directed against either region result in loss of mechanosensitivity. Co-transfection of syntrophin gamma 2 with SCN5A in HEK293 cells markedly shifts the activation kinetics of SCN5A and reduces the availability of Na(+) current. We propose that syntrophin gamma 2 is an essential Na(+) channel-interacting protein required for the full expression of the Na(+) current and that the SCN5A-syntrophin gamma 2 interaction determines mechanosensitivity and current availability.