Changes of intra-mitochondrial Ca2+ in adult ventricular cardiomyocytes examined using a novel fluorescent Ca2+ indicator targeted to mitochondria.

In this study a Ca(2+) sensitive protein was targeted to the mitochondria of adult rabbit ventricular cardiomyocytes using an adenovirus transfection technique. The probe (Mitycam) was a Ca(2+)-sensitive inverse pericam fused to subunit VIII of human cytochrome c oxidase. Mitycam expression pattern and Ca(2+) sensitivity was characterized in HeLa cells and isolated adult rabbit cardiomyocytes.

Identification of immunogenic proteins associated with protection against haemorrhagic septicaemia after vaccination of calves with a live-attenuated aroA derivative of Pasteurella multocida B:2.

Pasteurella multocida serotype B:2 is the causative agent of haemorrhagic septicaemia (HS), a fatal disease of cattle and buffaloes. As a step towards the identification of individual antigens that may protect against HS, proteins present in a sonicated cell extract (SCE) and outer-membrane protein (OMP) preparation of a wild-type P. multocida serotype B:2 were investigated by immunoblotting with sera from calves which had been protected against challenge with a virulent strain of P.

Motivated proteins: a web application for studying small three-dimensional protein motifs.

Background: Small loop-shaped motifs are common constituents of the three-dimensional structure of proteins. Typically they comprise between three and seven amino acid residues, and are defined by a combination of dihedral angles and hydrogen bonding partners. The most abundant of these are alphabeta-motifs, asx-motifs, asx-turns, beta-bulges, beta-bulge loops, beta-turns, nests, niches, Schellmann loops, ST-motifs, ST-staples and ST-turns.

Absorption, metabolism, and excretion of cider dihydrochalcones in healthy humans and subjects with an ileostomy.

The phloretin-O-glycosides, phloretin-2'-O-glucoside and phloretin-2'-O-(2''-O-xylosyl)glucoside, are thought to be unique to apples and apple products. To investigate the metabolism and bioavailability of these compounds, nine healthy and five ileostomy human subjects consumed 500 mL of Thatchers Redstreak apple cider containing 46 micromol of phloretin-O-glycosides. Over the ensuing 24 h period, plasma, urine, and ileal fluid were collected prior to analysis by high-performance liquid chromatography-mass spectrometry (HPLC-MS).

Regulation by c-Myc of ncRNA expression.

Deregulated activity of the proto-oncogene product c-Myc is instrumental in promoting many human cancers. As it is a transcription factor, priority has been given to identifying the genes that it regulates. Until recently, all the attention was focused on protein-encoding genes.

Penetration of antifungal agents through Candida biofilms.

A filter disk assay is described, which measures the penetration of antifungal agents through Candida biofilms. The technique involves forming a colony biofilm on a polycarbonate membrane filter, and capping it with a second, smaller membrane filter followed by a wetted paper disk of the type used in zone-of-inhibition assays. The entire assembly is transferred to agar medium containing the antifungal agent of interest.

Negative regulation of syntaxin4/SNAP-23/VAMP2-mediated membrane fusion by Munc18c in vitro.

Background: Translocation of the facilitative glucose transporter GLUT4 from an intracellular store to the plasma membrane is responsible for the increased rate of glucose transport into fat and muscle cells in response to insulin. This represents a specialised form of regulated membrane trafficking. Intracellular membrane traffic is subject to multiple levels of regulation by conserved families of proteins in all eukaryotic cells.

Differential in-gel electrophoresis (DIGE) analysis of human bone marrow osteoprogenitor cell contact guidance.

We have used a recent comparative proteomics technique, differential in-gel electrophoresis (DIGE), to study osteoprogenitor cell response to contact guidance in grooves. In order to increase protein output from small sample sizes, we used bioreactor culture before protein extraction and gel electrophoresis. Mass spectroscopy was used for protein identification.

Proteomic analysis of human osteoprogenitor response to disordered nanotopography.

Previous studies have shown that microgroove-initiated contact guidance can induce bone formation in osteoprogenitor cells (OPGs) and produce changes in the cell proteome. For proteomic analysis, differential in-gel electrophoresis (DIGE) can be used as a powerful diagnostic method to provide comparable data between the proteomic profiles of cells cultured in different conditions. This study focuses on the response of OPGs to a novel nanoscale pit topography with osteoinductive properties compared with planar controls.

Excitatory and inhibitory intermediate zone interneurons in pathways from feline group I and II afferents: differences in axonal projections and input.

The aim of the present study was to compare properties of excitatory and inhibitory spinal intermediate zone interneurons in pathways from group I and II muscle afferents in the cat. Interneurons were labelled intracellularly and their transmitter phenotypes were defined by using immunocytochemistry. In total 14 glutamatergic, 22 glycinergic and 2 GABAergic/glycinergic interneurons were retrieved.

A novel main chain motif in proteins bridged by cationic groups: the niche.

We have surveyed the bridging of pairs of main chain carbonyl oxygens by cations or by delta(+) hydrogens within hydrogen bonding groups. A three to four residue motif, which we call the niche, with characteristic phi,psi angles, is by far the commonest feature with this property. The niche accommodates atoms or groups that offer delta(+) charges, including water molecules or metal ions, as well as amines, guanidines, and other NH(2) groups.

Kynurenine metabolites and inflammation markers in depressed patients treated with fluoxetine or counselling.

1. Depression could result from changes in tryptophan availability caused by activation of the kynurenine pathway as a result of inflammation. In the present study, we examined patients newly diagnosed with depression to determine whether kynurenines and related factors change in parallel with improvements in mood.

Functional and structural studies on different forms of the adenylate cyclase toxin of Bordetella pertussis.

A comparison was made of the cytotoxic activity and secondary structural features of four recombinant forms of adenylate cyclase toxin (CyaA). These forms were fully functional CyaA, CyaA lacking adenylate cyclase enzymatic activity (CyaA*), and non-acylated forms of these toxins, proCyaA and proCyaA*. At a toxin concentration>1 microg/ml, CyaA* was as cytotoxic towards J774.

Fluid and electrolyte balance in elite gaelic football players.

The aim of this study was to investigate fluid and electrolyte balance in elite Gaelic Football players (n=20) during a typical training session in a warm environment (16 to 18 degrees C, 82-88% humidity). Pre-training urine samples were used to determine hydration status. Sweat sodium concentration was collected from four body site locations using absorbent patches.

Interactions with nanoscale topography: adhesion quantification and signal transduction in cells of osteogenic and multipotent lineage.

Polymeric medical devices widely used in orthopedic surgery play key roles in fracture fixation and orthopedic implant design. Topographical modification and surface micro-roughness of these devices regulate cellular adhesion, a process fundamental in the initiation of osteoinduction and osteogenesis. Advances in fabrication techniques have evolved the field of surface modification; in particular, nanotechnology has allowed the development of nanoscale substrates for the investigation into cell-nanofeature interactions.

Codon and mRNA sequence optimization of microdystrophin transgenes improves expression and physiological outcome in dystrophic mdx mice following AAV2/8 gene transfer.

Duchenne muscular dystrophy is a fatal muscle-wasting disorder. Lack of dystrophin compromises the integrity of the sarcolemma and results in myofibers that are highly prone to contraction-induced injury. Recombinant adeno-associated virus (rAAV)-mediated dystrophin gene transfer strategies to muscle for the treatment of Duchenne muscular dystrophy (DMD) have been limited by the small cloning capacity of rAAV vectors and high titers necessary to achieve efficient systemic gene transfer.

The effects of short-term antioxidant supplementation on oxidative stress and flight performance in adult budgerigars Melopsittacus undulatus.

Antioxidants are known to play an important role in quenching reactive oxygen species (ROS), thus ameliorating oxidative stress. Since increased metabolism associated with exercise can increase oxidative stress, dietary antioxidants may be a limiting factor in determining aspects of physical performance. Here we tested whether oxidative stress associated with flight exercise of captive adult budgerigars, Melopsittacus undulatus differed after they received a diet containing either enhanced (EQ) or reduced levels (RQ) of a nutritional supplement (Nutrivit) rich in antioxidants for 4 weeks.

Propidium iodide-based methods for monitoring drug action in the kinetoplastidae: comparison with the Alamar Blue assay.

The urgent need for new drug development for African trypanosomiasis is widely recognized. This requires reliable and informative high-throughput assays. Currently, drug action is determined with a fluorimetric/colorimetric assay based on the metabolism of the dye Alamar Blue (resazurin) by live cells.

Time-dependent evolution of tissue markers by MALDI-MS imaging.

We have used MALDI-MS imaging (MALDI-MSI) to monitor the time dependent appearance and loss of signals when tissue slices are brought rapidly to room temperature for short to medium periods of time. Sections from mouse brain were cut in a cryostat microtome, placed on a MALDI target and allowed to warm to room temperature for 30 s to 3 h. Sections were then refrozen, fixed by ethanol treatment and analysed by MALDI-MSI.

Delayed dispersal as a route to breeding: territorial inheritance, safe havens, and ecological constraints.

The relative roles of ecological constraints, the benefits of philopatry, and the role of life history continue to be debated in the evolution of natal philopatry and cooperative breeding. We compare three routes to breeding: departing to search for territories as a floater, staying and queuing to inherit the natal territory, or queuing and eventually shifting to a neighboring vacancy. Our model assumed a dominance-structured population.

Are unusually colored eggs a signal to potential conspecific brood parasites?

It has previously been suggested that some species of birds make the last egg in their clutch pale as a signal to potential conspecific brood parasites that incubation has commenced. Here, we use game theory to show that the signaling function of pale eggs can be evolutionarily stable and resistant to cheating and to demonstrate that such a signal can only be maintained under strict conditions. The key conditions are, first, that there is a cost associated with the production of pale eggs (in particular, the cost of a pale egg produced early in the clutch must be more expensive than the cost of one produced later in the clutch) and, second, that the cost of making the last egg pale is not too great (relative to the costs of parasitism).

Effect of transmission setting and mixed species infections on clinical measures of malaria in Malawi.

Background: In malaria endemic regions people are commonly infected with multiple species of malaria parasites but the clinical impact of these Plasmodium co-infections is unclear. Differences in transmission seasonality and transmission intensity between endemic regions have been suggested as important factors in determining the effect of multiple species co-infections.

Principal Findings: In order to investigate the impact of multiple-species infections on clinical measures of malaria we carried out a cross-sectional community survey in Malawi, in 2002.

Effects of preservation methods on stable isotope signatures in bird tissues.

Increasing use is being made of stable isotopes as indicators of habitat use and trophic ecology of animals. Preservation of tissues can alter stable isotope signatures. We investigated the effects of addition of ethanol and NaCl solution (hereafter 'salt'), and of freezing and drying, on carbon and nitrogen isotopic values in blood of the spectacled petrel Procellaria conspicillata, and compared these with those from simultaneously growing feathers.

Bioavailability of polyphenon E flavan-3-ols in humans with an ileostomy.

To investigate the degree of absorption of flavan-3-ols in the small intestine, human subjects with an ileostomy ingested 200 mg of Polyphenon E, a green tea extract, after which ileal fluid and urine, collected over a 24-h period, were analyzed by high-performance liquid chromatography with photodiode array and mass spectrometric detection. The data obtained indicated that although approximately 40% of flavan-3-ol intake is recovered in ileal fluid, substantial quantities are absorbed in the small intestine. Moreover, 14 urinary metabolites, comprising sulfates, glucuronide, and methylated derivatives, were identified and quantified.

Focal adhesion interactions with topographical structures: a novel method for immuno-SEM labelling of focal adhesions in S-phase cells.

Current understanding of the mechanisms involved in osseointegration following implantation of a biomaterial has led to adhesion quantification being implemented as an assay of cytocompatibility. Such measurement can be hindered by intra-sample variation owing to morphological changes associated with the cell cycle. Here we report on a new scanning electron microscopical method for the simultaneous immunogold labelling of cellular focal adhesions and S-phase nuclei identified by BrdU incorporation.