Metabonomic investigation of human Schistosoma mansoni infection

Schistosomiasis is a parasitic infection that is endemic in many developing countries in the tropics and subtropics afflicting more than 207 million people primarily in rural areas. After malaria, it is the second most important parasitic infection in terms of socio-economic and public health. Investigation of the host-parasite interaction at the molecular level and identification of biomarkers of infection and infection-related morbidity would be of value for improved strategies for treatment and morbidity control. To this end, we conducted a nuclear magnetic resonance (NMR) based metabonomics study involving a well-characterized cohort of 447 individuals from a rural area in Uganda near Lake Victoria with a high prevalence of Schistosoma mansoni, a species predominantly occurring in Africa including Madagascar and parts of South America. Cohort samples were collected from individuals at five time-points, before and after (one or two times) chemotherapy with praziquantel (PZQ). Using supervised multivariate statistical analysis of the recorded one-dimensional (1D) NMR spectra, we were able to discriminate infected from uninfected individuals in two age groups (children and adults) based on differences in their urinary profiles. The potential molecular markers of S. mansoni infection were found to be primarily linked to changes in gut microflora, energy metabolism and liver function. These findings are in agreement with data from earlier studies on S. mansoni infection in experimental animals and thus provide corroborating evidence for the existence of metabolic response specific for this infection.     

Salivary PYY: a putative bypass to satiety

Peptide YY(3-36) is a satiation hormone released postprandially into the bloodstream from L-endocrine cells in the gut epithelia. In the current report, we demonstrate PYY(3-36) is also present in murine as well as in human saliva. In mice, salivary PYY(3-36) derives from plasma and is also synthesized in the taste cells in taste buds of the tongue. Moreover, the cognate receptor Y2R is abundantly expressed in the basal layer of the progenitor cells of the tongue epithelia and von Ebner's gland. The acute augmentation of salivary PYY(3-36) induced stronger satiation as demonstrated in feeding behavioral studies. The effect is mediated through the activation of the specific Y2 receptor expressed in the lingual epithelial cells. In a long-term study involving diet-induced obese (DIO) mice, a sustained increase in PYY(3-36) was achieved using viral vector-mediated gene delivery targeting salivary glands. The chronic increase in salivary PYY(3-36) resulted in a significant long-term reduction in food intake (FI) and body weight (BW). Thus this study provides evidence for new functions of the previously characterized gut peptide PYY(3-36) suggesting a potential simple and efficient alternative therapeutic approach for the treatment of obesity.     

New approach to isolate mesenchymal stem cell (MSC) from human umbilical cord blood

HUCB (human umbilical cord blood) has been frequently used in clinical allogeneic HSC (haemopoietic stem cell) transplant. However, HUCB is poorly recognized as a rich source of MSC (mesenchymal stem cell). The aim of this study has been to establish a new method for isolating large number of MSC from HUCB to recognize it as a good source of MSC. HUCB samples were collected from women following their elective caesarean section. The new method (Clot Spot method) was carried out by explanting HUCB samples in mesencult complete medium and maintained in 37°C, in a 5% CO2 and air incubator. MSC presence was established by quantitative and qualitative immunophenotyping of cells and using FITC attached to MSC phenotypic markers (CD29, CD73, CD44 and CD105). Haematopoietic antibodies (CD34 and CD45) were used as negative control. MSC differentiation was examined in neurogenic and adipogenic media. Immunocytochemistry was carried out for the embryonic markers: SOX2 (sex determining region Y-box 2), OLIG-4 (oligodendrocyte-4) and FABP-4 (fatty acid binding protein-4). The new method was compared with the conventional Rosset Sep method. MSC cultures using the Clot Spot method showed 3-fold increase in proliferation rate compared with conventional method. Also, the cells showed high expression of MSC markers CD29, CD73, CD44 and CD105, but lacked the expression of specific HSC markers (CD34 and CD45). The isolated MSC showed some differentiation by expressing the neurogenic (SOX2 and Olig4) and adipogenic (FABP-4) markers respectively. In conclusion, HUCB is a good source of MSC using this new technique.     

Electrical detection of human immunoglobulins G from human serum using a microbiosensor

A biosensor for the electrical detection of human antibodies from serum has been fabricated and experimentally demonstrated. The device is based on the immobilization of proteins used as probes between a set of microelectrodes. Incubation with diluted human serum was followed by incubation with anti-human secondary antibodies labeled with gold nanoparticles (GNPs) and then precipitation of silver on the nanoparticles. The output of the device was defined as the percentage of short-circuited microelectrodes after silver deposition independently of the gap conductance. Two model probes were studied: protein A and goat antibodies. The effects of the microgap spacing (5, 10, 15 or 20 microm) and the duration of the silver treatment were examined. The data obtained showed that a large spacing (20 microm) led to poor sensitivity. Alternately, 5 microm gaps led to high sensitivity and saturation of the signal. Interestingly, 10-15 microm gaps enabled a non-saturated and distinct signal for both probes that was correlated with the GNP density between the microgaps as determined by atomic force microscopy. Different capture efficiencies could be easily distinguished. The biosensor described here is easy to use and thus can be applied to real detection experiments.     

2,3-Dihydroxy-quinoxaline induces ATPase activity of Herpes Simplex Virus thymidine kinase

2,3-Dihydroxy-quinoxaline, a small molecule that promotes ATPase catalytic activity of Herpes Simplex Virus thymidine kinase (HSV-TK), was identified by virtual screening. This compound competitively inhibited HSV-TK catalyzed phosphorylation of acyclovir with K_i = 250 μM (95% CI: 106–405 μM) and dose-dependently increased the rate of the ATP hydrolysis with K_M = 112 μM (95% CI: 28–195 μM). The kinetic scheme consistent with this experimental data is proposed.     

Homologous recombination and RecA protein: towards a new generation of tools for genome manipulations

Homologous recombination (HR) is one of the central processes of DNA metabolism, combining roles in both cell housekeeping and the evolution of genomes. In eukaryotes, HR underlies meiosis and ensures genome stability. The complete sequencing of numerous bacterial genomes has shown that HR has a substantial role in the evolution of microorganisms, especially pathogens. HR systems from different species and their isolated components are finding an expanding field of applications in modern genetic engineering and bio- and nanotechnologies. Recently, much progress has been made in our understanding of HR mechanisms in eukaryotes and the practical applications of HR systems.