Performance of the Interferon Gamma Release Assays in Tuberculosis Disease in Children Five Years Old or Less

Interferon Gamma Release Assays (IGRAs) were developed for the indirect or immunologic diagnosis of tuberculosis infection; however, they have also been used to assist in difficult to diagnose cases of tuberculosis disease in adults, and to a lesser extent, in children, especially in those under 5 years old. We evaluated the utility of using an IGRA in pediatric tuberculosis in younger children in a hospital setting. The diagnostic accuracy of T-SPOT.TB and TST was assessed in 117 children with active tuberculosis and 413 children with respiratory tract infection. Sensitivity and specificity were calculated for the tests used individually and together. Concordance was also calculated. Sensitivity of T-SPOT.TB (82.9%) was higher than TST (78.6% using a 5mm cut-off), especially in children confirmed to have TB. T-SPOT.TB was more specific than TST using a 5mm cut-off (96.1% vs. 70.9%). Combining T-SPOT.TB and TST results improved the sensitivity to 96.6%. In conclusion, the results of the current study indicate that T-SPOT.TB has good sensitivity and specificity, supporting its use among patients of this age. A combination of IGRA and TST would be useful additions to assist in the diagnosis of childhood TB.     

Aminopyrazine CB1 receptor inverse agonists

A series of 5,6-diaryl-2-amino-pyrazines were prepared and found to have antagonist-like properties at the CB1 receptor. Subsequent SAR studies optimized both receptor potency and drug-like properties including solubility and Cytochrome-P450 inhibition potential. Optimized compounds were demonstrated to be inverse agonists and compared in vivo with rimonabant for their ability to inhibit food intake, to occupy central CB1 receptors and to influence hormonal markers associated with obesity.     

Changes of Photosystem Ⅱ Electron Transport in the ChlorophyⅡ-deficient Oilseed Rape Mutant Studied by ChlorophyⅡ Fluorescence and Thermoluminescence

The photosystem Ⅱ （PSII） complex of photosynthetic membranes comprises a number of chlorophyll-binding proteins that are important to the electron flow. Here we report that the chlorophyll b-deficient mutant has decreased the amount of light-harvesting complexes with an increased amount of some core polypeptldes of PSII, including CP43 and CP47. By means of chlorophyll fluorescence and thermolumlnescence, we found that the ratio of Fv/Fm, qP and electron transport rate in the chlorophyll b-deficient mutant was higher compared to the wild type. In the chlorophyll lPdeflclent mutant, the decay of the primary electron acceptor quinones （QA-） reoxidation was decreased, measured by the fluorescence. Furthermore, the thermoluminescence studies in the chlorophyll bdeficient mutant showed that the B band （S2/S3QB-） decreased slightly and shifted up towards higher temperatures. In the presence of dlchlorophenyl-dlmethylurea, which is inhibited in the electron flow to the second electron acceptor quinines （QB） at the PSll acceptor side, the maximum of the Q band （S2QA-） was decreased slightly and shifted down to lower temperatures, compared to the wild type. Thus, the electron flow within PSll of the chlorophyⅡ b-deficient mutant was down-regulated and characterized by faster oxidation of the primary electron acceptor quinine QA-via forward electron flow and slower reduction of the oxidation S states.     

Construction of the industrial ethanol-producing strain of Saccharomyces cerevisiae able to ferment cellobiose and melibiose

The gene mel1, encoding alpha-galactosidase in Schizosaccharomyces pombe, and the gene bgl2, encoding and beta-glucosidase in Trichoderma reesei, were isolated and co-expressed in the industrial ethanol-producing strain of Saccharomyces cerevisiae. The resulting strains were able to grow on cellobiose and melibiose through simultaneous production of sufficient extracellular alpha-galactosidase and beta-glucosidase activity. Under aerobic conditions, the growth rate of the recombinant strain GC 1 co-expressing 2 genes could achieve 0.29 OD600 h(-1) and a biomass yield up to 7.8 g l(-1) dry cell weight on medium containing 10.0 g l(-1) cellobiose and 10.0 g l(-1) melibiose as sole carbohydrate source. Meanwhile, the new strain of S. cerevisiae CG 1 demonstrated the ability to directly produce ethanol from microcrystalline cellulose during simultaneous saccharification and fermentation process. Approximately 36.5 g l(-1) ethanol was produced from 100 g of cellulose supplied with 5 g l(-1) melibose within 60 h. The yield (g of ethanol produced/g of carbohydrate consumed) was 0.44 g/g, which corresponds to 88.0% of the theoretical yield.     

Layer-by-layer construction of graphene-based microbial fuel cell for improved power generation and methyl orange removal

Development of highly efficient anode is critical for enhancing the power output of microbial fuel cells (MFCs). The aim of this work is to investigate whether modification of carbon paper (CP) anode with graphene (GR) via layer-by-layer assembly technique is an effective approach to promote the electricity generation and methyl orange removal in MFCs. Using cyclic voltammetry and electrochemical impedance spectroscopy, the GR/CP electrode exhibited better electrochemical behavior. Scanning electron microscopy results revealed that the surface roughness of GR/CP increased, which was favorable for more bacteria to attach to the anode surface. The MFCs equipped with GR/CP anode achieved a stable maximum power density of 368 mW m^?2 under 1,000 Ω external resistance and a start time for the initial maximum voltage of 180 h, which were, respectively, 51 % higher and 31 % shorter than the corresponding values of the MFCs with blank anode. The anode and cathode polarization curves revealed negligible difference in cathode potentials but obviously difference in anode potentials, indicating that the GR-modified anode other than the cathode was responsible for the performance improvement of MFC. Meanwhile, compared with MFCs with blank anode, 11 % higher decolorization efficiency and 16 % higher the chemical oxygen demand removal rate were achieved in MFC with GR-modified anode during electricity generation. This study might provide an effective way to modify the anode for enhanced electricity generation and efficient removal of azo dye in MFCs.     

Small Activating RNA Restores the Activity of the Tumor Suppressor HIC-1 on Breast Cancer

HIC-1 is a gene that is hypermethylated in cancer, and commonly downregulated in human breast cancer. However, the precise mechanisms and molecular pathways regulated by HIC-1 remain unclear. We assessed HIC-1 expression on a tissue microarray containing 80 cases of breast cancer. We also analyzed its biological function by restoring HIC-1 expression using 5-aza-2′ deoxycytidine (5-CdR) and small-activating RNAs for the reversal of HIC-1 tumor suppressive effects on MCF-7 and MDA-MB-231 cell lines. An Agilent Q44h global expressing microarray was probed after restoring the expression of HIC-1. Data demonstrated that HIC-1 expression was reduced significantly in breast cancer tissues. HIC-1 immunohistochemistry resulted in mean staining scores in cancer tissue and normal ductal epithelia of 3.54 and 8.2, respectively (p<0.01). 5-CdR partially reversed HIC-1 expression, and modulated cell growth and apoptosis. dsHIC1-2998, an saRNA, showed activating efficacy in breast cancer cells. A group of differentially expressed genes were characterized by cDNA microarray. Upon saRNA treatment, genes upregulated included those involved in immune activation, cell cycle interference, the induction of apoptosis, anti-metastasis, and cell differentiation. Downregulated genes included oncogenes and those that play roles in cell invasion, cell growth, and cell division. Our findings may provide valuable resources not only for gene functional studies, but also for potential clinical applications to develop novel drug targets.     

FRET detection of calmodulin binding to the cardiac RyR2 calcium release channel

Calmodulin (CaM) binding to the type 2 ryanodine receptor (RyR2) regulates Ca release from the cardiac sarcoplasmic reticulum (SR). However, the structural basis of CaM regulation of the RyR2 is poorly defined, and the presence of other potential CaM binding partners in cardiac myocytes complicates resolution of CaM's regulatory interactions with RyR2. Here, we show that a fluorescence-resonance-energy-transfer (FRET)-based approach can effectively resolve RyR2 CaM binding, both in isolated SR membrane vesicles and in permeabilized ventricular myocytes. A small FRET donor was targeted to the RyR2 cytoplasmic assembly via fluorescent labeling of the FKBP12.6 subunit. Acceptor fluorophore was attached at discrete positions within either the N- or the C-lobe of CaM. FRET between FKBP12.6 and CaM bound to SR vesicles indicated CaM binding at a single high-affinity site within 60 Å of FKBP12.6. Micromolar Ca increased the apparent affinity of CaM binding and slowed CaM dissociation, but did not significantly affect maximal FRET efficiency at saturating CaM. FRET was strongest when the acceptor was attached at either of two positions within CaM's N-lobe versus sites in CaM's C-lobe, providing CaM orientation information. In permeabilized ventricular myocytes, FKBP12.6 and CaM colocalized to Z-lines, and the efficiency of energy transfer to both the N- and C-lobes of CaM was comparable to that observed in SR vesicle experiments. Results also indicate that both the location and orientation of CaM binding on the RyR2 are very similar to the skeletal muscle RyR1 isoform. Specific binding of CaM to functional RyR2 channels in the cardiac myocyte environment can be monitored using FKBP biosensors and FRET.