Adeno-associated virus type 2-mediated gene transfer: role of epidermal growth factor receptor protein tyrosine kinase in transgene expression

Adeno-associated virus type 2 (AAV), a single-stranded, DNA-containing, nonpathogenic human parvovirus, has gained attention as a potentially useful vector for human gene therapy. However, the transduction efficiency of AAV vectors varies greatly in different cells and tissues in vitro and in vivo. We have recently documented that a cellular tyrosine phosphoprotein, designated the single-stranded D-sequence-binding protein (ssD-BP), plays an important role in AAV-mediated transgene expression (K. Y. Qing et al., Proc. Natl. Acad. Sci. USA 94:10879-10884, 1997) and that a strong correlation exists between the phosphorylation state of the ssD-BP and AAV transduction efficiency in vitro as well as in vivo (K. Y. Qing et al., J. Virol. 72:1593-1599, 1998). In this report, we document that treatment of cells with specific inhibitors of the epidermal growth factor receptor protein tyrosine kinase (EGF-R PTK) activity, such as tyrphostin, leads to significant augmentation of AAV transduction efficiency, and phosphorylation of the ssD-BP is mediated by the EGF-R PTK. Treatment of cells with EGF results in phosphorylation of the ssD-BP, whereas treatment with tyrphostin causes dephosphorylation of the ssD-BP and consequently leads to increased expression of the transgene. Furthermore, AAV transduction efficiency inversely correlates with expression of the EGF-R in different cell types, and stable transfection of the EGF-R cDNA causes phosphorylation of the ssD-BP, leading to significant inhibition in AAV-mediated transgene expression which can be overcome by the tyrphostin treatment. These data suggest that the PTK activity of the EGF-R is a crucial determinant in the life cycle of AAV and that further studies on the interaction between the EGF-R and the ssD-BP may yield new insights not only into its role in the host cell but also in the successful use of AAV vectors in human gene therapy.     

Simultaneous and rapid analysis of chemical preservatives in processed animal products by ultra-performance liquid chromatography

Food Science and Biotechnology - An ultra-performance liquid chromatography-tunable ultraviolet method was optimized and validated for the simultaneous analysis of nine chemical preservatives in...     

Low temperature produced calcium-doped zinc oxide thick film via screen printing technique as thermal interface material in LED application

Pure and calcium-doped zinc oxide thick films were deposited on Aluminium substrate by screen printing technique using nanocrystalline powder synthesized from co-precipitation method. Shear thinning phenomenon with increment of shear rate was observed during the rheological analysis for all pastes. X-ray diffraction results confirmed the formation of ZnO with preferred orientation along (101) plane. Peak shifting to lower angle was observed upon increment of doping concentration of calcium. Crystallite size of doped ZnO powder decreased in the range (from 35.4 to 42.4 nm) from 116.1 nm of pure ZnO. Surface morphology analysed by FESEM had revealed the reduction of voids with increasing doping concentration up to 7 wt% of doping, followed by a slightly increase in the number of voids at 9 wt% doping. AFM analysis showed that the surface roughness of films exhibited a decreasing trend with the increase of calcium dopant until 7 wt% but became rougher at 9 wt%. Peaks shifting of ZnO to lower wavenumber revealed by FTIR study indicated that doping had affected the lattice structure of ZnO in the films. Thermal characterization showed the introduction of calcium dopant had increased the thermal resistance of the thick films. This led to a better junction temperature (T_j) of LED of 46.4?°C when compared with T_j of pure ZnO film at 47.3?°C.     

Determination of process parameters in stereolithography using neural network

For stereolithography process, accuracy of prototypes is related to laser power, scan speed, scan width, scan pattern, layer thickness, resin characteristics and etc. An accurate prototype is obtained by using appropriate process parameters. In order to determine these parameters, the stereolithography (SLA) machine using neural network was developed and efficiency of the developed SLA machine was compared with that of the traditional SLA. Optimum values for scan speed, hatching spacing and layer thickness improved the surface roughness and build time for the developed SLA.     

A study on the photo-reductive precipitation rate of europium sulfate from rare earth mixture by addition of hydrogen peroxide

The influence of H₂O₂ on the photo-reductive precipitation of Eu³⁺ from a solution containing Sm³⁺, Eu³⁺ and Gd³⁺ was investigated. The solution contains isopropanol as a reduction agent, ammonium sulfate as a precipitation agent, and hydrogen peroxide as an acceleration agent of precipitation rate. A mercury lamp emitting a wavelength of 254 nm was used as a light source. Adding a small amount of H₂O₂ considerably increased the photo-reductive precipitation rate of Eu³⁺. OH radicals obtained from the photodecomposition of H₂O₂ oxidized isopropanol into the radical (CH₃)₂C’OH and the resulting radicals reduced Eu³⁺ rapidly. It was found that the organic radical has the ability to reduce only Eu³⁺ in the solution containing Sm³⁺, Eu³⁺ and Gd³⁺. The precipitation yields of Eu, Sm and Gd were about 99%, 10% and 4%, respectively. It was also found that the Sm and Gd were not photo-reduced and co-precipitated with Eu.     

Ultrasound-assisted extraction of natural antioxidants from betel leaves (Piper betle): Extraction kinetics and modeling

Present study investigated the extraction kinetics of antioxidants from Piper betle by ultrasound-assisted extraction for three extraction variables: temperature, solute to solvent ratio and ethanol concentration. Based on the results of yield, total phenolic content and antioxidant activity, the optimum results were obtained at 50°C, 1:20 g/mL and 80% ethanol, respectively. Statistical coefficients of R² ≥ 0.961 and RMSE ≤ 0.508 for two-site kinetic model confirm the use of proposed models for simulation and prediction purpose. Comparison with ascorbic acid and butylated hydroxytoluene solidifies the use of Piper betle as feasible source of natural antioxidants. The presence of hydroxychavicol and eugenol was affirmed by high-performance liquid chromatography assay.     

Structural States of Hdm2 and HdmX: X-ray Elucidation of Adaptations and Binding Interactions for Different Chemical Compound Classes

Hdm2 (human MDM2, human double minute 2 homologue) counteracts p53 function by direct binding to p53 and by ubiquitin-dependent p53 protein degradation. Activation of p53 by inhibitors of the p53–Hdm2 interaction is being pursued as a therapeutic strategy in p53 wild-type cancers. In addition, HdmX (human MDMX, human MDM4) was also identified as an important therapeutic target to efficiently reactivate p53, and it is likely that dual inhibition of Hdm2 and HdmX is beneficial. Herein we report four new X-ray structures for Hdm2 and five new X-ray structures for HdmX complexes, involving different classes of synthetic compounds (including the worldwide highest resolutions for Hdm2 and HdmX, at 1.13 and 1.20 Å, respectively). We also reveal the key additive 18-crown-ether, which we discovered to enable HdmX crystallization and show its stabilization of various Lys residues. In addition, we report the previously unpublished details of X-ray structure determinations for eight further Hdm2 complexes, including the clinical trial compounds NVP-CGM097 and NVP-HDM201. An analysis of all compound binding modes reveals new and deepened insight into the possible adaptations and structural states of Hdm2 (e.g., flip of F55, flip of Y67, reorientation of H96) and HdmX (e.g., flip of H55, dimer induction), enabling key binding interactions for different compound classes. To facilitate comparisons, we used the same numbering for Hdm2 (as in Q00987) and HdmX (as in O15151, but minus 1). Taken together, these structural insights should prove useful for the design and optimization of further selective and/or dual Hdm2/HdmX inhibitors.     

Eutectic Composition in the Pseudobinary of Y4Al2O9 and Y2O3

The eutectic composition between Y₄Al₂O₉ and Y₂O₃ was determined using electron probe microanalysis (EPMA) on directionally solidified specimens with hypo- and hypereutectic compositions. The microstructures of the specimens as a function of composition differ considerably with small deviation from the eutectic composition (70.5 mol% Y₂O₃ and 29.5 mol% Al₂O₃). Based on the current results and other published data, the pseudobinary system between Al₂O₃ and Y₂O₃ is revised.