Identification of molecular target of AMP-activated protein kinase activator by affinity purification and mass spectrometry

We show an efficient method to identify molecular targets of small molecular compounds by affinity purification and mass spectrometry. Binding proteins were isolated from target cell lysate using affinity columns, which immobilized the active and inactive compounds. All proteins bound to these affinity columns were eluted by digestion using trypsin and then were identified by mass spectrometry. The specific binding proteins to the active compound, a candidate for molecular targets, were determined by subtracting the identified proteins in an inactive compound-immobilized affinity column from that in an active compound-immobilized affinity column. This method was applied to identification of molecular targets of D942, a furancarboxylic acid derivative, which increases glucose uptake in L6 myocytes through AMP-activated protein kinase (AMPK) activation. To elucidate the mechanism of AMPK activation by D942, affinity columns that immobilized D942 and its inactive derivative, D768, were prepared, and the binding proteins were purified from L6 cell lysate. NAD(P)H dehydrogenase [quinone] 1 (complex I), which was shown as one of the specific binding proteins to D942 by subtracting the binding proteins to D768, was partially inhibited by D942, not D768. Because inhibition of complex I activity led to a decrease in the ATP/AMP ratio, and the change in the ATP/AMP ratio triggered AMPK activation, we identified complex I as a potential protein target of AMPK activation by D942. This result shows our approach can provide crucial information about the molecular targets of small molecular compounds, especially target proteins not yet identified.     

C60 thin-film transistors with high field-effect mobility,fabricated by molecular beam deposition

We report performance of C₆₀ thin-film field-effect transistors and characterizations of C₆₀ thin-films on SiO₂ substrates fabricated by molecular beam deposition. Devices, fabricated and characterized under high vacuum without exposing to air, routinely showed current on/off ratios >10⁸ and field-effect mobility in the range of 0.5–0.3 cm²/V s. The obtained mobility is comparable to the highest value among n-type organic thin-film transistors and close to that derived from the photocurrent measurements on C₆₀ thin-films.The grain size of C₆₀ thin-film, condensed in an fcc solid, increases with the substrate temperature, while themobility did not exhibit a clear relation with substrate temperature.

© 2003 Elsevier Ltd. All rights reserved.     

Current Issues of Intercalation and Superconductivity in Solid State Fullerenes

Abstract

Co-intercalation of alkali metals and neutral molecules into C₆₀ have been investigated. Successful synthesis of this class of materials provides us a new insight into superconductivity and other physical properties; such as criterion for the occurrence of superconductivity and metal-insulator transitions. Based on the recently synthesized ammoniated fullerides, we discuss on the relation between the structure, and electronic properties.     

One-process surface texturing of a large area by electrochemical machining with short voltage pulses

Micro texturing is performed over a large area of a metal surface by an appropriate electrochemical machining. A micro texture pattern with a size of several tens of micrometers is fabricated on a millimeter-scale tool electrode surface and is simultaneously replicated on the workpiece in one process. A shorter pulse width is required for better machining accuracy, and then branched electric feeding is adopted because the minimum machinable pulse width is limited due to the inductance of the power supply circuit. Micro textures smaller than a hundred micrometers could be achieved with a machining area of several tens of square millimeters.     

X-Ray Observation of Gas Evolution, Flotation, and Emulsification of Molten Carbon Steel Immersed in Mold Flux

At two interfacial-tension measurement experiments with the same experimental conditions, steel samples and mold flux samples of the same compositions were melted in crucibles from the same batch. During the first experiment, the steel drop melted far below its liquidus and then was emulsified. At the second experiment, the steel melted at the expected temperature but did not emulsify. The difference that can be identified is the mass of the steel samples.     

High‐tensile‐strength polyvinyl alcohol films prepared from freeze/thaw cycled gels

The mechanical properties and molecular structure of a poly(vinyl alcohol) (PVA) film, which was obtained by eliminating water from a PVA hydrogel using repeated freeze/thaw cycles, were investigated by tensile tests, thermal analysis, and X‐ray diffraction measurements. The mechanical properties of PVA with 99.9% saponification were measured as a function of the number of freeze/thaw cycles performed. The tensile strength and Young's modulus increased and the elongation at break decreased with increasing freeze/thaw cycles. The tensile strength and Young's modulus of PVA films obtained after seven freeze/thaw cycles were as high as 255 MPa and 13.5 GPa after annealing at 130°C. Thermal analysis and X‐ray diffraction measurements revealed that this is because of a high crystallinity and a large crystallite size. A good relationship between the tensile strength and the glass transition temperature was obtained, regardless of the degree of saponification and annealing conditions. © 2014 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2014 , 131, 40578.     

大钢模板在地下室外墙中应用的技术分析

目前,大型公建项目的地下室外墙模板主要采用木模板及小钢模进行支设,但两者都有各自的优缺点.以京东方运营与研发中心工程为例,根据其特点,对木模板、小钢模板和大钢模板进行了对比分析,并着重介绍了地下室外墙模板采用了大钢模板的设计与方法,希望为后续类似工程地下室外墙模板的施工起到借鉴的作用.     

Rupture Strength of some Mould Flux Slag Films relevant to Swedish Continuous Casting Practice

In this paper, the high temperature rupture strengths of some solid mould flux slag films in Swedish continuous casting processes are presented. The apparatus, designed and constructed in the present laboratory is based on the principle that the solidified film of the mould flux slag is subjected to elongation stress at temperatures below the solidus points of the fluxes. The rupture strength tests were conducted in air. The results indicate that the rupture strength is dependent on the extent of polymerization of the silicate network. The results of Raman spectra studies of the mould flux slags used in the present work were corroborated by estimations of the NBO/T ratios of the slags. It was found that lower NBO/T ratios in the slag led to higher rupture strengths. Earlier measurements of the viscosities, thermal diffusivities as well as stress relaxation results for the same slags followed a similar trend indicating the dependency of these properties on the slag structure and a clear link between thermophysical and mechanical properties of slags. The results are expected to provide some basic guidelines to develop the next‐generation mould flux and modelling of the continuous casting process.     

In Situ PD-L1 Expression in Oral Squamous Cell Carcinoma Is Induced by Heterogeneous Mechanisms among Patients

The expression of programmed death ligand-1 (PD-L1) is controlled by complex mechanisms. The elucidation of the molecular mechanisms of PD-L1 expression is important for the exploration of new insights into PD-1 blockade therapy. Detailed mechanisms of the in situ expression of PD-L1 in tissues of oral squamous cell carcinomas (OSCCs) have not yet been clarified. We examined the mechanisms of PD-L1 expression focusing on the phosphorylation of downstream molecules of epidermal growth factor (EGF) and interferon gamma (IFN-γ) signaling in vitro and in vivo by immunoblotting and multi-fluorescence immunohistochemistry (MF-IHC), respectively. The in vitro experiments demonstrated that PD-L1 expression in OSCC cell lines is upregulated by EGF via the EGF receptor (EGFR)/PI3K/AKT pathway, the EGFR/STAT1 pathway, and the EGFR/MEK/ERK pathway, and by IFN-γ via the JAK2/STAT1 pathway. MF-IHC demonstrated that STAT1 and EGFR phosphorylation was frequently shown in PD-L1-positive cases and STAT1 phosphorylation was correlated with lymphocyte infiltration and EGFR phosphorylation. Moreover, the phosphorylation pattern of the related molecules in PD-L1-positive cells differed among the cases investigated. These findings indicate that PD-L1 expression mechanisms differ depending on the tissue environment and suggest that the examination of the tissue environment and molecular alterations of cancer cells affecting PD-L1 expression make it necessary for each patient to choose the appropriate combination drugs for PD-1 blockade cancer treatment.