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Sample records for pathway direct afm

  1. Segmental calibration for commercial AFM in vertical direction

    NASA Astrophysics Data System (ADS)

    Shi, Yushu; Gao, Sitian; Lu, Mingzhen; Li, Wei; Xu, Xuefang

    2013-01-01

    Atomic force microscopy (AFM) is most widely applied in scientific research and industrial production. AFM is a scanning probe imaging and measuring device, useful for physical and chemical studies. Depends on its basic structure, microscopic surface pattern can be measured and captured by mechanically scanning. Its vertical and horizon resolution can reach to 0.01nm and 0.1nm. Commonly the measurement values of commercial AFM are directly from scanning piezoelectric tube, so that it not a traceable value. In order to solve the problem of commercial AFM's traceability, step height standard references are applied to calibrate the piezoelectric ceramic housing in scanning tube. All of the serial of step height standard references, covering the commercial AFM vertical scale, are calibrated by Metrology AFM developed by National Institute of Metrology (NIM), China. Three interferometers have been assembled in its XYZ axis, therefore the measurement value can directly trace to laser wavelength. Because of nonlinear characteristic of PZT, the method of segmental calibration is proposed. The measurement scale can be divided into several subsections corresponding to the calibrated values of the series of step height standards references. By this method the accuracy of measurements can be ensured in each segment measurement scale and the calibration level of the whole instrument can be promoted. In order to get a standard step shape by commercial AFM, substrate removal method is applied to deal with the bow shape problem.

  2. Mapping site-specific endonuclease binding to DNA by direct imaging with AFM

    SciTech Connect

    Allison, D.P.; Thundat, T.; Doktycz, M.J.; Kerper, P.S.; Warmack, R.J.; Modrich, P.; Isfort, R.J.

    1995-12-31

    Physical mapping of DNA can be accomplished by direct AFM imaging of site specific proteins bound to DNA molecules. Using Gln-111, a mutant of EcoRI endonuclease with a specific affinity for EcoRI sites 1,000 times greater than wild type enzyme but with cleavage rate constants reduced by a factor of 10{sup 4}, the authors demonstrate site-specific mapping by direct AFM imaging. Images are presented showing specific-site binding of Gln-111 to plasmids having either one (pBS{sup +}) or two (pMP{sup 32}) EcoRI sites. Identification of the Gln-111/DNA complex is greatly enhanced by biotinylation of the complex followed by reaction with streptavidin gold prior to imaging. Image enhancement coupled with improvements in the preparation techniques for imaging large DNA molecules, such as lambda DNA (47 kb), has the potential to contribute to direct AFM restriction mapping of cosmid-sized genomic DNAs.

  3. Direct Measurement of Optical Force Induced by Near-Field Plasmonic Cavity Using Dynamic Mode AFM

    PubMed Central

    Guan, Dongshi; Hang, Zhi Hong; Marcet, Zsolt; Liu, Hui; Kravchenko, I. I.; Chan, C. T.; Chan, H. B.; Tong, Penger

    2015-01-01

    Plasmonic nanostructures have attracted much attention in recent years because of their potential applications in optical manipulation through near-field enhancement. Continuing experimental efforts have been made to develop accurate techniques to directly measure the near-field optical force induced by the plasmonic nanostructures in the visible frequency range. In this work, we report a new application of dynamic mode atomic force microscopy (DM-AFM) in the measurement of the enhanced optical force acting on a nano-structured plasmonic resonant cavity. The plasmonic cavity is made of an upper gold-coated glass sphere and a lower quartz substrate patterned with an array of subwavelength gold disks. In the near-field when the sphere is positioned close to the disk array, plasmonic resonance is excited in the cavity and the induced force by a 1550 nm infrared laser is found to be increased by an order of magnitude compared with the photon pressure generated by the same laser light. The experiment demonstrates that DM-AFM is a powerful tool for the study of light induced forces and their enhancement in plasmonic nanostructures. PMID:26586455

  4. Direct measurement of optical force induced by near-field plasmonic cavity using dynamic mode AFM

    SciTech Connect

    Guan, Dongshi; Hang, Zhi Hong; Marset, Zsolt; Liu, Hui; Kravchenko, Ivan I.; Chan, Ho Bun; Chan, C. T.; Tong, Penger

    2015-11-20

    Plasmonic nanostructures have attracted much attention in recent years because of their potential applications in optical manipulation through near-field enhancement. Continuing experimental efforts have been made to develop accurate techniques to directly measure the near-field optical force induced by the plasmonic nanostructures in the visible frequency range. In this work, we report a new application of dynamic mode atomic force microscopy (DM-AFM) in the measurement of the enhanced optical force acting on a nano-structured plasmonic resonant cavity. The plasmonic cavity is made of an upper gold-coated glass sphere and a lower quartz substrate patterned with an array of subwavelength gold disks. In the near-field when the sphere is positioned close to the disk array, plasmonic resonance is excited in the cavity and the induced force by a 1550 nm infrared laser is found to be increased by an order of magnitude compared with the photon pressure generated by the same laser light. Lastly, the experiment demonstrates that DM-AFM is a powerful tool for the study of light induced forces and their enhancement in plasmonic nanostructures.

  5. Direct measurement of optical force induced by near-field plasmonic cavity using dynamic mode AFM

    DOE PAGESBeta

    Guan, Dongshi; Hang, Zhi Hong; Marset, Zsolt; Liu, Hui; Kravchenko, Ivan I.; Chan, Ho Bun; Chan, C. T.; Tong, Penger

    2015-11-20

    Plasmonic nanostructures have attracted much attention in recent years because of their potential applications in optical manipulation through near-field enhancement. Continuing experimental efforts have been made to develop accurate techniques to directly measure the near-field optical force induced by the plasmonic nanostructures in the visible frequency range. In this work, we report a new application of dynamic mode atomic force microscopy (DM-AFM) in the measurement of the enhanced optical force acting on a nano-structured plasmonic resonant cavity. The plasmonic cavity is made of an upper gold-coated glass sphere and a lower quartz substrate patterned with an array of subwavelength goldmore » disks. In the near-field when the sphere is positioned close to the disk array, plasmonic resonance is excited in the cavity and the induced force by a 1550 nm infrared laser is found to be increased by an order of magnitude compared with the photon pressure generated by the same laser light. Lastly, the experiment demonstrates that DM-AFM is a powerful tool for the study of light induced forces and their enhancement in plasmonic nanostructures.« less

  6. Direct Measurement of Optical Force Induced by Near-Field Plasmonic Cavity Using Dynamic Mode AFM

    NASA Astrophysics Data System (ADS)

    Guan, Dongshi; Hang, Zhi Hong; Marcet, Zsolt; Liu, Hui; Kravchenko, I. I.; Chan, C. T.; Chan, H. B.; Tong, Penger

    2015-11-01

    Plasmonic nanostructures have attracted much attention in recent years because of their potential applications in optical manipulation through near-field enhancement. Continuing experimental efforts have been made to develop accurate techniques to directly measure the near-field optical force induced by the plasmonic nanostructures in the visible frequency range. In this work, we report a new application of dynamic mode atomic force microscopy (DM-AFM) in the measurement of the enhanced optical force acting on a nano-structured plasmonic resonant cavity. The plasmonic cavity is made of an upper gold-coated glass sphere and a lower quartz substrate patterned with an array of subwavelength gold disks. In the near-field when the sphere is positioned close to the disk array, plasmonic resonance is excited in the cavity and the induced force by a 1550 nm infrared laser is found to be increased by an order of magnitude compared with the photon pressure generated by the same laser light. The experiment demonstrates that DM-AFM is a powerful tool for the study of light induced forces and their enhancement in plasmonic nanostructures.

  7. Direct Measurement of Optical Force Induced by Near-Field Plasmonic Cavity Using Dynamic Mode AFM.

    PubMed

    Guan, Dongshi; Hang, Zhi Hong; Marcet, Zsolt; Liu, Hui; Kravchenko, I I; Chan, C T; Chan, H B; Tong, Penger

    2015-01-01

    Plasmonic nanostructures have attracted much attention in recent years because of their potential applications in optical manipulation through near-field enhancement. Continuing experimental efforts have been made to develop accurate techniques to directly measure the near-field optical force induced by the plasmonic nanostructures in the visible frequency range. In this work, we report a new application of dynamic mode atomic force microscopy (DM-AFM) in the measurement of the enhanced optical force acting on a nano-structured plasmonic resonant cavity. The plasmonic cavity is made of an upper gold-coated glass sphere and a lower quartz substrate patterned with an array of subwavelength gold disks. In the near-field when the sphere is positioned close to the disk array, plasmonic resonance is excited in the cavity and the induced force by a 1550 nm infrared laser is found to be increased by an order of magnitude compared with the photon pressure generated by the same laser light. The experiment demonstrates that DM-AFM is a powerful tool for the study of light induced forces and their enhancement in plasmonic nanostructures. PMID:26586455

  8. Direct electrochemical and AFM detection of amyloid-β peptide aggregation on basal plane HOPG

    NASA Astrophysics Data System (ADS)

    Lopes, Paula; Xu, Meng; Zhang, Min; Zhou, Ting; Yang, Yanlian; Wang, Chen; Ferapontova, Elena E.

    2014-06-01

    Amyloidogenesis is associated with more than 30 human diseases, including Alzheimer's which is related to aggregation of β-amyloid peptide (Aβ). Here, consecutive stages of Aβ42 aggregation and amyloid fibril formation were followed electrochemically via oxidation of tyrosines in Aβ42 adsorbed on the basal plane graphite electrode and directly correlated with Aβ42 morphological changes observed by atomic force microscopy of the same substrate. The results offer new tools for analysis of mechanisms of Aβ aggregation.Amyloidogenesis is associated with more than 30 human diseases, including Alzheimer's which is related to aggregation of β-amyloid peptide (Aβ). Here, consecutive stages of Aβ42 aggregation and amyloid fibril formation were followed electrochemically via oxidation of tyrosines in Aβ42 adsorbed on the basal plane graphite electrode and directly correlated with Aβ42 morphological changes observed by atomic force microscopy of the same substrate. The results offer new tools for analysis of mechanisms of Aβ aggregation. Electronic supplementary information (ESI) available: Experimental details: procedures for Aβ42 aggregation and electrode modification, DPV/AFM measurements and analysis. See DOI: 10.1039/c4nr02413c

  9. Non-reciprocal directional dichroism in the AFM phase of BiFeO3 at THz frequencies

    NASA Astrophysics Data System (ADS)

    Nagel, Urmas; Rõõm, T.; Farkas, D.; Szaller, D.; Bordács, S.; Kézsmárki, I.; Engelkamp, H.; Ozaki, Y.; Tomiaki, Y.; Ito, T.; Fishman, Randy S.

    We did THz absorption spectroscopy of BiFeO3 single crystals in the AFM phase, where the spin cycloid is destroyed in magnetic fields between 18 T and 32 T in Voigt geometry at 1.6 K. If B0 ∥ [ 1 1 0 ] , we see strong directional dichroism (DD) of absorption of the magnon mode with light propagating along the direction of the ferroelectric polarization k ∥ P ∥ [ 111 ] and eω ∥ [ 1 1 0 ] , bω ∥ [ 1 1 2 ] . The sign of DD can be reversed (i) by reversing the direction of B0 or (ii) by flipping the sample, thus reversing the propagation direction of light. The observed effect is caused by the strong magneto-electric coupling in the collinear AFM phase. Research sponsored by the Estonian Ministry of Education and Research (IUT23-3).

  10. Direct visualization of the trimeric structure of the ASIC1a channel, using AFM imaging

    SciTech Connect

    Carnally, Stewart M.; Dev, Harveer S.; Stewart, Andrew P.; Barrera, Nelson P.; Van Bemmelen, Miguel X.; Schild, Laurent; Henderson, Robert M.; Edwardson, J.Michael

    2008-08-08

    There has been confusion about the subunit stoichiometry of the degenerin family of ion channels. Recently, a crystal structure of acid-sensing ion channel (ASIC) 1a revealed that it assembles as a trimer. Here, we used atomic force microscopy (AFM) to image unprocessed ASIC1a bound to mica. We detected a mixture of subunit monomers, dimers and trimers. In some cases, triple-subunit clusters were clearly visible, confirming the trimeric structure of the channel, and indicating that the trimer sometimes disaggregated after adhesion to the mica surface. This AFM-based technique will now enable us to determine the subunit arrangement within heteromeric ASICs.

  11. Direct comparison of AFM and SEM measurements on the same set of nanoparticles

    NASA Astrophysics Data System (ADS)

    Delvallée, A.; Feltin, N.; Ducourtieux, S.; Trabelsi, M.; Hochepied, J. F.

    2015-08-01

    This article is the first step in the development of a hybrid metrology combining AFM and SEM techniques for measuring the dimensions of a nanoparticle population in 3D space (X,Y,Z). This method exploits the strengths of each technique on the same set of nanoparticles. AFM is used for measuring the nanoparticle height and the measurements along X and Y axes are deduced from SEM images. A sampling method is proposed in order to obtain the best deposition conditions of SiO2 and gold nanoparticles on mica or silicon substrates. Only the isolated nanoparticles are taken into account in the histogram of size distribution. Moreover, a semi-automatic Matlab routine has also been developed to process the AFM and SEM images, measure and count the nanoparticles. This routine allows the user to exclusively select the isolated nanoparticles through a control interface. The measurements have been performed on spherical-like nanoparticles to test the method by comparing the results obtained with both techniques.

  12. Accuracy of AFM force distance curves via direct solution of the Euler-Bernoulli equation

    NASA Astrophysics Data System (ADS)

    Eppell, Steven J.; Liu, Yehe; Zypman, Fredy R.

    2016-03-01

    In an effort to improve the accuracy of force-separation curves obtained from atomic force microscope data, we compare force-separation curves computed using two methods to solve the Euler-Bernoulli equation. A recently introduced method using a direct sequential forward solution, Causal Time-Domain Analysis, is compared against a previously introduced Tikhonov Regularization method. Using the direct solution as a benchmark, it is found that the regularization technique is unable to reproduce accurate curve shapes. Using L-curve analysis and adjusting the regularization parameter, λ, to match either the depth or the full width at half maximum of the force curves, the two techniques are contrasted. Matched depths result in full width at half maxima that are off by an average of 27% and matched full width at half maxima produce depths that are off by an average of 109%.

  13. Diversifying Carotenoid Biosynthetic Pathways by Directed Evolution

    PubMed Central

    Umeno, Daisuke; Tobias, Alexander V.; Arnold, Frances H.

    2005-01-01

    Microorganisms and plants synthesize a diverse array of natural products, many of which have proven indispensable to human health and well-being. Although many thousands of these have been characterized, the space of possible natural products—those that could be made biosynthetically—remains largely unexplored. For decades, this space has largely been the domain of chemists, who have synthesized scores of natural product analogs and have found many with improved or novel functions. New natural products have also been made in recombinant organisms, via engineered biosynthetic pathways. Recently, methods inspired by natural evolution have begun to be applied to the search for new natural products. These methods force pathways to evolve in convenient laboratory organisms, where the products of new pathways can be identified and characterized in high-throughput screening programs. Carotenoid biosynthetic pathways have served as a convenient experimental system with which to demonstrate these ideas. Researchers have mixed, matched, and mutated carotenoid biosynthetic enzymes and screened libraries of these “evolved” pathways for the emergence of new carotenoid products. This has led to dozens of new pathway products not previously known to be made by the assembled enzymes. These new products include whole families of carotenoids built from backbones not found in nature. This review details the strategies and specific methods that have been employed to generate new carotenoid biosynthetic pathways in the laboratory. The potential application of laboratory evolution to other biosynthetic pathways is also discussed. PMID:15755953

  14. Germ direct observation by AFM under crystallization of self-organized assemblies of mono-protonated meso-tetraphenylporphine dimers

    NASA Astrophysics Data System (ADS)

    Udal'tsov, Alexander V.

    2016-08-01

    Assemblies consisting of mono-protonated meso-tetraphenylporphine dimers and water have been investigated by UV-vis spectroscopy in solution and atomic force microscopy (AFM) in thin layers. These assemblies self-organized into domains produce microcrystals in thin layer. Morphology of the microcrystals and characteristic features of crystallization germ on the top found by contact AFM indicate that surface tension of an aqueous layer on the domain generates the crystallization process. Estimations of the pressure producing the germ and bulk modulus (Bm) of microcrystals give 26.3±2.6 MPa and 3.72 GPa and Bm=12.7 GPa obtained for dried thin films. The former modulus is comparable with bulk modulus of water (2.174 GPa) that implies liquid crystals formation. Absorptions of longitudinal optical (LO) phonons with ћωLO=0.3761 and 0.3577 eV, which are arisen because of hole polaron moving through water, are found in the electronic spectra of the assemblies. The crystallization is suggested to occur due to Zundel cation (H5O2+) operation like the water-porphyrin matrix self-organization found earlier.

  15. The role of confinement and corona crystallinity on the bending modulus of copolymer micelles measured directly by AFM flexural tests.

    PubMed

    Jennings, L; Glazer, P; Laan, A C; de Kruijff, R M; Waton, G; Schosseler, F; Mendes, E

    2016-09-21

    We present an approach which makes it possible to directly determine the bending modulus of single elongated block copolymer micelles. This is done by forming arrays of suspended micelles onto microfabricated substrates and by performing three-point bending flexural tests, using an atomic force microscope, on their suspended portions. By coupling the direct atomic force microscopy measurements with differential scanning calorimetry data, we show that the presence of a crystalline corona strongly increases the modulus of the copolymer elongated micelles. This large increase suggests that crystallites in the corona are larger and more uniformly oriented due to confinement effects. Our findings together with this hypothesis open new interesting avenues for the preparation of core-templated polymer fibres with enhanced mechanical properties. PMID:27506248

  16. Chemical Shifts to Metabolic Pathways: Identifying Metabolic Pathways Directly from a Single 2D NMR Spectrum.

    PubMed

    Dubey, Abhinav; Rangarajan, Annapoorni; Pal, Debnath; Atreya, Hanudatta S

    2015-12-15

    Identifying cellular processes in terms of metabolic pathways is one of the avowed goals of metabolomics studies. Currently, this is done after relevant metabolites are identified to allow their mapping onto specific pathways. This task is daunting due to the complex nature of cellular processes and the difficulty in establishing the identity of individual metabolites. We propose here a new method: ChemSMP (Chemical Shifts to Metabolic Pathways), which facilitates rapid analysis by identifying the active metabolic pathways directly from chemical shifts obtained from a single two-dimensional (2D) [(13)C-(1)H] correlation NMR spectrum without the need for identification and assignment of individual metabolites. ChemSMP uses a novel indexing and scoring system comprised of a "uniqueness score" and a "coverage score". Our method is demonstrated on metabolic pathways data from the Small Molecule Pathway Database (SMPDB) and chemical shifts from the Human Metabolome Database (HMDB). Benchmarks show that ChemSMP has a positive prediction rate of >90% in the presence of decluttered data and can sustain the same at 60-70% even in the presence of noise, such as deletions of peaks and chemical shift deviations. The method tested on NMR data acquired for a mixture of 20 amino acids shows a success rate of 93% in correct recovery of pathways. When used on data obtained from the cell lysate of an unexplored oncogenic cell line, it revealed active metabolic pathways responsible for regulating energy homeostasis of cancer cells. Our unique tool is thus expected to significantly enhance analysis of NMR-based metabolomics data by reducing existing impediments. PMID:26556218

  17. Direct and indirect dorsolateral striatum pathways reinforce different action strategies

    PubMed Central

    Vicente, Ana M.; Galvão-Ferreira, Pedro; Tecuapetla, Fatuel; Costa, Rui M.

    2016-01-01

    Summary The basal ganglia, and the striatum in particular, are critical for action reinforcement 1, 2. The dorsal striatum, which can be further subdivided into dorsomedial (DMS) and dorsolateral (DLS) striatum, is mainly composed of two subpopulations of striatal medium spiny projection neurons (MSNs): dopamine D1 receptor-expressing MSNs that constitute the striatonigral or direct pathway (dMSNs); and dopamine D2 receptor-expressing MSNs that constitute the striatopallidal or indirect pathway (iMSNs) [3]. It has been suggested that each pathway has opposing roles in reinforcement, with dMSNs being important to learn positive reinforcement and iMSNs to learn to avoid undesired actions (Go/No-Go) [1]. Furthermore, optogenetic self-stimulation of dMSNs in DMS leads to reinforcement of actions, while self-stimulation of iMSNs leads to avoidance of actions [2]. However, in DLS, which has been implicated in the consolidation of well-trained actions and habits in mice 4, 5, both pathways are active during lever-pressing for reward [6]. Furthermore, extensive skill training leads to long-lasting potentiation of glutamatergic inputs into both dMSNs and iMSNs [4]. We report here that, in DLS, both dMSNs and iMSNs are involved in positive reinforcement, but support different action strategies. PMID:27046807

  18. Direct and indirect dorsolateral striatum pathways reinforce different action strategies.

    PubMed

    Vicente, Ana M; Galvão-Ferreira, Pedro; Tecuapetla, Fatuel; Costa, Rui M

    2016-04-01

    The basal ganglia, and the striatum in particular, are critical for action reinforcement [1,2]. The dorsal striatum, which can be further subdivided into dorsomedial (DMS) and dorsolateral (DLS) striatum, is mainly composed of two subpopulations of striatal medium spiny projection neurons (MSNs): dopamine D1 receptor-expressing MSNs that constitute the striatonigral or direct pathway (dMSNs); and dopamine D2 receptor-expressing MSNs that constitute the striatopallidal or indirect pathway (iMSNs) [3]. It has been suggested that each pathway has opposing roles in reinforcement, with dMSNs being important to learn positive reinforcement and iMSNs to learn to avoid undesired actions (Go/No-Go) [1]. Furthermore, optogenetic self-stimulation of dMSNs in DMS leads to reinforcement of actions, while self-stimulation of iMSNs leads to avoidance of actions [2]. However, in DLS, which has been implicated in the consolidation of well-trained actions and habits in mice [4,5], both pathways are active during lever-pressing for reward [6]. Furthermore, extensive skill training leads to long-lasting potentiation of glutamatergic inputs into both dMSNs and iMSNs [4]. We report here that, in DLS, both dMSNs and iMSNs are involved in positive reinforcement, but support different action strategies. PMID:27046807

  19. Qplus AFM driven nanostencil.

    PubMed

    Grévin, B; Fakir, M; Hayton, J; Brun, M; Demadrille, R; Faure-Vincent, J

    2011-06-01

    We describe the development of a novel setup, in which large stencils with suspended silicon nitride membranes are combined with atomic force microscopy (AFM) regulation by using tuning forks. This system offers the possibility to perform separate AFM and nanostencil operations, as well as combined modes when using stencil chips with integrated tips. The flexibility and performances are demonstrated through a series of examples, including wide AFM scans in closed loop mode, probe positioning repeatability of a few tens of nanometer, simultaneous evaporation of large (several hundred of micron square) and nanoscopic metals and fullerene patterns in static, multistep, and dynamic modes. This approach paves the way for further developments, as it fully combines the advantages of conventional stenciling with the ones of an AFM driven shadow mask. PMID:21721701

  20. Developmental Pathways Direct Pancreatic Cancer Initiation from Its Cellular Origin

    PubMed Central

    Reichert, Maximilian; Blume, Karin; Kleger, Alexander; Hartmann, Daniel; von Figura, Guido

    2016-01-01

    Pancreatic ductal adenocarcinoma (PDA) is characterized by an extremely poor prognosis, since it is usually diagnosed at advanced stages. In order to employ tools for early detection, a better understanding of the early stages of PDA development from its main precursors, pancreatic intraepithelial neoplasia (PanIN), and intraductal papillary mucinous neoplasm (IPMN) is needed. Recent studies on murine PDA models have identified a different exocrine origin for PanINs and IPMNs. In both processes, developmental pathways direct the initiation of PDA precursors from their cellular ancestors. In this review, the current understanding of early PDA development is summarized. PMID:26681957

  1. Direct corticospinal pathways contribute to neuromuscular control of perturbed stance.

    PubMed

    Taube, Wolfgang; Schubert, Martin; Gruber, Markus; Beck, Sandra; Faist, Michael; Gollhofer, Albert

    2006-08-01

    The antigravity soleus muscle (Sol) is crucial for compensation of stance perturbation. A corticospinal contribution to the compensatory response of the Sol is under debate. The present study assessed spinal, corticospinal, and cortical excitability at the peaks of short- (SLR), medium- (MLR), and long-latency responses (LLR) after posterior translation of the feet. Transcranial magnetic stimulation (TMS) and peripheral nerve stimulation were individually adjusted so that the peaks of either motor evoked potential (MEP) or H reflex coincided with peaks of SLR, MLR, and LLR, respectively. The influence of specific, presumably direct, corticospinal pathways was investigated by H-reflex conditioning. When TMS was triggered so that the MEP arrived in the Sol at the same time as the peaks of SLR and MLR, EMG remained unaffected. Enhanced EMG was observed when the MEP coincided with the LLR peak (P < 0.001). Similarly, conditioning of the H reflex by subthreshold TMS facilitated H reflexes only at LLR (P < 0.001). The earliest facilitation after perturbation occurred after 86 ms. The TMS-induced H-reflex facilitation at LLR suggests that increased cortical excitability contributes to the augmentation of the LLR peaks. This provides evidence that the LLR in the Sol muscle is at least partly transcortical, involving direct corticospinal pathways. Additionally, these results demonstrate that approximately 86 ms after perturbation, postural compensatory responses are cortically mediated. PMID:16601305

  2. Identification of a direct GABAergic pallidocortical pathway in rodents

    PubMed Central

    Chen, Michael C.; Ferrari, Loris; Sacchet, Matthew D.; Foland-Ross, Lara C.; Qiu, Mei-Hong; Gotlib, Ian H.; Fuller, Patrick M.; Arrigoni, Elda; Lu, Jun

    2014-01-01

    The basal ganglia, interacting with the cortex, play a critical role in a range of behaviors. Output from the basal ganglia to the cortex is thought to relay through the thalamus, yet an intriguing alternative is that the basal ganglia may directly project to, and communicate with, the cortex. We explored an efferent projection from the globus pallidus externa (GPe), a key hub in the basal ganglia system, to the cortex of rats and mice. Anterograde and retrograde tracing revealed projections to the frontal premotor cortex, especially the deep projecting layers, originating from GPe neurons that receive axonal inputs from the dorsal striatum. Cre-dependent anterograde tracing in GPe Vgat-ires-cre mice confirmed that the pallidocortical projection is GABAergic, and in vitro optogenetic stimulation in the cortex of these projections produced a fast inhibitory postsynaptic current in targeted cells that was abolished by bicucculine. The pallidocortical projections targeted GABAergic interneurons and, to a lesser extent, pyramidal neurons. This GABAergic pallidocortical pathway directly links the basal ganglia and cortex and may play a key role in behavior and cognition in normal and disease states. PMID:25581560

  3. Molecular pathways for defect annihilation in directed self-assembly.

    PubMed

    Hur, Su-Mi; Thapar, Vikram; Ramírez-Hernández, Abelardo; Khaira, Gurdaman; Segal-Peretz, Tamar; Rincon-Delgadillo, Paulina A; Li, Weihua; Müller, Marcus; Nealey, Paul F; de Pablo, Juan J

    2015-11-17

    Over the last few years, the directed self-assembly of block copolymers by surface patterns has transitioned from academic curiosity to viable contender for commercial fabrication of next-generation nanocircuits by lithography. Recently, it has become apparent that kinetics, and not only thermodynamics, plays a key role for the ability of a polymeric material to self-assemble into a perfect, defect-free ordered state. Perfection, in this context, implies not more than one defect, with characteristic dimensions on the order of 5 nm, over a sample area as large as 100 cm(2). In this work, we identify the key pathways and the corresponding free energy barriers for eliminating defects, and we demonstrate that an extraordinarily large thermodynamic driving force is not necessarily sufficient for their removal. By adopting a concerted computational and experimental approach, we explain the molecular origins of these barriers and how they depend on material characteristics, and we propose strategies designed to overcome them. The validity of our conclusions for industrially relevant patterning processes is established by relying on instruments and assembly lines that are only available at state-of-the-art fabrication facilities, and, through this confluence of fundamental and applied research, we are able to discern the evolution of morphology at the smallest relevant length scales-a handful of nanometers-and present a view of defect annihilation in directed self-assembly at an unprecedented level of detail. PMID:26515095

  4. Molecular pathways for defect annihilation in directed self-assembly

    PubMed Central

    Hur, Su-Mi; Thapar, Vikram; Ramírez-Hernández, Abelardo; Khaira, Gurdaman; Segal-Peretz, Tamar; Rincon-Delgadillo, Paulina A.; Li, Weihua; Müller, Marcus; Nealey, Paul F.; de Pablo, Juan J.

    2015-01-01

    Over the last few years, the directed self-assembly of block copolymers by surface patterns has transitioned from academic curiosity to viable contender for commercial fabrication of next-generation nanocircuits by lithography. Recently, it has become apparent that kinetics, and not only thermodynamics, plays a key role for the ability of a polymeric material to self-assemble into a perfect, defect-free ordered state. Perfection, in this context, implies not more than one defect, with characteristic dimensions on the order of 5 nm, over a sample area as large as 100 cm2. In this work, we identify the key pathways and the corresponding free energy barriers for eliminating defects, and we demonstrate that an extraordinarily large thermodynamic driving force is not necessarily sufficient for their removal. By adopting a concerted computational and experimental approach, we explain the molecular origins of these barriers and how they depend on material characteristics, and we propose strategies designed to overcome them. The validity of our conclusions for industrially relevant patterning processes is established by relying on instruments and assembly lines that are only available at state-of-the-art fabrication facilities, and, through this confluence of fundamental and applied research, we are able to discern the evolution of morphology at the smallest relevant length scales—a handful of nanometers—and present a view of defect annihilation in directed self-assembly at an unprecedented level of detail. PMID:26515095

  5. Molecular pathways for defect annihilation in directed self-assembly.

    SciTech Connect

    Hur, Su-Mi; Thapar, Vikram; Ramirez-Hernandez, Abelardo; Khaira, Gurdaman S.; Segal-Peretz, Tamar; Rincon-Delgadillo, Paulina A.; Li, Weihua; Muller, Marcus; Nealey, Paul F.; de Pablo, Juan J.

    2015-11-17

    Over the last few years, the directed self-assembly of block copolymers by surface patterns has transitioned from academic curiosity to viable contender for commercial fabrication of next-generation nanocircuits by lithography. Recently, it has become apparent that kinetics, and not only thermodynamics, plays a key role for the ability of a polymeric material to self-assemble into a perfect, defect-free ordered state. Perfection, in this context, implies not more than one defect, with characteristic dimensions on the order of 5 nm, over a sample area as large as 100 cm2. In this work, we identify the key pathways and the corresponding free-energy barriers for eliminating defects, and we demonstrate that an extraordinarily large thermodynamic driving force is not necessarily sufficient for their removal. By adopting a concerted computational and experimental approach, we explain the molecular origins of these barriers, how they depend on material characteristics, and we propose strategies designed to over-come them. The validity of our conclusions for industrially-relevant patterning processes is established by relying on instruments and assembly lines that are only available at state-of-the-art fabrication facilities and, through this confluence of fundamental and applied research, we are able to discern the evolution of morphology at the smallest relevant length scales - a handful of nanometers -, and present a view of defect annihilation in directed self-assembly at an unprecedented level of detail.

  6. Molecular pathways for defect annihilation in directed self-assembly.

    DOE PAGESBeta

    Hur, Su-Mi; Thapar, Vikram; Ramirez-Hernandez, Abelardo; Khaira, Gurdaman S.; Segal-Peretz, Tamar; Rincon-Delgadillo, Paulina A.; Li, Weihua; Muller, Marcus; Nealey, Paul F.; de Pablo, Juan J.

    2015-11-17

    Over the last few years, the directed self-assembly of block copolymers by surface patterns has transitioned from academic curiosity to viable contender for commercial fabrication of next-generation nanocircuits by lithography. Recently, it has become apparent that kinetics, and not only thermodynamics, plays a key role for the ability of a polymeric material to self-assemble into a perfect, defect-free ordered state. Perfection, in this context, implies not more than one defect, with characteristic dimensions on the order of 5 nm, over a sample area as large as 100 cm2. In this work, we identify the key pathways and the corresponding free-energymore » barriers for eliminating defects, and we demonstrate that an extraordinarily large thermodynamic driving force is not necessarily sufficient for their removal. By adopting a concerted computational and experimental approach, we explain the molecular origins of these barriers, how they depend on material characteristics, and we propose strategies designed to over-come them. The validity of our conclusions for industrially-relevant patterning processes is established by relying on instruments and assembly lines that are only available at state-of-the-art fabrication facilities and, through this confluence of fundamental and applied research, we are able to discern the evolution of morphology at the smallest relevant length scales - a handful of nanometers -, and present a view of defect annihilation in directed self-assembly at an unprecedented level of detail.« less

  7. AFM indentation study of breast cancer cells

    SciTech Connect

    Li, Q.S.; Lee, G.Y.H.; Ong, C.N.; Lim, C.T.

    2008-10-03

    Mechanical properties of individual living cells are known to be closely related to the health and function of the human body. Here, atomic force microscopy (AFM) indentation using a micro-sized spherical probe was carried out to characterize the elasticity of benign (MCF-10A) and cancerous (MCF-7) human breast epithelial cells. AFM imaging and confocal fluorescence imaging were also used to investigate their corresponding sub-membrane cytoskeletal structures. Malignant (MCF-7) breast cells were found to have an apparent Young's modulus significantly lower (1.4-1.8 times) than that of their non-malignant (MCF-10A) counterparts at physiological temperature (37 deg. C), and their apparent Young's modulus increase with loading rate. Both confocal and AFM images showed a significant difference in the organization of their sub-membrane actin structures which directly contribute to their difference in cell elasticity. This change may have facilitated easy migration and invasion of malignant cells during metastasis.

  8. Direct Observation of Kinetic Pathways of Biomolecular Recognition.

    PubMed

    Choudhury, Susobhan; Batabyal, Subrata; Mondal, Prasanna Kumar; Singh, Priya; Lemmens, Peter; Pal, Samir Kumar

    2015-11-01

    The pathways of molecular recognition, which is a central event in all biological processes, belong to the most important subjects of contemporary research in biomolecular science. By using fluorescence spectroscopy in a microfluidics channel, it can be determined that molecular recognition of α-chymotrypsin in hydrous surroundings at two different pH values (3.6 and 6.3) follows two distinctly different pathways. Whereas one corroborates an induced-fit model (pH 3.6), the other one (pH 6.3) is consistent with the selected-fit model of biomolecular recognition. The role of massive structural perturbations of differential recognition pathways could be ruled out by earlier XRD studies, rather was consistent with the femtosecond-resolved observation of dynamic flexibility of the protein at different pH values. At low concentrations of ligands, the selected-fit model dominates, whereas increasing the ligand concentration leads to the induced-fit model. From molecular modelling and experimental results, the timescale associated with the conformational flexibility of the protein plays a key role in the selection of a pathway in biomolecular recognition. PMID:26367136

  9. Current perspectives of the signaling pathways directing neural crest induction.

    PubMed

    Stuhlmiller, Timothy J; García-Castro, Martín I

    2012-11-01

    The neural crest is a migratory population of embryonic cells with a tremendous potential to differentiate and contribute to nearly every organ system in the adult body. Over the past two decades, an incredible amount of research has given us a reasonable understanding of how these cells are generated. Neural crest induction involves the combinatorial input of multiple signaling pathways and transcription factors, and is thought to occur in two phases from gastrulation to neurulation. In the first phase, FGF and Wnt signaling induce NC progenitors at the border of the neural plate, activating the expression of members of the Msx, Pax, and Zic families, among others. In the second phase, BMP, Wnt, and Notch signaling maintain these progenitors and bring about the expression of definitive NC markers including Snail2, FoxD3, and Sox9/10. In recent years, additional signaling molecules and modulators of these pathways have been uncovered, creating an increasingly complex regulatory network. In this work, we provide a comprehensive review of the major signaling pathways that participate in neural crest induction, with a focus on recent developments and current perspectives. We provide a simplified model of early neural crest development and stress similarities and differences between four major model organisms: Xenopus, chick, zebrafish, and mouse. PMID:22547091

  10. Direct Observation of the Formation Pathway of [Mo132] Keplerates.

    PubMed

    Biswas, Subharanjan; Melgar, Dolores; Srimany, Amitava; Rodríguez-Fortea, Antonio; Pradeep, Thalappil; Bo, Carles; Poblet, Josep M; Roy, Soumyajit

    2016-09-01

    The formation pathway of a closed spherical cluster [Mo132], starting from a library of building blocks of molybdate anions, has been reported. Electrospray ionization mass spectrometry, Raman spectroscopy, and theoretical studies describe the formation of such a complex cluster from a reduced and acidified aqueous solution of molybdate. Understanding the emergence of such an enormous spherical model cluster may lead to the design of new clusters in the future. Formation of such a highly symmetric cluster is principally controlled by charge balance and the emergence of more symmetric structures at the expense of less symmetric ones. PMID:27560767

  11. [Distinct roles of the direct and indirect pathways in the basal ganglia circuit mechanism].

    PubMed

    Morita, Makiko; Hikida, Takatoshi

    2015-11-01

    The basal ganglia are key neural substrates that control not only motor balance but also emotion, motivation, cognition, learning, and decision-making. Dysfunction of the basal ganglia leads to neurodegenerative diseases (e.g. Parkinson's disease and Huntington's disease) and psychiatric disorders (e.g. drug addiction, schizophrenia, and depression). In the basal ganglia circuit, there are two important pathways: the direct and indirect striatal pathways. Recently, new molecular techniques that activate or inactive selectively the direct or indirect pathway neurons have revealed the function of each pathway. Here we review the distinct roles of the direct and indirect striatal pathways in brain function and drug addiction. We have developed a reversible neurotransmission blocking technique, in which transmission of each pathway is selectively blocked by specific expression of transmission-blocking tetanus toxin, and revealed that the activation of D1 receptors in the direct pathway is critical for reward learning/cocaine addiction, and that the inactivation of D2 receptors is critical for aversive learning/learning flexibility. We propose a new circuit mechanism by which the dopaminergic input from the ventral tegmental area can switch the direct and indirect pathways in the nucleus accumbens. These basal ganglia circuit mechanisms will give us insights into the pathophysiology of mental diseases. PMID:26785520

  12. Determination of the aflatoxin M1 (AFM1) from milk by direct analysis in real time - mass spectrometry (DART-MS)

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Certain fungi that grow on crops can produce aflatoxins, which are highly carcinogenic. One of these, aflatoxin B1 can be metabolized by mammals to aflatoxin M1, a form that retains potent carcinogenicity and which can be excreted into milk. Direct analysis in real time (DART) ionization coupled to ...

  13. Direct evidence for a xylose metabolic pathway in Saccharomyces cerevisiae

    SciTech Connect

    Batt, C.A.; Carvallo, S.; Easson, D.D.; Akedo, M.; Sinskey, A.J.

    1986-04-01

    Xylose transport, xylose reductase, and xylitol dehydrogenase activities are demonstrated in Saccharomyces cerevisiae. The enzymes in the xylose catabolic pathway necessary for the conversion of xylose xylulose are present, although S. cerevisiae cannot grow on xylose as a sole carbon source. Xylose transport is less efficient than glucose transport, and its rate is dependent upon aeration. Xylose reductase appears to be a xylose inducible enzyme and xylitol dehydrogenase activity is constitutive, although both are repressed by glucose. Both xylose reductase and xylitol dehydrogenase activities are five- to tenfold lower in S. cerevisie as compared to Candida utilis. In vivo conversion of /sup 14/C-xylose in S. cerevisiage is demonstrated and xylitol is detected, although no significant levels of any other /sup 14/C-labeled metabolites (e.g., ethanol) are observed. 22 references.

  14. Bidirectional Plasticity in Striatonigral Synapses: A Switch to Balance Direct and Indirect Basal Ganglia Pathways

    ERIC Educational Resources Information Center

    Aceves, Jose J.; Rueda-Orozco, Pavel E.; Hernandez-Martinez, Ricardo; Galarraga, Elvira; Bargas, Jose

    2011-01-01

    There is no hypothesis to explain how direct and indirect basal ganglia (BG) pathways interact to reach a balance during the learning of motor procedures. Both pathways converge in the substantia nigra pars reticulata (SNr) carrying the result of striatal processing. Unfortunately, the mechanisms that regulate synaptic plasticity in striatonigral…

  15. Concurrent Activation of Striatal Direct and Indirect Pathways During Action Initiation

    PubMed Central

    Cui, Guohong; Jun, Sang Beom; Jin, Xin; Pham, Michael D.

    2014-01-01

    Summary The basal ganglia are subcortical nuclei that control voluntary actions, and are affected by a number of debilitating neurological disorders1–4. The prevailing model of basal ganglia function proposes that two orthogonal projection circuits originating from distinct populations of spiny projection neurons (SPNs) in the striatum5,6 - the so-called direct and indirect pathways - have opposing effects on movement: while activity of direct-pathway SPNs purportedly facilitates movement, activity of indirect-pathway SPNs inhibits movement1,2. This model has been difficult to test due to the lack of methods to selectively measure the activity of direct- and indirect-pathway SPNs in freely moving animals. We developed a novel in-vivo method that allowed us to specifically measure direct- and indirect-pathway SPN activity using Cre-dependent viral expression of the genetically encoded calcium indicator (GECI) GCAMP3 in the dorsal striatum of D1-Cre (direct-pathway specific6,7) and A2A-Cre (indirect-pathway specific8,9) mice10. Using fiber optics and time-correlated single photon counting (TCSPC) in mice performing an operant task, we observed transient increases in neural activity in both direct- and indirect-pathway SPNs when animals initiated actions, but not when they were inactive. Concurrent activation of SPNs from both pathways in one hemisphere preceded the initiation of contraversive movements, and predicted the occurrence of specific movements within 500 ms. These observations challenge the classical view of basal ganglia function, and may have implications for understanding the origin of motor symptoms in basal ganglia disorders. PMID:23354054

  16. Direct AFM force measurements between air bubbles in aqueous polydisperse sodium poly(styrene sulfonate) solutions: effect of collision speed, polyelectrolyte concentration and molar mass.

    PubMed

    Browne, Christine; Tabor, Rico F; Grieser, Franz; Dagastine, Raymond R

    2015-07-01

    Interactions between colliding air bubbles in aqueous solutions of polydisperse sodium poly(styrene sulfonate) (NaPSS) using direct force measurements were studied. The forces measured with deformable interfaces were shown to be more sensitive to the presence of the polyelectrolytes when compared to similar measurements using rigid interfaces. The experimental factors that were examined were NaPSS concentration, bubble collision velocity and polyelectrolyte molar mass. These measurements were then compared with an analytical model based on polyelectrolyte scaling theory in order to explain the effects of concentration and bubble deformation on the interaction between bubbles. Typically structural forces from the presence of monodisperse polyelectrolyte between interacting surfaces may be expected, however, it was found that the polydispersity in molar mass resulted in the structural forces to be smoothed and only a depletion interaction was able to be measured between interacting bubbles. It was found that an increase in number density of NaPSS molecules resulted in an increase in the magnitude of the depletion interaction. Conversely this interaction was overwhelmed by an increase in the fluid flow in the system at higher bubble collision velocities. Polymer molar mass dispersity plays a significant role in the interactions present between the bubbles and has implications that also affect the polyelectrolyte overlap concentration of the solution. Further understanding of these implications can be expected to play a role in the improvement in operations in such fields as water treatment and mineral processing where polyelectrolytes are used extensively. PMID:25596872

  17. Differential innervation of direct- and indirect-pathway striatal projection neurons

    PubMed Central

    Wall, Nicholas R.; De La Parra, Mauricio; Callaway, Edward M.; Kreitzer, Anatol C.

    2013-01-01

    SUMMARY The striatum integrates information from multiple brain regions to shape motor learning. The two major projection cell types in striatum target different downstream basal ganglia targets and have opposing effects on motivated behavior, yet differential innervation of these neuronal subtypes is not well understood. To examine whether input specificity provides a substrate for information segregation in these circuits, we used a monosynaptic rabies virus system to generate brain-wide maps of neurons that form synapses with direct- or indirect-pathway striatal projection neurons. We discovered that sensory cortical and limbic structures preferentially innervated the direct pathway, whereas motor cortex preferentially targeted the indirect pathway. Thalamostriatal input, dopaminergic input, as well as input from specific cortical layers, was similar onto both pathways. We also confirm synaptic innervation of striatal projection neurons by the raphe and pedunculopontine nuclei. Together, these findings provide a framework for guiding future studies of basal ganglia circuit function. PMID:23810541

  18. Atomic force microscopy combined with optical tweezers (AFM/OT)

    NASA Astrophysics Data System (ADS)

    Pierini, F.; Zembrzycki, K.; Nakielski, P.; Pawłowska, S.; Kowalewski, T. A.

    2016-02-01

    The role of mechanical properties is essential to understand molecular, biological materials, and nanostructures dynamics and interaction processes. Atomic force microscopy (AFM) is the most commonly used method of direct force evaluation, but due to its technical limitations this single probe technique is unable to detect forces with femtonewton resolution. In this paper we present the development of a combined atomic force microscopy and optical tweezers (AFM/OT) instrument. The focused laser beam, on which optical tweezers are based, provides us with the ability to manipulate small dielectric objects and to use it as a high spatial and temporal resolution displacement and force sensor in the same AFM scanning zone. We demonstrate the possibility to develop a combined instrument with high potential in nanomechanics, molecules manipulation and biological studies. AFM/OT equipment is described and characterized by studying the ability to trap dielectric objects and quantifying the detectable and applicable forces. Finally, optical tweezers calibration methods and instrument applications are given.

  19. Different Roles of Direct and Indirect Frontoparietal Pathways for Individual Working Memory Capacity.

    PubMed

    Ekman, Matthias; Fiebach, Christian J; Melzer, Corina; Tittgemeyer, Marc; Derrfuss, Jan

    2016-03-01

    The ability to temporarily store and manipulate information in working memory is a hallmark of human intelligence and differs considerably across individuals, but the structural brain correlates underlying these differences in working memory capacity (WMC) are only poorly understood. In two separate studies, diffusion MRI data and WMC scores were collected for 70 and 109 healthy individuals. Using a combination of probabilistic tractography and network analysis of the white matter tracts, we examined whether structural brain network properties were predictive of individual WMC. Converging evidence from both studies showed that lateral prefrontal cortex and posterior parietal cortex of high-capacity individuals are more densely connected compared with low-capacity individuals. Importantly, our network approach was further able to dissociate putative functional roles associated with two different pathways connecting frontal and parietal regions: a corticocortical pathway and a subcortical pathway. In Study 1, where participants were required to maintain and update working memory items, the connectivity of the direct and indirect pathway was predictive of WMC. In contrast, in Study 2, where participants were required to maintain working memory items without updating, only the connectivity of the direct pathway was predictive of individual WMC. Our results suggest an important dissociation in the circuitry connecting frontal and parietal regions, where direct frontoparietal connections might support storage and maintenance, whereas subcortically mediated connections support the flexible updating of working memory content. PMID:26961945

  20. A Bayesian Approach to Pathway Analysis by Integrating Gene–Gene Functional Directions and Microarray Data

    PubMed Central

    Zhao, Yifang; Chen, Ming-Hui; Pei, Baikang; Rowe, David; Shin, Dong-Guk; Xie, Wangang; Yu, Fang; Kuo, Lynn

    2012-01-01

    Many statistical methods have been developed to screen for differentially expressed genes associated with specific phenotypes in the microarray data. However, it remains a major challenge to synthesize the observed expression patterns with abundant biological knowledge for more complete understanding of the biological functions among genes. Various methods including clustering analysis on genes, neural network, Bayesian network and pathway analysis have been developed toward this goal. In most of these procedures, the activation and inhibition relationships among genes have hardly been utilized in the modeling steps. We propose two novel Bayesian models to integrate the microarray data with the putative pathway structures obtained from the KEGG database and the directional gene–gene interactions in the medical literature. We define the symmetric Kullback–Leibler divergence of a pathway, and use it to identify the pathway(s) most supported by the microarray data. Monte Carlo Markov Chain sampling algorithm is given for posterior computation in the hierarchical model. The proposed method is shown to select the most supported pathway in an illustrative example. Finally, we apply the methodology to a real microarray data set to understand the gene expression profile of osteoblast lineage at defined stages of differentiation. We observe that our method correctly identifies the pathways that are reported to play essential roles in modulating bone mass. PMID:23482678

  1. Direct Visualization of the Perforant Pathway in the Human Brain with Ex Vivo Diffusion Tensor Imaging

    PubMed Central

    Augustinack, Jean C.; Helmer, Karl; Huber, Kristen E.; Kakunoori, Sita; Zöllei, Lilla; Fischl, Bruce

    2010-01-01

    Ex vivo magnetic resonance imaging yields high resolution images that reveal detailed cerebral anatomy and explicit cytoarchitecture in the cerebral cortex, subcortical structures, and white matter in the human brain. Our data illustrate neuroanatomical correlates of limbic circuitry with high resolution images at high field. In this report, we have studied ex vivo medial temporal lobe samples in high resolution structural MRI and high resolution diffusion MRI. Structural and diffusion MRIs were registered to each other and to histological sections stained for myelin for validation of the perforant pathway. We demonstrate probability maps and fiber tracking from diffusion tensor data that allows the direct visualization of the perforant pathway. Although it is not possible to validate the DTI data with invasive measures, results described here provide an additional line of evidence of the perforant pathway trajectory in the human brain and that the perforant pathway may cross the hippocampal sulcus. PMID:20577631

  2. PREFACE: Non-contact AFM Non-contact AFM

    NASA Astrophysics Data System (ADS)

    Giessibl, Franz J.; Morita, Seizo

    2012-02-01

    This special issue is focussed on high resolution non-contact atomic force microscopy (AFM). Non-contact atomic force microscopy was established approximately 15 years ago as a tool to image conducting and insulating surfaces with atomic resolution. Since 1998, an annual international conference has taken place, and although the proceedings of these conferences are a useful source of information, several key developments warrant devoting a special issue to this subject. In the theoretic field, the possibility of supplementing established techniques such as scanning tunneling microscopy (STM) and Kelvin probe microscopy with atomically resolved force micrsoscopy poses many challenges in the calculation of contrast and contrast reversal. The surface science of insulators, self-assembled monolayers and adsorbates on insulators is a fruitful field for the application of non-contact AFM: several articles in this issue are devoted to these subjects. Atomic imaging and manipulation have been pioneered using STM, but because AFM allows the measurement of forces, AFM has had a profound impact in this field as well. Three-dimensional force spectroscopy has allowed many important insights into surface science. In this issue a combined 3D tunneling and force microscopy is introduced. Non-contact AFM typically uses frequency modulation to measure force gradients and was initially used mainly in a vacuum. As can be seen in this issue, frequency modulation is now also used in ambient conditions, allowing better spatial and force resolution. We thank all of the contributors for their time and efforts in making this special issue possible. We are also very grateful to the staff of IOP Publishing for handling the administrative aspects and for steering the refereeing process. Non-contact AFM contents Relation between the chemical force and the tunnelling current in atomic point contacts: a simple model Pavel Jelínek, Martin Ondrácek and Fernando Flores Theoretical simulation of

  3. Modality-Based Organization of Ascending Somatosensory Axons in the Direct Dorsal Column Pathway

    PubMed Central

    Niu, Jingwen; Ding, Long; Li, Jian J.; Kim, Hyukmin; Liu, Jiakun; Li, Haipeng; Moberly, Andrew; Badea, Tudor C.; Duncan, Ian D.; Son, Young-Jin; Scherer, Steven S.

    2013-01-01

    The long-standing doctrine regarding the functional organization of the direct dorsal column (DDC) pathway is the “somatotopic map” model, which suggests that somatosensory afferents are primarily organized by receptive field instead of modality. Using modality-specific genetic tracing, here we show that ascending mechanosensory and proprioceptive axons, two main types of the DDC afferents, are largely segregated into a medial–lateral pattern in the mouse dorsal column and medulla. In addition, we found that this modality-based organization is likely to be conserved in other mammalian species, including human. Furthermore, we identified key morphological differences between these two types of afferents, which explains how modality segregation is formed and why a rough “somatotopic map” was previously detected. Collectively, our results establish a new functional organization model for the mammalian direct dorsal column pathway and provide insight into how somatotopic and modality-based organization coexist in the central somatosensory pathway. PMID:24198362

  4. Identification of a Specific Maleate Hydratase in the Direct Hydrolysis Route of the Gentisate Pathway

    PubMed Central

    Liu, Kun; Xu, Ying

    2015-01-01

    In contrast to the well-characterized and more common maleylpyruvate isomerization route of the gentisate pathway, the direct hydrolysis route occurs rarely and remains unsolved. In Pseudomonas alcaligenes NCIMB 9867, two gene clusters, xln and hbz, were previously proposed to be involved in gentisate catabolism, and HbzF was characterized as a maleylpyruvate hydrolase converting maleylpyruvate to maleate and pyruvate. However, the complete degradation pathway of gentisate through direct hydrolysis has not been characterized. In this study, we obtained from the NCIMB culture collection a Pseudomonas alcaligenes spontaneous mutant strain that lacked the xln cluster and designated the mutant strain SponMu. The hbz cluster in strain SponMu was resequenced, revealing the correct location of the stop codon for hbzI and identifying a new gene, hbzG. HbzIJ was demonstrated to be a maleate hydratase consisting of large and small subunits, stoichiometrically converting maleate to enantiomerically pure d-malate. HbzG is a glutathione-dependent maleylpyruvate isomerase, indicating the possible presence of two alternative pathways of maleylpyruvate catabolism. However, the hbzF-disrupted mutant could still grow on gentisate, while disruption of hbzG prevented this ability, indicating that the direct hydrolysis route was not a complete pathway in strain SponMu. Subsequently, a d-malate dehydrogenase gene was introduced into the hbzG-disrupted mutant, and the engineered strain was able to grow on gentisate via the direct hydrolysis route. This fills a gap in our understanding of the direct hydrolysis route of the gentisate pathway and provides an explanation for the high yield of d-malate from maleate by this d-malate dehydrogenase-deficient natural mutant. PMID:26070679

  5. Critical Roles of the Direct GABAergic Pallido-cortical Pathway in Controlling Absence Seizures.

    PubMed

    Chen, Mingming; Guo, Daqing; Li, Min; Ma, Tao; Wu, Shengdun; Ma, Jingling; Cui, Yan; Xia, Yang; Xu, Peng; Yao, Dezhong

    2015-10-01

    The basal ganglia (BG), serving as an intermediate bridge between the cerebral cortex and thalamus, are believed to play crucial roles in controlling absence seizure activities generated by the pathological corticothalamic system. Inspired by recent experiments, here we systematically investigate the contribution of a novel identified GABAergic pallido-cortical pathway, projecting from the globus pallidus externa (GPe) in the BG to the cerebral cortex, to the control of absence seizures. By computational modelling, we find that both increasing the activation of GPe neurons and enhancing the coupling strength of the inhibitory pallido-cortical pathway can suppress the bilaterally synchronous 2-4 Hz spike and wave discharges (SWDs) during absence seizures. Appropriate tuning of several GPe-related pathways may also trigger the SWD suppression, through modulating the activation level of GPe neurons. Furthermore, we show that the previously discovered bidirectional control of absence seizures due to the competition between other two BG output pathways also exists in our established model. Importantly, such bidirectional control is shaped by the coupling strength of this direct GABAergic pallido-cortical pathway. Our work suggests that the novel identified pallido-cortical pathway has a functional role in controlling absence seizures and the presented results might provide testable hypotheses for future experimental studies. PMID:26496656

  6. Critical Roles of the Direct GABAergic Pallido-cortical Pathway in Controlling Absence Seizures

    PubMed Central

    Li, Min; Ma, Tao; Wu, Shengdun; Ma, Jingling; Cui, Yan; Xia, Yang; Xu, Peng; Yao, Dezhong

    2015-01-01

    The basal ganglia (BG), serving as an intermediate bridge between the cerebral cortex and thalamus, are believed to play crucial roles in controlling absence seizure activities generated by the pathological corticothalamic system. Inspired by recent experiments, here we systematically investigate the contribution of a novel identified GABAergic pallido-cortical pathway, projecting from the globus pallidus externa (GPe) in the BG to the cerebral cortex, to the control of absence seizures. By computational modelling, we find that both increasing the activation of GPe neurons and enhancing the coupling strength of the inhibitory pallido-cortical pathway can suppress the bilaterally synchronous 2–4 Hz spike and wave discharges (SWDs) during absence seizures. Appropriate tuning of several GPe-related pathways may also trigger the SWD suppression, through modulating the activation level of GPe neurons. Furthermore, we show that the previously discovered bidirectional control of absence seizures due to the competition between other two BG output pathways also exists in our established model. Importantly, such bidirectional control is shaped by the coupling strength of this direct GABAergic pallido-cortical pathway. Our work suggests that the novel identified pallido-cortical pathway has a functional role in controlling absence seizures and the presented results might provide testable hypotheses for future experimental studies. PMID:26496656

  7. Liquid contact resonance AFM: analytical models, experiments, and limitations

    NASA Astrophysics Data System (ADS)

    Parlak, Zehra; Tu, Qing; Zauscher, Stefan

    2014-11-01

    Contact resonance AFM (CR-AFM) is a scanning probe microscopy technique that utilizes the contact resonances of the AFM cantilever for concurrent imaging of topography and surface stiffness. The technique has not been used in liquid until recently due to analytical and experimental difficulties, associated with viscous damping of cantilever vibrations and fluid loading effects. To address these difficulties, (i) an analytical approach for contact resonances in liquid is developed, and (ii) direct excitation of the contact resonances is demonstrated by actuating the cantilever directly in a magnetic field. By implementing the analytical approach and the direct actuation through magnetic particles, quantitative stiffness imaging on surfaces with a wide range of stiffness can be achieved in liquid with soft cantilevers and low contact forces.

  8. Quantitative Profiling of DNA Damage and Apoptotic Pathways in UV Damaged Cells Using PTMScan Direct

    PubMed Central

    Stokes, Matthew P.; Silva, Jeffrey C.; Jia, Xiaoying; Lee, Kimberly A.; Polakiewicz, Roberto D.; Comb, Michael J.

    2013-01-01

    Traditional methods for analysis of peptides using liquid chromatography and tandem mass spectrometry (LC-MS/MS) lack the specificity to comprehensively monitor specific biological processes due to the inherent duty cycle limitations of the MS instrument and the stochastic nature of the analytical platform. PTMScan Direct is a novel, antibody-based method that allows quantitative LC-MS/MS profiling of specific peptides from proteins that reside in the same signaling pathway. New PTMScan Direct reagents have been produced that target peptides from proteins involved in DNA Damage/Cell Cycle and Apoptosis/Autophagy pathways. Together, the reagents provide access to 438 sites on 237 proteins in these signaling cascades. These reagents have been used to profile the response to UV damage of DNA in human cell lines. UV damage was shown to activate canonical DNA damage response pathways through ATM/ATR-dependent signaling, stress response pathways and induce the initiation of apoptosis, as assessed by an increase in the abundance of peptides corresponding to cleaved, activated caspases. These data demonstrate the utility of PTMScan Direct as a multiplexed assay for profiling specific cellular responses to various stimuli, such as UV damage of DNA. PMID:23344034

  9. Directional Regulation of Enzyme Pathways through the Control of Substrate Channeling on a DNA Origami Scaffold.

    PubMed

    Ke, Guoliang; Liu, Minghui; Jiang, Shuoxing; Qi, Xiaodong; Yang, Yuhe Renee; Wootten, Shaun; Zhang, Fei; Zhu, Zhi; Liu, Yan; Yang, Chaoyong James; Yan, Hao

    2016-06-20

    Artificial multi-enzyme systems with precise and dynamic control over the enzyme pathway activity are of great significance in bionanotechnology and synthetic biology. Herein, we exploit a spatially addressable DNA nanoplatform for the directional regulation of two enzyme pathways (G6pDH-MDH and G6pDH-LDH) through the control of NAD(+) substrate channeling by specifically shifting NAD(+) between the two enzyme pairs. We believe that this concept will be useful for the design of regulatory biological circuits for synthetic biology and biomedicine. PMID:27159899

  10. Turning over renal osteodystrophy dogma: direct actions of FGF23 on osteoblast β-catenin pathway.

    PubMed

    Schiavi, Susan C; Moysés, Rosa M A

    2016-07-01

    Although recognized as a major complication of chronic kidney disease (CKD), the pathophysiology of the CKD-related mineral and bone disorder (CKD-MBD) is not completely understood. Recently, the inhibition of Wnt/β-catenin pathway in osteocytes by sclerostin has been shown to play a role in CKD-MBD. The study by Carrilo-Lopez et al. confirms this inhibition in an experimental model of CKD. Moreover, they describe direct actions of FGF23-Klotho on osteoblasts, increasing the expression of DKK1, another Wnt/β-catenin pathway inhibitor. PMID:27312441

  11. Confirmatory evidence for a direct parasympathetic pathway to internal eye structures.

    PubMed Central

    Parelman, J J; Fay, M T; Burde, R M

    1984-01-01

    The hypothesis that a direct neural pathway exists between the midbrain and the intrinsic musculature was investigated. Wheat germ agglutinin labeled with HRP and WG-HRP were injected intraocularly into the ciliary muscle of rabbits and cynomolgus monkeys, and their retrograde movement was monitored. Sparse, but definite labeling was found in the AM nuclei, the EW nuclei, and Perlia's nucleus of the monkey, confirming the existence of a nonsynapsing pathway between intraocular structures and the midbrain in the primate. Images FIGURE 1 A FIGURE 1 B FIGURE 1 C PMID:6549517

  12. Direct observation of multimer stabilization in the mechanical unfolding pathway of a protein undergoing oligomerization.

    PubMed

    Scholl, Zackary N; Yang, Weitao; Marszalek, Piotr E

    2015-02-24

    Understanding how protein oligomerization affects the stability of monomers in self-assembled structures is crucial to the development of new protein-based nanomaterials and protein cages for drug delivery. Here, we use single-molecule force spectroscopy (AFM-SMFS), protein engineering, and computer simulations to evaluate how dimerization and tetramerization affects the stability of the monomer of Streptavidin, a model homotetrameric protein. The unfolding force directly relates to the folding stability, and we find that monomer of Streptavidin is mechanically stabilized by 40% upon dimerization, and that it is stabilized an additional 24% upon tetramerization. We also find that biotin binding increases stability by another 50% as compared to the apo-tetrameric form. We used the distribution of unfolding forces to extract properties of the underlying energy landscape and found that the distance to the transition state is decreased and the barrier height is increased upon multimerization. Finally, we investigated the origin of the strengthening by ligand binding. We found that, rather than being strengthened through intramolecular contacts, it is strengthened due to the contacts provided by the biotin-binding loop that crosses the interface between the dimers. PMID:25639698

  13. Prenatal alcohol and other early childhood adverse exposures: Direct and indirect pathways to adolescent drinking.

    PubMed

    Cornelius, Marie D; De Genna, Natacha M; Goldschmidt, Lidush; Larkby, Cynthia; Day, Nancy L

    2016-01-01

    We examined direct and indirect pathways between adverse environmental exposures during gestation and childhood and drinking in mid-adolescence. Mothers and their offspring (n=917 mother/child dyads) were followed prospectively from second trimester to a 16-year follow-up assessment. Interim assessments occurred at delivery, 6, 10, and 14years. Adverse environmental factors included gestational exposures to alcohol, tobacco, and marijuana, exposures to childhood maltreatment and violence, maternal psychological symptoms, parenting practices, economic and home environments, and demographic characteristics of the mother and child. Indirect effects of early child behavioral characteristics including externalizing, internalizing activity, attention, and impulsivity were also examined. Polytomous logistic regression analyses were used to evaluate direct effects of adverse environmental exposures with level of adolescent drinking. Structural equation modeling (SEM) was applied to simultaneously estimate the relation between early adversity variables, childhood characteristics, and drinking level at age 16 while controlling for significant covariates. Level of drinking among the adolescent offspring was directly predicted by prenatal exposure to alcohol, less parental strictness, and exposures to maltreatment and violence during childhood. Whites and offspring with older mothers were more likely to drink at higher levels. There was a significant indirect effect between childhood exposure to violence and adolescent drinking via childhood externalizing behavior problems. All other hypothesized indirect pathways were not significant. Thus most of the early adversity measures directly predicted adolescent drinking and did not operate via childhood behavioral dysregulation characteristics. These results highlight the importance of adverse environmental exposures on pathways to adolescent drinking. PMID:26994529

  14. Prenatal alcohol and other early childhood adverse exposures: Direct and indirect pathways to adolescent drinking

    PubMed Central

    Cornelius, Marie D.; De Genna, Natacha M.; Goldschmidt, Lidush; Larkby, Cynthia; Day, Nancy L.

    2016-01-01

    We examined direct and indirect pathways between adverse environmental exposures during gestation and childhood and drinking in mid-adolescence. Mothers and their offspring (n = 917 mother/child dyads) were followed prospectively from second trimester to a 16-year follow-up assessment. Interim assessments occurred at delivery, 6, 10, and 14 years. Adverse environmental factors included gestational exposures to alcohol, tobacco, and marijuana, exposures to childhood maltreatment and violence, maternal psychological symptoms, parenting practices, economic and home environments, and demographic characteristics of the mother and child. Indirect effects of early child behavioral characteristics including externalizing, internalizing activity, attention, and impulsivity were also examined. Polytomous logistic regression analyses were used to evaluate direct effects of adverse environmental exposures with level of adolescent drinking. Structural equation modeling (SEM) was applied to simultaneously estimate the relation between early adversity variables, childhood characteristics, and drinking level at age 16 while controlling for significant covariates. Level of drinking among the adolescent offspring was directly predicted by prenatal exposure to alcohol, less parental strictness, and exposures to maltreatment and violence during childhood. Whites and offspring with older mothers were more likely to drink at higher levels. There was a significant indirect effect between childhood exposure to violence and adolescent drinking via childhood externalizing behavior problems. All other hypothesized indirect pathways were not significant. Thus most of the early adversity measures directly predicted adolescent drinking and did not operate via childhood behavioral dysregulation characteristics. These results highlight the importance of adverse environmental exposures on pathways to adolescent drinking. PMID:26994529

  15. [Direct cryothermal ablation eliminates conduction of the slow pathway without inducing ectopic rhythms].

    PubMed

    Márquez, Manlio F; Colín, Luis; Iturralde, Pedro; Nava, Santiago; González, Eric; Rodríguez, Gerardo; Gómez, Jorge; Salica, Gabriel; Cossío, Jorge; Hermosillo, Antonio G; Cárdenas, Manuel

    2005-01-01

    Radiofrequency catheter ablation of atrioventricular nodal reentrant tachycardia is based on the elimination of conduction of slow or fast intranodal pathway. To avoid potential atrioventricular (AV) block, a new technology has been developed, cryothermal ablation. We report a case of AV nodal reentrant tachycardia in whom direct cryoablation, without previous ice mapping, was successfully performed. Interestingly and as previously described, cryotherapy did not induce ectopic rhythms, the conventional surrogate during radiofrequency ablation. PMID:15909749

  16. Homology-directed Fanconi anemia pathway crosslink repair is dependent on DNA replication

    PubMed Central

    Nakanishi, Koji; Cavallo, Francesca; Perrouault, Loïc; Giovannangeli, Carine; Moynahan, Mary Ellen; Barchi, Marco; Brunet, Erika; Jasin, Maria

    2012-01-01

    Homologous recombination (also termed homology-directed repair, HDR) is a major pathway for the repair of DNA interstrand crosslinks (ICLs) in mammalian cells. Cells from Fanconi anemia (FA) patients are characterized by extreme ICL sensitivity, but their reported defect in HDR is mild. Here, we examined ICL-induced HDR using a GFP reporter and observed a profound defect in ICL-induced HDR in FA cells, but only when the reporter could replicate. PMID:21423196

  17. Exploration of molecular pathways mediating electric field-directed Schwann cell migration by RNA-seq.

    PubMed

    Yao, Li; Li, Yongchao; Knapp, Jennifer; Smith, Peter

    2015-07-01

    In peripheral nervous systems, Schwann cells wrap around axons of motor and sensory neurons to form the myelin sheath. Following spinal cord injury, Schwann cells regenerate and migrate to the lesion and are involved in the spinal cord regeneration process. Transplantation of Schwann cells into injured neural tissue results in enhanced spinal axonal regeneration. Effective directional migration of Schwann cells is critical in the neural regeneration process. In this study, we report that Schwann cells migrate anodally in an applied electric field (EF). The directedness and displacement of anodal migration increased significantly when the strength of the EF increased from 50 mV/mm to 200 mV/mm. The EF did not significantly affect the cell migration speed. To explore the genes and signaling pathways that regulate cell migration in EFs, we performed a comparative analysis of differential gene expression between cells stimulated with an EF (100 mV/mm) and those without using next-generation RNA sequencing, verified by RT-qPCR. Based on the cut-off criteria (FC > 1.2, q < 0.05), we identified 1,045 up-regulated and 1,636 down-regulated genes in control cells versus EF-stimulated cells. A Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis found that compared to the control group, 21 pathways are down-regulated, while 10 pathways are up-regulated. Differentially expressed genes participate in multiple cellular signaling pathways involved in the regulation of cell migration, including pathways of regulation of actin cytoskeleton, focal adhesion, and PI3K-Akt. PMID:25557037

  18. Exploring the transfer of recent plant photosynthates to soil microbes: mycorrhizal pathway vs direct root exudation

    PubMed Central

    Kaiser, Christina; Kilburn, Matt R; Clode, Peta L; Fuchslueger, Lucia; Koranda, Marianne; Cliff, John B; Solaiman, Zakaria M; Murphy, Daniel V

    2015-01-01

    Plants rapidly release photoassimilated carbon (C) to the soil via direct root exudation and associated mycorrhizal fungi, with both pathways promoting plant nutrient availability. This study aimed to explore these pathways from the root's vascular bundle to soil microbial communities. Using nanoscale secondary ion mass spectrometry (NanoSIMS) imaging and 13C-phospho- and neutral lipid fatty acids, we traced in-situ flows of recently photoassimilated C of 13CO2-exposed wheat (Triticum aestivum) through arbuscular mycorrhiza (AM) into root- and hyphae-associated soil microbial communities. Intraradical hyphae of AM fungi were significantly 13C-enriched compared to other root-cortex areas after 8 h of labelling. Immature fine root areas close to the root tip, where AM features were absent, showed signs of passive C loss and co-location of photoassimilates with nitrogen taken up from the soil solution. A significant and exclusively fresh proportion of 13C-photosynthates was delivered through the AM pathway and was utilised by different microbial groups compared to C directly released by roots. Our results indicate that a major release of recent photosynthates into soil leave plant roots via AM intraradical hyphae already upstream of passive root exudations. AM fungi may act as a rapid hub for translocating fresh plant C to soil microbes. PMID:25382456

  19. AFM Structural Characterization of Drinking Water Biofilm under Physiological Conditions

    EPA Science Inventory

    Due to the complexity of mixed culture drinking water biofilm, direct visual observation under in situ conditions has been challenging. In this study, atomic force microscopy (AFM) revealed the three dimensional morphology and arrangement of drinking water relevant biofilm in air...

  20. GPIM AF-M315E Propulsion System

    NASA Technical Reports Server (NTRS)

    Spores, Ronald A.; Masse, Robert; Kimbrel, Scott; McLean, Chris

    2014-01-01

    The NASA Space Technology mission Directorate's (STMD) Green Propellant Infusion Mission (GPIM) Technology Demonstration Mission (TDM) will demonstrate an operational AF-M315E green propellant propulsion system. Aerojet-Rocketdyne is responsible for the development of the propulsion system payload. This paper statuses the propulsion system module development, including thruster design and system design; Initial test results for the 1N engineering model thruster are presented. The culmination of this program will be high-performance, green AF-M315E propulsion system technology at TRL 7+, with components demonstrated to TRL 9, ready for direct infusion to a wide range of applications for the space user community.

  1. Direct Hospital Cost of Outcome Pathways in Implant-Based Reconstruction with Acellular Dermal Matrices

    PubMed Central

    Qureshi, Ali A.; Broderick, Kristen; Funk, Susan; Reaven, Nancy; Tenenbaum, Marissa M.

    2016-01-01

    Background: Current cost data on tissue expansion followed by exchange for permanent implant (TE/I) reconstruction lack a necessary assessment of the experience of a heterogenous breast cancer patient population and their multiple outcome pathways. We extend our previous analysis to that of direct hospital cost as bundling of payments is likely to follow the changing centralization of cancer care at the hospital level. Methods: We performed a retrospective analysis (2003–2009) of TE/I reconstructions with or without an acellular dermal matrix (ADM), namely Alloderm RTM. Postreconstructive events were analyzed and organized into outcome pathways as previously described. Aggregated and normalized inpatient and outpatient hospital direct costs and physician reimbursement were generated for each outcome pathway with or without ADM. Results: Three hundred sixty-seven patients were analyzed. The average 2-year hospital direct cost per TE/I breast reconstruction patient was $11,862 in the +ADM and $12,319 in the −ADM groups (P > 0.05). Initial reconstructions were costlier in the +ADM ($6,868) than in the −ADM ($5,615) group, but the average cost of subsequent postreconstructive events within 2 years was significantly lower in +ADM ($5,176) than −ADM ($6,704) patients (P < 0.05). When a complication occurred, but reconstruction was still completed within 2 years, greater costs were incurred in the −ADM than in the +ADM group for most scenarios, leading to a net equalization of cost between study groups. Conclusion: Although direct hospital cost is an important factor for resource and fund allocation, it should not remain the sole factor when deciding to use ADM in TE/I reconstruction.

  2. A Direct Cortico-Nigral Pathway as Revealed by Constrained Spherical Deconvolution Tractography in Humans

    PubMed Central

    Cacciola, Alberto; Milardi, Demetrio; Anastasi, Giuseppe P.; Basile, Gianpaolo A.; Ciolli, Pietro; Irrera, Mariangela; Cutroneo, Giuseppina; Bruschetta, Daniele; Rizzo, Giuseppina; Mondello, Stefania; Bramanti, Placido; Quartarone, Angelo

    2016-01-01

    Substantia nigra is an important neuronal structure, located in the ventral midbrain, that exerts a regulatory function within the basal ganglia circuitry through the nigro-striatal pathway. Although its subcortical connections are relatively well-known in human brain, little is known about its cortical connections. The existence of a direct cortico-nigral pathway has been demonstrated in rodents and primates but only hypothesized in humans. In this study, we aimed at evaluating cortical connections of substantia nigra in vivo in human brain by using probabilistic constrained spherical deconvolution (CSD) tractography on magnetic resonance diffusion weighted imaging data. We found that substantia nigra is connected with cerebral cortex as a whole, with the most representative connections involving prefrontal cortex, precentral and postcentral gyri and superior parietal lobule. These results may be relevant for the comprehension of the pathophysiology of several neurological disorders involving substantia nigra, such as parkinson's disease, schizophrenia, and pathological addictions. PMID:27507940

  3. A Direct Cortico-Nigral Pathway as Revealed by Constrained Spherical Deconvolution Tractography in Humans.

    PubMed

    Cacciola, Alberto; Milardi, Demetrio; Anastasi, Giuseppe P; Basile, Gianpaolo A; Ciolli, Pietro; Irrera, Mariangela; Cutroneo, Giuseppina; Bruschetta, Daniele; Rizzo, Giuseppina; Mondello, Stefania; Bramanti, Placido; Quartarone, Angelo

    2016-01-01

    Substantia nigra is an important neuronal structure, located in the ventral midbrain, that exerts a regulatory function within the basal ganglia circuitry through the nigro-striatal pathway. Although its subcortical connections are relatively well-known in human brain, little is known about its cortical connections. The existence of a direct cortico-nigral pathway has been demonstrated in rodents and primates but only hypothesized in humans. In this study, we aimed at evaluating cortical connections of substantia nigra in vivo in human brain by using probabilistic constrained spherical deconvolution (CSD) tractography on magnetic resonance diffusion weighted imaging data. We found that substantia nigra is connected with cerebral cortex as a whole, with the most representative connections involving prefrontal cortex, precentral and postcentral gyri and superior parietal lobule. These results may be relevant for the comprehension of the pathophysiology of several neurological disorders involving substantia nigra, such as parkinson's disease, schizophrenia, and pathological addictions. PMID:27507940

  4. BOREAS AFM-06 Mean Wind Profile Data

    NASA Technical Reports Server (NTRS)

    Wilczak, James; Hall, Forrest G. (Editor); Newcomer, Jeffrey A. (Editor); Smith, David E. (Technical Monitor)

    2000-01-01

    The Boreal Ecosystem-Atmosphere Study (BOREAS) Airborne Fluxes and Meteorology (AFM)-6 team from the National Oceanic and Atmospheric Administration/Environment Technology Laboratory (NOAA/ETL) operated a 915-MHz wind/Radio Acoustic Sounding System (RASS) profiler system in the Southern Study Area (SSA) near the Old Jack Pine (OJP) tower from 21 May 1994 to 20 Sep 1994. The data set provides wind profiles at 38 heights, containing the variables of wind speed; wind direction; and the u-, v-, and w-components of the total wind. The data are stored in tabular ASCII files. The mean wind profile data are available from the Earth Observing System Data and Information System (EOSDIS) Oak Ridge National Laboratory (ORNL) Distributed Active Archive Center (DAAC). The data files are available on a CD-ROM (see document number 20010000884).

  5. Chimerogenesis in estimation of specificity and pathway directions for cytochrome p45017alpha catalyzed reactions.

    PubMed

    Gilep, A A; Estabrook, R W; Usanov, S A

    2004-04-01

    Cytochrome P45017alpha is a key enzyme in steroid hormone biosynthesis. It catalyzes the reaction of 17alpha-hydroxylation of progesterone (P4) and pregnenolone (P5) and the 17,20-lyase reaction resulting in side chain cleavage of C21 steroids to form C19 steroids. Depending on the activity of cytochrome P45017alpha, steroid hormone biosynthesis pathways are directed either for biosynthesis of mineralocorticoids and glucocorticoids or sex hormones. The formation of sex hormones starts from biosynthesis of androstenedione. Androstenedione formation is a result of two reactions: 17,20-lyase reaction of 17alpha-hydroxyprogesterone (Delta4-pathway) and 3beta-hydroxysteroid dehydrogenase/Delta4,Delta5-isomerase reaction using dehydroepiandrosterone as substrate (Delta5-pathway). In case of exclusive direction of the 17,20-lyase reaction either through the Delta4- or the Delta5-pathway, the formation of sex hormones depends more on specificity and activity of 3beta-hydroxysteroid-dehydrogenase/Delta4,Delta5-isomerase. Depending on species, the cytochromes P45017alpha can utilize as a substrate for 17,20-lyase activity Delta4-steroids, Delta5-steroids, or both types of steroids. To identify the structural elements of cytochrome P45017alpha responsible for substrate recognition, in the present work we used exchange of homologous fragments of cytochrome P45017alpha having different types of activities. We engineered more than 10 different types of chimeric cytochrome P45017alpha. Chimeric cytochromes P45017alpha have been expressed in E. coli and purified. The expression of chimeric cytochrome P45017alpha with the point of exchange between exons III and IV results in inability of the recombinant hemeprotein to properly bind heme. The determination of activity of chimeric cytochromes P45017alpha shows that the structural element responsible for switching activity between Delta4- or Delta5-pathway is located in the region of polypeptide chain coded by exons II-V of CYP17 gene

  6. Direct visualization of the murine dorsal cochlear nucleus for optogenetic stimulation of the auditory pathway.

    PubMed

    Kozin, Elliott D; Darrow, Keith N; Hight, Ariel E; Lehmann, Ashton E; Kaplan, Alyson B; Brown, M Christian; Lee, Daniel J

    2015-01-01

    Investigation into the use of virus-mediated gene transfer to arrest or reverse hearing loss has largely been relegated to the peripheral auditory system. Few studies have examined gene transfer to the central auditory system. The dorsal cochlear nucleus (DCN) of the brainstem, which contains second order neurons of the auditory pathway, is a potential site for gene transfer. In this protocol, a technique for direct and maximal exposure of the murine DCN via a posterior fossa approach is demonstrated. This approach allows for either acute or survival surgery. Following direct visualization of the DCN, a host of experiments are possible, including injection of opsins into the cochlear nucleus and subsequent stimulation by an optical fiber coupled to a blue light laser. Other neurophysiology experiments, such as electrical stimulation and neural injector tracings are also feasible. The level of visualization and the duration of stimulation achievable make this approach applicable to a wide range of experiments. PMID:25650555

  7. Direct Visualization of the Murine Dorsal Cochlear Nucleus for Optogenetic Stimulation of the Auditory Pathway

    PubMed Central

    Lehmann, Ashton E.; Kaplan, Alyson B.; Brown, M. Christian; Lee, Daniel J.

    2015-01-01

    Investigation into the use of virus-mediated gene transfer to arrest or reverse hearing loss has largely been relegated to the peripheral auditory system. Few studies have examined gene transfer to the central auditory system. The dorsal cochlear nucleus (DCN) of the brainstem, which contains second order neurons of the auditory pathway, is a potential site for gene transfer. In this protocol, a technique for direct and maximal exposure of the murine DCN via a posterior fossa approach is demonstrated. This approach allows for either acute or survival surgery. Following direct visualization of the DCN, a host of experiments are possible, including injection of opsins into the cochlear nucleus and subsequent stimulation by an optical fiber coupled to a blue light laser. Other neurophysiology experiments, such as electrical stimulation and neural injector tracings are also feasible. The level of visualization and the duration of stimulation achievable make this approach applicable to a wide range of experiments. PMID:25650555

  8. Cyanobacterial production of 1,3-propanediol directly from carbon dioxide using a synthetic metabolic pathway.

    PubMed

    Hirokawa, Yasutaka; Maki, Yuki; Tatsuke, Tsuneyuki; Hanai, Taizo

    2016-03-01

    Production of chemicals directly from carbon dioxide using light energy is an attractive option for a sustainable future. The 1,3-propanediol (1,3-PDO) production directly from carbon dioxide was achieved by engineered Synechococcus elongatus PCC 7942 with a synthetic metabolic pathway. Glycerol dehydratase catalyzing the conversion of glycerol to 3-hydroxypropionaldehyde in a coenzyme B12-dependent manner worked in S. elongatus PCC 7942 without addition of vitamin B12, suggesting that the intrinsic pseudovitamin B12 served as a substitute of coenzyme B12. The highest titers of 1,3-PDO (3.79±0.23 mM; 288±17.7 mg/L) and glycerol (12.62±1.55 mM; 1.16±0.14 g/L), precursor of 1,3-PDO, were reached after 14 days of culture under optimized conditions in this study. PMID:26769097

  9. Single ricin detection by AFM chemomechanical mapping

    Technology Transfer Automated Retrieval System (TEKTRAN)

    This research reports a method of detecting ricin molecules immobilized on chemically modified gold (Au;111) surface by chemomechanically mapping the molecular interactions with a chemically modified Atomic Force Microscope (AFM) tip. AFM images resolved the different fold-up conformations of single...

  10. BPA Directly Decreases GnRH Neuronal Activity via Noncanonical Pathway.

    PubMed

    Klenke, Ulrike; Constantin, Stephanie; Wray, Susan

    2016-05-01

    Peripheral feedback of gonadal estrogen to the hypothalamus is critical for reproduction. Bisphenol A (BPA), an environmental pollutant with estrogenic actions, can disrupt this feedback and lead to infertility in both humans and animals. GnRH neurons are essential for reproduction, serving as an important link between brain, pituitary, and gonads. Because GnRH neurons express several receptors that bind estrogen, they are potential targets for endocrine disruptors. However, to date, direct effects of BPA on GnRH neurons have not been shown. This study investigated the effects of BPA on GnRH neuronal activity using an explant model in which large numbers of primary GnRH neurons are maintained and express many of the receptors found in vivo. Because oscillations in intracellular calcium have been shown to correlate with electrical activity in GnRH neurons, calcium imaging was used to assay the effects of BPA. Exposure to 50μM BPA significantly decreased GnRH calcium activity. Blockage of γ-aminobutyric acid ergic and glutamatergic input did not abrogate the inhibitory BPA effect, suggesting direct regulation of GnRH neurons by BPA. In addition to estrogen receptor-β, single-cell RT-PCR analysis confirmed that GnRH neurons express G protein-coupled receptor 30 (G protein-coupled estrogen receptor 1) and estrogen-related receptor-γ, all potential targets for BPA. Perturbation studies of the signaling pathway revealed that the BPA-mediated inhibition of GnRH neuronal activity occurred independent of estrogen receptors, GPER, or estrogen-related receptor-γ, via a noncanonical pathway. These results provide the first evidence of a direct effect of BPA on GnRH neurons. PMID:26934298

  11. Nano-Wilhelmy investigation of dynamic wetting properties of AFM tips through tip-nanobubble interaction

    PubMed Central

    Wang, Yuliang; Wang, Huimin; Bi, Shusheng; Guo, Bin

    2016-01-01

    The dynamic wetting properties of atomic force microscopy (AFM) tips are of much concern in many AFM-related measurement, fabrication, and manipulation applications. In this study, the wetting properties of silicon and silicon nitride AFM tips are investigated through dynamic contact angle measurement using a nano-Wilhelmy balance based method. This is done by capillary force measurement during extension and retraction motion of AFM tips relative to interfacial nanobubbles. The working principle of the proposed method and mathematic models for dynamic contact angle measurement are presented. Geometric models of AFM tips were constructed using scanning electronic microscopy (SEM) images taken from different view directions. The detailed process of tip-nanobubble interaction was investigated using force-distance curves of AFM on nanobubbles. Several parameters including nanobubble height, adhesion and capillary force between tip and nanobubbles are extracted. The variation of these parameters was studied over nanobubble surfaces. The dynamic contact angles of the AFM tips were calculated from the capillary force measurements. The proposed method provides direct measurement of dynamic contact angles for AFM tips and can also be taken as a general approach for nanoscale dynamic wetting property investigation. PMID:27452115

  12. Nano-Wilhelmy investigation of dynamic wetting properties of AFM tips through tip-nanobubble interaction

    NASA Astrophysics Data System (ADS)

    Wang, Yuliang; Wang, Huimin; Bi, Shusheng; Guo, Bin

    2016-07-01

    The dynamic wetting properties of atomic force microscopy (AFM) tips are of much concern in many AFM-related measurement, fabrication, and manipulation applications. In this study, the wetting properties of silicon and silicon nitride AFM tips are investigated through dynamic contact angle measurement using a nano-Wilhelmy balance based method. This is done by capillary force measurement during extension and retraction motion of AFM tips relative to interfacial nanobubbles. The working principle of the proposed method and mathematic models for dynamic contact angle measurement are presented. Geometric models of AFM tips were constructed using scanning electronic microscopy (SEM) images taken from different view directions. The detailed process of tip-nanobubble interaction was investigated using force-distance curves of AFM on nanobubbles. Several parameters including nanobubble height, adhesion and capillary force between tip and nanobubbles are extracted. The variation of these parameters was studied over nanobubble surfaces. The dynamic contact angles of the AFM tips were calculated from the capillary force measurements. The proposed method provides direct measurement of dynamic contact angles for AFM tips and can also be taken as a general approach for nanoscale dynamic wetting property investigation.

  13. Nano-Wilhelmy investigation of dynamic wetting properties of AFM tips through tip-nanobubble interaction.

    PubMed

    Wang, Yuliang; Wang, Huimin; Bi, Shusheng; Guo, Bin

    2016-01-01

    The dynamic wetting properties of atomic force microscopy (AFM) tips are of much concern in many AFM-related measurement, fabrication, and manipulation applications. In this study, the wetting properties of silicon and silicon nitride AFM tips are investigated through dynamic contact angle measurement using a nano-Wilhelmy balance based method. This is done by capillary force measurement during extension and retraction motion of AFM tips relative to interfacial nanobubbles. The working principle of the proposed method and mathematic models for dynamic contact angle measurement are presented. Geometric models of AFM tips were constructed using scanning electronic microscopy (SEM) images taken from different view directions. The detailed process of tip-nanobubble interaction was investigated using force-distance curves of AFM on nanobubbles. Several parameters including nanobubble height, adhesion and capillary force between tip and nanobubbles are extracted. The variation of these parameters was studied over nanobubble surfaces. The dynamic contact angles of the AFM tips were calculated from the capillary force measurements. The proposed method provides direct measurement of dynamic contact angles for AFM tips and can also be taken as a general approach for nanoscale dynamic wetting property investigation. PMID:27452115

  14. A direct anterior cingulate pathway to the primate primary olfactory cortex may control attention to olfaction.

    PubMed

    García-Cabezas, Miguel Á; Barbas, Helen

    2014-09-01

    Behavioral and functional studies in humans suggest that attention plays a key role in activating the primary olfactory cortex through an unknown circuit mechanism. We report that a novel pathway from the anterior cingulate cortex, an area which has a key role in attention, projects directly to the primary olfactory cortex in rhesus monkeys, innervating mostly the anterior olfactory nucleus. Axons from the anterior cingulate cortex formed synapses mostly with spines of putative excitatory pyramidal neurons and with a small proportion of a neurochemical class of inhibitory neurons that are thought to have disinhibitory effect on excitatory neurons. This novel pathway from the anterior cingulate is poised to exert a powerful excitatory effect on the anterior olfactory nucleus, which is a critical hub for odorant processing via extensive bilateral connections with primary olfactory cortices and the olfactory bulb. Acting on the anterior olfactory nucleus, the anterior cingulate may activate the entire primary olfactory cortex to mediate the process of rapid attention to olfactory stimuli. PMID:23797208

  15. Directing folding pathways for multi-component DNA origami nanostructures with complex topology

    NASA Astrophysics Data System (ADS)

    Marras, A. E.; Zhou, L.; Kolliopoulos, V.; Su, H.-J.; Castro, C. E.

    2016-05-01

    Molecular self-assembly has become a well-established technique to design complex nanostructures and hierarchical mesoscale assemblies. The typical approach is to design binding complementarity into nucleotide or amino acid sequences to achieve the desired final geometry. However, with an increasing interest in dynamic nanodevices, the need to design structures with motion has necessitated the development of multi-component structures. While this has been achieved through hierarchical assembly of similar structural units, here we focus on the assembly of topologically complex structures, specifically with concentric components, where post-folding assembly is not feasible. We exploit the ability to direct folding pathways to program the sequence of assembly and present a novel approach of designing the strand topology of intermediate folding states to program the topology of the final structure, in this case a DNA origami slider structure that functions much like a piston-cylinder assembly in an engine. The ability to program the sequence and control orientation and topology of multi-component DNA origami nanostructures provides a foundation for a new class of structures with internal and external moving parts and complex scaffold topology. Furthermore, this work provides critical insight to guide the design of intermediate states along a DNA origami folding pathway and to further understand the details of DNA origami self-assembly to more broadly control folding states and landscapes.

  16. Activated pathways for the directed insertion of patterned nanoparticles into polymer membranes.

    PubMed

    Ting, Christina L; Frischknecht, Amalie L

    2013-10-28

    We combine the string method with self-consistent field theory to compute the most probable transition pathway, i.e. the minimum free energy path, for the insertion of Janus and protein-like nanoparticles into a polymer membrane bilayer. The method makes no assumptions in the reaction coordinate and overcomes the long timescales challenge associated with simulating rare events. Our study suggests that one approach to building functional polymer–nanoparticle composite membranes with oriented nanoparticles is through electrostatic interactions. In particular, hydrophobic Janus nanoparticles with an asymmetric charge distribution can be made to directionally insert into charged membranes. This process is kinetically driven, and involves overcoming a thermally surmountable activation barrier, which requires favorable interactions between the nanoparticle and the hydrophilic block of the membrane. In contrast, the insertion of protein-like nanoparticles with alternating hydrophilic–hydrophobic–hydrophilic domains into polymer membranes does not occur as a thermally activated event. PMID:26029770

  17. Heading in the right direction: thermodynamics-based network analysis and pathway engineering.

    PubMed

    Ataman, Meric; Hatzimanikatis, Vassily

    2015-12-01

    Thermodynamics-based network analysis through the introduction of thermodynamic constraints in metabolic models allows a deeper analysis of metabolism and guides pathway engineering. The number and the areas of applications of thermodynamics-based network analysis methods have been increasing in the last ten years. We review recent applications of these methods and we identify the areas that such analysis can contribute significantly, and the needs for future developments. We find that organisms with multiple compartments and extremophiles present challenges for modeling and thermodynamics-based flux analysis. The evolution of current and new methods must also address the issues of the multiple alternatives in flux directionalities and the uncertainties and partial information from analytical methods. PMID:26360871

  18. Degradation pathways of lamotrigine under advanced treatment by direct UV photolysis, hydroxyl radicals, and ozone.

    PubMed

    Keen, Olya S; Ferrer, Imma; Michael Thurman, E; Linden, Karl G

    2014-12-01

    Lamotrigine is recently recognized as a persistent pharmaceutical in the water environment and wastewater effluents. Its degradation was studied under UV and ozone advanced oxidation treatments with reaction kinetics of lamotrigine with ozone (≈4 M(-1)s(-1)), hydroxyl radical [(2.1 ± 0.3) × 10(9)M(-1)s(-1)] and by UV photolysis with low and medium pressure mercury vapor lamps [quantum yields ≈0 and (2.7 ± 0.4)× 10(-4) respectively] determined. All constants were measured at pH 6 and at temperature ≈20°C. The results indicate that lamotrigine is slow to respond to direct photolysis or oxidation by ozone and no attenuation of the contaminant is expected in UV or ozone disinfection applications. The compound reacts rapidly with hydroxyl radicals indicating that advanced oxidation processes would be effective for its treatment. Degradation products were identified under each treatment process using accurate mass time-of-flight spectrometry and pathways of decay were proposed. The main transformation pathways in each process were: dechlorination of the benzene ring during direct photolysis; hydroxyl group addition to the benzene ring during the reaction with hydroxyl radicals; and triazine ring opening after reaction with ozone. Different products that form in each process may be to a varying degree less environmentally stable than the parent lamotrigine. In addition, a novel method of ozone quenching without addition of salts is presented. The new quenching method would allow subsequent mass spectrometry analysis without a solid phase extraction clean-up step. The method involves raising the pH of the sample to approximately 10 for a few seconds and lowering it back and is therefore limited to applications for which temporary pH change is not expected to affect the outcome of the analysis. PMID:25150682

  19. STM and AFM; Which is Better for Surface Structural Analysis? Non- contact AFM Studies on Ge/Si(105) Surface

    NASA Astrophysics Data System (ADS)

    Hasegawa, Yukio

    2006-03-01

    Scanning tunneling microscopy (STM) has been utilized to determine surface atomic structure with its highly resolved images. Probing surface electronic states near the Fermi energy (EF), STM images, however, do not necessarily represent the atomic structure of surfaces. It has been believed that atomic force microscopy (AFM) provides us surface topographic images without being disturbed by the electronic states. In order to prove the surpassing performance, we performed noncontact (nc) AFM on the Ge/Si(105) surface [1], which is a facet plane of the ?hut? clusters formed on Ge-deposited Si(001) surface. It is found that STM images taken on the surface, either filled- or empty-state images, do not show all surface atoms because of the electronic effect; some surface atoms have dangling bond states below EF and other surface atoms have states above EF. [2]. In a nc-AFM image, on the other hand, all surface atoms having a dangling bond are observed [3], directly representing an atomic structure of the surface. Electronic information can also be obtained in AFM by using a Kelvin-probe method. From atomically resolved potential profile we obtained, charge transfer among the dangling bond states is directly demonstrated. These results clearly demonstrate that highly-resolved nc-AFM with a Kelvin-probe method is an ideal tool for analysis of atomic structures and electronic properties of surfaces. This work was done in collaboration with T. Eguchi, K. Akiyama, T. An, and M. Ono, ISSP, Univ. Tokyo and JST, Y. Fujikawa and T. Sakurai, IMR. Tohoku Univ. T. Hashimoto, AIST, Y. Morikawa, ISIR, Osaka Univ. K. Terakura, Hokkaido Univ., and M.G. Lagally, University of Wisconsin-Madison. [1] T. Eguchi et al., Phys. Rev. Lett. 93, 266102 (2004). [2] Y. Fujikawa et al., Phys. Rev. Lett. 88, 176101 (2002). [3] T. Eguchi and Y. Hasegawa, Phys. Rev. Lett. 89, 256105 (2002)

  20. Local and distal effects of arbuscular mycorrhizal colonization on direct pathway Pi uptake and root growth in Medicago truncatula.

    PubMed

    Watts-Williams, Stephanie J; Jakobsen, Iver; Cavagnaro, Timothy R; Grønlund, Mette

    2015-07-01

    Two pathways exist for plant Pi uptake from soil: via root epidermal cells (direct pathway) or via associations with arbuscular mycorrhizal (AM) fungi, and the two pathways interact in a complex manner. This study investigated distal and local effects of AM colonization on direct root Pi uptake and root growth, at different soil P levels. Medicago truncatula was grown at three soil P levels in split-pots with or without AM fungal inoculation and where one root half grew into soil labelled with (33)P. Plant genotypes included the A17 wild type and the mtpt4 mutant. The mtpt4 mutant, colonized by AM fungi, but with no functional mycorrhizal pathway for Pi uptake, was included to better understand effects of AM colonization per se. Colonization by AM fungi decreased expression of direct Pi transporter genes locally, but not distally in the wild type. In mtpt4 mutant plants, direct Pi transporter genes and the Pi starvation-induced gene Mt4 were more highly expressed than in wild-type roots. In wild-type plants, less Pi was taken up via the direct pathway by non-colonized roots when the other root half was colonized by AM fungi, compared with non-mycorrhizal plants. Colonization by AM fungi strongly influenced root growth locally and distally, and direct root Pi uptake activity locally, but had only a weak influence on distal direct pathway activity. The responses to AM colonization in the mtpt4 mutant suggested that in the wild type, the increased P concentration of colonized roots was a major factor driving the effects of AM colonization on direct root Pi uptake. PMID:25944927

  1. Towards direct synthesis of alane: A predicted defect-mediated pathway confirmed experimentally

    DOE PAGESBeta

    Wang, Lin -Lin; Herwadkar, Aditi; Reich, Jason M.; Johnson, Duane D.; House, Stephen D.; Pena-Martin, Pamela; Rockett, Angus A.; Robertson, Ian M.; Gupta, Shalabh; Pecharsky, Vitalij K.

    2016-08-18

    Here, alane (AlH3) is a unique energetic material that has not found a broad practical use for over 70 years because it is difficult to synthesize directly from its elements. Using density functional theory, we examine the defect-mediated formation of alane monomers on Al(111) in a two-step process: (1) dissociative adsorption of H2 and (2) alane formation, which are both endothermic on a clean surface. Only with Ti dopant to facilitate H2 dissociation and vacancies to provide Al adatoms, both processes become exothermic. In agreement, in situ scanning tunneling microscopy showed that during H2 exposure, alane monomers and clusters formmore » primarily in the vicinity of Al vacancies and Ti atoms. Moreover, ball milling of the Al samples with Ti (providing necessary defects) showed a 10 % conversion of Al into AlH3 or closely related species at 344 bar H2, indicating that the predicted pathway may lead to the direct synthesis of alane from elements at pressures much lower than the 104 bar expected from bulk thermodynamics.« less

  2. Towards Direct Synthesis of Alane: A Predicted Defect-Mediated Pathway Confirmed Experimentally.

    PubMed

    Wang, Lin-Lin; Herwadkar, Aditi; Reich, Jason M; Johnson, Duane D; House, Stephen D; Peña-Martin, Pamela; Rockett, Angus A; Robertson, Ian M; Gupta, Shalabh; Pecharsky, Vitalij K

    2016-09-01

    Alane (AlH3 ) is a unique energetic material that has not found a broad practical use for over 70 years because it is difficult to synthesize directly from its elements. Using density functional theory, we examine the defect-mediated formation of alane monomers on Al(111) in a two-step process: (1) dissociative adsorption of H2 and (2) alane formation, which are both endothermic on a clean surface. Only with Ti dopant to facilitate H2 dissociation and vacancies to provide Al adatoms, both processes become exothermic. In agreement, in situ scanning tunneling microscopy showed that during H2 exposure, alane monomers and clusters form primarily in the vicinity of Al vacancies and Ti atoms. Moreover, ball milling of the Al samples with Ti (providing necessary defects) showed a 10 % conversion of Al into AlH3 or closely related species at 344 bar H2 , indicating that the predicted pathway may lead to the direct synthesis of alane from elements at pressures much lower than the 10(4)  bar expected from bulk thermodynamics. PMID:27535100

  3. MOS-based nanocapacitor using C-AFM

    NASA Astrophysics Data System (ADS)

    Hill, Daniel; Sadewasser, Sascha; Aymerich, Xavier

    2003-04-01

    This report details the attempts made to realise nanocapacitors for nanoscale MOS based integrated circuits by AFM anodic oxidation, and therefore isolation, of nano-sized squares of poly-silicon, titanium and aluminium on Si/SiO2. Conductive AFM (C-AFM) was used to perform topographical and electrical characterisation. The experiments were performed with contact mode C-AFM, in ambient air, using Pt-Ir, Co-Cr and Ti coated (20nm) n-type silicon cantilevers. Each sample consisted of a 3-5nm thick conductor deposited on 6nm of SiO2, which was thermally grown on Phosphorus doped (1019 cm-3) n-type Si(100) substrates. Standard cleaning and passivation processes were used. Poly-silicon was immediately found to be too rough to oxidise. Initial current-voltage measurements inside of the titanium-oxide squares suggest initial isolation followed by degradation through Fowler-Nordheim tunnelling. Measurement inconsistencies seen suggest charge storage on the surface or tip with the barrier height of the native titanium oxide thought to be responsible. Al has a thicker natural oxide. To overcome this we designed a series of structures consisting of a Ti finger on SiO2, that is connected to a Ti bond pad, allowing direct probing by a semiconductor parameter analyser. AFM anodic oxidation was performed upon these Ti fingers to reduce their in-plane dimensions towards the nanoscale. To confirm the existence of a nanocapacitor topographical and electrical measurements were then done on and around them.

  4. ezAFM: A low cost Atomic Force Microscope(AFM)

    NASA Astrophysics Data System (ADS)

    Celik, Umit; Celik, Kubra; Aslan, Husnu; Kehribar, Ihsan; Dede, Munir; Ozgur Ozer, H.; Oral, Ahmet

    2012-02-01

    A low cost AFM, ezAFM is developed for educational purposes as well as research. Optical beam deflection method is used to measure the deflection of cantilever. ezAFM scanner is built using voice coil motors (VCM) with ˜50x50x6 μm scan area. The microscope uses alignment free cantilevers, which minimizes setup times. FPGA based AFM feedback Control electronics is developed. FPGA technology allows us to drive all peripherals in parallel. ezAFM Controller is connected to PC by USB 2.0 interface as well as Wi-Fi. We have achieved <5nm lateral and ˜0.01nm vertical resolution. ezAFM can image single atomic steps in HOPG and mica. An optical microscope with <3 μm resolution is also integrated into the system. ezAFM supports different AFM operation modes such as dynamic mode, contact mode, lateral force microscopy. Advanced modes like magnetic force microscopy and electric force microscopy will be implemented later on. The new ezAFM system provides, short learning times for student labs, quick setup and easy to transport for portable applications with the best price/performance ratio. The cost of the system starts from 15,000, with system performance comparable with the traditional AFM systems.

  5. Identification of Thiotetronic Acid Antibiotic Biosynthetic Pathways by Target-directed Genome Mining.

    PubMed

    Tang, Xiaoyu; Li, Jie; Millán-Aguiñaga, Natalie; Zhang, Jia Jia; O'Neill, Ellis C; Ugalde, Juan A; Jensen, Paul R; Mantovani, Simone M; Moore, Bradley S

    2015-12-18

    Recent genome sequencing efforts have led to the rapid accumulation of uncharacterized or "orphaned" secondary metabolic biosynthesis gene clusters (BGCs) in public databases. This increase in DNA-sequenced big data has given rise to significant challenges in the applied field of natural product genome mining, including (i) how to prioritize the characterization of orphan BGCs and (ii) how to rapidly connect genes to biosynthesized small molecules. Here, we show that by correlating putative antibiotic resistance genes that encode target-modified proteins with orphan BGCs, we predict the biological function of pathway specific small molecules before they have been revealed in a process we call target-directed genome mining. By querying the pan-genome of 86 Salinispora bacterial genomes for duplicated house-keeping genes colocalized with natural product BGCs, we prioritized an orphan polyketide synthase-nonribosomal peptide synthetase hybrid BGC (tlm) with a putative fatty acid synthase resistance gene. We employed a new synthetic double-stranded DNA-mediated cloning strategy based on transformation-associated recombination to efficiently capture tlm and the related ttm BGCs directly from genomic DNA and to heterologously express them in Streptomyces hosts. We show the production of a group of unusual thiotetronic acid natural products, including the well-known fatty acid synthase inhibitor thiolactomycin that was first described over 30 years ago, yet never at the genetic level in regards to biosynthesis and autoresistance. This finding not only validates the target-directed genome mining strategy for the discovery of antibiotic producing gene clusters without a priori knowledge of the molecule synthesized but also paves the way for the investigation of novel enzymology involved in thiotetronic acid natural product biosynthesis. PMID:26458099

  6. Developmental Pathway of the MPER-Directed HIV-1-Neutralizing Antibody 10E8

    PubMed Central

    Zhang, Baoshan; McKee, Krisha; Longo, Nancy S.; Yang, Yongping; Huang, Jinghe; Parks, Robert; Eudailey, Joshua; Lloyd, Krissey E.; Alam, S. Munir; Haynes, Barton F.; Mullikin, James C.; Connors, Mark; Mascola, John R.; Shapiro, Lawrence; Kwong, Peter D.

    2016-01-01

    Antibody 10E8 targets the membrane-proximal external region (MPER) of HIV-1 gp41, neutralizes >97% of HIV-1 isolates, and lacks the auto-reactivity often associated with MPER-directed antibodies. The developmental pathway of 10E8 might therefore serve as a promising template for vaccine design, but samples from time-of-infection—often used to infer the B cell record—are unavailable. In this study, we used crystallography, next-generation sequencing (NGS), and functional assessments to infer the 10E8 developmental pathway from a single time point. Mutational analysis indicated somatic hypermutation of the 2nd-heavy chain-complementarity determining region (CDR H2) to be critical for neutralization, and structures of 10E8 variants with V-gene regions reverted to genomic origin for heavy-and-light chains or heavy chain-only showed structural differences >2 Å relative to mature 10E8 in the CDR H2 and H3. To understand these developmental changes, we used bioinformatic sieving, maximum likelihood, and parsimony analyses of immunoglobulin transcripts to identify 10E8-lineage members, to infer the 10E8-unmutated common ancestor (UCA), and to calculate 10E8-developmental intermediates. We were assisted in this analysis by the preservation of a critical D-gene segment, which was unmutated in most 10E8-lineage sequences. UCA and early intermediates weakly bound a 26-residue-MPER peptide, whereas HIV-1 neutralization and epitope recognition in liposomes were only observed with late intermediates. Antibody 10E8 thus develops from a UCA with weak MPER affinity and substantial differences in CDR H2 and H3 from the mature 10E8; only after extensive somatic hypermutation do 10E8-lineage members gain recognition in the context of membrane and HIV-1 neutralization. PMID:27299673

  7. Fsh and Lh direct conserved and specific pathways during flatfish semicystic spermatogenesis.

    PubMed

    Chauvigné, François; Zapater, Cinta; Crespo, Diego; Planas, Josep V; Cerdà, Joan

    2014-10-01

    The current view of the control of spermatogenesis by Fsh and Lh in non-mammalian vertebrates is largely based on studies carried out in teleosts with cystic and cyclic spermatogenesis. Much less is known concerning the specific actions of gonadotropins during semicystic germ cell development, a type of spermatogenesis in which germ cells are released into the tubular lumen where they transform into spermatozoa. In this study, using homologous gonadotropins and a candidate gene approach, for which the genes' testicular cell-type-specific expression was established, we investigated the regulatory effects of Fsh and Lh on gene expression during spermatogenesis in Senegalese sole (Solea senegalensis), a flatfish with asynchronous and semicystic germ cell development. During early spermatogenesis, Fsh and Lh upregulated steroidogenesis-related genes and nuclear steroid receptors, expressed in both somatic and germ cells, through steroid-dependent pathways, although Lh preferentially stimulated the expression of downstream genes involved in androgen and progestin syntheses. In addition, Lh specifically promoted the expression of spermatid-specific genes encoding spermatozoan flagellar proteins through direct interaction with the Lh receptor in these cells. Interestingly, at this spermatogenic stage, Fsh primarily regulated genes encoding Sertoli cell growth factors with potentially antagonistic effects on germ cell proliferation and differentiation through steroid mediation. During late spermatogenesis, fewer genes were regulated by Fsh or Lh, which was associated with a translational and posttranslational downregulation of the Fsh receptor in different testicular compartments. These results reveal that conserved and specialized gonadotropic pathways regulate semicystic spermatogenesis in flatfish, which may spatially adjust cell germ development to maintain a continuous reservoir of spermatids in the testis. PMID:25024405

  8. Carcinogen-specific mutations in preferred Ras-Raf pathway oncogenes directed by strand bias.

    PubMed

    Keller, Ross R; Gestl, Shelley A; Lu, Amy Q; Hoke, Alicia; Feith, David J; Gunther, Edward J

    2016-08-01

    Carcinogen exposures inscribe mutation patterns on cancer genomes and sometimes bias the acquisition of driver mutations toward preferred oncogenes, potentially dictating sensitivity to targeted agents. Whether and how carcinogen-specific mutation patterns direct activation of preferred oncogenes remains poorly understood. Here, mouse models of breast cancer were exploited to uncover a mechanistic link between strand-biased mutagenesis and oncogene preference. When chemical carcinogens were employed during Wnt1-initiated mammary tumorigenesis, exposure to either 7,12-dimethylbenz(a)anthracene (DMBA) or N-ethyl-N-nitrosourea (ENU) dramatically accelerated tumor onset. Mammary tumors that followed DMBA exposure nearly always activated the Ras pathway via somatic Hras(CAA61CTA) mutations. Surprisingly, mammary tumors that followed ENU exposure typically lacked Hras mutations, and instead activated the Ras pathway downstream via Braf(GTG636GAG) mutations. Hras(CAA61CTA) mutations involve an A-to-T change on the sense strand, whereas Braf(GTG636GAG) mutations involve an inverse T-to-A change, suggesting that strand-biased mutagenesis may determine oncogene preference. To examine this possibility further, we turned to an alternative Wnt-driven tumor model in which carcinogen exposures augment a latent mammary tumor predisposition in Apc(min) mice. DMBA and ENU each accelerated mammary tumor onset in Apc(min) mice by introducing somatic, "second-hit" Apc mutations. Consistent with our strand bias model, DMBA and ENU generated strikingly distinct Apc mutation patterns, including stringently strand-inverse mutation signatures at A:T sites. Crucially, these contrasting signatures precisely match those proposed to confer bias toward Hras(CAA61CTA) versus Braf(GTG636GAG) mutations in the original tumor sets. Our findings highlight a novel mechanism whereby exposure history acts through strand-biased mutagenesis to specify activation of preferred oncogenes. PMID:27207659

  9. A direct descending pathway informing locomotor networks about tactile sensor movement.

    PubMed

    Ache, Jan M; Haupt, S Shuichi; Dürr, Volker

    2015-03-01

    Much like visually impaired humans use a white-cane, nocturnal insects and mammals use antennae or whiskers for near-range orientation. Stick insects, for example, rely heavily on antennal tactile cues to find footholds and detect obstacles. Antennal contacts can even induce aimed reaching movements. Because tactile sensors are essentially one-dimensional, they must be moved to probe the surrounding space. Sensor movement is thus an essential cue for tactile sensing, which needs to be integrated by thoracic networks for generating appropriate adaptive leg movements. Based on single and double recordings, we describe a descending neural pathway comprising three identified ON- and OFF-type neurons that convey complementary, unambiguous, and short-latency information about antennal movement to thoracic networks in the stick insect. The neurons are sensitive to the velocity of antennal movements across the entire range covered by natural movements, regardless of movement direction and joint angle. Intriguingly, none of them originates from the brain. Instead, they descend from the gnathal ganglion and receive input from antennal mechanoreceptors in this lower region of the CNS. From there, they convey information about antennal movement to the thorax. One of the descending neurons, which is additionally sensitive to substrate vibration, feeds this information back to the brain via an ascending branch. We conclude that descending interneurons with complementary tuning characteristics, gains, input and output regions convey detailed information about antennal movement to thoracic networks. This pathway bypasses higher processing centers in the brain and thus constitutes a shortcut between tactile sensors on the head and the thorax. PMID:25740535

  10. Diffusion tensor imaging of dolphin brains reveals direct auditory pathway to temporal lobe

    PubMed Central

    Berns, Gregory S.; Cook, Peter F.; Foxley, Sean; Jbabdi, Saad; Miller, Karla L.; Marino, Lori

    2015-01-01

    The brains of odontocetes (toothed whales) look grossly different from their terrestrial relatives. Because of their adaptation to the aquatic environment and their reliance on echolocation, the odontocetes' auditory system is both unique and crucial to their survival. Yet, scant data exist about the functional organization of the cetacean auditory system. A predominant hypothesis is that the primary auditory cortex lies in the suprasylvian gyrus along the vertex of the hemispheres, with this position induced by expansion of ‘associative′ regions in lateral and caudal directions. However, the precise location of the auditory cortex and its connections are still unknown. Here, we used a novel diffusion tensor imaging (DTI) sequence in archival post-mortem brains of a common dolphin (Delphinus delphis) and a pantropical dolphin (Stenella attenuata) to map their sensory and motor systems. Using thalamic parcellation based on traditionally defined regions for the primary visual (V1) and auditory cortex (A1), we found distinct regions of the thalamus connected to V1 and A1. But in addition to suprasylvian-A1, we report here, for the first time, the auditory cortex also exists in the temporal lobe, in a region near cetacean-A2 and possibly analogous to the primary auditory cortex in related terrestrial mammals (Artiodactyla). Using probabilistic tract tracing, we found a direct pathway from the inferior colliculus to the medial geniculate nucleus to the temporal lobe near the sylvian fissure. Our results demonstrate the feasibility of post-mortem DTI in archival specimens to answer basic questions in comparative neurobiology in a way that has not previously been possible and shows a link between the cetacean auditory system and those of terrestrial mammals. Given that fresh cetacean specimens are relatively rare, the ability to measure connectivity in archival specimens opens up a plethora of possibilities for investigating neuroanatomy in cetaceans and other species

  11. Diffusion tensor imaging of dolphin brains reveals direct auditory pathway to temporal lobe.

    PubMed

    Berns, Gregory S; Cook, Peter F; Foxley, Sean; Jbabdi, Saad; Miller, Karla L; Marino, Lori

    2015-07-22

    The brains of odontocetes (toothed whales) look grossly different from their terrestrial relatives. Because of their adaptation to the aquatic environment and their reliance on echolocation, the odontocetes' auditory system is both unique and crucial to their survival. Yet, scant data exist about the functional organization of the cetacean auditory system. A predominant hypothesis is that the primary auditory cortex lies in the suprasylvian gyrus along the vertex of the hemispheres, with this position induced by expansion of 'associative' regions in lateral and caudal directions. However, the precise location of the auditory cortex and its connections are still unknown. Here, we used a novel diffusion tensor imaging (DTI) sequence in archival post-mortem brains of a common dolphin (Delphinus delphis) and a pantropical dolphin (Stenella attenuata) to map their sensory and motor systems. Using thalamic parcellation based on traditionally defined regions for the primary visual (V1) and auditory cortex (A1), we found distinct regions of the thalamus connected to V1 and A1. But in addition to suprasylvian-A1, we report here, for the first time, the auditory cortex also exists in the temporal lobe, in a region near cetacean-A2 and possibly analogous to the primary auditory cortex in related terrestrial mammals (Artiodactyla). Using probabilistic tract tracing, we found a direct pathway from the inferior colliculus to the medial geniculate nucleus to the temporal lobe near the sylvian fissure. Our results demonstrate the feasibility of post-mortem DTI in archival specimens to answer basic questions in comparative neurobiology in a way that has not previously been possible and shows a link between the cetacean auditory system and those of terrestrial mammals. Given that fresh cetacean specimens are relatively rare, the ability to measure connectivity in archival specimens opens up a plethora of possibilities for investigating neuroanatomy in cetaceans and other species

  12. Tuning the resonance of a photonic crystal microcavity with an AFM probe.

    PubMed

    Märki, Iwan; Salt, Martin; Herzig, Hans Peter

    2006-04-01

    We present theoretical and experimental results on switching and tuning of a two-dimensional photonic crystal resonant microcavity by means of a silicon AFM tip, probing the highly localized optical field in the vicinity of the cavity. On-off switching and modulation of the transmission signal in the kHz range is achieved by bringing an AFM tip onto the center of the microcavity, inducing a damping effect on the transmission resonance. Tuning of the resonant wavelength in the order of several nanometers becomes possible by inserting the AFM tip into one of the holes of the Bragg mirror forming the microcavity in the propagation direction. PMID:19516436

  13. Developmental pathway for potent V1V2-directed HIV-neutralizing antibodies.

    PubMed

    Doria-Rose, Nicole A; Schramm, Chaim A; Gorman, Jason; Moore, Penny L; Bhiman, Jinal N; DeKosky, Brandon J; Ernandes, Michael J; Georgiev, Ivelin S; Kim, Helen J; Pancera, Marie; Staupe, Ryan P; Altae-Tran, Han R; Bailer, Robert T; Crooks, Ema T; Cupo, Albert; Druz, Aliaksandr; Garrett, Nigel J; Hoi, Kam H; Kong, Rui; Louder, Mark K; Longo, Nancy S; McKee, Krisha; Nonyane, Molati; O'Dell, Sijy; Roark, Ryan S; Rudicell, Rebecca S; Schmidt, Stephen D; Sheward, Daniel J; Soto, Cinque; Wibmer, Constantinos Kurt; Yang, Yongping; Zhang, Zhenhai; Mullikin, James C; Binley, James M; Sanders, Rogier W; Wilson, Ian A; Moore, John P; Ward, Andrew B; Georgiou, George; Williamson, Carolyn; Abdool Karim, Salim S; Morris, Lynn; Kwong, Peter D; Shapiro, Lawrence; Mascola, John R

    2014-05-01

    Antibodies capable of neutralizing HIV-1 often target variable regions 1 and 2 (V1V2) of the HIV-1 envelope, but the mechanism of their elicitation has been unclear. Here we define the developmental pathway by which such antibodies are generated and acquire the requisite molecular characteristics for neutralization. Twelve somatically related neutralizing antibodies (CAP256-VRC26.01-12) were isolated from donor CAP256 (from the Centre for the AIDS Programme of Research in South Africa (CAPRISA)); each antibody contained the protruding tyrosine-sulphated, anionic antigen-binding loop (complementarity-determining region (CDR) H3) characteristic of this category of antibodies. Their unmutated ancestor emerged between weeks 30-38 post-infection with a 35-residue CDR H3, and neutralized the virus that superinfected this individual 15 weeks after initial infection. Improved neutralization breadth and potency occurred by week 59 with modest affinity maturation, and was preceded by extensive diversification of the virus population. HIV-1 V1V2-directed neutralizing antibodies can thus develop relatively rapidly through initial selection of B cells with a long CDR H3, and limited subsequent somatic hypermutation. These data provide important insights relevant to HIV-1 vaccine development. PMID:24590074

  14. Prep1 directly regulates the intrinsic apoptotic pathway by controlling Bcl-XL levels.

    PubMed

    Micali, Nicola; Ferrai, Carmelo; Fernandez-Diaz, Luis C; Blasi, Francesco; Crippa, Massimo P

    2009-03-01

    The Prep1 homeodomain transcription factor is essential in embryonic development. Prep1 hypomorphic mutant mouse (Prep1(i/i)) embryos (embryonic day 9.5) display an increased terminal deoxynucleotidyltransferase-mediated dUTP-biotin nick end labeling reaction compared to wild-type (WT) littermates. Prep1(i/i) mouse embryo fibroblasts (MEFs) show an increased basal level of annexin V binding activity, reduction of the mitochondrial-membrane potential, and increased caspase 9 and 3 activation, indicating increased apoptosis. Prep1(i/i) MEFs also respond faster than WT MEFs to genotoxic stress, indicating increased activation of the intrinsic apoptotic pathways. We did not observe an increase in p53 or an abnormal p53 response to apoptotic stimuli. However, hypomorphic MEFs have decreased endogenous levels of antiapoptotic Bcl-X(L) mRNA and protein, and Bcl-x overexpression rescues the defect of Prep1(i/i) MEFs. Using transient transfections and chromatin immunoprecipitation, we identified the Bcl-x promoter as a novel target of Prep1. Thus, Prep1 directly controls mitochondrial homeostasis (and the apoptotic potential) by modulating Bcl-x gene expression. PMID:19103748

  15. Unraveling the microscopic pathway of homogeneous water crystallization at supercooled conditions from direct simulations

    NASA Astrophysics Data System (ADS)

    Martelli, Fausto; Palmer, Jeremy; Singh, Rakesh; Debenedetti, Pablo; Car, Roberto

    By means of unbiased classical molecular dynamics simulations, we identify the microscopic pathways of spontaneous homogeneous crystallization in supercooled ST2 water. By introducing a new order parameter, we are able to monitor formation/disruption of locally ordered regions characterized by small ice clusters with intermediate range order. When two of these regions are close each other, they percolate and form a larger ordered region. The process is slow enough to allow for polymorphic selection in favor of cubic ice (Ic). The formation of an ice nucleus requires percolation of many small clusters so that the transformations at the interface of the nucleus do not involve its core, thus guaranteeing the stability of the nucleus. The growth of the crystalline nucleus is fast and involves direct transformation of interfacial liquid molecules as well as percolation of small Ic/Ih clusters. The growth is too fast to allow conversion of Ih into Ic sites, originating the formation of a stacking fault in the final crystal. We recognize Euclidean structures in the oxygen configuration of the second shell in Ic and Ih clusters. This new point of view allows us to explain the source of the ordered stacking fault geometry.

  16. Nano Mechanical Machining Using AFM Probe

    NASA Astrophysics Data System (ADS)

    Mostofa, Md. Golam

    Complex miniaturized components with high form accuracy will play key roles in the future development of many products, as they provide portability, disposability, lower material consumption in production, low power consumption during operation, lower sample requirements for testing, and higher heat transfer due to their very high surface-to-volume ratio. Given the high market demand for such micro and nano featured components, different manufacturing methods have been developed for their fabrication. Some of the common technologies in micro/nano fabrication are photolithography, electron beam lithography, X-ray lithography and other semiconductor processing techniques. Although these methods are capable of fabricating micro/nano structures with a resolution of less than a few nanometers, some of the shortcomings associated with these methods, such as high production costs for customized products, limited material choices, necessitate the development of other fabricating techniques. Micro/nano mechanical machining, such an atomic force microscope (AFM) probe based nano fabrication, has, therefore, been used to overcome some the major restrictions of the traditional processes. This technique removes material from the workpiece by engaging micro/nano size cutting tool (i.e. AFM probe) and is applicable on a wider range of materials compared to the photolithographic process. In spite of the unique benefits of nano mechanical machining, there are also some challenges with this technique, since the scale is reduced, such as size effects, burr formations, chip adhesions, fragility of tools and tool wear. Moreover, AFM based machining does not have any rotational movement, which makes fabrication of 3D features more difficult. Thus, vibration-assisted machining is introduced into AFM probe based nano mechanical machining to overcome the limitations associated with the conventional AFM probe based scratching method. Vibration-assisted machining reduced the cutting forces

  17. Insights into Epoxy Network Nanostructural Heterogeneity Using AFM-IR.

    PubMed

    Morsch, Suzanne; Liu, Yanwen; Lyon, Stuart B; Gibbon, Simon R

    2016-01-13

    The first direct observation of a chemically heterogeneous nanostructure within an epoxy resin is reported. Epoxy resins comprise the matrix component of many high performance composites, coatings and adhesives, yet the molecular network structure that underpins the performance of these industrially essential materials is not well understood. Internal nodular morphologies have repeatedly been reported for epoxy resins analyzed using SEM or AFM, yet the origin of these features remains a contentious subject, and epoxies are still commonly assumed to be chemically homogeneous. Uniquely, in this contribution we use the recently developed AFM-IR technique to eliminate previous differences in interpretation, and establish that nodule features correspond to heterogeneous network connectivity within an epoxy phenolic formulation. PMID:26694687

  18. SUMO regulates somatic cyst stem cell maintenance and directly targets the Hedgehog pathway in adult Drosophila testis.

    PubMed

    Lv, Xiangdong; Pan, Chenyu; Zhang, Zhao; Xia, Yuanxin; Chen, Hao; Zhang, Shuo; Guo, Tong; Han, Hui; Song, Haiyun; Zhang, Lei; Zhao, Yun

    2016-05-15

    SUMO (Small ubiquitin-related modifier) modification (SUMOylation) is a highly dynamic post-translational modification (PTM) that plays important roles in tissue development and disease progression. However, its function in adult stem cell maintenance is largely unknown. Here, we report the function of SUMOylation in somatic cyst stem cell (CySC) self-renewal in adult Drosophila testis. The SUMO pathway cell-autonomously regulates CySC maintenance. Reduction of SUMOylation promotes premature differentiation of CySCs and impedes the proliferation of CySCs, which leads to a reduction in the number of CySCs. Consistent with this, CySC clones carrying a mutation of the SUMO-conjugating enzyme are rapidly lost. Furthermore, inhibition of the SUMO pathway phenocopies disruption of the Hedgehog (Hh) pathway, and can block the proliferation of CySCs induced by Hh activation. Importantly, the SUMO pathway directly regulates the SUMOylation of Hh pathway transcription factor Cubitus interruptus (Ci), which is required for promoting CySC proliferation. Thus, we conclude that SUMO directly targets the Hh pathway and regulates CySC maintenance in adult Drosophila testis. PMID:27013244

  19. Key Modulatory Role of Presynaptic Adenosine A2A Receptors in Cortical Neurotransmission to the Striatal Direct Pathway

    PubMed Central

    Quiroz, César; Luján, Rafael; Uchigashima, Motokazu; Simoes, Ana Patrícia; Lerner, Talia N.; Borycz, Janusz; Kachroo, Anil; Canas, Paula M.; Orru, Marco; Schwarzschild, Michael A.; Rosin, Diane L.; Kreitzer, Anatol C.; Cunha, Rodrigo A.; Watanabe, Masahiko; Ferré, Sergi

    2010-01-01

    Basal ganglia processing results from a balanced activation of direct and indirect striatal efferent pathways, which are controlled by dopamine D1 and D2 receptors, respectively. Adenosine A2A receptors are considered novel anti-parkinsonian targets, based on their selective postsynaptic localization in the indirect pathway, where they modulate D2 receptor function. The present study provides evidence for the existence of an additional functionally significant segregation of A2A receptors at the presynaptic level. Using integrated anatomical, electrophysiological and biochemical approaches, we demonstrate that presynaptic A2A receptors are preferentially localized in cortical glutamatergic terminals that contact striatal neurons of the direct pathway, where they exert a selective modulation of cortico-striatal neurotransmission. Presynaptic striatal A2A receptors could provide a new target for the treatment of neuropsychiatric disorders. PMID:19936569

  20. Work Stressors, Social Support, and Burnout in Junior Doctors: Exploring Direct and Indirect Pathways

    ERIC Educational Resources Information Center

    Sochos, Antigonos; Bowers, Alexis; Kinman, Gail

    2012-01-01

    The study tested a pathway model linking different occupational stressors, different sources of social support, and burnout. A sample of 184 junior medical doctors was used. Pathway analysis suggested that doctors who experienced increased time demands, organizational constraints, and a lack of personal confidence perceived their consultants as…

  1. Developmental Pathways to Conduct Disorder: Implications for Future Directions in Research, Assessment, and Treatment

    ERIC Educational Resources Information Center

    Frick, Paul J.

    2012-01-01

    Research has indicated that there are several common pathways through which children and adolescents develop conduct disorder, each with different risk factors and each with different underlying developmental mechanisms leading to the child's aggressive and antisocial behavior. The current article briefly summarizes research on these pathways,…

  2. Direct Visualization of Ebola Virus Fusion Triggering in the Endocytic Pathway

    PubMed Central

    Spence, Jennifer S.; Krause, Tyler B.; Mittler, Eva; Jangra, Rohit K.

    2016-01-01

    ABSTRACT Ebola virus (EBOV) makes extensive and intricate use of host factors in the cellular endosomal/lysosomal pathway to release its genome into the cytoplasm and initiate infection. Following viral internalization into endosomes, host cysteine proteases cleave the EBOV fusion glycoprotein (GP) to unmask the binding site for its intracellular receptor, the cholesterol transporter Niemann-Pick C1 (NPC1). GP-NPC1 interaction is required for viral entry. Despite these and other recent discoveries, late events in EBOV entry following GP-NPC1 binding and culminating in GP-catalyzed fusion between viral and cellular lipid bilayers remain enigmatic. A mechanistic understanding of EBOV membrane fusion has been hampered by the failure of previous efforts to reconstitute fusion in vitro or at the cell surface. This report describes an assay to monitor initial steps directly in EBOV membrane fusion—triggering of GP and virus-cell lipid mixing—by single virions in live cells. Fusogenic triggering of GP occurs predominantly in Rab7-positive (Rab7+) endosomes, absolutely requires interaction between proteolytically primed GP and NPC1, and is blocked by key GP-specific neutralizing antibodies with therapeutic potential. Unexpectedly, cysteine protease inhibitors do not inhibit lipid mixing by virions bearing precleaved GP, even though they completely block cytoplasmic entry by these viruses, as shown previously. These results point to distinct cellular requirements for different steps in EBOV membrane fusion and suggest a model in which host cysteine proteases are dispensable for GP fusion triggering after NPC1 binding but are required for the formation of fusion pores that permit genome delivery. PMID:26861015

  3. AFM imaging of functionalized carbon nanotubes on biological membranes

    NASA Astrophysics Data System (ADS)

    Lamprecht, C.; Liashkovich, I.; Neves, V.; Danzberger, J.; Heister, E.; Rangl, M.; Coley, H. M.; McFadden, J.; Flahaut, E.; Gruber, H. J.; Hinterdorfer, P.; Kienberger, F.; Ebner, A.

    2009-10-01

    Multifunctional carbon nanotubes are promising for biomedical applications as their nano-size, together with their physical stability, gives access into the cell and various cellular compartments including the nucleus. However, the direct and label-free detection of carbon nanotube uptake into cells is a challenging task. The atomic force microscope (AFM) is capable of resolving details of cellular surfaces at the nanometer scale and thus allows following of the docking of carbon nanotubes to biological membranes. Here we present topographical AFM images of non-covalently functionalized single walled (SWNT) and double walled carbon nanotubes (DWNT) immobilized on different biological membranes, such as plasma membranes and nuclear envelopes, as well as on a monolayer of avidin molecules. We were able to visualize DWNT on the nuclear membrane while at the same time resolving individual nuclear pore complexes. Furthermore, we succeeded in localizing individual SWNT at the border of incubated cells and in identifying bundles of DWNT on cell surfaces by AFM imaging.

  4. Hydration states of AFm cement phases

    SciTech Connect

    Baquerizo, Luis G.; Matschei, Thomas; Scrivener, Karen L.; Saeidpour, Mahsa; Wadsö, Lars

    2015-07-15

    The AFm phase, one of the main products formed during the hydration of Portland and calcium aluminate cement based systems, belongs to the layered double hydrate (LDH) family having positively charged layers and water plus charge-balancing anions in the interlayer. It is known that these phases present different hydration states (i.e. varying water content) depending on the relative humidity (RH), temperature and anion type, which might be linked to volume changes (swelling and shrinkage). Unfortunately the stability conditions of these phases are insufficiently reported. This paper presents novel experimental results on the different hydration states of the most important AFm phases: monocarboaluminate, hemicarboaluminate, strätlingite, hydroxy-AFm and monosulfoaluminate, and the thermodynamic properties associated with changes in their water content during absorption/desorption. This data opens the possibility to model the response of cementitious systems during drying and wetting and to engineer systems more resistant to harsh external conditions.

  5. LET spectrum measurements in Cr-39 PNTD with AFM

    SciTech Connect

    Johnson, Carl Edward; De Witt, Joel M; Benton, Eric R; Yasuda, Nakahiro; Benton, Eugene V

    2010-01-01

    Energetic protons, neutrons, and heavy ions undergoing collisions with target nuclei of varying Z can produce residual heavy recoil fragments via intra-nuclear cascade/evaporation reactions. The particles produced in these non-elastic collisions generally have such extremely short range ({approx}< 10 {mu}m) that they cannot be directly observed by conventional detection methods including CR-39 plastic nuclear track detector (PNTD) that has been chemically etched for analysis by standard visible light microscopy. However, high-LET recoil fragments having range on the order of several cell diameters can be produced in tissue during radiotherapy using proton and carbon beams. We have developed a method to analyze short-range, high-LET tracks in CR-39 plastic nuclear track detector (PNTD) using short duration chemical etching ({approx}< 1 {mu}m) followed by automated atomic force microscope (AFM) scanning. The post-scan data processing used in this work was based on semi-automated matrix analysis opposed to traditional grey-scale image analysis. This method takes advantage of the 3-D data obtained via AFM to achieve robust discrimination of nuclear tracks from other features. Through automation of AFM scanning, sufficient AFM scan frames were obtained to attain an LET spectrum spanning the LET range from 200-1500 keV/{mu}m. In addition to our experiments, simulations were carried out with the Monte Carlo transport code, FLUKA. To demonstrate this method, CR-39 PNTD was exposed to the proton therapy beam at Loma Linda University Medical Center (LLUMC) at 60 and 230 MeV. Additionally, detectors were exposed to I GeV protons at the NASA Space Radiation Laboratory (NSRL) at Brookhaven National Laboratory (BNL). For these exposures CR-39 PNTD, Al and Cu target foils were used between detector layers.

  6. LET Spectrum Measurements In CR-39 PNTD With AFM

    NASA Astrophysics Data System (ADS)

    Johnson, C. E.; DeWitt, J. M.; Benton, E. R.; Yasuda, N.; Benton, E. V.

    2011-06-01

    Energetic protons, neutrons, and heavy ions undergoing collisions with target nuclei of varying Z can produce residual heavy recoil fragments via intra-nuclear cascade/evaporation reactions. The particles produced in these non-elastic collisions generally have such extremely short range (˜<10 μm) that they cannot be directly observed by conventional detection methods including CR-39 plastic nuclear track detector (PNTD) that has been chemically etched for analysis by standard visible light microscopy. However, high-LET recoil fragments having range on the order of several cell diameters can be produced in tissue during radiotherapy using proton and carbon beams. We have developed a method to analyze short-range, high-LET tracks in CR-39 plastic nuclear track detector (PNTD) using short duration chemical etching (˜<1 μm) following by automated atomic force microscope (AFM) scanning. The post-scan data processing used in this work was based on semi-automated matrix analysis opposed to traditional grey-scale image analysis. This method takes advantage of the 3-D data obtained via AFM to achieve robust discrimination of nuclear tracks from other features inherently present in the post-etch detector surface. Through automation of AFM scanning, sufficient AFM scan frames were obtained to attain an LET spectrum spanning the LET range from 200-1500 keV/μm. In addition to our experiments, simulations were carried out with the Monte Carlo transport code, FLUKA. To demonstrate this method, CR-39 PNTD was exposed to the proton therapy beam at Loma Linda University Medical Center (LLUMC) at 60 and 230 MeV. Additionally, detectors were exposed to 1 GeV protons at the NASA Space Radiation Laboratory (NSRL) at Brookhaven National Laboratory (BNL). For these exposures CR-39 PNTD, Al and Cu target foils were used between detector layers.

  7. LET Spectrum Measurements In CR-39 PNTD With AFM

    SciTech Connect

    Johnson, C. E.; DeWitt, J. M.; Benton, E. R.; Yasuda, N.; Benton, E. V.

    2011-06-01

    Energetic protons, neutrons, and heavy ions undergoing collisions with target nuclei of varying Z can produce residual heavy recoil fragments via intra-nuclear cascade/evaporation reactions. The particles produced in these non-elastic collisions generally have such extremely short range ({approx}<10 {mu}m) that they cannot be directly observed by conventional detection methods including CR-39 plastic nuclear track detector (PNTD) that has been chemically etched for analysis by standard visible light microscopy. However, high-LET recoil fragments having range on the order of several cell diameters can be produced in tissue during radiotherapy using proton and carbon beams. We have developed a method to analyze short-range, high-LET tracks in CR-39 plastic nuclear track detector (PNTD) using short duration chemical etching ({approx}<1 {mu}m) following by automated atomic force microscope (AFM) scanning. The post-scan data processing used in this work was based on semi-automated matrix analysis opposed to traditional grey-scale image analysis. This method takes advantage of the 3-D data obtained via AFM to achieve robust discrimination of nuclear tracks from other features inherently present in the post-etch detector surface. Through automation of AFM scanning, sufficient AFM scan frames were obtained to attain an LET spectrum spanning the LET range from 200-1500 keV/{mu}m. In addition to our experiments, simulations were carried out with the Monte Carlo transport code, FLUKA. To demonstrate this method, CR-39 PNTD was exposed to the proton therapy beam at Loma Linda University Medical Center (LLUMC) at 60 and 230 MeV. Additionally, detectors were exposed to 1 GeV protons at the NASA Space Radiation Laboratory (NSRL) at Brookhaven National Laboratory (BNL). For these exposures CR-39 PNTD, Al and Cu target foils were used between detector layers.

  8. An adenylyl cyclase signaling pathway predicts direct dopaminergic input to vestibular hair cells.

    PubMed

    Drescher, M J; Cho, W J; Folbe, A J; Selvakumar, D; Kewson, D T; Abu-Hamdan, M D; Oh, C K; Ramakrishnan, N A; Hatfield, J S; Khan, K M; Anne, S; Harpool, E C; Drescher, D G

    2010-12-29

    Adenylyl cyclase (AC) signaling pathways have been identified in a model hair cell preparation from the trout saccule, for which the hair cell is the only intact cell type. The use of degenerate primers targeting cDNA sequence conserved across AC isoforms, and reverse transcription-polymerase chain reaction (RT-PCR), coupled with cloning of amplification products, indicated expression of AC9, AC7 and AC5/6, with cloning efficiencies of 11:5:2. AC9 and AC5/6 are inhibited by Ca(2+), the former in conjunction with calcineurin, and message for calcineurin has also been identified in the trout saccular hair cell layer. AC7 is independent of Ca(2+). Given the lack of detection of calcium/calmodulin-activated isoforms previously suggested to mediate AC activation in the absence of Gαs in mammalian cochlear hair cells, the issue of hair-cell Gαs mRNA expression was re-examined in the teleost vestibular hair cell model. Two full-length coding sequences were obtained for Gαs/olf in the vestibular type II-like hair cells of the trout saccule. Two messages for Gαi have also been detected in the hair cell layer, one with homology to Gαi1 and the second with homology to Gαi3 of higher vertebrates. Both Gαs/olf protein and Gαi1/Gαi3 protein were immunolocalized to stereocilia and to the base of the hair cell, the latter consistent with sites of efferent input. Although a signaling event coupling to Gαs/olf and Gαi1/Gαi3 in the stereocilia is currently unknown, signaling with Gαs/olf, Gαi3, and AC5/6 at the base of the hair cell would be consistent with transduction pathways activated by dopaminergic efferent input. mRNA for dopamine receptors D1A4 and five forms of dopamine D2 were found to be expressed in the teleost saccular hair cell layer, representing information on vestibular hair cell expression not directly available for higher vertebrates. Dopamine D1A receptor would couple to Gαolf and activation of AC5/6. Co-expression with dopamine D2 receptor, which

  9. Genome-Wide Transcriptome Directed Pathway Analysis of Maternal Pre-Eclampsia Susceptibility Genes

    PubMed Central

    Yong, Hannah E. J.; Melton, Phillip E.; Johnson, Matthew P.; Freed, Katy A.; Kalionis, Bill; Murthi, Padma; Brennecke, Shaun P.; Keogh, Rosemary J.; Moses, Eric K.

    2015-01-01

    Background Preeclampsia (PE) is a serious hypertensive pregnancy disorder with a significant genetic component. Numerous genetic studies, including our own, have yielded many susceptibility genes from distinct functional groups. Additionally, transcriptome profiling of tissues at the maternal-fetal interface has likewise yielded many differentially expressed genes. Often there is little overlap between these two approaches, although genes identified in both approaches are significantly associated with PE. We have thus taken a novel integrative bioinformatics approach of analysing pathways common to the susceptibility genes and the PE transcriptome. Methods Using Illumina Human Ht12v4 and Wg6v3 BeadChips, transcriptome profiling was conducted on n = 65 normotensive and n = 60 PE decidua basalis tissues collected at delivery. The R software package libraries lumi and limma were used to preprocess transcript data for pathway analysis. Pathways were analysed and constructed using Pathway Studio. We examined ten candidate genes, which are from these functional groups: activin/inhibin signalling—ACVR1, ACVR1C, ACVR2A, INHA, INHBB; structural components—COL4A1, COL4A2 and M1 family aminopeptidases—ERAP1, ERAP2 and LNPEP. Results/Conclusion Major common regulators/targets of these susceptibility genes identified were AGT, IFNG, IL6, INHBA, SERPINE1, TGFB1 and VEGFA. The top two categories of pathways associated with the susceptibility genes, which were significantly altered in the PE decidual transcriptome, were apoptosis and cell signaling (p < 0.001). Thus, susceptibility genes from distinct functional groups share similar downstream pathways through common regulators/targets, some of which are altered in PE. This study contributes to a better understanding of how susceptibility genes may interact in the development of PE. With this knowledge, more targeted functional analyses of PE susceptibility genes in these key pathways can be performed to examine their

  10. Conductance of AFM Deformed Carbon Nanotubes

    NASA Technical Reports Server (NTRS)

    Svizhenko, Alexei; Maiti, Amitesh; Anatram, M. P.; Biegel, Bryan (Technical Monitor)

    2002-01-01

    This viewgraph presentation provides information on the electrical conductivity of carbon nanotubes upon deformation by atomic force microscopy (AFM). The density of states and conductance were computed using four orbital tight-binding method with various parameterizations. Different chiralities develop bandgap that varies with chirality.

  11. A Batch Fabricated SECM-AFM Probe

    NASA Astrophysics Data System (ADS)

    Dobson, P. S.; Macpherson, J. V.; Holder, M.; Weaver, J. M. R.

    2003-12-01

    A scheme for the fabrication of combined Scanning Electrochemical Microscopy — Atomic Force Microscopy (SECM-AFM) probes is presented for both silicon nitride and silicon cantilevers. The advantages over exsisting methods used for their production is explained. The process flow is described and initial results from electrodeposition of silver are presented.

  12. Contact nanomechanical measurements with the AFM

    NASA Astrophysics Data System (ADS)

    Geisse, Nicholas

    2013-03-01

    The atomic force microscope (AFM) has found broad use in the biological sciences largely due to its ability to make measurements on unfixed and unstained samples under liquid. In addition to imaging at multiple spatial scales ranging from micro- to nanometer, AFMs are commonly used as nanomechanical probes. This is pertinent for cell biology, as it has been demonstrated that the geometrical and mechanical properties of the extracellular microenvironment are important in such processes as cancer, cardiovascular disease, muscular dystrophy, and even the control of cell life and death. Indeed, the ability to control and quantify these external geometrical and mechanical parameters arises as a key issue in the field. Because AFM can quantitatively measure the mechanical properties of various biological samples, novel insights to cell function and to cell-substrate interactions are now possible. As the application of AFM to these types of problems is widened, it is important to understand the performance envelope of the technique and its associated data analyses. This talk will discuss the important issues that must be considered when mechanical models are applied to real-world data. Examples of the effect of different model assumptions on our understanding of the measured material properties will be shown. Furthermore, specific examples of the importance of mechanical stimuli and the micromechanical environment to the structure and function of biological materials will be presented.

  13. Regulatory punctuated equilibrium and convergence in the evolution of developmental pathways in direct-developing sea urchins.

    PubMed

    Raff, Elizabeth C; Popodi, Ellen M; Kauffman, Jeffery S; Sly, Belinda J; Turner, F Rudolf; Morris, Valerie B; Raff, Rudolf A

    2003-01-01

    We made hybrid crosses between closely and distantly related sea urchin species to test two hypotheses about the evolution of gene regulatory systems in the evolution of ontogenetic pathways and larval form. The first hypothesis is that gene regulatory systems governing development evolve in a punctuational manner during periods of rapid morphological evolution but are relatively stable over long periods of slow morphological evolution. We compared hybrids between direct and indirect developers from closely and distantly related families. Hybrids between eggs of the direct developer Heliocidaris erythrogramma and sperm of the 4-million year distant species H. tuberculata, an indirect developer, restored feeding larval structures and paternal gene expression that were lost in the evolution of the direct-developing maternal parent. Hybrids resulting from the cross between eggs of H. erythrogramma and sperm of the 40-million year distant indirect-developer Pseudoboletia maculata are strikingly similar to hybrids between the congeneric hybrids. The marked similarities in ontogenetic trajectory and morphological outcome in crosses of involving either closely or distantly related indirect developing species indicates that their regulatory mechanisms interact with those of H. erythrogramma in the same way, supporting remarkable conservation of molecular control pathways among indirect developers. Second, we tested the hypothesis that convergent developmental pathways in independently evolved direct developers reflect convergence of the underlying regulatory systems. Crosses between two independently evolved direct-developing species from two 70-million year distant families, H. erythrogramma and Holopneustes purpurescens, produced harmoniously developing hybrid larvae that maintained the direct mode of development and did not exhibit any obvious restoration of indirect-developing features. These results are consistent with parallel evolution of direct-developing features

  14. KynR, a Lrp/AsnC-type transcriptional regulator, directly controls the kynurenine pathway in Pseudomonas aeruginosa.

    PubMed

    Knoten, Claire A; Hudson, L Lynn; Coleman, James P; Farrow, John M; Pesci, Everett C

    2011-12-01

    The opportunistic pathogen Pseudomonas aeruginosa can utilize a variety of carbon sources and produces many secondary metabolites to help survive harsh environments. P. aeruginosa is part of a small group of bacteria that use the kynurenine pathway to catabolize tryptophan. Through the kynurenine pathway, tryptophan is broken down into anthranilate, which is further degraded into tricarboxylic acid cycle intermediates or utilized to make numerous aromatic compounds, including the Pseudomonas quinolone signal (PQS). We have previously shown that the kynurenine pathway is a critical source of anthranilate for PQS synthesis and that the kynurenine pathway genes (kynA and kynBU) are upregulated in the presence of kynurenine. A putative Lrp/AsnC-type transcriptional regulator (gene PA2082, here called kynR), is divergently transcribed from the kynBU operon and is highly conserved in gram-negative bacteria that harbor the kynurenine pathway. We show that a mutation in kynR renders P. aeruginosa unable to utilize L-tryptophan as a sole carbon source and decreases PQS production. In addition, we found that the increase of kynA and kynB transcriptional activity in response to kynurenine was completely abolished in a kynR mutant, further indicating that KynR mediates the kynurenine-dependent expression of the kynurenine pathway genes. Finally, we found that purified KynR specifically bound the kynA promoter in the presence of kynurenine and bound the kynB promoter in the absence or presence of kynurenine. Taken together, our data show that KynR directly regulates the kynurenine pathway genes. PMID:21965577

  15. Directed random walks and constraint programming reveal active pathways in hepatocyte growth factor signaling.

    PubMed

    Kittas, Aristotelis; Delobelle, Aurélien; Schmitt, Sabrina; Breuhahn, Kai; Guziolowski, Carito; Grabe, Niels

    2016-01-01

    An effective means to analyze mRNA expression data is to take advantage of established knowledge from pathway databases, using methods such as pathway-enrichment analyses. However, pathway databases are not case-specific and expression data could be used to infer gene-regulation patterns in the context of specific pathways. In addition, canonical pathways may not always describe the signaling mechanisms properly, because interactions can frequently occur between genes in different pathways. Relatively few methods have been proposed to date for generating and analyzing such networks, preserving the causality between gene interactions and reasoning over the qualitative logic of regulatory effects. We present an algorithm (MCWalk) integrated with a logic programming approach, to discover subgraphs in large-scale signaling networks by random walks in a fully automated pipeline. As an exemplary application, we uncover the signal transduction mechanisms in a gene interaction network describing hepatocyte growth factor-stimulated cell migration and proliferation from gene-expression measured with microarray and RT-qPCR using in-house perturbation experiments in a keratinocyte-fibroblast co-culture. The resulting subgraphs illustrate possible associations of hepatocyte growth factor receptor c-Met nodes, differentially expressed genes and cellular states. Using perturbation experiments and Answer Set programming, we are able to select those which are more consistent with the experimental data. We discover key regulator nodes by measuring the frequency with which they are traversed when connecting signaling between receptors and significantly regulated genes and predict their expression-shift consistently with the measured data. The Java implementation of MCWalk is publicly available under the MIT license at: https://bitbucket.org/akittas/biosubg. PMID:26518250

  16. Starter unit specificity directs genome mining of polyketide synthase pathways in fungi

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Search of the protein database with the aflatoxin pathway polyketide synthase (PKS) revealed putative PKSs in the pathogenic fungi Coccidioides immitis and Coccidioides posadasii that could require partnerships with a pair of fatty acid synthase (FAS) subunits for the biosynthesis of fatty acid-poly...

  17. Diversity in Pathways to Parenthood: Patterns, Implications, and Emerging Research Directions

    ERIC Educational Resources Information Center

    Smock, Pamela J.; Greenland, Fiona Rose

    2010-01-01

    This review examines and synthesizes recent research on pathways to parenthood. We begin by providing basic information about patterns, differentials, and trends and discussing adoption and new reproductive technologies. We next turn to several areas of inquiry that became particularly prominent in the last decade: the continued "decoupling" of…

  18. An approach towards 3D sensitive AFM cantilevers

    NASA Astrophysics Data System (ADS)

    Koops, Richard; Fokkema, Vincent

    2014-04-01

    The atomic force microscope (AFM) tapping mode is a highly sensitive local probing technique that is very useful to study and measure surface properties down to the atomic scale. The tapping mode is mostly implemented using the resonance of the first bending mode of the cantilever and therefore provides sensitivity mainly along the direction of this oscillation. Driven by the semiconductor industry, there is an increasing need for accurate measurements of nanoscale structures for side wall characterization by AFM that requires additional sensitivity in the lateral direction. The conventional tapping mode has been augmented by various authors, for example by tilting the cantilever system (Cho et al 2011 Rev. Sci. Instrum. 82 023707) to access the sidewall or using a torsion mode (Dai et al 2011 Meas. Sci. Technol. 22 094009) of the cantilever to provide additional lateral sensitivity. These approaches however trade lateral sensitivity for vertical sensitivity or still lack sensitivity in the remaining lateral direction. We present an approach towards true 3D sensitivity for AFM cantilevers based on simultaneous excitation and optical detection of multiple cantilever resonance modes along three axes. Tuning the excitation of the cantilever to specific frequencies provides a mechanism to select only those cantilever modes that have the desired characteristics. Additionally, cantilever engineering has been used to design and create a substructure within the cantilever that has been optimized for specific resonance behavior around 4 MHz. In contrast to the conventional approach of using a piezo to actuate the cantilever modulation, we present results on photo-thermal excitation using an intensity modulated low-power laser source. By tightly focusing the excitation spot on the cantilever we were able to attain a deflection efficiency of 0.7 nm µW-1 for the first bending mode. The presented approach results in an efficient all optical excitation and deflection detection

  19. The dual pathway inhibitor rigosertib is effective in direct-patient tumor xenografts of head and neck squamous cell carcinomas

    PubMed Central

    Anderson, Ryan T.; Keysar, Stephen B.; Bowles, Daniel W.; Glogowska, Magdalena J.; Astling, David P.; Morton, J. Jason; Le, Phuong; Umpierrez, Adrian; Eagles-Soukup, Justin; Gan, Gregory N.; Vogler, Brian W.; Sehrt, Daniel; Takimoto, Sarah M.; Aisner, Dara L.; Wilhelm, Francois; Frederick, Barbara A.; Varella-Garcia, Marileila; Tan, Aik-Choon; Jimeno, Antonio

    2013-01-01

    The dual pathway inhibitor rigosertib inhibits phosphoinositide 3-kinase (PI3K) pathway activation as well as polo-like kinase 1 (PLK1) activity across a broad spectrum of cancer cell lines. The importance of PIK3CA alterations in head and neck squamous cell cancer (HNSCC) has raised interest in exploring agents targeting PI3K, the product of PIK3CA. The genetic and molecular basis of rigosertib treatment response was investigated in a panel of 16 HNSCC cell lines, and direct patient tumor xenografts from 8 HNSCC patients (4 HPV16-positive). HNSCC cell lines and xenografts were characterized by pathway enrichment gene expression analysis, exon sequencing, gene copy number, western blotting, and IHC. Rigosertib had potent antiproliferative effects on 11 of the 16 HPV− HNSCC cell lines. Treatment sensitivity was confirmed in two cell lines using an orthotopic in vivo xenograft model. Growth reduction after rigosertib treatment was observed in 3/8 HNSCC direct patient tumor lines. The responsive tumor lines carried a combination of a PI3KCA activating event (amplification or mutation) and a p53 inactivating event (either HPV16-mediated or mutation-mediated TP53 inactivation). In this study, we evaluated the in vitro and in vivo efficacy of rigosertib in both HPV+ and HPV− HNSCCs focusing on inhibition of the PI3K pathway. Although consistent inhibition of the PI3K pathway was not evident in HNSCC, we identified a combination of PI3K/TP53 events necessary, but not sufficient for rigosertib-sensitivity. PMID:23873848

  20. Insight into mechanics of AFM tip-based nanomachining: bending of cantilevers and machined grooves.

    PubMed

    Al-Musawi, R S J; Brousseau, E B; Geng, Y; Borodich, F M

    2016-09-23

    Atomic force microscope (AFM) tip-based nanomachining is currently the object of intense research investigations. Values of the load applied to the tip at the free end of the AFM cantilever probe used for nanomachining are always large enough to induce plastic deformation on the specimen surface contrary to the small load values used for the conventional contact mode AFM imaging. This study describes an important phenomenon specific for AFM nanomachining in the forward direction: under certain processing conditions, the deformed shape of the cantilever probe may change from a convex to a concave orientation. The phenomenon can principally change the depth and width of grooves machined, e.g. the grooves machined on a single crystal copper specimen may increase by 50% on average following such a change in the deformed shape of the cantilever. It is argued that this phenomenon can take place even when the AFM-based tool is operated in the so-called force-controlled mode. The study involves the refined theoretical analysis of cantilever probe bending, the analysis of experimental signals monitored during the backward and forward AFM tip-based machining and the inspection of the topography of produced grooves. PMID:27532247

  1. Probing ternary solvent effect in high Voc polymer solar cells using advanced AFM techniques

    DOE PAGESBeta

    Li, Chao; Soleman, Mikhael; Lorenzo, Josie; Dhasmana, Nitesh; Chantharasupawong, Panit; Ievlev, Anton; Gesquiere, Andre; Tetard, Laurene; Thomas, Jayan

    2016-01-25

    This work describes a simple method to develop a high Voc low band gap PSCs. In addition, two new atomic force microscopy (AFM)-based nanoscale characterization techniques to study the surface morphology and physical properties of the structured active layer are introduced. With the help of ternary solvent processing of the active layer and C60 buffer layer, a bulk heterojunction PSC with Voc more than 0.9 V and conversion efficiency 7.5% is developed. In order to understand the fundamental properties of the materials ruling the performance of the PSCs tested, AFM-based nanoscale characterization techniques including Pulsed-Force-Mode AFM (PFM-AFM) and Mode-Synthesizing AFMmore » (MSAFM) are introduced. Interestingly, MSAFM exhibits high sensitivity for direct visualization of the donor–acceptor phases in the active layer of the PSCs. Lastly, conductive-AFM (cAFM) studies reveal local variations in conductivity in the donor and acceptor phases as well as a significant increase in photocurrent in the PTB7:ICBA sample obtained with the ternary solvent processing.« less

  2. Role of Direct vs. Indirect Pathways from the Motor Cortex to Spinal Motoneurons in the Control of Hand Dexterity

    PubMed Central

    Isa, Tadashi; Kinoshita, Masaharu; Nishimura, Yukio

    2013-01-01

    Evolutionally, development of the direct connection from the motor cortex to spinal motoneurons [corticomotoneuronal (CM) pathway] parallels the ability of hand dexterity. Damage to the corticofugal fibers in higher primates resulted in deficit of fractionated digit movements. Based on such observations, it was generally believed that the CM pathway plays a critical role in the control of hand dexterity. On the other hand, a number of “phylogenetically older” indirect pathways from the motor cortex to motoneurons still exist in primates. The indirect pathways are mediated by intercalated neurons such as segmental interneurons (sINs), propriospinal neurons (PNs) reticulospinal neurons (RSNs), or rubrospinal neurons (RuSNs). However, their contribution to hand dexterity remains elusive. Lesion of the brainstem pyramid sparing the transmission through the RuSNs and RSNs, resulted in permanent deficit of fractionated digit movements in macaque monkeys. On the other hand, in our recent study, after lesion of the dorsolateral funiculus (DLF) at the C5 segment, which removed the lateral corticospinal tract (l-CST) including the CM pathway and the transmission through sINs and RuSNs but spared the processing through the PNs and RSNs, fractionated digit movements recovered within several weeks. These results suggest that the PNs can be involved in the recovery of fractionated digit movements, but the RSNs and RuSNs have less capacity in this regard. However, on closer inspection, it was found that the activation pattern of hand and arm muscles considerably changed after the C5 lesion, suggesting limitation of PNs for the compensation of hand dexterity. Altogether, it is suggested that PNs, RSNs RuSNs, and the CM pathway (plus sINs) make a different contribution to the hand dexterity and appearance of motor deficit of the hand dexterity caused by damage to the corticofugal fibers and potential of recovery varies depending on the rostrocaudal level of the lesion. PMID

  3. A novel pathway of direct methane production and emission by eukaryotes including plants, animals and fungi: An overview

    NASA Astrophysics Data System (ADS)

    Liu, Jiangong; Chen, Huai; Zhu, Qiuan; Shen, Yan; Wang, Xue; Wang, Meng; Peng, Changhui

    2015-08-01

    Methane (CH4) is a powerful greenhouse gas with a global warming potential 28 times that of carbon dioxide (CO2). CH4 is responsible for approximately 20% of the Earth's warming since pre-industrial times. Knowledge of the sources of CH4 is crucial due to the recent substantial interannual variability of growth rates and uncertainties regarding individual sources. The prevailing paradigm is that methanogenesis carried out by methanogenic archaea occurs primarily under strictly anaerobic conditions. However, in the past decade, studies have confirmed direct CH4 release from three important kingdoms of eukaryotes-Plantae, Animalia and Fungi-even in the presence of oxygen. This novel CH4 production pathway has been aptly termed "aerobic CH4 production" to distinguish it from the well-known anaerobic CH4 production pathway, which involves catalytic activity by methanogenic archaeal enzymes. In this review, we collated recent experimental evidence from the published literature and documented this novel pathway of direct CH4 production and emission by eukaryotes. The mechanisms involved in this pathway may be related to protective strategies of eukaryotes in response to changing environmental stresses, with CH4 a by-product or end-product during or at the end of the process(es) that originates from organic methyl-type compounds. Based on the existing, albeit uncertain estimates, plants seem to contribute less to the global CH4 budget (3-24%) compared to previous estimates (10-37%). We still lack estimates of CH4 emissions by animals and fungi. Overall, there is an urgent need to identify the precursors for this novel CH4 source and improve our understanding of the mechanisms of direct CH4 production and the impacts of environmental stresses. An estimate of this new CH4 source, which was not considered as a CH4 source by the Intergovernmental Panel on Climate Change (IPCC) (2013), could be useful for better quantitation of the global CH4 budget.

  4. Detection of Pathogens Using AFM and SPR

    NASA Astrophysics Data System (ADS)

    Vaseashta, Ashok

    2005-03-01

    A priori detection of pathogens in food and water has become a subject of paramount importance. Several recent incidents have resulted in the government passing stringent regulations for tolerable amounts of contamination of food products. Identification and/or monitoring of bacterial contamination in food are critical. The conventional methods of pathogen detection require time-consuming steps to arrive disembark at meaningful measurement in a timely manner as the detection time exceeds the time in which perishable food recycles through the food chain distribution. The aim of this presentation is to outline surface plasmon resonance (SPR) and atomic force microscopy (AFM) as two methods for fast detect6ion of pathogens. Theoretical basis of SPR and experimental results of SPR and AFM on E. coli O157:H7 and prion are presented.

  5. Direct accumulation pathway of radioactive cesium to fruit-bodies of edible mushroom from contaminated wood logs

    PubMed Central

    Ohnuki, Toshihiko; Aiba, Yukitoshi; Sakamoto, Fuminori; Kozai, Naofumi; Niizato, Tadafumi; Sasaki, Yoshito

    2016-01-01

    This paper presents the accumulation process of radioactive Cs in edible mushrooms. We here first report the direct accumulation pathway of radioactive Cs from contaminated wood logs to the fruit-bodies of shiitake mushrooms through the basal portion of the stipe. In this pathway, radioactive Cs is not transported through the hyphae. This pathway results in a high accumulation of radioactive Cs in the fruit-body, more by the excess accumulation of radioactive Cs from the wood logs than that through the hyphae. We grew the fruit-bodies of Shiitake mushroom from radioactive-Cs-contaminated wood logs. The spatial distributions of radioactive Cs and Prussian blue as a tracer of interstitial water in the cross section of the wood log measured after the harvest of the fruit-body from the inoculated sawdust spawn area indicated that some fraction of the radioactive Cs and Prussian blue were transported directly to the basal portion of the stipe during the growth of the fruit-bodies. PMID:27430163

  6. Direct accumulation pathway of radioactive cesium to fruit-bodies of edible mushroom from contaminated wood logs

    NASA Astrophysics Data System (ADS)

    Ohnuki, Toshihiko; Aiba, Yukitoshi; Sakamoto, Fuminori; Kozai, Naofumi; Niizato, Tadafumi; Sasaki, Yoshito

    2016-07-01

    This paper presents the accumulation process of radioactive Cs in edible mushrooms. We here first report the direct accumulation pathway of radioactive Cs from contaminated wood logs to the fruit-bodies of shiitake mushrooms through the basal portion of the stipe. In this pathway, radioactive Cs is not transported through the hyphae. This pathway results in a high accumulation of radioactive Cs in the fruit-body, more by the excess accumulation of radioactive Cs from the wood logs than that through the hyphae. We grew the fruit-bodies of Shiitake mushroom from radioactive-Cs-contaminated wood logs. The spatial distributions of radioactive Cs and Prussian blue as a tracer of interstitial water in the cross section of the wood log measured after the harvest of the fruit-body from the inoculated sawdust spawn area indicated that some fraction of the radioactive Cs and Prussian blue were transported directly to the basal portion of the stipe during the growth of the fruit-bodies.

  7. Direct accumulation pathway of radioactive cesium to fruit-bodies of edible mushroom from contaminated wood logs.

    PubMed

    Ohnuki, Toshihiko; Aiba, Yukitoshi; Sakamoto, Fuminori; Kozai, Naofumi; Niizato, Tadafumi; Sasaki, Yoshito

    2016-01-01

    This paper presents the accumulation process of radioactive Cs in edible mushrooms. We here first report the direct accumulation pathway of radioactive Cs from contaminated wood logs to the fruit-bodies of shiitake mushrooms through the basal portion of the stipe. In this pathway, radioactive Cs is not transported through the hyphae. This pathway results in a high accumulation of radioactive Cs in the fruit-body, more by the excess accumulation of radioactive Cs from the wood logs than that through the hyphae. We grew the fruit-bodies of Shiitake mushroom from radioactive-Cs-contaminated wood logs. The spatial distributions of radioactive Cs and Prussian blue as a tracer of interstitial water in the cross section of the wood log measured after the harvest of the fruit-body from the inoculated sawdust spawn area indicated that some fraction of the radioactive Cs and Prussian blue were transported directly to the basal portion of the stipe during the growth of the fruit-bodies. PMID:27430163

  8. Morpholine Degradation Pathway of Mycobacterium aurum MO1: Direct Evidence of Intermediates by In Situ 1H Nuclear Magnetic Resonance

    PubMed Central

    Combourieu, B.; Besse, P.; Sancelme, M.; Veschambre, H.; Delort, A. M.; Poupin, P.; Truffaut, N.

    1998-01-01

    Resting Mycobacterium aurum MO1 cells were incubated with morpholine, a waste from the chemical industry. The kinetics of biodegradation was monitored by using in situ nuclear magnetic resonance (NMR). The incubation medium was directly analyzed by 1H NMR. This technique allowed the unambiguous identification of two intermediates of the metabolic pathway involved in the biodegradation process, glycolate and 2-(2-aminoethoxy)acetate. The latter compound, which was not commercially available, was synthesized, in three steps, from 2-(2-aminoethoxy)ethanol. Quantitative analysis of the kinetics of degradation of morpholine was performed by integrating the signals of the different metabolites in 1H-NMR spectra. Morpholine was degraded within 10 h. The intermediates increased during the first 10 h and finally disappeared after 20 h incubation. Assays of degradation were also carried out with glycolate and ethanolamine, hypothetical intermediates of the morpholine degradation pathway. They were degraded within 4 and 8 h, respectively. Until now, no tool for direct detection of intermediates or even morpholine has been available, consequently, only hypothetical pathways have been proposed. The approach described here gives both qualitative and quantitative information about the metabolic routes used in morpholine degradation by M. aurum MO1. It could be used to investigate many biodegradative processes. PMID:9435073

  9. Characterizing Cell Mechanics with AFM and Microfluidics

    NASA Astrophysics Data System (ADS)

    Walter, N.; Micoulet, A.; Suresh, S.; Spatz, J. P.

    2007-03-01

    Cell mechanical properties and functionality are mainly determined by the cytoskeleton, besides the cell membrane, the nucleus and the cytosol, and depend on various parameters e.g. surface chemistry and rigidity, surface area and time available for cell spreading, nutrients and drugs provided in the culture medium. Human epithelial pancreatic and mammary cancer cells and their keratin intermediate filaments are the main focus of our work. We use Atomic Force Microscopy (AFM) to study cells adhering to substrates and Microfluidic Channels to probe cells in suspension, respectively. Local and global properties are extracted by varying AFM probe tip size and the available adhesion area for cells. Depth-sensing, instrumented indentation tests with AFM show a clear difference in contact stiffness for cells that are spread of controlled substrates and those that are loosely attached. Microfluidic Channels are utilized in parallel to evaluate cell deformation and ``flow resistance'', which are dependent on channel cross section, flow rate, cell nucleus size and the mechanical properties of cytoskeleton and membrane. The results from the study are used to provide some broad and quantitative assessments of the connections between cellular/subcellular mechanics and biochemical origins of disease states.

  10. Resveratrol directly targets DDX5 resulting in suppression of the mTORC1 pathway in prostate cancer

    PubMed Central

    Taniguchi, T; Iizumi, Y; Watanabe, M; Masuda, M; Morita, M; Aono, Y; Toriyama, S; Oishi, M; Goi, W; Sakai, T

    2016-01-01

    Resveratrol has various attractive bioactivities, such as prevention of cancer, neurodegenerative disorders, and obesity-related diseases. Therefore, identifying its direct binding proteins is expected to discover druggable targets. Sirtuin 1 and phosphodiesterases have so far been found as the direct molecular targets of resveratrol. We herein identified 11 novel resveratrol-binding proteins, including the DEAD (Asp-Glu-Ala-Asp) box helicase 5 (DDX5, also known as p68), using resveratrol-immobilized beads. Treatment with resveratrol induced degradation of DDX5 in prostate cancer cells. Depletion of DDX5 caused apoptosis by inhibiting mammalian target of rapamycin complex 1 (mTORC1) signaling. Moreover, knockdown of DDX5 attenuated the inhibitory activities of resveratrol against mTORC1 signaling and cancer cell growth. These data show that resveratrol directly targets DDX5 and induces cancer cell death by inhibiting the mTORC1 pathway. PMID:27148684

  11. Microbial production of natural and non-natural flavonoids: Pathway engineering, directed evolution and systems/synthetic biology.

    PubMed

    Pandey, Ramesh Prasad; Parajuli, Prakash; Koffas, Mattheos A G; Sohng, Jae Kyung

    2016-01-01

    In this review, we address recent advances made in pathway engineering, directed evolution, and systems/synthetic biology approaches employed in the production and modification of flavonoids from microbial cells. The review is divided into two major parts. In the first, various metabolic engineering and system/synthetic biology approaches used for production of flavonoids and derivatives are discussed broadly. All the manipulations/engineering accomplished on the microorganisms since 2000 are described in detail along with the biosynthetic pathway enzymes, their sources, structures of the compounds, and yield of each product. In the second part of the review, post-modifications of flavonoids by four major reactions, namely glycosylations, methylations, hydroxylations and prenylations using recombinant strains are described. PMID:26946281

  12. AMP-activated Protein Kinase Directly Phosphorylates and Destabilizes Hedgehog Pathway Transcription Factor GLI1 in Medulloblastoma

    PubMed Central

    Li, Yen-Hsing; Luo, Jia; Mosley, Yung-Yi C.; Hedrick, Victoria E.; Paul, Lake N.; Chang, Julia; Zhang, GuangJun; Wang, Yu-Kuo; Banko, Max R.; Brunet, Anne; Kuang, Shihuan; Wu, Jen-Leih; Chang, Chun-Ju; Scott, Matthew P.; Yang, Jer-Yen

    2015-01-01

    Summary The Hedgehog (Hh) pathway regulates cell differentiation and proliferation during development by controlling the Gli transcription factors. Cell fate decisions and progression toward organ and tissue maturity must be coordinated and how energy sensor regulates Hh pathway is not clear. AMP-activated Protein Kinase (AMPK) is an important sensor of energy stores that controls protein synthesis and other energy-intensive processes. AMPK is directly responsive to intracellular AMP levels, inhibiting a wide range of cell activities if ATP is low and AMP is high. Thus, AMPK can affect development by influencing protein synthesis and other processes needed for growth and differentiation. Activation of AMPK reduces GLI1 protein levels and stability, thus blocking Sonic hedgehog-induced transcriptional activity. AMPK phosphorylates GLI1 at serines 102 and 408 and threonine 1074. Mutation of these three sites into alanine prevents phosphorylation by AMPK. This in turn leads to increased GLI1 protein stability, transcriptional activity, and oncogenic potency. PMID:26190112

  13. Role of the direct and indirect pathways for glycogen synthesis in rat liver in the postprandial state

    SciTech Connect

    Huang, M.T.; Veech, R.L.

    1988-03-01

    The pathway for hepatic glycogen synthesis in the postprandial state was studied in meal-fed rats chronically cannulated in the portal vein. Plasma glucose concentration in the portal vein was found to be 4.50 +/- 1.01 mM (mean +/- SE; n = 3) before a meal and 11.54 +/- 0.70 mM (mean +/- SE; n = 4) after a meal in rats meal-fed a diet consisting of 100% commercial rat chow for 7 d. The hepatic-portal difference of plasma glucose concentration showed that liver released glucose in the fasted state and either extracted or released glucose after feeding depending on plasma glucose concentration in the portal vein. The concentration of portal vein glucose at which liver changes from glucose releasing to glucose uptake was 8 mM, the Km of glucokinase. The rate of glycogen synthesis in liver during meal-feeding was found to be approximately 1 mumol glucosyl U/g wet wt/min in rats meal-fed a 50% glucose supplemented chow diet. The relative importance of the direct vs. indirect pathway for the replenishment of hepatic glycogen was determined by the incorporation of (3-/sup 3/H,U-/sup 14/C)glucose into liver glycogen. Labeled glucose was injected into the portal vein at the end of meal-feeding. The ratio of /sup 3/H//sup 14/C in the glucosyl units of glycogen was found to be 83-92% of the ratio in liver free glucose six minutes after the injection, indicating that the majority of exogenous glucose incorporated into glycogen did not go through glycolysis. The percent contribution of the direct versus indirect pathway was quantitated from the difference in the relative specific activity (RSA) of (/sup 3/H) and (/sup 14/C)-glycogen in rats infused with (3-/sup 3/H,U-/sup 14/C)glucose. No significant difference was found between the RSA of (/sup 3/H)glycogen and (/sup 14/C)glycogen, indicating further that the pathway for glycogen synthesis in liver from exogenous glucose is from the direct pathway.

  14. A versatile pathway for the direct assembly of organo-functional mesostructures from sodium silicate.

    PubMed

    Shah, Jainisha; Kim, Seong-Su; Pinnavaia, Thomas J

    2004-03-01

    Silica mesostructures with well-expressed hexagonal and wormhole framework structures and up to 50% organo-functionalization of the framework silicon sites have been directly assembled from sodium silicate. PMID:14973613

  15. A family of transketolases that directs isoprenoid biosynthesis via a mevalonate-independent pathway.

    PubMed

    Lange, B M; Wildung, M R; McCaskill, D; Croteau, R

    1998-03-01

    Isopentenyl diphosphate, the common precursor of all isoprenoids, has been widely assumed to be synthesized by the acetate/mevalonate pathway in all organisms. However, based on in vivo feeding experiments, isopentenyl diphosphate formation in several eubacteria, a green alga, and plant chloroplasts has been demonstrated very recently to originate via a mevalonate-independent route from pyruvate and glyceraldehyde 3-phosphate as precursors. Here we describe the cloning from peppermint (Mentha x piperita) and heterologous expression in Escherichia coli of 1-deoxy-D-xylulose-5-phosphate synthase, the enzyme that catalyzes the first reaction of this pyruvate/glyceraldehyde 3-phosphate pathway. This synthase gene contains an ORF of 2,172 base pairs. When the proposed plastid targeting sequence is excluded, the deduced amino acid sequence indicates the peppermint synthase to be about 650 residues in length, corresponding to a native size of roughly 71 kDa. The enzyme appears to represent a novel class of highly conserved transketolases and likely plays a key role in the biosynthesis of plastid-derived isoprenoids essential for growth, development, and defense in plants. PMID:9482845

  16. New Direction in Hydrogeochemical Transport Modeling: Incorporating Multiple Kinetic and Equilibrium Reaction Pathways

    SciTech Connect

    Steefel, C.I.

    2000-02-02

    At least two distinct kinds of hydrogeochemical models have evolved historically for use in analyzing contaminant transport, but each has important limitations. One kind, focusing on organic contaminants, treats biodegradation reactions as parts of relatively simple kinetic reaction networks with no or limited coupling to aqueous and surface complexation and mineral dissolution/precipitation reactions. A second kind, evolving out of the speciation and reaction path codes, is capable of handling a comprehensive suite of multicomponent complexation (aqueous and surface) and mineral precipitation and dissolution reactions, but has not been able to treat reaction networks characterized by partial redox disequilibrium and multiple kinetic pathways. More recently, various investigators have begun to consider biodegradation reactions in the context of comprehensive equilibrium and kinetic reaction networks (e.g. Hunter et al. 1998, Mayer 1999). Here we explore two examples of multiple equilibrium and kinetic reaction pathways using the reactive transport code GIMRT98 (Steefel, in prep.): (1) a computational example involving the generation of acid mine drainage due to oxidation of pyrite, and (2) a computational/field example where the rates of chlorinated VOC degradation are linked to the rates of major redox processes occurring in organic-rich wetland sediments overlying a contaminated aerobic aquifer.

  17. Direct contact between dust and HBCD-treated fabrics is an important pathway of source-to-dust transfer.

    PubMed

    Rauert, Cassandra; Kuribara, Isamu; Kataoka, Toshiyuki; Wada, Takeharu; Kajiwara, Natsuko; Suzuki, Go; Takigami, Hidetaka; Harrad, Stuart

    2016-03-01

    Hexabromocyclododecanes (HBCDs) are a class of brominated flame retardant that have found extensive application in consumer products used widely in indoor environments. Although uncertainty remains about the human health impacts of HBCDs, ingestion of HBCD-contaminated indoor dust has been shown to be a particularly significant exposure pathway for young children. Despite this, understanding of the mechanisms via which HBCD transfer from products to indoor dust remains incomplete. In this study, an in-house test chamber was used to investigate transfer of HBCDs from a treated textile sample to indoor dust via direct textile:dust contact. Results were compared with previous data using the same test chamber to examine other pathways via which HBCDs transfer from products to dust, and highlighted HBCD transfer via direct source:dust contact as being particularly important. This novel finding was corroborated by complementary experiments that examined HBCD transfer via direct contact, from other treated textiles to three major components of indoor dust: artificial indoor dust, soil particles, and cotton linters. PMID:26745295

  18. A combinatorial F box protein directed pathway controls TRAF adaptor stability to regulate inflammation.

    PubMed

    Chen, Bill B; Coon, Tiffany A; Glasser, Jennifer R; McVerry, Bryan J; Zhao, Jing; Zhao, Yutong; Zou, Chunbin; Ellis, Bryon; Sciurba, Frank C; Zhang, Yingze; Mallampalli, Rama K

    2013-05-01

    Uncontrolled activation of tumor necrosis factor receptor-associated factor (TRAF) proteins may result in profound tissue injury by linking surface signals to cytokine release. Here we show that a ubiquitin E3 ligase component, Fbxo3, potently stimulates cytokine secretion from human inflammatory cells by destabilizing a sentinel TRAF inhibitor, Fbxl2. Fbxo3 and TRAF protein in circulation positively correlated with cytokine responses in subjects with sepsis, and we identified a polymorphism in human Fbxo3, with one variant being hypofunctional. A small-molecule inhibitor targeting Fbxo3 was sufficient to lessen severity of cytokine-driven inflammation in several mouse disease models. These studies identified a pathway of innate immunity that may be useful to detect subjects with altered immune responses during critical illness or provide a basis for therapeutic intervention targeting TRAF protein abundance. PMID:23542741

  19. The Spn4 gene of Drosophila encodes a potent furin-directed secretory pathway serpin

    PubMed Central

    Richer, Martin J.; Keays, Clairessa A.; Waterhouse, Jennifer; Minhas, Jessey; Hashimoto, Carl; Jean, François

    2004-01-01

    Proprotein convertases (PCs) are an important class of host-cell serine endoproteases implicated in many physiological and pathological processes. Owing to their expanding roles in the proteolytic events required for generating infectious microbial pathogens and for tumor growth and invasiveness, there is increasing interest in identifying endogenous PC inhibitors. Here we report the identification of Spn4A, a previously uncharacterized secretory pathway serine protease inhibitor (serpin) from Drosophila melanogaster that contains a consensus furin cleavage site, -ArgP4-Arg-Lys-ArgP1↓-, in its reactive site loop (RSL). Our biochemical and kinetics analysis revealed that recombinant Spn4A inhibits human furin (Ki, 13 pM; kass, 3.2 × 107 M–1·s–1) and Drosophila PC2 (Ki, 3.5 nM; kass, 9.2 × 104 M–1·s–1) by a slow-binding mechanism characteristic of serpin molecules and forms a kinetically trapped SDS-stable complex with each enzyme. For both PCs, the stoichiometry of inhibition by Spn4A is nearly 1, which is characteristic of known physiological serpin–protease interactions. Mass analysis of furin–Spn4A reaction products identified the actual reactive site center of Spn4A to be -ArgP4-Arg-Lys-ArgP1↓-. Moreover, we demonstrate that Spn4A's highly effective PC inhibition properties are critically dependent on the unusual length of its RSL, which is composed of 18 aa instead of the typical 17-residue RSL found in most other inhibitory serpins. The identification of Spn4A, the most potent and effective natural serpin of PCs identified to date, suggests that Spn4A could be a prototype of endogenous serpins involved in the precise regulation of PC-dependent proteolytic cleavage events in the secretory pathway of eukaryotic cells. PMID:15247425

  20. Dual AFM probes alignment based on vision guidance

    NASA Astrophysics Data System (ADS)

    Zhang, Hua-kun; Gao, Si-tian; Lu, Ming-zhen; Wang, Long-long

    2013-10-01

    Atomic force microscope (AFM) with dual probes that operate together can measure both side walls excellently at the same time, which virtually eliminates the prevalent effect of probe width that contributes a large component of uncertainty in measurement results and finally obtains the critical dimension (CD)(e.g. the linewidth) through data synthesis. In calibration process, the dual probes must contact each other in advance, which realizes the alignment in the three dimensions, to establish a zero reference point and ensure the accuracy of measurement. Because nowadays the optical resolution of advanced lens have exceeded micrometer range, and the size of probes is within micro level, it is possible to acquire dual probes images in both horizontal and vertical directions, through which the movement of the probes can be controlled in time. In order to further enhance the alignment precision, sub-pixel edge detection method based on Zernike orthogonal moment is used to obtain relative position between these two probes, which helps the tips alignment attains sub-micron range. Piezoelectric nanopositioning stages calibrated by laser interferometer are used to implement fine movement of the probes to verify the accuracy of the experimental results. To simplify the system, novel self-sensing and self-actuating probe based on a quartz tuning fork combined with a micromachined cantilever is used for dynamic mode AFM. In this case, an external optical detection system is not needed, so the system is simple and small.

  1. BOREAS AFM-6 Surface Meteorological Data

    NASA Technical Reports Server (NTRS)

    Wilczak, James; Hall, Forrest G. (Editor); Newcomer, Jeffrey A. (Editor); Smith, David E. (Technical Monitor)

    2000-01-01

    The Boreal Ecosystem-Atmosphere Study (BOREAS) Airborne Fluxes and Meteorology (AFM)-6 team from the National Oceanic and Atmospheric Adminsitration/Environment Technology Laboratory (NOAA/ETL) collected surface meteorological data from 21 May to 20 Sep 1994 near the Southern Study Area-Old Jack Pine (SSA-OJP) tower site. The data are in tabular ASCII files. The surface meteorological data are available from the Earth Observing System Data and Information System (EOSDIS) Oak Ridge National Laboratory (ORNL) Distributed Active Archive Center (DAAC). The data files are available on a CD-ROM (see document number 20010000884).

  2. Involvement of the Striatal Medium Spiny Neurons of the Direct Pathway in the Motor Stimulant Effects of Phencyclidine

    PubMed Central

    Bonito-Oliva, Alessandra; DuPont, Caitlin; Madjid, Nather; Ögren, Sven Ove

    2016-01-01

    Background: The psychotomimetic phencyclidine (PCP) produces behavioral symptoms similar to those observed in schizophrenia, accompanied by increased motor activity. The dopamine and adenosine 3’,5’-cyclic monophosphate-regulated phosphoprotein of 32kDa (DARPP-32) is enriched in the medium spiny neurons (MSNs) of the striatum and has been implicated in the actions of PCP. We examined the effects of deletion of DARPP-32 in distinct populations of striatal MSNs, on the ability of PCP to induce motor activation and memory deficit. Methods: The effects of PCP were examined in mice with conditional knockout of DARPP-32 in the MSNs of the direct, or indirect pathway. DARPP-32 phosphorylation was determined by Western blotting. The motor stimulant effects of PCP were determined by measuring locomotion following acute and chronic administration. Memory deficit was evaluated using the passive avoidance test. Results: Loss of DARPP-32 in direct MSNs prevents PCP-induced phosphorylation and abolishes the motor stimulation effects of PCP. In contrast, lack of DARPP-32 in indirect MSNs does not affect the ability of PCP to promote DARPP-32 phosphorylation and to increase motor activity. The impairment in passive avoidance induced by PCP is independent of the expression of DARPP-32 in direct or indirect MSNs. Conclusions: The increase in DARPP-32 phosphorylation induced by PCP occurs selectively in the MSNs of the direct pathway, which are also specifically involved in the motor stimulant effects of this drug. The memory deficit induced by PCP is not linked to the expression of DARPP-32 in striatal MSNs. PMID:26657176

  3. Mechanisms of direct inhibition of the respiratory sulfate-reduction pathway by (per)chlorate and nitrate

    PubMed Central

    Carlson, Hans K; Kuehl, Jennifer V; Hazra, Amrita B; Justice, Nicholas B; Stoeva, Magdalena K; Sczesnak, Andrew; Mullan, Mark R; Iavarone, Anthony T; Engelbrektson, Anna; Price, Morgan N; Deutschbauer, Adam M; Arkin, Adam P; Coates, John D

    2015-01-01

    We investigated perchlorate (ClO4−) and chlorate (ClO3−) (collectively (per)chlorate) in comparison with nitrate as potential inhibitors of sulfide (H2S) production by mesophilic sulfate-reducing microorganisms (SRMs). We demonstrate the specificity and potency of (per)chlorate as direct SRM inhibitors in both pure cultures and undefined sulfidogenic communities. We demonstrate that (per)chlorate and nitrate are antagonistic inhibitors and resistance is cross-inducible implying that these compounds share at least one common mechanism of resistance. Using tagged-transposon pools we identified genes responsible for sensitivity and resistance in Desulfovibrio alaskensis G20. We found that mutants in Dde_2702 (Rex), a repressor of the central sulfate-reduction pathway were resistant to both (per)chlorate and nitrate. In general, Rex derepresses its regulon in response to increasing intracellular NADH:NAD+ ratios. In cells in which respiratory sulfate reduction is inhibited, NADH:NAD+ ratios should increase leading to derepression of the sulfate-reduction pathway. In support of this, in (per)chlorate or nitrate-stressed wild-type G20 we observed higher NADH:NAD+ ratios, increased transcripts and increased peptide counts for genes in the core Rex regulon. We conclude that one mode of (per)chlorate and nitrate toxicity is as direct inhibitors of the central sulfate-reduction pathway. Our results demonstrate that (per)chlorate are more potent inhibitors than nitrate in both pure cultures and communities, implying that they represent an attractive alternative for controlling sulfidogenesis in industrial ecosystems. Of these, perchlorate offers better application logistics because of its inhibitory potency, solubility, relative chemical stability, low affinity for mineral cations and high mobility in environmental systems. PMID:25405978

  4. Mechanisms of direct inhibition of the respiratory sulfate-reduction pathway by (per)chlorate and nitrate.

    PubMed

    Carlson, Hans K; Kuehl, Jennifer V; Hazra, Amrita B; Justice, Nicholas B; Stoeva, Magdalena K; Sczesnak, Andrew; Mullan, Mark R; Iavarone, Anthony T; Engelbrektson, Anna; Price, Morgan N; Deutschbauer, Adam M; Arkin, Adam P; Coates, John D

    2015-06-01

    We investigated perchlorate (ClO(4)(-)) and chlorate (ClO(3)(-)) (collectively (per)chlorate) in comparison with nitrate as potential inhibitors of sulfide (H(2)S) production by mesophilic sulfate-reducing microorganisms (SRMs). We demonstrate the specificity and potency of (per)chlorate as direct SRM inhibitors in both pure cultures and undefined sulfidogenic communities. We demonstrate that (per)chlorate and nitrate are antagonistic inhibitors and resistance is cross-inducible implying that these compounds share at least one common mechanism of resistance. Using tagged-transposon pools we identified genes responsible for sensitivity and resistance in Desulfovibrio alaskensis G20. We found that mutants in Dde_2702 (Rex), a repressor of the central sulfate-reduction pathway were resistant to both (per)chlorate and nitrate. In general, Rex derepresses its regulon in response to increasing intracellular NADH:NAD(+) ratios. In cells in which respiratory sulfate reduction is inhibited, NADH:NAD(+) ratios should increase leading to derepression of the sulfate-reduction pathway. In support of this, in (per)chlorate or nitrate-stressed wild-type G20 we observed higher NADH:NAD(+) ratios, increased transcripts and increased peptide counts for genes in the core Rex regulon. We conclude that one mode of (per)chlorate and nitrate toxicity is as direct inhibitors of the central sulfate-reduction pathway. Our results demonstrate that (per)chlorate are more potent inhibitors than nitrate in both pure cultures and communities, implying that they represent an attractive alternative for controlling sulfidogenesis in industrial ecosystems. Of these, perchlorate offers better application logistics because of its inhibitory potency, solubility, relative chemical stability, low affinity for mineral cations and high mobility in environmental systems. PMID:25405978

  5. Targeted Expression of Mu-Opioid Receptors in a Subset of Striatal Direct-Pathway Neurons Restores Opiate Reward

    PubMed Central

    Cui, Yijun; Ostlund, Sean B.; James, Alex; Park, Chang Sin; Ge, Weihong; Roberts, Kristofer W.; Mittal, Nitish; Murphy, Niall P.; Cepeda, Carlos; Kieffer, Brigitte L.; Levine, Michael S.; Jentsch, J. David; Walwyn, Wendy M.; Sun, Yi E.; Evans, Christopher J.; Maidment, Nigel T.; Yang, X. William

    2014-01-01

    SUMMARY Mu-Opioid Receptors (MOR) are necessary for the analgesic and addictive effects of opioids such as morphine, but the MOR-expressing neuronal populations that mediate the distinct opiate effects remain elusive. Here we devised a novel conditional BAC rescue strategy to show that mice with targeted MOR expression in a subpopulation of striatal direct-pathway neurons enriched in the striosome and nucleus accumbens, in an otherwise MOR-null background, restore opiate reward, opiate-induced striatal dopamine release, and partially restore motivation to self-administer opiates. However, they lack opiate analgesia or withdrawal. Importantly, we used Cre-mediated deletion of the rescued MOR transgene to establish that striatal, rather than a few extrastriatal sites of MOR transgene expression, is needed for the restoration of opiate reward. Together, our study demonstrates that a subpopulation of striatal direct-pathway neurons is sufficient to support opiate reward-driven behaviors and provides a novel intersectional genetic approach to dissect neurocircuit-specific gene function in vivo. PMID:24413699

  6. Directed Evolution of Metabolic Pathways in Microbial Populations. I. Modification of the Acid Phosphatase Ph Optimum in S. CEREVISIAE

    PubMed Central

    Francis, J. C.; Hansche, P. E.

    1972-01-01

    An experimental system for directing the evolution of enzymes and metabolic pathways in microbial populations is proposed and an initial test of its power is provided.—The test involved an attempt to genetically enhance certain functional properties of the enzyme acid phosphatase in S. cerevisiae by constructing an environment in which the functional changes desired would be "adaptive". Naturally occurring mutations in a population of 109 cells were automatically and continuously screened, over 1,000 generations, for their effect on the efficiency (Km) and activity of acid phosphatase at pH 6, and for their effect on the efficiency of orthophosphate metabolism.—The first adaptation observed, M1, was due to a single mutational event that effected a 30% increase in the efficiency of orthophosphate metabolism. The second, M2, effected an adaptive shift in the pH optimum of acid phosphatase and an increase in its activity over a wide range of pH values (an increment of 60% at pH 6). M2 was shown to result from a single mutational event in the region of the acid phosphatase structural gene. The third, M3, effected cell clumping, an adaptation to the culture apparatus that had no effect on phosphate metabolism.—The power of this system for directing the evolution of enzymes and of metabolic pathways is discussed in terms of the kinetic properties of the experimental system and in terms of the results obtained. PMID:4552227

  7. The GABAergic septohippocampal pathway is directly involved in internal processes related to operant reward learning.

    PubMed

    Vega-Flores, Germán; Rubio, Sara E; Jurado-Parras, M Teresa; Gómez-Climent, María Ángeles; Hampe, Christiane S; Manto, Mario; Soriano, Eduardo; Pascual, Marta; Gruart, Agnès; Delgado-García, José M

    2014-08-01

    We studied the role of γ-aminobutyric acid (GABA)ergic septohippocampal projections in medial septum (MS) self-stimulation of behaving mice. Self-stimulation was evoked in wild-type (WT) mice using instrumental conditioning procedures and in J20 mutant mice, a type of mouse with a significant deficit in GABAergic septohippocampal projections. J20 mice showed a significant modification in hippocampal activities, including a different response for input/output curves and the paired-pulse test, a larger long-term potentiation (LTP), and a delayed acquisition and lower performance in the MS self-stimulation task. LTP evoked at the CA3-CA1 synapse further decreased self-stimulation performance in J20, but not in WT, mice. MS self-stimulation evoked a decrease in the amplitude of field excitatory postsynaptic potentials (fEPSPs) at the CA3-CA1 synapse in WT, but not in J20, mice. This self-stimulation-dependent decrease in the amplitude of fEPSPs was also observed in the presence of another positive reinforcer (food collected during an operant task) and was canceled by the local administration of an antibody-inhibiting glutamate decarboxylase 65 (GAD65). LTP evoked in the GAD65Ab-treated group was also larger than in controls. The hippocampus has a different susceptibility to septal GABAergic inputs depending on ongoing cognitive processes, and the GABAergic septohippocampal pathway is involved in consummatory processes related to operant rewards. PMID:23479403

  8. The GABAergic Septohippocampal Pathway Is Directly Involved in Internal Processes Related to Operant Reward Learning

    PubMed Central

    Vega-Flores, Germán; Rubio, Sara E.; Jurado-Parras, M. Teresa; Gómez-Climent, María Ángeles; Hampe, Christiane S.; Manto, Mario; Soriano, Eduardo; Pascual, Marta; Gruart, Agnès; Delgado-García, José M.

    2014-01-01

    We studied the role of γ-aminobutyric acid (GABA)ergic septohippocampal projections in medial septum (MS) self-stimulation of behaving mice. Self-stimulation was evoked in wild-type (WT) mice using instrumental conditioning procedures and in J20 mutant mice, a type of mouse with a significant deficit in GABAergic septohippocampal projections. J20 mice showed a significant modification in hippocampal activities, including a different response for input/output curves and the paired-pulse test, a larger long-term potentiation (LTP), and a delayed acquisition and lower performance in the MS self-stimulation task. LTP evoked at the CA3–CA1 synapse further decreased self-stimulation performance in J20, but not in WT, mice. MS self-stimulation evoked a decrease in the amplitude of field excitatory postsynaptic potentials (fEPSPs) at the CA3–CA1 synapse in WT, but not in J20, mice. This self-stimulation-dependent decrease in the amplitude of fEPSPs was also observed in the presence of another positive reinforcer (food collected during an operant task) and was canceled by the local administration of an antibody-inhibiting glutamate decarboxylase 65 (GAD65). LTP evoked in the GAD65Ab-treated group was also larger than in controls. The hippocampus has a different susceptibility to septal GABAergic inputs depending on ongoing cognitive processes, and the GABAergic septohippocampal pathway is involved in consummatory processes related to operant rewards. PMID:23479403

  9. A bacterial signal peptide is functional in plants and directs proteins to the secretory pathway

    PubMed Central

    Moeller, Lorena; Gan, Qinglei; Wang, Kan

    2009-01-01

    The Escherichia coli heat-labile enterotoxin B subunit (LT-B) has been used as a model antigen for the production of plant-derived high-valued proteins in maize. LT-B with its native signal peptide (BSP) has been shown to accumulate in starch granules of transgenic maize kernels. To elucidate the targeting properties of the bacterial LT-B protein and BSP in plant systems, the subcellular localization of visual marker green fluorescent protein (GFP) fused to LT-B and various combinations of signal peptides was examined in Arabidopsis protoplasts and transgenic maize. Biochemical analysis indicates that the LT-B::GFP fusion proteins can assemble and fold properly retaining both the antigenicity of LT-B and the fluorescing properties of GFP. Maize kernel fractionation revealed that transgenic lines carrying BSP result in recombinant protein association with fibre and starch fractions. Confocal microscopy analysis indicates that the fusion proteins accumulate in the endomembrane system of plant cells in a signal peptide-dependent fashion. This is the first report providing evidence of the ability of a bacterial signal peptide to target proteins to the plant secretory pathway. The results provide important insights for further understanding the heterologous protein trafficking mechanisms and for developing effective strategies in molecular farming. PMID:19491306

  10. Delineating the pathways for the site-directed synthesis of individual nanoparticles on surfaces

    PubMed Central

    Liu, Guoliang; Eichelsdoerfer, Daniel J.; Rasin, Boris; Zhou, Yu; Brown, Keith A.; Liao, Xing; Mirkin, Chad A.

    2013-01-01

    Although nanoparticles with exquisite properties have been synthesized for a variety of applications, their incorporation into functional devices is challenging owing to the difficulty in positioning them at specified sites on surfaces. In contrast with the conventional synthesis-then-assembly paradigm, scanning probe block copolymer lithography can pattern precursor materials embedded in a polymer matrix and synthesize desired nanoparticles on site, offering great promise for incorporating nanoparticles into devices. This technique, however, is extremely limited from a materials standpoint. To develop a materials-general method for synthesizing nanoparticles on surfaces for broader applications, a mechanistic understanding of polymer-mediated nanoparticle formation is crucial. Here, we design a four-step synthetic process that enables independent study of the two most critical steps for synthesizing single nanoparticles on surfaces: phase separation of precursors and particle formation. Using this process, we elucidate the importance of the polymer matrix in the diffusion of metal precursors to form a single nanoparticle and the three pathways that the precursors undergo to form nanoparticles. Based on this mechanistic understanding, the synthetic process is generalized to create metal (Au, Ag, Pt, and Pd), metal oxide (Fe2O3, Co2O3, NiO, and CuO), and alloy (AuAg) nanoparticles. This mechanistic understanding and resulting process represent a major advance in scanning probe lithography as a tool to generate patterns of tailored nanoparticles for integration with solid-state devices. PMID:23277538

  11. Harnessing the PD-1 Pathway in Renal Cell Carcinoma: Current Evidence and Future Directions

    PubMed Central

    Tripathi, Abhishek; Drake, Charles G.

    2016-01-01

    Programmed cell death-1 (PD-1) is a recognized immune checkpoint. It is frequently upregulated on the T cells that infiltrate tumors, providing an inhibitory signal, which may facilitate immune escape. Blocking antibodies have been developed to interrupt the interaction of PD-1 with its ligands PD-L1/PD-L2, with the goal of increasing the host antitumor immune response. Initial results have been encouraging, with durable responses in both treatment-naive and pretreated patients, along with an acceptable toxicity profile. This tolerability makes PD-1 blockade an excellent potential partner for combination strategies with the approved targeted agents, such as tyrosine kinase inhibitors (TKIs) and anti-vascular endothelial growth factor (anti-VEGF) antibodies, as well as other investigational immune checkpoint inhibitors or agonist antibodies that may costimulate an immune response. PD-L1 expression on tumor cells and tumor-infiltrating immune cells is also being evaluated as a predictive biomarker of response to treatment. This review summarizes the biological basis, preclinical studies, ongoing trials, and future challenges associated with targeting the PD-1 pathway in renal cell carcinoma. PMID:25445176

  12. Nuclear disconnection within the amygdala reveals a direct pathway to fear.

    PubMed

    Jimenez, Stephanie A; Maren, Stephen

    2009-12-01

    It is widely believed that a descending serial circuit consisting of neural projections from the basolateral complex (BLA) to the central nucleus (CEA) of the amygdala mediates fear expression. Here we directly test this hypothesis and show that disconnecting the BLA and CEA with asymmetric neurotoxic lesions after Pavlovian fear conditioning in rats completely abolishes the expression of conditional freezing. These results demonstrate that neural projections from the BLA to CEA are essential for the expression of learned fear responses. PMID:19933881

  13. AFM1 in Milk: Physical, Biological, and Prophylactic Methods to Mitigate Contamination

    PubMed Central

    Giovati, Laura; Magliani, Walter; Ciociola, Tecla; Santinoli, Claudia; Conti, Stefania; Polonelli, Luciano

    2015-01-01

    Aflatoxins (AFs) are toxic, carcinogenic, immunosuppressive secondary metabolites produced by some Aspergillus species which colonize crops, including many dietary staple foods and feed components. AFB1 is the prevalent and most toxic among AFs. In the liver, it is biotransformed into AFM1, which is then excreted into the milk of lactating mammals, including dairy animals. AFM1 has been shown to be cause of both acute and chronic toxicoses. The presence of AFM1 in milk and dairy products represents a worldwide concern since even small amounts of this metabolite may be of importance as long-term exposure is concerned. Contamination of milk may be mitigated either directly, decreasing the AFM1 content in contaminated milk, or indirectly, decreasing AFB1 contamination in the feed of dairy animals. Current strategies for AFM1 mitigation include good agricultural practices in pre-harvest and post-harvest management of feed crops (including storage) and physical or chemical decontamination of feed and milk. However, no single strategy offers a complete solution to the issue. PMID:26512694

  14. Gene fusions for the directed modification of the carotenoid biosynthesis pathway in Mucor circinelloides.

    PubMed

    Iturriaga, Enrique A; Papp, Tamás; Alvarez, María Isabel; Eslava, Arturo P

    2012-01-01

    Several fungal species, particularly some included in the Mucorales, have been used to develop fermentation processes for the production of β-carotene. Oxygenated derivatives of β-carotene are more valuable products, and the preference by the market of carotenoids from biological sources has increased the research in different carotenoid-producing organisms. We currently use Mucor circinelloides as a model organism to develop strains able to produce new, more valuable, and with an increased content of carotenoids. In this chapter we describe part of our efforts to construct active gene fusions which could advance in the diversification of carotenoid production by this fungus. The main carotenoid accumulated by M. circinelloides is β-carotene, although it has some hydroxylase activity and produces low amounts of zeaxanthin. Two enzymatic activities are required for the production of astaxanthin from β-carotene: a hydroxylase and a ketolase. We used the ctrW gene of Paracoccus sp. N81106, encoding a bacterial β-carotene ketolase, to construct gene fusions with two fungal genes essential for the modification of the pathway in M. circinelloides. First we fused it to the carRP gene of M. circinelloides, which is responsible for the phytoene synthase and lycopene cyclase activities in this fungus. The expected activity of this fusion gene would be the accumulation by M. circinelloides of canthaxanthin and probably some astaxanthin. A second construction was the fusion of the crtW gene of Paracoccus sp. to the crtS gene of Xanthophyllomyces dendrorhous, responsible for the synthesis of astaxanthin from β-carotene in this fungus, but which was shown to have only hydroxylase activity in M. circinelloides. The expected result in M. circinelloides transformants was the accumulation of astaxanthin. Here we describe a detailed and empirically tested protocol for the construction of these gene fusions. PMID:22711120

  15. Tissue-Specific Whole Transcriptome Sequencing in Castor, Directed at Understanding Triacylglycerol Lipid Biosynthetic Pathways

    PubMed Central

    Swarbreck, David; Febrer, Melanie; Larson, Tony R.; Graham, Ian A.; Caccamo, Mario; Slabas, Antoni R.

    2012-01-01

    Background Storage triacylglycerols in castor bean seeds are enriched in the hydroxylated fatty acid ricinoleate. Extensive tissue-specific RNA-Seq transcriptome and lipid analysis will help identify components important for its biosynthesis. Methodology/Findings Storage triacylglycerols (TAGs) in the endosperm of developing castor (Ricinus communis) seeds are highly enriched in ricinoleic acid (18:1-OH). We have analysed neutral lipid fractions from other castor tissues using TLC, GLC and mass spectrometry. Cotyledons, like the endosperm, contain high levels of 18:1-OH in TAG. Pollen and male developing flowers accumulate TAG but do not contain 18:1-OH and leaves do not contain TAG or 18:1-OH. Analysis of acyl-CoAs in developing endosperm shows that ricinoleoyl-CoA is not the dominant acyl-CoA, indicating that either metabolic channelling or enzyme substrate selectivity are important in the synthesis of tri-ricinolein in this tissue. RNA-Seq transcriptomic analysis, using Illumina sequencing by synthesis technology, has been performed on mRNA isolated from two stages of developing seeds, germinating seeds, leaf and pollen-producing male flowers in order to identify differences in lipid-metabolic pathways and enzyme isoforms which could be important in the biosynthesis of TAG enriched in 18:1-OH. This study gives comprehensive coverage of gene expression in a variety of different castor tissues. The potential role of differentially expressed genes is discussed against a background of proteins identified in the endoplasmic reticulum, which is the site of TAG biosynthesis, and transgenic studies aimed at increasing the ricinoleic acid content of TAG. Conclusions/Significance Several of the genes identified in this tissue-specific whole transcriptome study have been used in transgenic plant research aimed at increasing the level of ricinoleic acid in TAG. New candidate genes have been identified which might further improve the level of ricinoleic acid in transgenic

  16. TAK-063, a PDE10A Inhibitor with Balanced Activation of Direct and Indirect Pathways, Provides Potent Antipsychotic-Like Effects in Multiple Paradigms.

    PubMed

    Suzuki, Kazunori; Harada, Akina; Suzuki, Hirobumi; Miyamoto, Maki; Kimura, Haruhide

    2016-08-01

    Phosphodiesterase 10A (PDE10A) inhibitors are expected to be novel drugs for schizophrenia through activation of both direct and indirect pathway medium spiny neurons. However, excess activation of the direct pathway by a dopamine D1 receptor agonist SKF82958 canceled antipsychotic-like effects of a dopamine D2 receptor antagonist haloperidol in methamphetamine (METH)-induced hyperactivity in rats. Thus, balanced activation of these pathways may be critical for PDE10A inhibitors. Current antipsychotics and the novel PDE10A inhibitor TAK-063, but not the selective PDE10A inhibitor MP-10, produced dose-dependent antipsychotic-like effects in METH-induced hyperactivity and prepulse inhibition in rodents. TAK-063 and MP-10 activated the indirect pathway to a similar extent; however, MP-10 caused greater activation of the direct pathway than did TAK-063. Interestingly, the off-rate of TAK-063 from PDE10A in rat brain sections was faster than that of MP-10, and a slower off-rate PDE10A inhibitor with TAK-063-like chemical structure showed an MP-10-like pharmacological profile. In general, faster off-rate enzyme inhibitors are more sensitive than slower off-rate inhibitors to binding inhibition by enzyme substrates. As expected, TAK-063 was more sensitive than MP-10 to binding inhibition by cyclic nucleotides. Moreover, an immunohistochemistry study suggested that cyclic adenosine monophosphate levels in the direct pathway were higher than those in the indirect pathway. These data can explain why TAK-063 showed partial activation of the direct pathway compared with MP-10. The findings presented here suggest that TAK-063's antipsychotic-like efficacy may be attributable to its unique pharmacological properties, resulting in balanced activation of the direct and indirect striatal pathways. PMID:26849714

  17. Synthesis of Bulk BC8 Silicon Allotrope by Direct Transformation and Reduced-Pressure Chemical Pathways.

    PubMed

    Kurakevych, Oleksandr O; Le Godec, Yann; Crichton, Wilson A; Guignard, Jérémy; Strobel, Timothy A; Zhang, Haidong; Liu, Hanyu; Coelho Diogo, Cristina; Polian, Alain; Menguy, Nicolas; Juhl, Stephen J; Gervais, Christel

    2016-09-01

    Phase-pure samples of a metastable allotrope of silicon, Si-III or BC8, were synthesized by direct elemental transformation at 14 GPa and ∼900 K and also at significantly reduced pressure in the Na-Si system at 9.5 GPa by quenching from high temperatures ∼1000 K. Pure sintered polycrystalline ingots with dimensions ranging from 0.5 to 2 mm can be easily recovered at ambient conditions. The chemical route also allowed us to decrease the synthetic pressures to as low as 7 GPa, while pressures required for direct phase transition in elemental silicon are significantly higher. In situ control of the synthetic protocol, using synchrotron radiation, allowed us to observe the underlying mechanism of chemical interactions and phase transformations in the Na-Si system. Detailed characterization of Si-III using X-ray diffraction, Raman spectroscopy, (29)Si NMR spectroscopy, and transmission electron microscopy are discussed. These large-volume syntheses at significantly reduced pressures extend the range of possible future bulk characterization methods and applications. PMID:27532223

  18. Localized electrografting of vinylic monomers on a conducting substrate by means of an integrated electrochemical AFM probe.

    PubMed

    Ghorbal, Achraf; Grisotto, Federico; Charlier, Julienne; Palacin, Serge; Goyer, Cédric; Demaille, Christophe

    2009-05-11

    Combinations of scanning electrochemical microscopy (SECM) with other scanning probe microscopy techniques, such as atomic force microscopy (AFM), show great promise for directing localized modification, which is of great interest for chemical, biochemical and technical applications. Herein, an atomic force scanning electrochemical microscope is used as a new electrochemical lithographic tool (L-AFM-SECM) to locally electrograft, with submicrometer resolution, a non-conducting organic coating on a conducting substrate. PMID:19308970

  19. Device level 3D characterization using PeakForce AFM

    NASA Astrophysics Data System (ADS)

    Timoney, Padraig; Zhang, Xiaoxiao; Vaid, Alok; Hand, Sean; Osborne, Jason; Milligan, Eric; Feinstein, Adam

    2016-03-01

    Traditional metrology solutions face a range of challenges at the 1X node such as three dimensional (3D) measurement capabilities, shrinking overlay and critical dimension (CD) error budgets driven by multi-patterning and via in trench CD measurements. With advent of advanced technology nodes and 3D processing, an increasing need is emerging for in-die metrology including across-structure and structure-to-structure characterization. A myriad of work has emerged in the past few years intending to address these challenges from various aspects; in-die OCD with reduced spot size and tilt beam on traditional critical dimension scanning electron microscopy (CDSEM) for height measurements. This paper explores the latest capability offered by PeakForceTM Tapping Atomic Force Microscopy (PFT-AFM). The use of traditional harmonic tapping mode for scanning high aspect ratio, and complex "3D" wafer structures, results in limited depth probing capability as well as excessive tip wear. These limitations arise due to the large tip-sample interaction volume in such confined spaces. PeakForce Tapping eliminates these limitations through direct real time control of the tip-sample interaction contact force. The ability of PeakForce to measure, and respond directly to tip- sample interaction forces results in more detailed feature resolution, reduced tip wear, and improved depth capability. In this work, the PFT-AFM tool was applied for multiple applications, including the 14nm fin and replacement metal gate (RMG) applications outlined below. Results from DOE wafers, detailed measurement precision studies and correlation to reference metrology are presented for validation of this methodology. With the fin application, precision of 0.3nm is demonstrated by measuring 5 dies with 10 consecutive runs. Capability to resolve within-die and localized within-macro height variation is also demonstrated. Results obtained from the fin measurements support the increasing trend that measurements

  20. Direct neural pathways convey distinct visual information to Drosophila mushroom bodies.

    PubMed

    Vogt, Katrin; Aso, Yoshinori; Hige, Toshihide; Knapek, Stephan; Ichinose, Toshiharu; Friedrich, Anja B; Turner, Glenn C; Rubin, Gerald M; Tanimoto, Hiromu

    2016-01-01

    Previously, we demonstrated that visual and olfactory associative memories of Drosophila share mushroom body (MB) circuits (Vogt et al., 2014). Unlike for odor representation, the MB circuit for visual information has not been characterized. Here, we show that a small subset of MB Kenyon cells (KCs) selectively responds to visual but not olfactory stimulation. The dendrites of these atypical KCs form a ventral accessory calyx (vAC), distinct from the main calyx that receives olfactory input. We identified two types of visual projection neurons (VPNs) directly connecting the optic lobes and the vAC. Strikingly, these VPNs are differentially required for visual memories of color and brightness. The segregation of visual and olfactory domains in the MB allows independent processing of distinct sensory memories and may be a conserved form of sensory representations among insects. PMID:27083044

  1. Long noncoding RNA hotair mediated angiogenesis in nasopharyngeal carcinoma by direct and indirect signaling pathways

    PubMed Central

    Hu, Bao-guang; Liang, Wei-cheng; Zhu, Xiao; Yang, Hai-di; Li, Gang; Zhang, Jin-fang

    2016-01-01

    Nasopharyngeal carcinoma (NPC), as a unique head and neck cancer type, is particularly prevalent in certain geographic areas such as eastern Asia. Until now, the therapeutic options have been restricted mainly to radiotherapy or chemotherapy. However, the clinical treatment effect remains unsatisfactory even if the combined radio-chemotherapies. Therefore, it is urgently needed to develop effective novel therapies against NPC. In this study, we discovered that lncRNA Hotair was extremely abundant in NPC cells and clinical NPC samples. Further studies showed that Hotair knockdown significantly attenuated both in vitro and in vivo tumor cell growth and angiogenesis. Our study also demonstrated that Hotair promoted angiogenesis through directly activating the transcription of angiogenic factor VEGFA as well as through GRP78-mediated upregulation of VEGFA and Ang2 expression. Therefore, Hotair may serve as a promising diagnostic marker and therapeutic target for NPC patients. PMID:26717040

  2. Red blood cells as a model to differentiate between direct and indirect oxidation pathways of peroxynitrite.

    PubMed

    Minetti, Maurizio; Pietraforte, Donatella; Straface, Elisabetta; Metere, Alessio; Matarrese, Paola; Malorni, Walter

    2008-01-01

    Red blood cells are the major physiological scavengers of reactive nitrogen species and have been proposed as real-time biomarkers of some vascular-related diseases. This chapter proposes that the erythrocyte is a suitable cell model for studying the modifications induced by peroxynitrite. Peroxynitrite decays both extra- and intracellularly as a function of cell density and CO(2) concentration, inducing the appearance of distinct cellular biomarkers, as well as the modulation of signaling and metabolism. Intracellular oxidations are due mostly to direct reactions of peroxynitrite with hemoglobin but also lead to the appearance of apoptotic biomarkers. Surface/membrane oxidations are due principally to indirect radical reactions generated by CO(2)-catalyzed peroxynitrite homolysis. PMID:18423223

  3. Direct neural pathways convey distinct visual information to Drosophila mushroom bodies

    PubMed Central

    Vogt, Katrin; Aso, Yoshinori; Hige, Toshihide; Knapek, Stephan; Ichinose, Toshiharu; Friedrich, Anja B; Turner, Glenn C; Rubin, Gerald M; Tanimoto, Hiromu

    2016-01-01

    Previously, we demonstrated that visual and olfactory associative memories of Drosophila share mushroom body (MB) circuits (Vogt et al., 2014). Unlike for odor representation, the MB circuit for visual information has not been characterized. Here, we show that a small subset of MB Kenyon cells (KCs) selectively responds to visual but not olfactory stimulation. The dendrites of these atypical KCs form a ventral accessory calyx (vAC), distinct from the main calyx that receives olfactory input. We identified two types of visual projection neurons (VPNs) directly connecting the optic lobes and the vAC. Strikingly, these VPNs are differentially required for visual memories of color and brightness. The segregation of visual and olfactory domains in the MB allows independent processing of distinct sensory memories and may be a conserved form of sensory representations among insects. DOI: http://dx.doi.org/10.7554/eLife.14009.001 PMID:27083044

  4. BOREAS AFM-04 Twin Otter Aircraft Sounding Data

    NASA Technical Reports Server (NTRS)

    MacPherson, J. Ian; Desjardins, Raymond L.; Hall, Forrest G. (Editor); Knapp, David E. (Editor); Smith, David E. (Technical Monitor)

    2000-01-01

    The Boreal Ecosystem-Atmosphere Study (BOREAS) Airborne Fluxes and Meteorology (AFM)-4 team used the National Research Council, Canada (NRC) Twin Otter aircraft to make sounding measurements through the boundary layer. These measurements included concentrations of carbon dioxide and ozone, atmospheric pressure, dry bulb temperature, potential temperature, dewpoint temperature, calculated mixing ratio, and wind speed and direction. Aircraft position, heading, and altitude were also recorded. Data were collected at both the Northern Study Area (NSA) and the Southern Study Area (SSA) in 1994 and 1996. These data are stored in tabular ASCII files. The Twin Otter aircraft sounding data are available from the Earth Observing System Data and Information System (EOSDIS) Oak Ridge National Laboratory (ORNL) Distributed Active Archive Center (DAAC). The data files also are available on a CD-ROM (see document number 20010000884).

  5. [AFM fishing of proteins under impulse electric field].

    PubMed

    Ivanov, Yu D; Pleshakova, T O; Malsagova, K A; Kaysheva, A L; Kopylov, A T; Izotov, A A; Tatur, V Yu; Vesnin, S G; Ivanova, N D; Ziborov, V S; Archakov, A I

    2016-05-01

    A combination of (atomic force microscopy)-based fishing (AFM-fishing) and mass spectrometry allows to capture protein molecules from solutions, concentrate and visualize them on an atomically flat surface of the AFM chip and identify by subsequent mass spectrometric analysis. In order to increase the AFM-fishing efficiency we have applied pulsed voltage with the rise time of the front of about 1 ns to the AFM chip. The AFM-chip was made using a conductive material, highly oriented pyrolytic graphite (HOPG). The increased efficiency of AFM-fishing has been demonstrated using detection of cytochrome b5 protein. Selection of the stimulating pulse with a rise time of 1 ns, corresponding to the GHz frequency range, by the effect of intrinsic emission from water observed in this frequency range during water injection into the cell. PMID:27562998

  6. Multiple Smaller Missions as a Direct Pathway to Mars Sample Return

    NASA Technical Reports Server (NTRS)

    Niles, P. B.; Draper, D. S.; Evans, C. A.; Gibson, E. K.; Graham, L. D.; Jones, J. H.; Lederer, S. M.; Ming, D.; Seaman, C. H.; Archer, P. D.; Andrews-Hanna, J.; Baldridge, A. M.; Bourke, M. C.; Crown, D. A.; Fries, M.; Knudson, A. T.; Michalski, J.; Dobrea, E. Noe; Vaniman, D.; Weitz, C. M.; Williams, R. M. E.; Bell, J. F., III; Knauth, L. P.

    2012-01-01

    Recent discoveries by the Mars Exploration Rovers, Mars Express, Mars Odyssey, and Mars Reconnaissance Orbiter spacecraft include multiple, tantalizing astrobiological targets representing both past and present environments on Mars. The most desirable path to Mars Sample Return (MSR) would be to collect and return samples from that site which provides the clearest examples of the variety of rock types considered a high priority for sample return (pristine igneous, sedimentary, and hydrothermal). Here we propose an MSR architecture in which the next steps (potentially launched in 2018) would entail a series of smaller missions, including caching, to multiple landing sites to verify the presence of high priority sample return targets through in situ analyses. This alternative architecture to one flagship-class sample caching mission to a single site would preserve a direct path to MSR as stipulated by the Planetary Decadal Survey, while permitting investigation of diverse deposit types and providing comparison of the site of returned samples to other aqueous environments on early Mars

  7. Bem1p contributes to secretory pathway polarization through a direct interaction with Exo70p

    PubMed Central

    Liu, Dongmei

    2014-01-01

    The exocyst serves to tether secretory vesicles to cortical sites specified by polarity determinants, in preparation for fusion with the plasma membrane. Although most exocyst components are brought to these sites by riding on secretory vesicles as they are actively transported along actin cables, Exo70p displays actin-independent localization to these sites, implying an interaction with a polarity determinant. Here we show that Exo70p directly and specifically binds to the polarity determinant scaffold protein Bem1p. The interaction involves multiple domains of both Exo70p and Bem1p. Mutations in Exo70p that disrupt its interaction with Bem1, without impairing its interactions with other known binding partners, lead to the loss of actin-independent localization. Synthetic genetic interactions confirm the importance of the Exo70p–Bem1p interaction, although there is some possible redundancy with Sec3p and Sec15p, other exocyst components that also interact with polarity determinants. Similar to Sec3p, the actin-independent localization of Exo70p requires a synergistic interaction with the phosphoinositide PI(4,5)P2. PMID:25313406

  8. S(mu) mutation patterns suggest different progression pathways in follicular lymphoma: early direct or late from FL progenitor cells.

    PubMed

    Ruminy, Philippe; Jardin, Fabrice; Picquenot, Jean-Michel; Parmentier, Françoise; Contentin, Nathalie; Buchonnet, Gérard; Tison, Sandrine; Rainville, Vinciane; Tilly, Hervé; Bastard, Christian

    2008-09-01

    Follicular lymphoma (FL) is a B-cell malignancy characterized by the t(14;18) translocation. Although sensitive to treatment, the disease remains incurable and the reason why tumor cells invariably evade treatment, leading to clinical relapse, is still unknown. Here, we tracked the clonal history of tumor cells by studying mutations introduced by activation-induced cytidine deaminase on the switch mu region of the der(14)t(14;18) during the early phase of the class-switch recombination (CSR) process. We observed frequent intraclonal variations, suggesting that CSR often remains active after the acquisition of the fully transformed phenotype. However, mutations only rarely accumulated over time, but instead showed complex evolutionary scenarios and 2 different progression pathways. The first pathway was a direct and rapid evolution from the dominant clone. The second was indirect, arising from earlier subclones usually after years of remission. A better understanding of these mechanisms might influence the future choice of treatment strategies. PMID:18515657

  9. BOREAS AFM-06 Mean Temperature Profile Data

    NASA Technical Reports Server (NTRS)

    Wilczak, James; Hall, Forrest G. (Editor); Newcomer, Jeffrey A. (Editor); Smith, David E. (Technical Monitor)

    2000-01-01

    The Boreal Ecosystem-Atmosphere Study (BOREAS) Airborne Fluxes and Meteorology (AFM)-6 team from the National Oceanic and Atmospheric Adminsitration/Environment Technology Laboratory (NOAA/ETL) operated a 915-MHz wind/Radio Acoustic Sounding System (RASS) profiler system in the Southern Study Area (SSA) near the Old Jack Pine (OJP) tower from 21 May 1994 to 20 Sep 1994. The data set provides temperature profiles at 15 heights, containing the variables of virtual temperature, vertical velocity, the speed of sound, and w-bar. The data are stored in tabular ASCII files. The mean temperature profile data are available from the Earth Observing System Data and Information System (EOSDIS) Oak Ridge National Laboratory (ORNL) Distributed Active Archive Center (DAAC). The data files are available on a CD-ROM (see document number 20010000884).

  10. BOREAS AFM-6 Boundary Layer Height Data

    NASA Technical Reports Server (NTRS)

    Wilczak, James; Hall, Forrest G. (Editor); Newcomer, Jeffrey A. (Editor); Smith, David E. (Technical Monitor)

    2000-01-01

    The Boreal Ecosystem-Atmosphere Study (BOREAS) Airborne Fluxes and Meteorology (AFM)-6 team from National Oceanic and Atmospheric Adminsitration/Environment Technology Laboratory (NOAA/ETL) operated a 915-MHz wind/Radio Acoustic Sounding System (RASS) profiler system in the Southern Study Area (SSA) near the Old Jack Pine (OJP) site. This data set provides boundary layer height information over the site. The data were collected from 21 May 1994 to 20 Sep 1994 and are stored in tabular ASCII files. The boundary layer height data are available from the Earth Observing System Data and Information System (EOSDIS) Oak Ridge National Laboratory (ORNL) Distributed Active Archive Center (DAAC). The data files are available on a CD-ROM (see document number 20010000884).

  11. Control of torque direction by spinal pathways at the cat ankle joint.

    PubMed

    Nichols, T R; Lawrence, J H; Bonasera, S J

    1993-01-01

    To study the biomechanics of the calcaneal tendon's complex insertion onto the calcaneus, we measured torque-time trajectories exerted by the triceps surae and tibialis anterior muscles in eight unanesthetized decerebrate cats using a multi-axis force-moment sensor placed at the ankle joint. The ankle was constrained to an angle of 110 degrees plantarflexion. Muscles were activated using crossed-extension (XER), flexion (FWR), and caudal cutaneous sural nerve (SNR) reflexes. Torque contributions of other muscles activated by these reflexes were eliminated by denervation or tenotomy. In two animals, miniature pressure transducers were implanted among tendon fibers from the lateral gastrocnemius (LG) muscle that insert straight into the calcaneus or among tendon fibers from the medial gastrocnemius (MG) that cross over and insert on the lateral aspect of calcaneus. Reflexively evoked torques had the following directions: FWR, dorsiflexion and adduction; SNR, plantarflexion and abduction; and XER, plantarflexion and modest abduction or adduction. The proportion of abduction torque to plantarflexion torque was always greater for SNR than XER; this difference was about 50% of the magnitude of abduction torque generated by tetanic stimulation of the peronei. During SNR, pressures were higher in regions of the calcaneal tendon originating from MG than regions originating from LG. Similarly, pressures within the MG portion of the calcaneal tendon were higher during SNR than during XER, although these two reflexes produced matched ankle plantarflexion forces. Selective tenotomies and electromyographic recordings further demonstrated that MG generated most of the torque in response to SNR, while soleus, LG, and MG all generated torques in response to XER.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:8150057

  12. The cAMP Signaling Pathway and Direct Protein Kinase A Phosphorylation Regulate Polycystin-2 (TRPP2) Channel Function.

    PubMed

    Cantero, María del Rocío; Velázquez, Irina F; Streets, Andrew J; Ong, Albert C M; Cantiello, Horacio F

    2015-09-25

    Polycystin-2 (PC2) is a TRP-type, Ca(2+)-permeable non-selective cation channel that plays an important role in Ca(2+) signaling in renal and non-renal cells. The effect(s) of the cAMP pathway and kinase mediated phosphorylation of PC2 seem to be relevant to PC2 trafficking and its interaction with polycystin-1. However, the role of PC2 phosphorylation in channel function is still poorly defined. Here we reconstituted apical membranes of term human syncytiotrophoblast (hST), containing endogenous PC2 (PC2hst), and in vitro translated channel protein (PC2iv). Addition of the catalytic subunit of PKA increased by 566% the spontaneous PC2hst channel activity in the presence of ATP. Interestingly, 8-Br-cAMP also stimulated spontaneous PC2hst channel activity in the absence of the exogenous kinase. Either stimulation was inhibited by addition of alkaline phosphatase, which in turn, was reversed by the phosphatase inhibitor vanadate. Neither maneuver modified the single channel conductance but instead increased channel mean open time. PKA directly phosphorylated PC2, which increased the mean open time but not the single channel conductance of the channel. PKA phosphorylation did not modify either R742X truncated or S829A-mutant PC2iv channel function. The data indicate that the cAMP pathway regulates PC2-mediated cation transport in the hST. The relevant PKA site for PC2 channel regulation centers on a single residue serine 829, in the carboxyl terminus. PMID:26269590

  13. Particle deformation induced by AFM tapping under different setpoint voltages

    NASA Astrophysics Data System (ADS)

    Wu, Chung-Lin; Farkas, Natalia; Dagata, John A.; He, Bo-Ching; Fu, Wei-En

    2014-09-01

    The measured height of polystyrene nanoparticles varies with setpoint voltage during atomic force microscopy (AFM) tapping-mode imaging. Nanoparticle height was strongly influenced by the magnitude of the deformation caused by the AFM tapping forces, which was determined by the setpoint voltage. This influence quantity was studied by controlling the operational AFM setpoint voltage. A test sample consisting of well-dispersed 60-nm polystyrene and gold nanoparticles co-adsorbed on poly-l-lysine-coated mica was studied in this research. Gold nanoparticles have not only better mechanical property than polystyrene nanoparticles, but also obvious facets in AFM phase image. By using this sample of mixed nanoparticles, it allows us to confirm that the deformation resulted from the effect of setpoint voltage, not noise. In tapping mode, the deformation of polystyrene nanoparticles increased with decreasing setpoint voltage. Similar behavior was observed with both open loop and closed loop AFM instruments.

  14. Microrheology using a custom-made AFM

    NASA Astrophysics Data System (ADS)

    Kosgodagan Acharige, Sebastien; Benzaquen, Michael; Steinberger, Audrey

    In the past few years, a new method was developed to measure local properties of liquids (X. Xiong et al., Phys. Rev. E 80, 2009). This method consists of gluing a micron-sized glass fiber at the tip of an AFM cantilever and probing the liquid with it. In ENS Lyon, this method was perfected (C. Devailly et al., EPL, 106 5, 2014) with the help of an interferometer developped in the same laboratory (L. Bellon et al., Opt. Commun. 207 49, 2002 and P. Paolino et al., Rev. Sci. Instrum. 84, 2013), which background noise can reach 10-14 m /√{ Hz } . This method allows us to measure a wide range of viscosities (1 mPa . s to 500 mPa . s) of transparent and opaque fluids using a small sample volume ( 5 mL). In this presentation, I will briefly describe the interferometer developped in ENS Lyon, then explain precisely the microrheology measurements and then compare the experimental results to a model developped by M. Benzaquen. This work is supported financially by the ANR project NANOFLUIDYN (Grant Number ANR-13-BS10-0009).

  15. Manufacturing process of nanofluidics using afm probe

    NASA Astrophysics Data System (ADS)

    Karingula, Varun Kumar

    A new process for fabricating a nano fluidic device that can be used in medical application is developed and demonstrated. Nano channels are fabricated using a nano tip in indentation mode on AFM (Atomic Force Microscopy). The nano channels are integrated between the micro channels and act as a filter to separate biomolecules. Nano channels of 4 to7 m in length, 80nm in width, and at varying depths from 100nm to 850 nm allow the resulting device to separate selected groups of lysosomes and other viruses. Sharply developed vertical micro channels are produced from a deep reaction ion etching followed by deposition of different materials, such as gold and polymers, on the top surface, allowing the study of alternative ways of manufacturing a nanofluidic device. PDMS (Polydimethylsiloxane) bonding is performed to close the top surface of the device. An experimental setup is used to test and validate the device by pouring fluid through the channels. A detailed cost evaluation is conducted to compare the economical merits of the proposed process. It is shown that there is a 47:7% manufacturing time savings and a 60:6% manufacturing cost savings.

  16. BOREAS AFM-07 SRC Surface Meteorological Data

    NASA Technical Reports Server (NTRS)

    Osborne, Heather; Hall, Forrest G. (Editor); Newcomer, Jeffrey A. (Editor); Young, Kim; Wittrock, Virginia; Shewchuck, Stan; Smith, David E. (Technical Monitor)

    2000-01-01

    The Saskatchewan Research Council (SRC) collected surface meteorological and radiation data from December 1993 until December 1996. The data set comprises Suite A (meteorological and energy balance measurements) and Suite B (diffuse solar and longwave measurements) components. Suite A measurements were taken at each of ten sites, and Suite B measurements were made at five of the Suite A sites. The data cover an approximate area of 500 km (North-South) by 1000 km (East-West) (a large portion of northern Manitoba and northern Saskatchewan). The measurement network was designed to provide researchers with a sufficient record of near-surface meteorological and radiation measurements. The data are provided in tabular ASCII files, and were collected by Aircraft Flux and Meteorology (AFM)-7. The surface meteorological and radiation data are available from the Earth Observing System Data and Information System (EOSDIS) Oak Ridge National Laboratory (ORNL) Distributed Active Archive Center (DAAC). The data files are available on a CD-ROM (see document number 20010000884).

  17. An AFM-based pit-measuring method for indirect measurements of cell-surface membrane vesicles

    SciTech Connect

    Zhang, Xiaojun; Chen, Yuan; Chen, Yong

    2014-03-28

    Highlights: • Air drying induced the transformation of cell-surface membrane vesicles into pits. • An AFM-based pit-measuring method was developed to measure cell-surface vesicles. • Our method detected at least two populations of cell-surface membrane vesicles. - Abstract: Circulating membrane vesicles, which are shed from many cell types, have multiple functions and have been correlated with many diseases. Although circulating membrane vesicles have been extensively characterized, the status of cell-surface membrane vesicles prior to their release is less understood due to the lack of effective measurement methods. Recently, as a powerful, micro- or nano-scale imaging tool, atomic force microscopy (AFM) has been applied in measuring circulating membrane vesicles. However, it seems very difficult for AFM to directly image/identify and measure cell-bound membrane vesicles due to the similarity of surface morphology between membrane vesicles and cell surfaces. Therefore, until now no AFM studies on cell-surface membrane vesicles have been reported. In this study, we found that air drying can induce the transformation of most cell-surface membrane vesicles into pits that are more readily detectable by AFM. Based on this, we developed an AFM-based pit-measuring method and, for the first time, used AFM to indirectly measure cell-surface membrane vesicles on cultured endothelial cells. Using this approach, we observed and quantitatively measured at least two populations of cell-surface membrane vesicles, a nanoscale population (<500 nm in diameter peaking at ∼250 nm) and a microscale population (from 500 nm to ∼2 μm peaking at ∼0.8 μm), whereas confocal microscopy only detected the microscale population. The AFM-based pit-measuring method is potentially useful for studying cell-surface membrane vesicles and for investigating the mechanisms of membrane vesicle formation/release.

  18. Controlled AFM detachments and movement of nanoparticles: gold clusters on HOPG at different temperatures

    NASA Astrophysics Data System (ADS)

    Tripathi, Manoj; Paolicelli, Guido; D'Addato, Sergio; Valeri, Sergio

    2012-06-01

    The effect of temperature on the onset of movement of gold nanoclusters (diameter 27 nm) deposited on highly oriented pyrolytic graphite (HOPG) has been studied by atomic force microscopy (AFM) techniques. Using the AFM with amplitude modulation (tapping mode AFM) we have stimulated and controlled the movement of individual clusters. We show how, at room temperature, controlled detachments and smooth movements can be obtained for clusters having dimensions comparable to or smaller than the tip radius. Displacement is practically visible in real time and it can be started and stopped easily by adjusting only one parameter, the tip amplitude oscillation. Analysing the energy dissipation signal at the onset of nanocluster sliding we evaluated a detachment threshold energy as a function of temperature in the range 300-413 K. We also analysed single cluster thermal induced displacement and combining this delicate procedure with AFM forced movement behaviour we conclude that detachment threshold energy is directly related to the activation energy of nanocluster diffusion and it scales linearly with temperature as expected for a single-particle thermally activated process.

  19. CDSEM AFM hybrid metrology for the characterization of gate-all-around silicon nano wires

    NASA Astrophysics Data System (ADS)

    Levi, Shimon; Schwarzband, Ishai; Weinberg, Yakov; Cornell, Roger; Adan, Ofer; Cohen, Guy M.; Gignac, Lynne; Bangsaruntip, Sarunya; Hand, Sean; Osborne, Jason; Feinstein, Adam

    2014-04-01

    In an ongoing study of the physical characterization of Gate-All-Around Silicon Nano Wires (GAASiNW), we found that the thin, suspended wires are prone to buckling as a function of their length and diameter. This buckling takes place between the fixed source and drain regions of the suspended wire, and can affect the device performance and therefore must be studied and controlled. For cylindrical SiNW, theory predicts that buckling has no directional preference. However, 3D CDSEM measurement results indicated that cylindrical wires prefer to buckle towards the wafer. To validate these results and to determine if the electron beam or charging is affecting our observations, we used 3D-AFM measurements to evaluate the buckling. To assure that the CDSEM and 3D-AFM measure the exact same locations, we developed a design based recipe generation approach to match the 3D-AFM and CDSEM coordinate systems. Measuring the exact same sites enables us to compare results and use 3D-AFM data to optimize CDSEM models. In this paper we will present a hybrid metrology approach to the characterization of GAASiNW for sub-nanometer variations, validating experimental results, and proposing methods to improve metrology capabilities.

  20. 3D-stacked carbon composites employing networked electrical intra-pathways for direct-printable, extremely stretchable conductors.

    PubMed

    Chae, Changju; Seo, Yeong-Hui; Jo, Yejin; Kim, Ki Woong; Song, Wooseok; An, Ki-Seok; Choi, Sungho; Choi, Youngmin; Lee, Sun Sook; Jeong, Sunho

    2015-02-25

    The newly designed materials for stretchable conductors meeting the demands for both electrical and mechanical stability upon morphological elongation have recently been of paramount interest in the applications of stretchable, wearable electronics. To date, carbon nanotube-elastomeric polymer mixtures have been mainly developed; however, the method of preparing such CNT-polymer mixtures as stretchable conductors has been limited to an ionic liquid-mediated approach. In this study, we suggest a simple wet-chemical method for producing newly designed, three-dimensionally stacked carbon composite materials that facilitate the stable morphological elongation up to a strain of 300% with normalized conductivity variation of only 0.34 under a strain of 300%. Through a comparative study with other control samples, it is demonstrated that the intraconnected electrical pathways in hierarchically structured composite materials enable the generation of highly stretchable conductors. Their direct patternability is also evaluated by printing on demand using a programmable disperser without the use of prepatterned masks. PMID:25647807

  1. Broadening of neutralization activity to directly block a dominant antibody-driven SARS-coronavirus evolution pathway.

    PubMed

    Sui, Jianhua; Aird, Daniel R; Tamin, Azaibi; Murakami, Akikazu; Yan, Meiying; Yammanuru, Anuradha; Jing, Huaiqi; Kan, Biao; Liu, Xin; Zhu, Quan; Yuan, Qing-An; Adams, Gregory P; Bellini, William J; Xu, Jianguo; Anderson, Larry J; Marasco, Wayne A

    2008-11-01

    Phylogenetic analyses have provided strong evidence that amino acid changes in spike (S) protein of animal and human SARS coronaviruses (SARS-CoVs) during and between two zoonotic transfers (2002/03 and 2003/04) are the result of positive selection. While several studies support that some amino acid changes between animal and human viruses are the result of inter-species adaptation, the role of neutralizing antibodies (nAbs) in driving SARS-CoV evolution, particularly during intra-species transmission, is unknown. A detailed examination of SARS-CoV infected animal and human convalescent sera could provide evidence of nAb pressure which, if found, may lead to strategies to effectively block virus evolution pathways by broadening the activity of nAbs. Here we show, by focusing on a dominant neutralization epitope, that contemporaneous- and cross-strain nAb responses against SARS-CoV spike protein exist during natural infection. In vitro immune pressure on this epitope using 2002/03 strain-specific nAb 80R recapitulated a dominant escape mutation that was present in all 2003/04 animal and human viruses. Strategies to block this nAb escape/naturally occurring evolution pathway by generating broad nAbs (BnAbs) with activity against 80R escape mutants and both 2002/03 and 2003/04 strains were explored. Structure-based amino acid changes in an activation-induced cytidine deaminase (AID) "hot spot" in a light chain CDR (complementarity determining region) alone, introduced through shuffling of naturally occurring non-immune human VL chain repertoire or by targeted mutagenesis, were successful in generating these BnAbs. These results demonstrate that nAb-mediated immune pressure is likely a driving force for positive selection during intra-species transmission of SARS-CoV. Somatic hypermutation (SHM) of a single VL CDR can markedly broaden the activity of a strain-specific nAb. The strategies investigated in this study, in particular the use of structural information in

  2. Dynamics of the nanoneedle probe in trolling mode AFM.

    PubMed

    Abdi, Ahmad; Pishkenari, Hossein Nejat; Keramati, Ramtin; Minary-Jolandan, Majid

    2015-05-22

    Atomic force microscopy (AFM), as an indispensable tool for nanoscale characterization, presents major drawbacks for operation in a liquid environment arising from the large hydrodynamic drag on the vibrating cantilever. The newly introduced 'Trolling mode' (TR-mode) AFM resolves this complication by using a specialized nanoneedle cantilever that keeps the cantilever outside of the liquid. Herein, a mechanical model with a lumped mass was developed to capture the dynamics of such a cantilever with a nanoneedle tip. This new developed model was applied to investigate the effects of the needle-liquid interface on the performance of the AFM, including the imaging capability in liquid. PMID:25915451

  3. Raman and AFM study of gamma irradiated plastic bottle sheets

    SciTech Connect

    Ali, Yasir; Kumar, Vijay; Dhaliwal, A. S.; Sonkawade, R. G.

    2013-02-05

    In this investigation, the effects of gamma irradiation on the structural properties of plastic bottle sheet are studied. The Plastic sheets were exposed with 1.25MeV {sup 60}Co gamma rays source at various dose levels within the range from 0-670 kGy. The induced modifications were followed by micro-Raman and atomic force microscopy (AFM). The Raman spectrum shows the decrease in Raman intensity and formation of unsaturated bonds with an increase in the gamma dose. AFM image displays rough surface morphology after irradiation. The detailed Raman analysis of plastic bottle sheets is presented here, and the results are correlated with the AFM observations.

  4. AFM Studies of Conformational Changes in Proteins and Peptides

    NASA Astrophysics Data System (ADS)

    Ploscariu, Nicoleta; Sukthankar, Pinakin; Tomich, John; Szoszkiewicz, Robert

    2015-03-01

    Here, we present estimates of molecular stiffness and mechanical energy dissipation factors for some examples of proteins and peptides. The results are obtained from AFM force spectroscopy measurements. To determine molecular stiffness and mechanical energy dissipation factors we developed a model based on measuring several resonance frequencies of an AFM cantilever in contact with either single protein molecule or peptides adsorbed on arbitrary surface. We used compliant AFM cantilevers with a small aspect ratio - a ratio of length to width - in air and in liquid, including biologically relevant phosphate buffered saline medium. Department of Physics.

  5. Production of resident fish benefits from experimental salmon subsidies via direct and indirect pathways across stream-riparian boundaries

    USGS Publications Warehouse

    Collins, Scott F.; Baxter, Colden V.; Marcarelli, Amy M.; Wipfli, Mark S.

    2016-01-01

    Artificial additions of nutrients of differing forms such as salmon carcasses and analog pellets (i.e. pasteurized fishmeal) have been proposed as a means of stimulating aquatic productivity and enhancing populations of anadromous and resident fishes. Nutrient mitigation to enhance fish production in stream ecosystems assumes that the central pathway by which effects occur is bottom-up, through aquatic primary and secondary production, with little consideration of reciprocal aquatic-terrestrial pathways. The net outcome (i.e. bottom-up vs. top-down) of adding salmon-derived materials to streams depend on whether or not these subsidies indirectly intensify predation on in situ prey via increases in a shared predator or alleviate such predation pressure. We conducted a 3-year experiment across nine tributaries of the N. Fork Boise River, Idaho, USA, consisting of 500-m stream reaches treated with salmon carcasses (n = 3), salmon carcass analog (n = 3), and untreated control reaches (n = 3). We observed 2–8 fold increases in streambed biofilms in the 2–6 weeks following additions of both salmon subsidy treatments in years 1 and 2 and a 1.5-fold increase in standing crop biomass of aquatic invertebrates to carcass additions in the second year of our experiment. The consumption of benthic invertebrates by stream fishes increased 110–140% and 44–66% in carcass and analog streams in the same time frame, which may have masked invertebrate standing crop responses in years 3 and 4. Resident trout directly consumed 10.0–24.0 g·m−2·yr−1 of salmon carcass and <1–11.0 g·m−2·yr−1 of analog material, which resulted in 1.2–2.9 g·m−2·yr−1 and 0.03–1.4 g·m−2·yr−1 of tissue produced. In addition, a feedback flux of terrestrial maggots to streams contributed 0.0–2.0 g·m−2·yr−1 to trout production. Overall, treatments increased annual trout production by 2–3 fold, though density and biomass were unaffected. Our results

  6. Asymmetric direct α-alkylation of 2-oxindoles with Michler's hydrol catalyzed by bis-cinchona alkaloid-Brønsted acid via an SN1-type pathway.

    PubMed

    Zhang, Tao; Qiao, Zhen; Wang, Yan; Zhong, Nengjun; Liu, Li; Wang, Dong; Chen, Yong-Jun

    2013-02-25

    An enantioselective direct α-alkylation of 2-oxindoles with Michler's hydrol via an S(N)1-type pathway in the non-covalent activation mode using the bis-cinchona alkaloid and Brønsted acid as a co-catalyst was developed and good to high yields and enantioselectivities were obtained. PMID:23340601

  7. Dopamine D1 Receptor-Mediated Transmission Maintains Information Flow Through the Cortico-Striato-Entopeduncular Direct Pathway to Release Movements

    PubMed Central

    Chiken, Satomi; Sato, Asako; Ohta, Chikara; Kurokawa, Makoto; Arai, Satoshi; Maeshima, Jun; Sunayama-Morita, Tomoko; Sasaoka, Toshikuni; Nambu, Atsushi

    2015-01-01

    In the basal ganglia (BG), dopamine plays a pivotal role in motor control, and dopamine deficiency results in severe motor dysfunctions as seen in Parkinson's disease. According to the well-accepted model of the BG, dopamine activates striatal direct pathway neurons that directly project to the output nuclei of the BG through D1 receptors (D1Rs), whereas dopamine inhibits striatal indirect pathway neurons that project to the external pallidum (GPe) through D2 receptors. To clarify the exact role of dopaminergic transmission via D1Rs in vivo, we developed novel D1R knockdown mice in which D1Rs can be conditionally and reversibly regulated. Suppression of D1R expression by doxycycline treatment decreased spontaneous motor activity and impaired motor ability in the mice. Neuronal activity in the entopeduncular nucleus (EPN), one of the output nuclei of the rodent BG, was recorded in awake conditions to examine the mechanism of motor deficits. Cortically evoked inhibition in the EPN mediated by the cortico-striato-EPN direct pathway was mostly lost during suppression of D1R expression, whereas spontaneous firing rates and patterns remained unchanged. On the other hand, GPe activity changed little. These results suggest that D1R-mediated dopaminergic transmission maintains the information flow through the direct pathway to appropriately release motor actions. PMID:26443442

  8. Effects of Tumor Suppressor Lysyl Oxidase Propeptide on Prostate Cancer Xenograft Growth and Its Direct Interactions with DNA Repair Pathways

    PubMed Central

    Bais, Manish V.; Ozdener, Gokhan Baris; Sonenshein, Gail E.; Trackman, Philip C.

    2014-01-01

    Lysyl oxidase (LOX) is a multifunctional protein required for normal collagen and elastin biosynthesis and maturation. In addition, LOX has complex roles in cancer in which the lysyl oxidase propeptide (LOX-PP) domain of secreted pro-LOX has tumor suppressor activity, while the active enzyme promotes metastasis. In prostate cancer cell lines, recombinant LOX-PP (rLOX-PP) inhibits the growth of PC3 cells in vitro by mechanisms which were not characterized, while in DU145 cells rLOX-PP targeted FGF signaling. Because rLOX-PP can enhance effects of a genotoxic chemotherapeutic on breast cancer cell apoptosis, we reasoned that rLOX-PP could target DNA repair pathways typically elevated in cancer. Here we demonstrate for the first time that rLOX-PP inhibits prostate xenograft growth in vivo and that activating phosphorylations of the key DNA repair molecules ATM and CHK2 are inhibited by rLOX-PP expression in vivo. In addition, in vitro studies showed that rLOX-PP inhibits radiation induced activating phosphorylations of ATM and CHK2, and that exogenously added rLOX-PP protein can localize to the nucleus in both DU145 and PC3 cells. rLOX-PP pull-down studies resulted in detection of a protein complex with the nuclear DNA repair regulator MRE11 in both cell lines, and rLOX-PP localized to radiation-induced nuclear DNA repair foci. Finally, rLOX-PP was shown to sensitize both DU145 and PC3 cells to radiation-induced cell death determined in colony formation assays. These data provide evidence that rLOX-PP has a nuclear mechanism of action in which it directly interacts with DNA repair proteins to sensitize prostate cancer cells to the effects of ionizing radiation. PMID:24882580

  9. Ability of Lactobacillus rhamnosus GAF01 to remove AFM1 in vitro and to counteract AFM1 immunotoxicity in vivo.

    PubMed

    Abbès, Samir; Salah-Abbès, Jalila Ben; Sharafi, Hakimeh; Jebali, Rania; Noghabi, Kambiz Akbari; Oueslati, Ridha

    2013-01-01

    Aflatoxin M1 (AFM1) has been detected in many parts of the world both in raw milk and many dairy products, causing great economic losses and human disease. Unfortunately, there are few studies dealing with AFM1 immunotoxicity/interactions with lactic acid bacteria for potential application as a natural preventive agent. The aim of this study was to isolate (from dairy products) food-grade probiotic bacteria able to degrade/bind AFM1 in vitro and evaluate whether the same organism(s) could impart a protective role against AFM1-induced immunotoxicity in exposed Balb/c mice. Bacteria (Lactobacillus plantarum MON03 and L. rhamnosus GAF01) were isolated from Tunisian artisanal butter and then tested for abilities to eliminate AFM1 from phosphate-buffered saline (PBS) and reconstituted milk (containing 0.05, 0.10, and 0.20 µg AFM1/ml) after 0, 6, and 24 h at 37°C. Results showed that the selected bacteria could 'remove' AFM1 both in PBS and skimmed milk. The binding abilities of AFM1 by L. plantarum MON03 and L. rhamnosus GAF01 strains (at 10(8) CFU/ml) in PBS and reconstituted milk ranged, respectively, from 16.1-78.6% and 15.3-95.1%; overall, L. rhamnosus showed a better potential for removal than L. plantarum. 'Removal' appeared to be by simple binding; the bacteria/AFM1 complex was stable and only a very small proportion of mycotoxin was released back into the solution. L. rhamnosus GAF01 had the highest binding capacity and was selected for use in the in vivo study. Those results indicated that use of the organism prevented AFM1-induced effects on total white and red blood cells, and lymphocyte subtypes, after 15 days of host treatment. These studies clearly indicated that L. rhamnosus GAF01 was able to bind AFM1 in vitro and-by mechanisms that might also be related to a binding effect-counteract AFM1-induced immunotoxicity. Moreover, by itself, this bacterium was not toxic and could potentially be used as an additive in dairy products and in biotechnology for

  10. BOREAS AFM-2 Wyoming King Air 1994 Aircraft Sounding Data

    NASA Technical Reports Server (NTRS)

    Kelly, Robert D.; Hall, Forrest G. (Editor); Knapp, David E. (Editor); Smith, David E. (Technical Monitor)

    2000-01-01

    The BOREAS AFM-2 team used the University of Wyoming King Air aircraft during IFCs 1, 2, and 3 in 1994 to collected pass-by-pass fluxes (and many other statistics) for the large number of level (constant altitude), straight-line passes used in a variety of flight patterns over the SSA and NSA and areas along the transect between these study areas. The data described here form a second set, namely soundings that were incorporated into nearly every research flight by the King Air in 1994. These soundings generally went from near the surface to above the inversion layer. Most were flown immediately after takeoff or immediately after finishing the last flux pattern of that particular day's flights. The parameters that were measured include wind direction, wind speed, west wind component (u), south wind component (v), static pressure, air dry bulb temperature, potential temperature, dewpoint, temperature, water vapor mixing ratio, and CO2 concentration. Data on the aircraft's location, attitude, and altitude during data collection are also provided. These data are stored in tabular ASCH files. The data files are available on a CD-ROM (see document number 20010000884) or from the Oak Ridge National Laboratory (ORNL) Distributed Active Archive Center (DAAC).

  11. BOREAS AFM-03-NCAR Electra 1994 Aircraft Sounding Data

    NASA Technical Reports Server (NTRS)

    Lenschow, Donald H.; Oncley, Steven P.; Hall, Forrest G. (Editor); Knapp, David E. (Editor); Smith, David E. (Technical Monitor)

    2000-01-01

    The Boreal Ecosystem-Atmosphere Study (BOREAS) Airborne Fluxes and Meteorology (AFM)-3 team used the National Center for Atmospheric Research's (NCAR) Electra aircraft to make sounding measurements to study the planetary boundary layer using in situ and remote-sensing measurements. Measurements were made of wind speed and direction, air pressure and temperature, potential temperature, dewpoint, mixing ratio of H, O, CO, concentration, and ozone concentration. Twenty-five research missions were flown over the Northern Study Area (NSA), Southern Study Area (SSA), and the transect during BOREAS Intensive Field Campaigns (IFCs) 1, 2, and 3 during 1994. All missions had from four to ten soundings through the top of the planetary boundary layer. This sounding data set contains all of the in situ vertical profiles through the boundary layer top that were made (with the exception of 'porpoise' maneuvers). Data were recorded in one-second time intervals. These data are stored in tabular ASCII files. The NCAR Electra 1994 aircraft sounding data are available from the Earth Observing System Data and Information System (EOSDIS) Oak Ridge National Laboratory (ORNL) Distributed Active Archive Center (DAAC). The data files are available on a CD-ROM (see document number 20010000884).

  12. The Conductance of Nanotubes Deformed by the AFM Tip

    NASA Technical Reports Server (NTRS)

    Svizhenko, Alexei; Maiti, Amitesh; Anantram, M. P.

    2003-01-01

    The conductance drop under AFM-tip deformation can be explained by stretching of the tube length. NT sensors can be built utilizing uniform stretching. Single sp3 bond cross section cannot block electrons, because another conducting path may exist. AFM tip which forms sp3 bonds with the tube will decrease conductance. In the "table experiment" a conductance drop of 2 orders of magnitude happened only after some bonds were broken.

  13. Nanoscale structural features determined by AFM for single virus particles.

    PubMed

    Chen, Shu-wen W; Odorico, Michael; Meillan, Matthieu; Vellutini, Luc; Teulon, Jean-Marie; Parot, Pierre; Bennetau, Bernard; Pellequer, Jean-Luc

    2013-11-21

    In this work, we propose "single-image analysis", as opposed to multi-image averaging, for extracting valuable information from AFM images of single bio-particles. This approach allows us to study molecular systems imaged by AFM under general circumstances without restrictions on their structural forms. As feature exhibition is a resolution correlation, we have performed AFM imaging on surfaces of tobacco mosaic virus (TMV) to demonstrate variations of structural patterns with probing resolution. Two AFM images were acquired with the same tip at different probing resolutions in terms of pixel width, i.e., 1.95 and 0.49 nm per pixel. For assessment, we have constructed an in silico topograph based on the three-dimensional crystal structure of TMV as a reference. The prominent artifacts observed in the AFM-determined shape of TMV were attributed to tip convolutions. The width of TMV rod was systematically overestimated by ~10 nm at both probing resolutions of AFM. Nevertheless, the effects of tip convolution were less severe in vertical orientation so that the estimated height of TMV by AFM imaging was in close agreement with the in silico X-ray topograph. Using dedicated image processing algorithms, we found that at low resolution (i.e., 1.95 nm per pixel), the extracted surface features of TMV can be interpreted as a partial or full helical repeat (three complete turns with ~7.0 nm in length), while individual protein subunits (~2.5 nm) were perceivable only at high resolution. The present study shows that the scales of revealed structural features in AFM images are subject to both probing resolution and processing algorithms for image analysis. PMID:24056758

  14. Characterization of the interaction between AFM tips and surface nanobubbles.

    PubMed

    Walczyk, Wiktoria; Schönherr, Holger

    2014-06-24

    While the presence of gaseous enclosures observed at various solid-water interfaces, the so-called "surface nanobubles", has been confirmed by many groups in recent years, their formation, properties, and stability have not been convincingly and exhaustively explained. Here we report on an atomic force microscopy (AFM) study of argon nanobubbles on highly oriented pyrolitic graphite (HOPG) in water to elucidate the properties of nanobubble surfaces and the mechanism of AFM tip-nanobubble interaction. In particular, the deformation of the nanobubble-water interface by the AFM tip and the question whether the AFM tip penetrates the nanobubble during scanning were addressed by this combined intermittent contact (tapping) mode and force volume AFM study. We found that the stiffness of nanobubbles was smaller than the cantilever spring constant and comparable with the surface tension of water. The interaction with the AFM tip resulted in severe quasi-linear deformation of the bubbles; however, in the case of tip-bubble attraction, the interface deformed toward the tip. We tested two models of tip-bubble interaction, namely, the capillary force and the dynamic interaction model, and found, depending on the tip properties, good agreement with experimental data. The results showed that the tip-bubble interaction strength and the magnitude of the bubble deformation depend strongly on tip and bubble geometry and on tip and substrate material, and are very sensitive to the presence of contaminations that alter the interfacial tension. In particular, nanobubbles interacted differently with hydrophilic and hydrophobic AFM tips, which resulted in qualitatively and quantitatively different force curves measured on the bubbles in the experiments. To minimize bubble deformation and obtain reliable AFM results, nanobubbles must be measured with a sharp hydrophilic tip and with a cantilever having a very low spring constant in a contamination-free system. PMID:24856074

  15. Modeling the Interaction between AFM Tips and Pinned Surface Nanobubbles.

    PubMed

    Guo, Zhenjiang; Liu, Yawei; Xiao, Qianxiang; Schönherr, Holger; Zhang, Xianren

    2016-01-26

    Although the morphology of surface nanobubbles has been studied widely with different AFM modes, AFM images may not reflect the real shapes of the nanobubbles due to AFM tip-nanobubble interactions. In addition, the interplay between surface nanobubble deformation and induced capillary force has not been well understood in this context. In our work we used constraint lattice density functional theory to investigate the interaction between AFM tips and pinned surface nanobubbles systematically, especially concentrating on the effects of tip hydrophilicity and shape. For a hydrophilic tip contacting a nanobubble, its hydrophilic nature facilitates its departure from the bubble surface, displaying a weak and intermediate-range attraction. However, when the tip squeezes the nanobubble during the approach process, the nanobubble shows an elastic effect that prevents the tip from penetrating the bubble, leading to a strong nanobubble deformation and repulsive interactions. On the contrary, a hydrophobic tip can easily pierce the vapor-liquid interface of the nanobubble during the approach process, leading to the disappearance of the repulsive force. In the retraction process, however, the adhesion between the tip and the nanobubble leads to a much stronger lengthening effect on nanobubble deformation and a strong long-range attractive force. The trends of force evolution from our simulations agree qualitatively well with recent experimental AFM observations. This favorable agreement demonstrates that our model catches the main intergradient of tip-nanobubble interactions for pinned surface nanobubbles and may therefore provide important insight into how to design minimally invasive AFM experiments. PMID:26751634

  16. Synthesis of tertiary arylamines: Lewis acid-catalyzed direct reductive N-alkylation of secondary amines with ketones through an alternative pathway.

    PubMed

    Nayal, Onkar S; Thakur, Maheshwar S; Bhatt, Vinod; Kumar, Manoranjan; Kumar, Neeraj; Singh, Bikram; Sharma, Upendra

    2016-08-11

    We report herein a highly efficient, tin(ii)/PMHS catalyzed reductive N-alkylation of arylamines with ketones affording tertiary arylamines. A very wide substrate scope was observed for the current catalytic method as all six permutations of ketones/aldehydes/heterocyclic carbonyls and primary/secondary/heterocyclic amines were well tolerated, enabling access to secondary, tertiary and heterocyclic amines. The method is also convenient for the synthesis of N-substituted isoindolinones and phthalazinones via a tandem amination-amidation sequence. Mechanistic investigations revealed a carbocationic pathway instead of an ordinary direct reductive amination pathway. PMID:27363507

  17. Nanoscopic polypyrrole AFM-SECM probes enabling force measurements under potential control

    NASA Astrophysics Data System (ADS)

    Knittel, P.; Higgins, M. J.; Kranz, C.

    2014-01-01

    Conductive polymers, and in particular polypyrrole, are frequently used as biomimetic interfaces facilitating growth and/or differentiation of cells and tissues. Hence, studying forces and local interactions between such polymer interfaces and cells at the nanoscale is of particular interest. Frequently, such force interactions are not directly accessible with high spatial resolution. Consequently, we have developed nanoscopic polypyrrole electrodes, which are integrated in AFM-SECM probes. Bifunctional AFM-SECM probes were modified via ion beam-induced deposition resulting in pyramidal conductive Pt-C composite electrodes. These nanoscopic electrodes then enabled localized polypyrrole deposition, thus resulting in polymer-modified AFM probes with a well-defined geometry. Furthermore, such probes may be reversibly switched from an insulating to a conductive state. In addition, the hydrophilicity of such polymer tips is dependent on the dopant, and hence, on the oxidation state. Force studies applying different tip potentials were performed at plasma-treated glass surfaces providing localized information on the associated force interactions, which are dependent on the applied potential and the dopant.Conductive polymers, and in particular polypyrrole, are frequently used as biomimetic interfaces facilitating growth and/or differentiation of cells and tissues. Hence, studying forces and local interactions between such polymer interfaces and cells at the nanoscale is of particular interest. Frequently, such force interactions are not directly accessible with high spatial resolution. Consequently, we have developed nanoscopic polypyrrole electrodes, which are integrated in AFM-SECM probes. Bifunctional AFM-SECM probes were modified via ion beam-induced deposition resulting in pyramidal conductive Pt-C composite electrodes. These nanoscopic electrodes then enabled localized polypyrrole deposition, thus resulting in polymer-modified AFM probes with a well

  18. Structure and permeability of ion-channels by integrated AFM and waveguide TIRF microscopy.

    PubMed

    Ramachandran, Srinivasan; Arce, Fernando Teran; Patel, Nirav R; Quist, Arjan P; Cohen, Daniel A; Lal, Ratnesh

    2014-01-01

    Membrane ion channels regulate key cellular functions and their activity is dependent on their 3D structure. Atomic force microscopy (AFM) images 3D structure of membrane channels placed on a solid substrate. Solid substrate prevents molecular transport through ion channels thus hindering any direct structure-function relationship analysis. Here we designed a ~70 nm nanopore to suspend a membrane, allowing fluidic access to both sides. We used these nanopores with AFM and total internal reflection fluorescence microscopy (TIRFM) for high resolution imaging and molecular transport measurement. Significantly, membranes over the nanopore were stable for repeated AFM imaging. We studied structure-activity relationship of gap junction hemichannels reconstituted in lipid bilayers. Individual hemichannels in the membrane overlying the nanopore were resolved and transport of hemichannel-permeant LY dye was visualized when the hemichannel was opened by lowering calcium in the medium. This integrated technique will allow direct structure-permeability relationship of many ion channels and receptors. PMID:24651823

  19. Crystallization of Probucol in Nanoparticles Revealed by AFM Analysis in Aqueous Solution.

    PubMed

    Egami, Kiichi; Higashi, Kenjirou; Yamamoto, Keiji; Moribe, Kunikazu

    2015-08-01

    The crystallization behavior of a pharmaceutical drug in nanoparticles was directly evaluated by atomic force microscopy (AFM) force curve measurements in aqueous solution. A ternary spray-dried sample (SPD) was prepared by spray drying the organic solvent containing probucol (PBC), hypromellose (HPMC), and sodium dodecyl sulfate (SDS). The amorphization of PBC in the ternary SPD was confirmed by powder X-ray diffraction (PXRD) and solid-state 13C NMR measurements. A nanosuspension containing quite small particles of 25 nm in size was successfully prepared immediately after dispersion of the ternary SPD into water. Furthermore, solution-state 1H NMR measurements revealed that a portion of HPMC coexisted with PBC as a mixed state in the freshly prepared nanosuspension particles. After storing the nanosuspension at 25 °C, a gradual increase in the size of the nanoparticles was observed, and the particle size changed to 93.9 nm after 7 days. AFM enabled the direct observation of the morphology and agglomeration behavior of the nanoparticles in water. Moreover, AFM force-distance curves were changed from (I) to (IV), depending on the storage period, as follows: (I) complete indentation within an applied force of 1 nN, (II) complete indentation with an applied force of 1-5 nN, (III) partial indentation with an applied force of 5 nN, and (IV) nearly no indentation with an applied force of 5 nN. This stiffness increase of the nanoparticles was attributed to gradual changes in the molecular state of PBC from the amorphous to the crystal state. Solid-state 13C NMR measurements of the freeze-dried samples demonstrated the presence of metastable PBC Form II crystals in the stored nanosuspension, strongly supporting the AFM results. PMID:26106951

  20. An Interferon Regulated MicroRNA Provides Broad Cell-Intrinsic Antiviral Immunity through Multihit Host-Directed Targeting of the Sterol Pathway

    PubMed Central

    Robertson, Kevin A.; Hsieh, Wei Yuan; Forster, Thorsten; Blanc, Mathieu; Lu, Hongjin; Crick, Peter J.; Yutuc, Eylan; Watterson, Steven; Martin, Kimberly; Griffiths, Samantha J.; Enright, Anton J.; Yamamoto, Mami; Pradeepa, Madapura M.; Lennox, Kimberly A.; Behlke, Mark A.; Talbot, Simon; Haas, Jürgen; Dölken, Lars; Griffiths, William J.; Wang, Yuqin; Angulo, Ana; Ghazal, Peter

    2016-01-01

    In invertebrates, small interfering RNAs are at the vanguard of cell-autonomous antiviral immunity. In contrast, antiviral mechanisms initiated by interferon (IFN) signaling predominate in mammals. Whilst mammalian IFN-induced miRNA are known to inhibit specific viruses, it is not known whether host-directed microRNAs, downstream of IFN-signaling, have a role in mediating broad antiviral resistance. By performing an integrative, systematic, global analysis of RNA turnover utilizing 4-thiouridine labeling of newly transcribed RNA and pri/pre-miRNA in IFN-activated macrophages, we identify a new post-transcriptional viral defense mechanism mediated by miR-342-5p. On the basis of ChIP and site-directed promoter mutagenesis experiments, we find the synthesis of miR-342-5p is coupled to the antiviral IFN response via the IFN-induced transcription factor, IRF1. Strikingly, we find miR-342-5p targets mevalonate-sterol biosynthesis using a multihit mechanism suppressing the pathway at different functional levels: transcriptionally via SREBF2, post-transcriptionally via miR-33, and enzymatically via IDI1 and SC4MOL. Mass spectrometry-based lipidomics and enzymatic assays demonstrate the targeting mechanisms reduce intermediate sterol pathway metabolites and total cholesterol in macrophages. These results reveal a previously unrecognized mechanism by which IFN regulates the sterol pathway. The sterol pathway is known to be an integral part of the macrophage IFN antiviral response, and we show that miR-342-5p exerts broad antiviral effects against multiple, unrelated pathogenic viruses such Cytomegalovirus and Influenza A (H1N1). Metabolic rescue experiments confirm the specificity of these effects and demonstrate that unrelated viruses have differential mevalonate and sterol pathway requirements for their replication. This study, therefore, advances the general concept of broad antiviral defense through multihit targeting of a single host pathway. PMID:26938778

  1. Evaluating legacy contaminants and emerging chemicals in marine environments using adverse outcome pathways and biological effects-directed analysis.

    PubMed

    Hutchinson, Thomas H; Lyons, Brett P; Thain, John E; Law, Robin J

    2013-09-30

    important scientific, economic and health challenges. In order to meet these challenges and pursue cost-effective scientific approaches that can provide evidence necessary to support policy needs (e.g. the European Marine Strategy Framework Directive), it is widely recognised that there is a need to (i) provide marine exposure assessments for priority contaminants using a range of validated models, passive samplers and biomarkers; (ii) integrate chemical monitoring data with biological effects data across spatial and temporal scales (including quality controls); and (iii) strengthen the evidence base to understand the relationship between exposure to complex chemical mixtures, biological and ecological impacts through integrated approaches and molecular data (e.g. genomics, proteomics and metabolomics). Additionally, we support the widely held view that (iv) that rather than increasing the analytical chemistry monitoring of large number of emerging contaminants, it will be important to target analytical chemistry towards key groups of chemicals of concern using effects-directed analysis. It is also important to evaluate to what extent existing biomarkers and bioassays can address various classes of emerging chemicals using the adverse outcome pathway (AOP) approach now being developed by the Organization for Economic Cooperation and Development (OECD) with respect to human toxicology and ecotoxicology. PMID:23820191

  2. Caudal medullary pathways to lumbosacral motoneuronal cell groups in the cat: evidence for direct projections possibly representing the final common pathway for lordosis.

    PubMed

    Vanderhorst, V G; Holstege, G

    1995-08-28

    The nucleus retroambiguus (NRA) projects to distinct brainstem and cervical and thoracic cord motoneuronal cell groups. The present paper describes NRA projections to distinct motoneuronal cell groups in the lumbar enlargement. Lumbosacral injections of wheat germ agglutinin-horseradish peroxidase (WGA-HRP) were made to localize and quantify the retrogradely labeled neurons in the caudal medullary lateral tegmentum. These injections were combined with spinal hemisections to distinguish between neurons having ipsi-or contralaterally descending axons. The NRA-lumbosacral fibers descend almost exclusively contralaterally, but neurons in areas surrounding the NRA project mainly ipsilaterally. In an anterograde tracing study, injections of WGA-HRP or tritiated leucine were made in the region of the NRA to determine the NRA targets in the lumbosarcral cord. Hemisections in C2 made it possible to distinguish between NRA projections and projections from neurons in the adjoining lateral tegmentum. The results show delicate NRA projections to distinct lumbosacral motoneuronal cell groups innervating specific hindlimb muscles (iliopsoas, adductors, and hamstrings) as well as axial muscles (medial longissimus and proximal tail muscles). The projection is bilateral, with a contralateral predominance. Ipsilaterally terminating fibers are derived from NRA neurons whose axons cross the midline at the level of the obex, descend through the contralateral spinal white matter, and recross at the level of termination. A conceptual description is presented in which the periaqueductal gray-NRA-lumbosacral projections form the final common pathway for lordosis in the cat. PMID:7499541

  3. Molecular dynamics study on the mechanism of AFM-based nanoscratching process with water-layer lubrication

    NASA Astrophysics Data System (ADS)

    Ren, Jiaqi; Zhao, Jinsheng; Dong, Zeguang; Liu, Pinkuan

    2015-08-01

    The atomic force microscopy (AFM) based direct nanoscratching has been thoroughly studied but the mechanism of nanoscratching with water-layer lubrication is yet to be well understood. In current study, three-dimensional molecular dynamics (MD) simulations are conducted to evaluate the effects of the water-layer lubrication on the AFM-based nanoscratching process on monocrystalline copper. Comparisons of workpiece deformation, scratching forces, and friction coefficients are made between the water-lubricated and dry scratching under various thickness of water layer, scratching depth and scratching velocity. Simulation results reveal that the water layer has positive impact on the surface quality and significant influence on the scratching forces (normal forces and tangential forces). The friction coefficients of the tip in water-lubricated nanoscratching are significantly bigger than those in the dry process. Our simulation results shed lights on a promising AFM-based nanofabrication method, which can assist to get nanoscale surface morphologies with higher quality than traditional approaches.

  4. AFM investigation of Martian soil simulants on micromachined Si substrates.

    PubMed

    Vijendran, S; Sykulska, H; Pike, W T

    2007-09-01

    The micro and nanostructures of Martian soil simulants with particles in the micrometre-size range have been studied using a combination of optical and atomic force microscopy (AFM) in preparation for the 2007 NASA Phoenix Mars Lander mission. The operation of an atomic force microscope on samples of micrometre-sized soil particles is a poorly investigated area where the unwanted interaction between the scanning tip and loose particles results in poor image quality and tip contamination by the sample. In order to mitigate these effects, etched silicon substrates with a variety of features have been used to facilitate the sorting and gripping of particles. From these experiments, a number of patterns were identified that were particularly good at isolating and immobilizing particles for AFM imaging. This data was used to guide the design of micromachined substrates for the Phoenix AFM. Both individual particles as well as aggregates were successfully imaged, and information on sizes, shapes and surface morphologies were obtained. This study highlights both the strengths and weaknesses of AFM for the potential in situ investigation of Martian soil and dust. Also presented are more general findings of the limiting operational constraints that exist when attempting the AFM of high aspect ratio particles with current technology. The performance of the final designs of the substrates incorporated on Phoenix will be described in a later paper. PMID:17760618

  5. A low-cost AFM setup with an interferometer for undergraduates and secondary-school students

    NASA Astrophysics Data System (ADS)

    Bergmann, Antje; Feigl, Daniela; Kuhn, David; Schaupp, Manuel; Quast, Günter; Busch, Kurt; Eichner, Ludwig; Schumacher, Jens

    2013-07-01

    Atomic force microscopy (AFM) is an important tool in nanotechnology. This method makes it possible to observe nanoscopic surfaces beyond the resolution of light microscopy. In order to provide undergraduate and secondary-school students with insights into this world, we have developed a very robust low-cost AFM setup with a Fabry-Perot interferometer as a detecting device. This setup is designed to be operated almost completely manually and its simplicity gives access to a profound understanding of the working principle. Our AFM is operated in a constant height mode, i.e. the topography of the sample surface is represented directly by the deflection of the cantilever. Thus, the measuring procedure can be understood even by secondary-school students; furthermore, it is the method with the lowest cost, totalling not more than 10-15 k Euros. Nevertheless, we are able to examine a large variety of sample topographies such as CD and DVD surfaces, IC structures, blood cells, butterfly wings or moth eyes. Furthermore, force-distance curves can be recorded and the tensile moduli of some materials can be evaluated. We present our setup in detail and describe its working principles. In addition, we show various experiments which have already been performed by students.

  6. Graphene Nanopore Support System for Simultaneous High-Resolution AFM Imaging and Conductance Measurements

    PubMed Central

    2015-01-01

    Accurately defining the nanoporous structure and sensing the ionic flow across nanoscale pores in thin films and membranes has a wide range of applications, including characterization of biological ion channels and receptors, DNA sequencing, molecule separation by nanoparticle films, sensing by block co-polymers films, and catalysis through metal–organic frameworks. Ionic conductance through nanopores is often regulated by their 3D structures, a relationship that can be accurately determined only by their simultaneous measurements. However, defining their structure–function relationships directly by any existing techniques is still not possible. Atomic force microscopy (AFM) can image the structures of these pores at high resolution in an aqueous environment, and electrophysiological techniques can measure ion flow through individual nanoscale pores. Combining these techniques is limited by the lack of nanoscale interfaces. We have designed a graphene-based single-nanopore support (∼5 nm thick with ∼20 nm pore diameter) and have integrated AFM imaging and ionic conductance recording using our newly designed double-chamber recording system to study an overlaid thin film. The functionality of this integrated system is demonstrated by electrical recording (<10 pS conductance) of suspended lipid bilayers spanning a nanopore and simultaneous AFM imaging of the bilayer. PMID:24581087

  7. Surface Morphological Studies on Nerve Cells by AFM

    NASA Astrophysics Data System (ADS)

    Durkaya, Goksel; Zhong, Lei; Rehder, Vincent; Dietz, Nikolaus

    2009-03-01

    Surface morphological properties of fixed and living nerve cells removed from the buccal ganglion of Helisoma trivolvis have been studied by using Atomic Force Microscopy (AFM). Identified, individual neurons were removed from the buccal ganglion of Helisoma trivolvis and plated into poly-L-lysine coated glass cover-slips. The growth of the nerve cells was stopped and fixed with 0.1% Glutaraldehyde and 4% Formaldehyde solution after extension of growth cones at the tip of the axons. Topography and softness of growth cone filopodia and overlying lamellopodium (veil) were probed by AFM. Information obtained from AFM's amplitude and phase channels have been used for determination of softness of the region probed. The results of structural studies on the cells are linked to their mechanical properties and internal molecular density distribution.

  8. Imaging resolution of AFM with probes modified with FIB.

    PubMed

    Skibinski, J; Rebis, J; Wejrzanowski, T; Rozniatowski, K; Pressard, K; Kurzydlowski, K J

    2014-11-01

    This study concerns imaging of the structure of materials using AFM tapping (TM) and phase imaging (PI) mode, using probes modified with focused ion beam (FIB). Three kinds of modifications were applied - thinning of the cantilever, sharpening of the tip and combination of these two modifications. Probes shaped in that way were used for AFM investigations with Bruker AFM Nanoscope 8. As a testing material, titanium roughness standard supplied by Bruker was used. The results show that performed modifications influence the oscillation of the probes. In particular thinning of the cantilever enables one to acquire higher self-resonant frequencies, which can be advantageous for improving the quality of imaging in PI mode. It was found that sharpening the tip improves imaging resolution in tapping mode, which is consistent with existing knowledge, but lowered the quality of high frequency topography images. In this paper the Finite Element Method (FEM) was used to explain the results obtained experimentally. PMID:25080273

  9. AFM of biological complexes: what can we learn?

    PubMed Central

    Gaczynska, Maria; Osmulski, Pawel A.

    2009-01-01

    The term “biological complexes” broadly encompasses particles as diverse as multisubunit enzymes, viral capsids, transport cages, molecular nets, ribosomes, nucleosomes, biological membrane components and amyloids. The complexes represent a broad range of stability and composition. Atomic force microscopy offers a wealth of structural and functional data about such assemblies. For this review, we choose to comment on the significance of AFM to study various aspects of biology of selected nonmembrane protein assemblies. Such particles are large enough to reveal many structural details under the AFM probe. Importantly, the specific advantages of the method allow for gathering dynamic information about their formation, stability or allosteric structural changes critical for their function. Some of them have already found their way to nanomedical or nanotechnological applications. Here we present examples of studies where the AFM provided pioneering information about the biology of complexes, and examples of studies where the simplicity of the method is used toward the development of potential diagnostic applications. PMID:19802337

  10. Charging C60 islands with the AFM tip.

    PubMed

    Hoff, Brice; Henry, Claude R; Barth, Clemens

    2016-01-01

    We show that electrons can be transferred on demand from an AFM tip into single bulk-like C60 islands, which are supported on the insulating NaCl(001) surface. We exemplify this by controlled charge-manipulation experiments conducted in ultrahigh vacuum by noncontact AFM (nc-AFM), electrostatic force microscopy (EFM) and Kelvin probe force microscopy (KPFM). KPFM shows a homogeneous contrast at the islands, which is a signature for an equal distribution of the electrons in the T1u band. The charge dissipates during half a day due to an interaction of the charged C60 islands with defects in the near surface region of NaCl. Our results open the perspective in photo-voltaics to study charge attachment, stability and charge exchange with the environment of any C60 bulk-like system. PMID:26617348

  11. Mounting of Escherichia coli spheroplasts for AFM imaging.

    SciTech Connect

    Sullivan, Claretta J; Morrell-Falvey, Jennifer L; Allison, David P; Doktycz, Mitchel John

    2005-11-01

    The cytoplasmic membrane of Escherichia coli (E. coli) is the location of numerous, chemically specific transporters and recognition elements. Investigation of this membrane in vivo by atomic force microscopy (AFM) requires removal of the cell wall and stable immobilization of the spheroplast. AFM images demonstrate that spheroplasts can be secured with warm gelatin applied to the mica substrate just before the addition of a spheroplast suspension. The resulting preparation can be repeatedly imaged by AFM over the course of several hours. Confocal fluorescence imaging confirms the association of the spheroplasts with the gelatin layer. Gelatin molecules are known to reorder into a network after heating. Entrapment within this gelatin network is believed to be responsible for the immobilization of spheroplasts on mica.

  12. Improvement in metrology on new 3D-AFM platform

    NASA Astrophysics Data System (ADS)

    Schmitz, Ingo; Osborn, Marc; Hand, Sean; Chen, Qi

    2008-10-01

    According to the 2007 edition of the ITRS roadmap, the requirement for CD uniformity of isolated lines on a binary or attenuated phase shift mask is 2.1nm (3σ) in 2008 and requires improvement to1.3 nm (3σ) in 2010. In order to meet the increasing demand for CD uniformity on photo masks, improved CD metrology is required. A next generation AFM, InSightTM 3DAFM, has been developed to meet these increased requirements for advanced photo mask metrology. The new system achieves 2X improvement in CD and depth precision on advanced photo masks features over the previous generation 3D-AFM. This paper provides measurement data including depth, CD, and sidewall angle metrology. In addition the unique capabilities of damage-free defect inspection and Nanoimprint characterization by 3D AFM are presented.

  13. Recent CD AFM probe developments for sub-45 nm technology nodes

    NASA Astrophysics Data System (ADS)

    Liu, Hao-Chih; Osborne, Jason R.; Dahlen, Gregory A.; Greschner, Johann; Bayer, Thomas; Kalt, Samuel; Fritz, Georg

    2008-03-01

    This paper reports on new developments of advanced CD AFM probes after the prior introduction of "trident probes" in SPIE Advanced Lithography 2007 [1]. Trident probes, having sharpened extensions in the tip apex region, make possible bottom CD measurements within a few nanometers of the feature bottom corner; an area where other CD probes have difficulties due to tip shape limitations. Moreover, new metrology applications of trident probes have been developed for novel devices such as FinFET and vertical read/write hard disk heads. For ever smaller technology nodes, new probes evolved from the design of the trident probe. For example, the number of sharpened tip flares was reduced from three (trident) to two (bi-pod) to prevent possible interference of the third leg in the slow scan direction, as shown in Figure 3. Maintaining tip lateral stiffness as the tip size shrinks to less than 30 nm is vital for successful scanning. Consequently, a significant recent improvement is the change of probe shank cross-sectional geometry in order to maintain tip vertical aspect ratio of 1:5 (and lateral stiffness > 1 N/m). Finally, modifications of probe substrate are proposed and evaluated for current and new CD AFM systems. Hydrophobic, self-assembled monolayer (SAM) coatings were applied on CD probes to reduced tip "pull-away" distance1 during CD AFM scanning. Test results show that the pull away distance can be reduced more than 5 times on average (in some cases, by a factor of 15). Consequently, use of hydrophobic SAM coatings on CD probes mitigates pull-away distance thus allowing narrow trench CD measurements. We discuss limitations of prior CD AFM probes and design considerations of new CD probes. The characterization of first prototypes and evaluation of scan performance are presented in this work.

  14. Charging C60 islands with the AFM tip

    NASA Astrophysics Data System (ADS)

    Hoff, Brice; Henry, Claude R.; Barth, Clemens

    2015-12-01

    We show that electrons can be transferred on demand from an AFM tip into single bulk-like C60 islands, which are supported on the insulating NaCl(001) surface. We exemplify this by controlled charge-manipulation experiments conducted in ultrahigh vacuum by noncontact AFM (nc-AFM), electrostatic force microscopy (EFM) and Kelvin probe force microscopy (KPFM). KPFM shows a homogeneous contrast at the islands, which is a signature for an equal distribution of the electrons in the T1u band. The charge dissipates during half a day due to an interaction of the charged C60 islands with defects in the near surface region of NaCl. Our results open the perspective in photo-voltaics to study charge attachment, stability and charge exchange with the environment of any C60 bulk-like system.We show that electrons can be transferred on demand from an AFM tip into single bulk-like C60 islands, which are supported on the insulating NaCl(001) surface. We exemplify this by controlled charge-manipulation experiments conducted in ultrahigh vacuum by noncontact AFM (nc-AFM), electrostatic force microscopy (EFM) and Kelvin probe force microscopy (KPFM). KPFM shows a homogeneous contrast at the islands, which is a signature for an equal distribution of the electrons in the T1u band. The charge dissipates during half a day due to an interaction of the charged C60 islands with defects in the near surface region of NaCl. Our results open the perspective in photo-voltaics to study charge attachment, stability and charge exchange with the environment of any C60 bulk-like system. Electronic supplementary information (ESI) available. See DOI: 10.1039/C5NR04541J

  15. BOREAS AFM-04 Twin Otter Aircraft Flux Data

    NASA Technical Reports Server (NTRS)

    MacPherson, J. Ian; Hall, Forrest G. (Editor); Knapp, David E. (Editor); Desjardins, Raymond L.; Smith, David E. (Technical Monitor)

    2000-01-01

    The BOREAS AFM-5 team collected and processed data from the numerous radiosonde flights during the project. The goals of the AFM-05 team were to provide large-scale definition of the atmosphere by supplementing the existing AES aerological network, both temporally and spatially. This data set includes basic upper-air parameters collected from the network of upper-air stations during the 1993, 1994, and 1996 field campaigns over the entire study region. The data are contained in tabular ASCII files. The data files are available on a CD-ROM (see document number 20010000884) or from the Oak Ridge National Laboratory (ORNL) Distributed Active Archive Center (DAAC).

  16. Directing dopaminergic fiber growth along a preformed molecular pathway from embryonic ventral mesencephalon transplants in the rat brain.

    PubMed

    Jin, Y; Zhang, C; Ziemba, K S; Goldstein, G A; Sullivan, P G; Smith, G M

    2011-05-01

    To identify guidance molecules to promote long-distance growth of dopaminergic axons from transplanted embryonic ventral mesencephalon (VM) tissue, three pathways were created by expressing green fluorescent protein (GFP), glial cell line-derived neurotrophic factor (GDNF), or a combination of GDNF/GDNF receptor α1 (GFRα1) along the corpus callosum. To generate the guidance pathway, adenovirus encoding these transcripts was injected at four positions along the corpus callosum. In all groups, GDNF adenovirus was also injected on the right side 2.5 mm from the midline at the desired transplant site. Four days later, a piece of VM tissue from embryonic day 14 rats was injected at the transplant site. All rats also received daily subcutaneous injections of N-acetyl-L-cysteinamide (NACA; 100 μg per rat) as well as chondroitinase ABC at transplant site (10 U/ml, 2 μl). Two weeks after transplantation, the rats were perfused and the brains dissected out. Coronal sections were cut and immunostained with antibody to tyrosine hydroxylase (TH) to identify and count dopaminergic fibers in the corpus callosum. In GFP-expressing pathways, TH(+) fibers grew out of the transplants for a short distance in the corpus callosum. Very few TH(+) fibers grew across the midline. However, pathways expressing GDNF supported more TH(+) fiber growth across the midline into the contralateral hemisphere. Significantly greater numbers of TH(+) fibers grew across the midline in animals expressing a combination of GDNF and GFRα1 in the corpus callosum. These data suggest that expression of GDNF or a combination of GDNF and GFRα1 can support the long-distance dopaminergic fiber growth from a VM transplant, with the combination having a superior effect. PMID:21337366

  17. A Direct Link between Abscisic Acid Sensing and the Chromatin-Remodeling ATPase BRAHMA via Core ABA Signaling Pathway Components.

    PubMed

    Peirats-Llobet, Marta; Han, Soon-Ki; Gonzalez-Guzman, Miguel; Jeong, Cheol Woong; Rodriguez, Lesia; Belda-Palazon, Borja; Wagner, Doris; Rodriguez, Pedro L

    2016-01-01

    Optimal response to drought is critical for plant survival and will affect biodiversity and crop performance during climate change. Mitotically heritable epigenetic or dynamic chromatin state changes have been implicated in the plant response to the drought stress hormone abscisic acid (ABA). The Arabidopsis SWI/SNF chromatin-remodeling ATPase BRAHMA (BRM) modulates response to ABA by preventing premature activation of stress response pathways during germination. We show that core ABA signaling pathway components physically interact with BRM and post-translationally modify BRM by phosphorylation/dephosphorylation. Genetic evidence suggests that BRM acts downstream of SnRK2.2/2.3 kinases, and biochemical studies identified phosphorylation sites in the C-terminal region of BRM at SnRK2 target sites that are evolutionarily conserved. Finally, the phosphomimetic BRM(S1760D S1762D) mutant displays ABA hypersensitivity. Prior studies showed that BRM resides at target loci in the ABA pathway in the presence and absence of the stimulus, but is only active in the absence of ABA. Our data suggest that SnRK2-dependent phosphorylation of BRM leads to its inhibition, and PP2CA-mediated dephosphorylation of BRM restores the ability of BRM to repress ABA response. These findings point to the presence of a rapid phosphorylation-based switch to control BRM activity; this property could be potentially harnessed to improve drought tolerance in plants. PMID:26499068

  18. Suppression of esophageal tumor growth and chemoresistance by directly targeting the PI3K/AKT pathway.

    PubMed

    Li, Bin; Li, Jin; Xu, Wen Wen; Guan, Xin Yuan; Qin, Yan Ru; Zhang, Li Yi; Law, Simon; Tsao, Sai Wah; Cheung, Annie L M

    2014-11-30

    Esophageal cancer is the sixth most common cause of cancer-related deaths worldwide. Novel therapeutic intervention is urgently needed for this deadly disease. The functional role of PI3K/AKT pathway in esophageal cancer is little known. In this study, our results from 49 pairs of human esophageal tumor and normal specimens demonstrated that AKT was constitutively active in the majority (75.5%) of esophageal tumors compared with corresponding normal tissues. Inhibition of the PI3K/AKT pathway with specific inhibitors, wortmannin and LY294002, significantly reduced Bcl-xL expression, induced caspase-3-dependent apoptosis, and repressed cell proliferation and tumor growth in vitro and in vivo without obvious toxic effects. Moreover, significantly higher expression level of p-AKT was observed in fluorouracil (5-FU)-resistant esophageal cancer cells. Inactivation of PI3K/AKT pathway markedly increased the sensitivity and even reversed acquired resistance of esophageal cancer cells to chemotherapeutic drugs in vitro. More importantly, the resistance of tumor xenografts derived from esophageal cancer cells with acquired 5-FU resistance to chemotherapeutic drugs was significantly abrogated by wortmannin treatment in animals. In summary, our data support PI3K/AKT as a valid therapeutic target and strongly suggest that PI3K/AKT inhibitors used in conjunction with conventional chemotherapy may be a potentially useful therapeutic strategy in treating esophageal cancer patients. PMID:25344912

  19. Conductive supports for combined AFM SECM on biological membranes

    NASA Astrophysics Data System (ADS)

    Frederix, Patrick L. T. M.; Bosshart, Patrick D.; Akiyama, Terunobu; Chami, Mohamed; Gullo, Maurizio R.; Blackstock, Jason J.; Dooleweerdt, Karin; de Rooij, Nico F.; Staufer, Urs; Engel, Andreas

    2008-09-01

    Four different conductive supports are analysed regarding their suitability for combined atomic force and scanning electrochemical microscopy (AFM-SECM) on biological membranes. Highly oriented pyrolytic graphite (HOPG), MoS2, template stripped gold, and template stripped platinum are compared as supports for high resolution imaging of reconstituted membrane proteins or native membranes, and as electrodes for transferring electrons from or to a redox molecule. We demonstrate that high resolution topographs of the bacterial outer membrane protein F can be recorded by contact mode AFM on all four supports. Electrochemical feedback experiments with conductive cantilevers that feature nanometre-scale electrodes showed fast re-oxidation of the redox couple Ru(NH3)63+/2+ with the two metal supports after prolonged immersion in electrolyte. In contrast, the re-oxidation rates decayed quickly to unpractical levels with HOPG or MoS2 under physiological conditions. On HOPG we observed heterogeneity in the re-oxidation rate of the redox molecules with higher feedback currents at step edges. The latter results demonstrate the capability of conductive cantilevers with small electrodes to measure minor variations in an SECM signal and to relate them to nanometre-scale features in a simultaneously recorded AFM topography. Rapid decay of re-oxidation rate and surface heterogeneity make HOPG or MoS2 less attractive for combined AFM-SECM experiments on biological membranes than template stripped gold or platinum supports.

  20. Hydrodynamic effects in fast AFM single-molecule force measurements.

    PubMed

    Janovjak, Harald; Struckmeier, Jens; Müller, Daniel J

    2005-02-01

    Atomic force microscopy (AFM) allows the critical forces that unfold single proteins and rupture individual receptor-ligand bonds to be measured. To derive the shape of the energy landscape, the dynamic strength of the system is probed at different force loading rates. This is usually achieved by varying the pulling speed between a few nm/s and a few microm/s, although for a more complete investigation of the kinetic properties higher speeds are desirable. Above 10 microm/s, the hydrodynamic drag force acting on the AFM cantilever reaches the same order of magnitude as the molecular forces. This has limited the maximum pulling speed in AFM single-molecule force spectroscopy experiments. Here, we present an approach for considering these hydrodynamic effects, thereby allowing a correct evaluation of AFM force measurements recorded over an extended range of pulling speeds (and thus loading rates). To support and illustrate our theoretical considerations, we experimentally evaluated the mechanical unfolding of a multi-domain protein recorded at 30 microm/s pulling speed. PMID:15257425

  1. AFM CHARACTERIZATION OF LASER INDUCED DAMAGE ON CDZNTE CRYSTAL SURFACES

    SciTech Connect

    Hawkins, S; Lucile Teague, L; Martine Duff, M; Eliel Villa-Aleman, E

    2008-06-10

    Semi-conducting CdZnTe (or CZT) crystals can be used in a variety of detector-type applications. CZT shows great promise for use as a gamma radiation spectrometer. However, its performance is adversely affected by point defects, structural and compositional heterogeneities within the crystals, such as twinning, pipes, grain boundaries (polycrystallinity), secondary phases and in some cases, damage caused by external forces. One example is damage that occurs during characterization of the surface by a laser during Raman spectroscopy. Even minimal laser power can cause Te enriched areas on the surface to appear. The Raman spectra resulting from measurements at moderate intensity laser power show large increases in peak intensity that is attributed to Te. Atomic Force Microscopy (AFM) was used to characterize the extent of damage to the CZT crystal surface following exposure to the Raman laser. AFM data reveal localized surface damage in the areas exposed to the Raman laser beam. The degree of surface damage to the crystal is dependent on the laser power, with the most observable damage occurring at high laser power. Moreover, intensity increases in the Te peaks of the Raman spectra are observed even at low laser power with little to no visible damage observed by AFM. AFM results also suggest that exposure to the same amount of laser power yields different amounts of surface damage depending on whether the exposed surface is the Te terminating face or the Cd terminating face of CZT.

  2. 3D Color Digital Elevation Map of AFM Sample

    NASA Technical Reports Server (NTRS)

    2008-01-01

    This color image is a three dimensional (3D) view of a digital elevation map of a sample collected by NASA's Phoenix Mars Lander's Atomic Force Microscope (AFM).

    The image shows four round pits, only 5 microns in depth, that were micromachined into the silicon substrate, which is the background plane shown in red. This image has been processed to reflect the levelness of the substrate.

    A Martian particle only one micrometer, or one millionth of a meter, across is held in the upper left pit.

    The rounded particle shown at the highest magnification ever seen from another world is a particle of the dust that cloaks Mars. Such dust particles color the Martian sky pink, feed storms that regularly envelop the planet and produce Mars' distinctive red soil.

    The particle was part of a sample informally called 'Sorceress' delivered to the AFM on the 38th Martian day, or sol, of the mission (July 2, 2008). The AFM is part of Phoenix's microscopic station called MECA, or the Microscopy, Electrochemistry, and Conductivity Analyzer.

    The AFM was developed by a Swiss-led consortium, with Imperial College London producing the silicon substrate that holds sampled particles.

    The Phoenix Mission is led by the University of Arizona, Tucson, on behalf of NASA. Project management of the mission is by NASA's Jet Propulsion Laboratory, Pasadena, Calif. Spacecraft development is by Lockheed Martin Space Systems, Denver.

  3. Structural investigations on native collagen type I fibrils using AFM

    SciTech Connect

    Strasser, Stefan; Zink, Albert; Janko, Marek; Heckl, Wolfgang M.; Thalhammer, Stefan . E-mail: stefan.thalhammer@gsf.de

    2007-03-02

    This study was carried out to determine the elastic properties of single collagen type I fibrils with the use of atomic force microscopy (AFM). Native collagen fibrils were formed by self-assembly in vitro characterized with the AFM. To confirm the inner assembly of the collagen fibrils, the AFM was used as a microdissection tool. Native collagen type I fibrils were dissected and the inner core uncovered. To determine the elastic properties of collagen fibrils the tip of the AFM was used as a nanoindentor by recording force-displacement curves. Measurements were done on the outer shell and in the core of the fibril. The structural investigations revealed the banding of the shell also in the core of native collagen fibrils. Nanoindentation experiments showed the same Young's modulus on the shell as well as in the core of the investigated native collagen fibrils. In addition, the measurements indicate a higher adhesion in the core of the collagen fibrils compared to the shell.

  4. Introduction to Atomic Force Microscopy (AFM) in Biology.

    PubMed

    Kreplak, Laurent

    2016-01-01

    The atomic force microscope (AFM) has the unique capability of imaging biological samples with molecular resolution in buffer solution over a wide range of time scales from milliseconds to hours. In addition to providing topographical images of surfaces with nanometer- to angstrom-scale resolution, forces between single molecules and mechanical properties of biological samples can be investigated from the nano-scale to the micro-scale. Importantly, the measurements are made in buffer solutions, allowing biological samples to "stay alive" within a physiological-like environment while temporal changes in structure are measured-e.g., before and after addition of chemical reagents. These qualities distinguish AFM from conventional imaging techniques of comparable resolution, e.g., electron microscopy (EM). This unit provides an introduction to AFM on biological systems and describes specific examples of AFM on proteins, cells, and tissues. The physical principles of the technique and methodological aspects of its practical use and applications are also described. © 2016 by John Wiley & Sons, Inc. PMID:27479503

  5. Microglial Janus kinase/signal transduction and activator of transcription 3 pathway activity directly impacts astrocyte and spinal neuron characteristics.

    PubMed

    Molet, Jenny; Mauborgne, Annie; Diallo, Mickael; Armand, Vincent; Geny, David; Villanueva, Luis; Boucher, Yves; Pohl, Michel

    2016-01-01

    After peripheral nerve injury microglial reactivity change in the spinal cord is associated with an early activation of Janus kinase (JAK)/STAT3 transduction pathway whose blockade attenuates local inflammation and pain hypersensitivity. However, the consequences of microglial JAK/STAT3-mediated signaling on neighboring cells are unknown. Using an in vitro paradigm we assessed the impact of microglial JAK/STAT3 activity on functional characteristics of astrocytes and spinal cord neurons. Purified rat primary microglia was stimulated with JAK/STAT3 classical activator interleukin-6 in the presence or absence of a selective STAT3 inhibitor and rat primary astrocytes or spinal cord neurons were exposed to microglia conditioned media (CM). JAK/STAT3 activity-generated microglial CM modulated both astrocyte and neuron characteristics. Beyond inducing mRNA expression changes in various targets of interest in astrocytes and neurons, microglia CM activated c-Jun N-terminal kinase, STAT3 and NF-κB intracellular pathways in astrocytes and promoted their proliferation. Without modifying neuronal excitability or survival, CM affected the nerve processes morphology and distribution of the post-synaptic density protein 95, a marker of glutamatergic synaptic contacts. These findings show that JAK/STAT3 activity in microglia impacts the functional characteristics of astrocytes and neurons. This suggests its participation in spinal cord tissue plasticity and remodeling occurring after peripheral nerve injury. We show that the activity of JAK/STAT3 pathway in microglial cells confers them a specific signaling modality toward neighboring cells, promoting astrocyte proliferation and changes in neuronal morphology. These in vitro data suggest that the early JAK/STAT3 activation in spinal cord microglia, associated with peripheral nerve injury, participates in functional alteration of various cell populations and in spinal tissue remodeling. PMID:26440453

  6. Angiosperms Are Unique among Land Plant Lineages in the Occurrence of Key Genes in the RNA-Directed DNA Methylation (RdDM) Pathway

    PubMed Central

    Ma, Lu; Hatlen, Andrea; Kelly, Laura J.; Becher, Hannes; Wang, Wencai; Kovarik, Ales; Leitch, Ilia J.; Leitch, Andrew R.

    2015-01-01

    The RNA-directed DNA methylation (RdDM) pathway can be divided into three phases: 1) small interfering RNA biogenesis, 2) de novo methylation, and 3) chromatin modification. To determine the degree of conservation of this pathway we searched for key genes among land plants. We used OrthoMCL and the OrthoMCL Viridiplantae database to analyze proteomes of species in bryophytes, lycophytes, monilophytes, gymnosperms, and angiosperms. We also analyzed small RNA size categories and, in two gymnosperms, cytosine methylation in ribosomal DNA. Six proteins were restricted to angiosperms, these being NRPD4/NRPE4, RDM1, DMS3 (defective in meristem silencing 3), SHH1 (SAWADEE homeodomain homolog 1), KTF1, and SUVR2, although we failed to find the latter three proteins in Fritillaria persica, a species with a giant genome. Small RNAs of 24 nt in length were abundant only in angiosperms. Phylogenetic analyses of Dicer-like (DCL) proteins showed that DCL2 was restricted to seed plants, although it was absent in Gnetum gnemon and Welwitschia mirabilis. The data suggest that phases (1) and (2) of the RdDM pathway, described for model angiosperms, evolved with angiosperms. The absence of some features of RdDM in F. persica may be associated with its large genome. Phase (3) is probably the most conserved part of the pathway across land plants. DCL2, involved in virus defense and interaction with the canonical RdDM pathway to facilitate methylation of CHH, is absent outside seed plants. Its absence in G. gnemon, and W. mirabilis coupled with distinctive patterns of CHH methylation, suggest a secondary loss of DCL2 following the divergence of Gnetales. PMID:26338185

  7. Profiling the HER3/PI3K Pathway in Breast Tumors Using Proximity-Directed Assays Identifies Correlations between Protein Complexes and Phosphoproteins

    PubMed Central

    Mukherjee, Ali; Badal, Youssouf; Nguyen, Xuan-Thao; Miller, Johanna; Chenna, Ahmed; Tahir, Hasan; Newton, Alicia; Parry, Gordon; Williams, Stephen

    2011-01-01

    Background The identification of patients for targeted antineoplastic therapies requires accurate measurement of therapeutic targets and associated signaling complexes. HER3 signaling through heterodimerization is an important growth-promoting mechanism in several tumor types and may be a principal resistance mechanism by which EGFR and HER2 expressing tumors elude targeted therapies. Current methods that can study these interactions are inadequate for formalin-fixed, paraffin-embedded (FFPE) tumor samples. Methodology and Principal Findings Herein, we describe a panel of proximity-directed assays capable of measuring protein-interactions and phosphorylation in FFPE samples in the HER3/PI3K/Akt pathway and examine the capability of these assays to inform on the functional state of the pathway. We used FFPE breast cancer cell line and tumor models for this study. In breast cancer cell lines we observe both ligand-dependent and independent activation of the pathway and strong correlations between measured activation of key analytes. When selected cell lines are treated with HER2 inhibitors, we not only observe the expected molecular effects based on mechanism of action knowledge, but also novel effects of HER2 inhibition on key targets in the HER receptor pathway. Significantly, in a xenograft model of delayed tumor fixation, HER3 phosphorylation is unstable, while alternate measures of pathway activation, such as formation of the HER3PI3K complex is preserved. Measurements in breast tumor samples showed correlations between HER3 phosphorylation and receptor interactions, obviating the need to use phosphorylation as a surrogate for HER3 activation. Significance This assay system is capable of quantitatively measuring therapeutically relevant responses and enables molecular profiling of receptor networks in both preclinical and tumor models. PMID:21297994

  8. Angiosperms Are Unique among Land Plant Lineages in the Occurrence of Key Genes in the RNA-Directed DNA Methylation (RdDM) Pathway.

    PubMed

    Ma, Lu; Hatlen, Andrea; Kelly, Laura J; Becher, Hannes; Wang, Wencai; Kovarik, Ales; Leitch, Ilia J; Leitch, Andrew R

    2015-09-01

    The RNA-directed DNA methylation (RdDM) pathway can be divided into three phases: 1) small interfering RNA biogenesis, 2) de novo methylation, and 3) chromatin modification. To determine the degree of conservation of this pathway we searched for key genes among land plants. We used OrthoMCL and the OrthoMCL Viridiplantae database to analyze proteomes of species in bryophytes, lycophytes, monilophytes, gymnosperms, and angiosperms. We also analyzed small RNA size categories and, in two gymnosperms, cytosine methylation in ribosomal DNA. Six proteins were restricted to angiosperms, these being NRPD4/NRPE4, RDM1, DMS3 (defective in meristem silencing 3), SHH1 (SAWADEE homeodomain homolog 1), KTF1, and SUVR2, although we failed to find the latter three proteins in Fritillaria persica, a species with a giant genome. Small RNAs of 24 nt in length were abundant only in angiosperms. Phylogenetic analyses of Dicer-like (DCL) proteins showed that DCL2 was restricted to seed plants, although it was absent in Gnetum gnemon and Welwitschia mirabilis. The data suggest that phases (1) and (2) of the RdDM pathway, described for model angiosperms, evolved with angiosperms. The absence of some features of RdDM in F. persica may be associated with its large genome. Phase (3) is probably the most conserved part of the pathway across land plants. DCL2, involved in virus defense and interaction with the canonical RdDM pathway to facilitate methylation of CHH, is absent outside seed plants. Its absence in G. gnemon, and W. mirabilis coupled with distinctive patterns of CHH methylation, suggest a secondary loss of DCL2 following the divergence of Gnetales. PMID:26338185

  9. Wnt/β-Catenin Signaling Pathway Is a Direct Enhancer of Thyroid Transcription Factor-1 in Human Papillary Thyroid Carcinoma Cells

    PubMed Central

    Gilbert-Sirieix, Marie; Makoukji, Joelle; Kimura, Shioko; Talbot, Monique; Caillou, Bernard; Massaad, Charbel; Massaad-Massade, Liliane

    2011-01-01

    The Wnt/β-catenin signaling pathway is involved in the normal development of thyroid gland, but its disregulation provokes the appearance of several types of cancers, including papillary thyroid carcinomas (PTC) which are the most common thyroid tumours. The follow-up of PTC patients is based on the monitoring of serum thyroglobulin levels which is regulated by the thyroid transcription factor 1 (TTF-1): a tissue-specific transcription factor essential for the differentiation of the thyroid. We investigated whether the Wnt/β-catenin pathway might regulate TTF-1 expression in a human PTC model and examined the molecular mechanisms underlying this regulation. Immunofluorescence analysis, real time RT-PCR and Western blot studies revealed that TTF-1 as well as the major Wnt pathway components are co-expressed in TPC-1 cells and human PTC tumours. Knocking-down the Wnt/β-catenin components by siRNAs inhibited both TTF-1 transcript and protein expression, while mimicking the activation of Wnt signaling by lithium chloride induced TTF-1 gene and protein expression. Functional promoter studies and ChIP analysis showed that the Wnt/β-catenin pathway exerts its effect by means of the binding of β-catenin to TCF/LEF transcription factors on the level of an active TCF/LEF response element at [−798, −792 bp] in TTF-1 promoter. In conclusion, we demonstrated that the Wnt/β-catenin pathway is a direct and forward driver of the TTF-1 expression. The localization of TCF-4 and TTF-1 in the same area of PTC tissues might be of clinical relevance, and justifies further examination of these factors in the papillary thyroid cancers follow-up. PMID:21814573

  10. Novel Polymer Linkers for Single Molecule AFM Force Spectroscopy

    PubMed Central

    Tong, Zenghan; Mikheikin, Andrey; Krasnoslobodtsev, Alexey; Lv, Zhengjian; Lyubchenko, Yuri L.

    2013-01-01

    Flexible polymer linkers play an important role in various imaging and probing techniques that require surface immobilization, including atomic force microscopy (AFM). In AFM force spectroscopy, polymer linkers are necessary for the covalent attachment of molecules of interest to the AFM tip and the surface. The polymer linkers tether the molecules and provide their proper orientation in probing experiments. Additionally, the linkers separate specific interactions from nonspecific short-range adhesion and serve as a reference point for the quantitative analysis of single molecule probing events. In this report, we present our results on the synthesis and testing of a novel polymer linker and the identification of a number of potential applications for its use in AFM force spectroscopy experiments. The synthesis of the linker is based on the well-developed phosphoramidate (PA) chemistry that allows the routine synthesis of linkers with predetermined lengths and PA composition. These linkers are homogeneous in length and can be terminated with various functional groups. PA linkers with different functional groups were synthesized and tested in experimental systems utilizing different immobilization chemistries. We probed interactions between complementary DNA oligonucleotides; DNA and protein complexes formed by the site-specific binding protein SfiI; and interactions between amyloid peptide (Aβ42). The results of the AFM force spectroscopy experiments validated the feasibility of the proposed approach for the linker design and synthesis. Furthermore, the properties of the tether (length, functional groups) can be adjusted to meet the specific requirements for different force spectroscopy experiments and system characteristics, suggesting that it could be used for a large number of various applications. PMID:23624104

  11. High-Speed AFM Images of Thermal Motion Provide Stiffness Map of Interfacial Membrane Protein Moieties

    PubMed Central

    2014-01-01

    The flexibilities of extracellular loops determine ligand binding and activation of membrane receptors. Arising from fluctuations in inter- and intraproteinaceous interactions, flexibility manifests in thermal motion. Here we demonstrate that quantitative flexibility values can be extracted from directly imaging the thermal motion of membrane protein moieties using high-speed atomic force microscopy (HS-AFM). Stiffness maps of the main periplasmic loops of single reconstituted water channels (AqpZ, GlpF) revealed the spatial and temporal organization of loop-stabilizing intraproteinaceous H-bonds and salt bridges. PMID:25516527

  12. Distinct Argonaute-mediated 22G-RNA pathways direct genome surveillance in the C. elegans germline

    PubMed Central

    Gu, Weifeng; Shirayama, Masaki; Conte, Darryl; Vasale, Jessica; Batista, Pedro J.; Claycomb, Julie M.; Moresco, James J.; Youngman, Elaine; Keys, Jennifer; Stoltz, Matthew J.; Chen, Chun-Chieh G.; Chaves, Daniel A.; Duan, Shenghua E.; Kasschau, Kristen D.; Falgren, Noah; Yates, John R.; Mitani, Shohei; Carrington, James C.; Mello, Craig C.

    2009-01-01

    Summary Endogenous small RNAs (endo-siRNAs) interact with Argonaute (AGO) proteins to mediate sequence-specific regulation of diverse biological processes. Here, we combine deep-sequencing and genetic approaches to explore the biogenesis and function of endo-siRNAs in C. elegans. We describe conditional alleles of the dicer-related helicase, drh-3, that abrogate both RNA interference and the biogenesis of endo-siRNAs, called 22G-RNAs. DRH-3 is a core component of RNA-dependent RNA polymerase (RdRP) complexes essential for several distinct 22G-RNA systems. We show that in the germ-line, one system is dependent on worm-specific AGOs, including WAGO-1, which localizes to germ-line nuage structures called P-granules. WAGO-1 silences certain genes, transposons, pseudogenes and cryptic loci. Finally, we demonstrate that components of the nonsense-mediated decay pathway function in at least one WAGO-mediated surveillance pathway. These findings broaden our understanding of the biogenesis and diversity of 22G-RNAs and suggest novel regulatory functions for small RNAs. PMID:19800275

  13. Nonclassical antigen-processing pathways are required for MHC class II-restricted direct tumor recognition by NY-ESO-1-specific CD4(+) T cells.

    PubMed

    Matsuzaki, Junko; Tsuji, Takemasa; Luescher, Immanuel; Old, Lloyd J; Shrikant, Protul; Gnjatic, Sacha; Odunsi, Kunle

    2014-04-01

    Tumor antigen-specific CD4(+) T cells that directly recognize cancer cells are important for orchestrating antitumor immune responses at the local tumor sites. However, the mechanisms of direct MHC class II (MHC-II) presentation of intracellular tumor antigen by cancer cells are poorly understood. We found that two functionally distinct subsets of CD4(+) T cells were expanded after HLA-DPB1*04 (DP04)-binding NY-ESO-1157-170 peptide vaccination in patients with ovarian cancer. Although both subsets recognized exogenous NY-ESO-1 protein pulsed on DP04(+) target cells, only one type recognized target cells with intracellular expression of NY-ESO-1. The tumor-recognizing CD4(+) T cells more efficiently recognized the short 8-9-mer peptides than the non-tumor-recognizing CD4(+) T cells. In addition to endosomal/lysosomal proteases that are typically involved in MHC-II antigen presentation, several pathways in the MHC class I presentation pathways, such as the proteasomal degradation and transporter-associated with antigen-processing-mediated peptide transport, were also involved in the presentation of intracellular NY-ESO-1 on MHC-II. The presentation was inhibited significantly by primaquine, a small molecule that inhibits endosomal recycling, consistent with findings that pharmacologic inhibition of new protein synthesis enhances antigen presentation. Together, our data demonstrate that cancer cells selectively present peptides from intracellular tumor antigens on MHC-II by multiple nonclassical antigen-processing pathways. Harnessing the direct tumor-recognizing ability of CD4(+) T cells could be a promising strategy to enhance antitumor immune responses in the immunosuppressive tumor microenvironment. PMID:24764581

  14. Non-classical antigen processing pathways are required for MHC class II-restricted direct tumor recognition by NY-ESO-1-specific CD4+ T cells

    PubMed Central

    Matsuzaki, Junko; Tsuji, Takemasa; Luescher, Immanuel; Old, Lloyd J.; Shrikant, Protul; Gnjatic, Sacha; Odunsi, Kunle

    2014-01-01

    Tumor antigen-specific CD4+ T cells that directly recognize cancer cells are important for orchestrating antitumor immune responses at the local tumor sites. However, the mechanisms of direct MHC class II (MHC-II) presentation of intracellular tumor antigen by cancer cells are poorly understood. We found that two functionally distinct subsets of CD4+ T cells were expanded after HLA-DPB1*04 (DP04)-binding NY-ESO-1157–170 peptide vaccination in ovarian cancer patients. While both subsets similarly recognized exogenous NY-ESO-1 protein pulsed on DP04+ target cells, only one type recognized target cells with intracellular expression of NY-ESO-1. The tumor-recognizing CD4+ T cells more efficiently recognized the short 8–9-mer peptides than the non-tumor-recognizing CD4+ T cells. In addition to endosomal/lysosomal proteases that are typically involved in MHC-II antigen presentation, several pathways in the MHC class I presentation pathways such as the proteasomal degradation and transporter-associated with antigen-processing (TAP)-mediated peptide transport were also involved in the presentation of intracellular NY-ESO-1 on MHC-II. The presentation was inhibited significantly by primaquine, a small molecule that inhibits endosomal recycling, consistent with findings that pharmacological inhibition of new protein synthesis enhances antigen presentation. Together, our data demonstrated that cancer cells selectively present peptides from intracellular tumor antigens on MHC-II by multiple non-classical antigen-processing pathways. Harnessing direct tumor-recognizing ability of CD4+ T cells could be a promising strategy to enhance antitumor immune responses in the immunosuppressive tumor microenvironment. PMID:24764581

  15. Possible enhancements of AFM spin-fluctuations in high-TC cuprates

    NASA Astrophysics Data System (ADS)

    Jarlborg, Thomas

    2009-03-01

    Ab-initio band calculations for high-TC cuprates, together with modelling based of a free electron like band, show a strong interaction between anti-ferromagnetic (AFM) spin waves and periodic lattice distortions as for phonons, even though this type of spin-phonon coupling (SPC) is underestimated in calculations using the local density approximation. The SPC has a direct influence on the properties of the HTC cuprates and it can explain many observations. The strongest effects are seen for modulated waves in the CuO bond direction, and a band gap is formed near the X,Y points, but unusal band dispersion (like ``waterfalls'') might also be induced below the Fermi energy (EF) in the diagonal direction. The band results are used to propose different ways of increasing AFM spin-fluctuations locally, and to have a higher density-of-states (DOS) at EF. Static potential modulations, via periodic distribution of dopants or lattice distortions, can be tuned to increase the DOS. This opens for possibilities to enhance coupling for spin fluctuations (λsf) and superconductivity. The exchange enhancement is in general increased near a surface, which suggests a tendency towards static spin configurations. The sensivity of the band results to corrections of the local density potential are discussed.

  16. Direct and indirect pathways to lamina I in the medulla oblongata and spinal cord of the cat

    NASA Technical Reports Server (NTRS)

    Holstege, Gert

    1988-01-01

    The pathways to lamina I in the medulla oblongata and spinal cord of the cat were traced using horse-radish-peroxidase (HRP) and autoradiographic techniques. The HRP results indicated that several neuronal cell groups in the brain stem and hypothalamus project to the spinal cord throughout its total length. The autoradiographic tracing results demonstrated that the strongest projections to lamina I are derived from the following four areas: the caudal nucleus raphe magnus (NRM), the ventral part of the caudal pontine and NRM, the contralaterally projecting lateral pontine or paralemniscal tegmentum, and the paraventricular nucleus of the hypothalamus. In addition, a limited, especially at lumbosacral levels, distinct projection to lamina I was found to originate in the most caudal part of the medullary tegmentum.

  17. Thaliporphine Preserves Cardiac Function of Endotoxemic Rabbits by Both Directly and Indirectly Attenuating NFκB Signaling Pathway

    PubMed Central

    Lee, A. S.; Chen, W. P.; Lee, S. S.; Su, M. J.

    2012-01-01

    Cardiac depression in sepsis is associated with the increased morbidity and mortality. Although myofilaments damage, autonomic dysfunction, and apoptosis play roles in sepsis-induced myocardial dysfunction, the underlying mechanism is not clear. All of these possible factors are related to NFκB signaling, which plays the main role in sepsis signaling. Thaliporphine was determined to possess anti-inflammatory and cardioprotective activity by suppressing NFκB signaling in rodents. The purpose of this study is to further prove this protective effect in larger septic animals, and try to find the underlying mechanisms. The systolic and diastolic functions were evaluated in vivo by pressure-volume analysis at different preloads. Both preload-dependent and -independent hemodynamic parameters were performed. Inflammatory factors of whole blood and serum samples were analyzed. Several sepsis-related signaling pathways were also determined at protein level. Changes detected by conductance catheter showed Thaliporphine could recover impaired left ventricular systolic function after 4 hours LPS injection. It could also reverse the LPS induced steeper EDPVR and gentler ESPVR, thus improve Ees, Ea, and PRSW. Thaliporphine may exert this protective effect by decreasing TNFα and caspase3 dependent cell apoptosis, which was consistent with the decreased serum cTnI and LDH concentration. Thaliporphine could protect sepsis-associated myocardial dysfunction in both preload-dependent and -independent ways. It may exert these protective effects by both increase of “good”-PI3K/Akt/mTOR and decrease of “bad”-p38/NFκB pathways, which followed by diminishing TNFα and caspase3 dependent cell apoptosis. PMID:22761733

  18. Quality Child Care Supports the Achievement of Low-Income Children: Direct and Indirect Pathways Through Caregiving and the Home Environment

    PubMed Central

    McCartney, Kathleen; Dearing, Eric; Taylor, Beck A.; Bub, Kristen L.

    2009-01-01

    Existing studies of child care have not been able to determine whether higher quality child care protects children from the effects of poverty, whether poverty and lower quality child care operate as dual risk factors, or whether both are true. The objective of the current study was to test two pathways through which child care may serve as a naturally occurring intervention for low-income children: a direct pathway through child care quality to child outcomes, and an indirect pathway through improvements in the home environment. Children were observed in their homes and child care settings at 6, 15, 24, and 36 months. An interaction between family income-to-needs ratio and child care quality predicted School Readiness, Receptive Language, and Expressive Language, as well as improvements in the home environment. Children from low-income families profited from observed learning supports in the form of sensitive care and stimulation of cognitive development, and their parents profited from unobserved informal and formal parent supports. Policy implications are discussed. PMID:19578561

  19. High aspect ratio AFM Probe processing by helium-ion-beam induced deposition.

    PubMed

    Onishi, Keiko; Guo, Hongxuan; Nagano, Syoko; Fujita, Daisuke

    2014-11-01

    A Scanning Helium Ion Microscope (SHIM) is a high resolution surface observation instrument similar to a Scanning Electron Microscope (SEM) since both instruments employ finely focused particle beams of ions or electrons [1]. The apparent difference is that SHIMs can be used not only for a sub-nanometer scale resolution microscopic research, but also for the applications of very fine fabrication and direct lithography of surfaces at the nanoscale dimensions. On the other hand, atomic force microscope (AFM) is another type of high resolution microscopy which can measure a three-dimensional surface morphology by tracing a fine probe with a sharp tip apex on a specimen's surface.In order to measure highly uneven and concavo-convex surfaces by AFM, the probe of a high aspect ratio with a sharp tip is much more necessary than the probe of a general quadrangular pyramid shape. In this paper we report the manufacture of the probe tip of the high aspect ratio by ion-beam induced gas deposition using a nanoscale helium ion beam of SHIM.Gas of platinum organic compound was injected into the sample surface neighborhood in the vacuum chamber of SHIM. The decomposition of the gas and the precipitation of the involved metal brought up a platinum nano-object in a pillar shape on the normal commercial AFM probe tip. A SHIM system (Carl Zeiss, Orion Plus) equipped with the gas injection system (OmniProbe, OmniGIS) was used for the research. While the vacuum being kept to work, we injected platinum organic compound ((CH3)3(CH3C5H4)Pt) into the sample neighborhood and irradiated the helium ion beam with the shape of a point on the apex of the AFM probe tip. It is found that we can control the length of the Pt nano-pillar by irradiation time of the helium ion beam. The AFM probe which brought up a Pt nano-pillar is shown in Figure 1. It is revealed that a high-aspect-ratio Pt nano-pillar of ∼40nm diameter and up to ∼2000 nm length can be grown. In addition, for possible heating

  20. PI3 kinase directly phosphorylates Akt1/2 at Ser473/474 in the insulin signal transduction pathway

    PubMed Central

    Tsuchiya, A; Kanno, T; Nishizaki, T

    2014-01-01

    Insulin stimulated translocation of the glucose transporter GLUT4 from the cytosol to the plasma membrane in a concentration (1 nM–1 μM)-dependent manner and increased glucose uptake in 3T3-L1 adipocytes. Insulin-induced GLUT4 translocation to the cell surface was prevented by the phosphoinositide 3 kinase (PI3K) inhibitor wortmannin, the 3-phosphoinositide-dependent protein kinase 1 (PDK1) inhibitor BX912 or the Akt1/2 inhibitor MK2206, and by knocking-down PI3K, PDK1 or Akt1/2. Insulin increased phosphorylation of Akt1/2 at Thr308/309 and Ser473/474, to activate Akt1/2, in the adipocytes. Insulin-induced phosphorylation of Akt1/2 was suppressed by wortmannin and knocking-down PI3K, while no significant inhibition of the phosphorylation was obtained with BX912 or knocking-down PDK1. In the cell-free Akt assay, PI3K phosphorylated Akt1 both at Thr308 and Ser473 and Akt2 at Ser474 alone. In contrast, PDK1 phosphorylates Akt1 at Thr308 and Akt2 at Thr309. The results of this study indicate that PI3K activates Akt1, independently of PDK1, and Akt2 by cooperating with PDK1 in the insulin signal transduction pathway linked to GLUT4 translocation. PMID:24169049

  1. Direct evidence of the cyclooxygenase pathway of prostaglandin synthesis in arthropods: genetic and biochemical characterization of two crustacean cyclooxygenases.

    PubMed

    Varvas, Külliki; Kurg, Reet; Hansen, Kristella; Järving, Reet; Järving, Ivar; Valmsen, Karin; Lõhelaid, Helike; Samel, Nigulas

    2009-12-01

    Prostaglandins, well-known lipid mediators in vertebrate animals, have also shown to play certain regulatory roles in insects and other arthropods acting on reproduction, immune system and ion transport. However, knowledge of their biosynthetic pathways in arthropods is lacking. In the present study, we report the cloning and expression of cyclooxygenase (COX) from amphipod crustaceans Gammarus spp and Caprella spp. The amphipod COX proteins contain key residues shown to be important for cyclooxygenase and peroxidase activities. Differently from all other known cyclooxygenases the N-terminal signal sequence of amphipod enzymes is not cleaved during protein expression in mammalian cells. The C-terminus of amphipod COX is shorter than that of mammalian isoforms and lacks the KDEL(STEL)-type endoplasmic reticulum retention/retrieval signal. Despite that, amphipod COX proteins are N-glycosylated and locate similarly to the vertebrate COX on the endoplasmic reticulum and nuclear envelope. Both amphipod COX mRNAs encode functional cyclooxygenases that catalyze the transformation of arachidonic acid into prostaglandins. Using bioinformatic analysis we identified a COX-like gene from the human body louse Pediculus humanus corporis genome that encodes a protein with about 30% sequence identity with human COX-1 and COX-2. Although the COX gene is known to be absent from genomes of Drosophila sp., Aedes aegypti, Bombyx mori, and other insects, our studies establish the existence of the COX gene in certain lineages within the insect world. PMID:19854273

  2. Hydrodynamic effects of the tip movement on surface nanobubbles: a combined tapping mode, lift mode and force volume mode AFM study.

    PubMed

    Walczyk, Wiktoria; Hain, Nicole; Schönherr, Holger

    2014-08-28

    We report on an Atomic Force Microscopy (AFM) study of AFM tip-nanobubble interactions in experiments conducted on argon surface nanobubbles on HOPG (highly oriented pyrolytic graphite) in water in tapping mode, lift mode and Force Volume (FV) mode AFM. By subsequent data acquisition on the same nanobubbles in these three different AFM modes, we could directly compare the effect of different tip-sample interactions. The tip-bubble interaction strength was found to depend on the vertical and horizontal position of the tip on the bubble with respect to the bubble center. The interaction forces measured experimentally were in good agreement with the forces calculated using the dynamic interaction model. The strength of the hydrodynamic effect was also found to depend on the direction of the tip movement. It was more pronounced in the FV mode, in which the tip approaches the bubble from the top, than in the lift mode, in which the tip approaches the bubble from the side. This result suggests that the direction of tip movement influences the bubble deformation. The effect should be taken into account when nanobubbles are analysed by AFM in various scanning modes. PMID:24988375

  3. AFM-based mechanical characterization of single nanofibres.

    PubMed

    Neugirg, Benedikt R; Koebley, Sean R; Schniepp, Hannes C; Fery, Andreas

    2016-04-28

    Nanofibres are found in a broad variety of hierarchical biological systems as fundamental structural units, and nanofibrillar components are playing an increasing role in the development of advanced functional materials. Accurate determination of the mechanical properties of single nanofibres is thus of great interest, yet measurement of these properties is challenging due to the intricate specimen handling and the exceptional force and deformation resolution that is required. The atomic force microscope (AFM) has emerged as an effective, reliable tool in the investigation of nanofibrillar mechanics, with the three most popular approaches-AFM-based tensile testing, three-point deformation testing, and nanoindentation-proving preferable to conventional tensile testing in many (but not all) cases. Here, we review the capabilities and limitations of each of these methods and give a comprehensive overview of the recent advances in this field. PMID:27055900

  4. Nano-Bio-Mechanics of Neuroblastoma Cells Using AFM

    NASA Astrophysics Data System (ADS)

    Bastatas, Lyndon; Matthews, James; Kang, Min; Park, Soyeun

    2011-10-01

    We have conducted an in vitro study to determine the elastic moduli of neurobalstoma cell lines using atomic force microscopy. Using a panel of cell lines established from neuroblastoma patients at different stages of disease progress and treatment, we have investigated the differences in elastic moduli during a course of cancer progression and chemotherapy. The cells were grown on the hard substrates that are chemically functionalized to enhance adhesion. We have performed the AFM indentation experiments with different applied forces from the AFM probe. For the purpose of the comparison between cell lines, the indentations were performed only on cell centers. The obtained force-distance curves were analyzed using the Hertz model in order to extract the elastic moduli. We have found that the elastic moduli of human neuroblastoma cells significantly varied during the disease progression. We postulate that the observed difference might be affected by the treatment and chemotherapy.

  5. Wedged AFM-cantilevers for parallel plate cell mechanics.

    PubMed

    Stewart, Martin P; Hodel, Adrian W; Spielhofer, Andreas; Cattin, Cedric J; Müller, Daniel J; Helenius, Jonne

    2013-04-01

    The combination of atomic force microscopy (AFM) and optical microscopy has gained popularity for mechanical analysis of living cells. In particular, recent AFM-based assays featuring tipless cantilevers and whole-cell deformation have yielded insights into cellular function, structure, and dynamics. However, in these assays the standard ≈10° tilt of the cantilever prevents uniaxial loading, which complicates assessment of cellular geometry and can cause cell sliding or loss of loosely adherent cells. Here, we describe an approach to modify tipless cantilevers with wedges and, thereby, achieve proper parallel plate mechanics. We provide guidance on material selection, the wedge production process, property and geometry assessment, and the calibration of wedged cantilevers. Furthermore, we demonstrate their ability to simplify the assessment of cell shape, prevent lateral displacement of round cells during compression, and improve the assessment of cell mechanical properties. PMID:23473778

  6. BOREAS AFM-5 Level-1 Upper Air Network Data

    NASA Technical Reports Server (NTRS)

    Barr, Alan; Hrynkiw, Charmaine; Newcomer, Jeffrey A. (Editor); Hall, Forrest G. (Editor); Smith, David E. (Technical Monitor)

    2000-01-01

    The Boreal Ecosystem-Atmosphere Study (BOREAS) Airborne Fluxes and Meteorology (AFM)-5 team collected and processed data from the numerous radiosonde flights during the project. The goals of the AFM-05 team were to provide large-scale definition of the atmosphere by supplementing the existing Atmospheric Environment Service (AES) aerological network, both temporally and spatially. This data set includes basic upper-air parameters collected from the network of upper-air stations during the 1993, 1994, and 1996 field campaigns over the entire study region. The data are contained in tabular ASCII files. The level-1 upper-air network data are available from the Earth Observing System Data and Information System (EOSDIS) Oak Ridge National Laboratory (ORNL) Distributed Active Archive Center (DAAC). The data files also are available on a CD-ROM (see document number 20010000884).

  7. Molecular modeling of enzyme attachment on AFM probes.

    PubMed

    Oliveira, Guedmiller S; Leite, Fabio L; Amarante, Adriano M; Franca, Eduardo F; Cunha, Richard A; Briggs, James M; Freitas, Luiz C G

    2013-09-01

    The immobilization of enzymes on atomic force microscope tip (AFM tip) surface is a crucial step in the development of nanobiosensors to be used in detection process. In this work, an atomistic modeling of the attachment of the acetyl coenzyme A carboxylase (ACC enzyme) on a functionalized AFM tip surface is proposed. Using electrostatic considerations, suitable enzyme-surface orientations with the active sites of the ACC enzyme available for interactions with bulk molecules were found. A 50 ns molecular dynamics trajectory in aqueous solution was obtained and surface contact area, hydrogen bonding and protein stability were analyzed. The enzyme-surface model proposed here with minor adjustment can be applied to study antigen-antibody interactions as well as enzyme immobilization on silica for chromatography applications. PMID:24029365

  8. AFM study of polymer lubricants on hard disk surfaces

    NASA Astrophysics Data System (ADS)

    Bao, G. W.; Troemel, M.; Li, S. F. Y.

    Thin liquid films of PFPE (perfluoropolyether) lubricants dip-coated on hard disk surfaces were imaged with non-contact mode AFM. Demnum lubricants with phosphazene additives exhibited strong interactions with a silicon tip due to the formation of liquid bridges between the lubricants and the tip, as indicated by a remarkable hysteresis loop between approach and retraction curves in force vs. distance measurements. Features resulting from capillary forces due to tip tapping to the lubricants were revealed, which demonstrated that the capillary forces could be used to lock the non-contacting tip at a certain separation from the substrate surface to obtain AFM images. Force vs. distance curves for Fomblin Z-dol lubricants showed negligible hysteresis effects and features corresponding to lateral distortion of the tip by the lubricants only were observed. In both cases, only when the tip was positioned far above the surfaces could the natural distributions of the lubricants be imaged.

  9. AFM-based mechanical characterization of single nanofibres

    NASA Astrophysics Data System (ADS)

    Neugirg, Benedikt R.; Koebley, Sean R.; Schniepp, Hannes C.; Fery, Andreas

    2016-04-01

    Nanofibres are found in a broad variety of hierarchical biological systems as fundamental structural units, and nanofibrillar components are playing an increasing role in the development of advanced functional materials. Accurate determination of the mechanical properties of single nanofibres is thus of great interest, yet measurement of these properties is challenging due to the intricate specimen handling and the exceptional force and deformation resolution that is required. The atomic force microscope (AFM) has emerged as an effective, reliable tool in the investigation of nanofibrillar mechanics, with the three most popular approaches--AFM-based tensile testing, three-point deformation testing, and nanoindentation--proving preferable to conventional tensile testing in many (but not all) cases. Here, we review the capabilities and limitations of each of these methods and give a comprehensive overview of the recent advances in this field.

  10. PbSe nanocrystal growth as nanocubes and nanorods on peptide nanotubes via different directed-assembly pathways

    NASA Astrophysics Data System (ADS)

    Shi, Menglu; Su, Wei; Matsui, Hiroshi

    2010-11-01

    Pb-binding TAR-1 peptides (Ile-Ser-Leu-Leu-His-Ser-Thr) were covalently conjugated on a bolaamphiphile peptide nanotube substrate and the precursors of PbSe were incubated at room temperature. This resulted in the growth of highly crystalline PbSe nanocubes on this biomimetic cylindrical substrate. The growth mechanism to generate nanocubes occurs via the directed self-assembly of nanoparticles and then nanoparticle fusion. The peptide conformation and the cylindrical peptide nanotube substrate play important roles in the mesoscopic crystallization of PbSe nanocubes. Changing the buffer for the peptide immobilization process from 2-(N-morpholino)ethanesulfonic acid to phosphate induces a transformation in the nanocrystal shape from nanocube to nanorods. The conformational change of the TAR-1 peptide on the nanotubes due to the change in the buffer seems to be responsible for aggregating intermediate nanoparticles in different directions for the directed fusion and mesoscopic crystallization of PbSe into the different shapes.