Modulated extrusion for textured 3D printing

This research utilises a Fused Deposition Modelling 3D Printer to investigate the aesthetics of 3D printing and it's broader applications. The presented research re-evaluates the 3D printer as a tool to manipulate materials, as opposed to a machine that discretely reproduces digital models at a fine resolution. The research questions the utility of automation, and attempts to find a level that permits materially expressive modes of fabrication. The exploration of aesthetics has uncovered a variety of unexpected textures and interesting material properties that may have wider use. For instance, rigid plastic has been extruded and manipulated finer than the extrusion nozzle diameter, which confers flexibility and fabric like qualities to the printed object. The discovered techniques for 3D printed aesthetics are reproducibly reliable and can be incorporated back into orthodox digital-model driven fabrication

Studies on the enzyme glucose oxidase from Penicillium amagasakiense

A short method of purification of P.amagasakiense glucose oxidase has been achieved. Attempts to prepare the apoenzyme were unsuccessful. Inhibition of activity of the holoenzyme by heavy metals was investigated. It was shown that inhibition was due to the metal cations rather than the undissociated metal salts. The effect of bisulphite on the enzyme was shown to be the same as for Aspergillus niger glucose oxidase. From the kinetic experiments with bisulphite, an ionizable group with a pK of 4.2 was found to be involved in the reaction. This pK was assigned to the amino group of adenine of the FAD moiety of the enzyme. Experiments on the binding of halide anions to the enzyme indicated the involvement of an ionizable group with the same pK. Photo-chemical experiments on the enzyme revealed three different phenomena. (1) Photo-oxidation with methylene blue or Rose Bengal as sensitizer, destroyed an ionizable group with pK of 7.2. This was assigned to a histidine residue in the protein. (2) At high pH in the presence of EDTA and with no oxygen present photo-reduction of the enzyme spectrum was obtained. The spectrum of the oxidised enzyme was obtained when air was admitted to the reduced species. (3) At high pH in the presence of EDTA, light at 450 nm wavelength caused the enzyme to lose activity. This activity loss was irreversible with respect to oxygen or glucose. The presence of glucose in the reaction mixture during illumination protected the enzyme from this photo-destruction

Photosynthesis dependent acidification of perialgal vacuoles in theParamedum bursaria/Chlorella symbiosis. Visualization by monensin

After treatment with the carboxylic ionophore monensin theChlorella containing perialgal vacuoles of the greenParamecium bursaria swell. TheParamecium cells remain motile at this concentration for at least one day. The swelling is only observed in illuminated cells and can be inhibited by DCMU. We assume that during photosynthesis the perialgal vacuoles are acidified and that monensin exchanges H+ ions against monovalent cations (here K+). In consequence the osmotic value of the vacuoles increases. The proton gradient is believed to drive the transport of maltose from the symbiont into the host. Another but light independent effect of the monensin treatment is the swelling of peripheral alveoles of the ciliates, likewise indicating that the alveolar membrane contains an active proton pump

Annexins-a family of proteins with distinctive tastes for cell signaling and membrane dynamics

Annexins are cytosolic proteins with conserved three-dimensional structures that bind acidic phospholipids in cellular membranes at elevated Ca2+ levels. Through this they act as Ca2+-regulated membrane binding modules that organize membrane lipids, facilitating cellular membrane transport but also displaying extracellular activities. Recent discoveries highlight annexins as sensors and regulators of cellular and organismal stress, controlling inflammatory reactions in mammals, environmental stress in plants, and cellular responses to plasma membrane rupture. Here, we describe the role of annexins as Ca2+-regulated membrane binding modules that sense and respond to cellular stress and share our view on future research directions in the field.Work in our laboratories was supported by grants from the German Research Foundation to V.G. (CRC1348/A04, GE514/6-3) and U.R. (CRC1009/A06, CRC1348/A11); the Interdisciplinary Center for Clinical Research of the Münster Medical School to U.R. (Re2/022/20); the Danish Council for Independent Research (6108-00378A, 9040-00252B), Scientific Committee Danish Cancer Society (R90-A5847-14-S2, R269-A15812), and the Novo Nordisk Foundation (NNF18OC0034936) to J.N.; the Royal Society and Wolfson Foundation (RSWF\R3\183001) to F.N.E.G.; the National Institute of Health (NIH R01AR055686) to J.K.J.; the Ara Parseghian Medical Research Fund (G217137) to T.G

Development of Timd2 as a reporter gene for MRI.

PURPOSE: To assess the potential of an MRI gene reporter based on the ferritin receptor Timd2 (T-cell immunoglobulin and mucin domain containing protein 2), using T1- and T2-weighted imaging. METHODS: Pellets of cells that had been modified to express the Timd2 transgene, and incubated with either iron-loaded or manganese-loaded ferritin, were imaged using T1- and T2-weighted MRI. Mice were also implanted subcutaneously with Timd2-expressing cells and the resulting xenograft tissue imaged following intravenous injection of ferritin using T2-weighted imaging. RESULTS: Timd2-expressing cells, but not control cells, showed a large increase in both R2 and R1 in vitro following incubation with iron-loaded and manganese-loaded ferritin, respectively. Expression of Timd2 had no effect on cell viability or proliferation; however, manganese-loaded ferritin, but not iron-loaded ferritin, was toxic to Timd2-expressing cells. Timd2-expressing xenografts in vivo showed much smaller changes in R2 following injection of iron-loaded ferritin than the same cells incubated in vitro with iron-loaded ferritin. CONCLUSION: Timd2 has demonstrated potential as an MRI reporter gene, producing large increases in R2 and R1 with ferritin and manganese-loaded ferritin respectively in vitro, although more modest changes in R2 in vivo. Manganese-loaded apoferritin was not used in vivo due to the toxicity observed in vitro. Magnetic Resonance in Medicine published by Wiley Periodicals, Inc. on behalf of International Society for Magnetic Resonance.This work was supported by the Medical Research Council and Cancer Research UK (CRUK) doctoral training grants (to P.S.P.) and a CRUK Program Grant to K.M.B. T.B.R. was in receipt of Intra-European Marie Curie and Long-Term European Molecular Biology Organization fellowships.This is the final published version. It first appeared at http://onlinelibrary.wiley.com/doi/10.1002/mrm.25750/abstract

Canine respiratory coronavirus employs caveolin-1-mediated pathway for internalization to HRT-18G cells

Canine respiratory coronavirus (CRCoV), identified in 2003, is a member of the Coronaviridae family. The virus is a betacoronavirus and a close relative of human coronavirus OC43 and bovine coronavirus. Here, we examined entry of CRCoV into human rectal tumor cells (HRT-18G cell line) by analyzing co-localization of single virus particles with cellular markers in the presence or absence of chemical inhibitors of pathways potentially involved in virus entry. We also targeted these pathways using siRNA. The results show that the virus hijacks caveolin-dependent endocytosis to enter cells via endocytic internalization

Glycoprotein analysis using protein microarrays and mass spectrometry

Protein glycosylation plays an important role in a multitude of biological processes such as cell–cell recognition, growth, differentiation, and cell death. It has been shown that specific glycosylation changes are key in disease progression and can have diagnostic value for a variety of disease types such as cancer and inflammation. The complexity of carbohydrate structures and their derivatives makes their study a real challenge. Improving the isolation, separation, and characterization of carbohydrates and their glycoproteins is a subject of increasing scientific interest. With the development of new stationary phases and molecules that have affinity properties for glycoproteins, the isolation and separation of these compounds have advanced significantly. In addition to detection with mass spectrometry, the microarray platform has become an essential tool to characterize glycan structure and to study glycosylation-related biological interactions, by using probes as a means to interrogate the spotted or captured glycosylated molecules on the arrays. Furthermore, the high-throughput and reproducible nature of microarray platforms have been highlighted by its extensive applications in the field of biomarker validation, where a large number of samples must be analyzed multiple times. This review covers a brief survey of the other experimental methodologies that are currently being developed and used to study glycosylation and emphasizes methodologies that involve the use of microarray platforms. This review describes recent advances in several options of microarray platforms used in glycoprotein analysis, including glycoprotein arrays, glycan arrays, lectin arrays, and antibody/lectin arrays. The translational use of these arrays in applications related to characterization of cells and biomarker discovery is also included. © 2010 Wiley Periodicals, Inc., Mass Spec Rev 29:830–844, 2010Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/77969/1/20269_ftp.pd

Acute-Phase-HDL Remodeling by Heparan Sulfate Generates a Novel Lipoprotein with Exceptional Cholesterol Efflux Activity from Macrophages

During episodes of acute-inflammation high-density lipoproteins (HDL), the carrier of so-called good cholesterol, experiences a major change in apolipoprotein composition and becomes acute-phase HDL (AP-HDL). This altered, but physiologically important, HDL has an increased binding affinity for macrophages that is dependent on cell surface heparan sulfate (HS). While exploring the properties of AP-HDL∶HS interactions we discovered that HS caused significant remodeling of AP-HDL. The physical nature of this change in structure and its potential importance for cholesterol efflux from cholesterol-loaded macrophages was therefore investigated. In the presence of heparin, or HS, AP-HDL solutions at pH 5.2 became turbid within minutes. Analysis by centrifugation and gel electrophoresis indicated that AP-HDL was remodeled generating novel lipid poor particles composed only of apolipoprotein AI, which we designate β2. This remodeling is dependent on pH, glycosaminoglycan type, is promoted by Ca2+ and is independent of protease or lipase activity. Compared to HDL and AP-HDL, remodeled AP-HDL (S-HDL-SAA), containing β2 particles, demonstrated a 3-fold greater cholesterol efflux activity from cholesterol-loaded macrophage. Because the identified conditions causing this change in AP-HDL structure and function can exist physiologically at the surface of the macrophage, or in its endosomes, we postulate that AP-HDL contains latent functionalities that become apparent and active when it associates with macrophage cell surface/endosomal HS. In this way initial steps in the reverse cholesterol transport pathway are focused at sites of injury to mobilize cholesterol from macrophages that are actively participating in the phagocytosis of damaged membranes rich in cholesterol. The mechanism may also be of relevance to aspects of atherogenesis