In situ apparatus for the study of clathrate hydrates relevant to solar system bodies using synchrotron X-ray diffraction and Raman spectroscopy

Clathrate hydrates are believed to play a significant role in various solar system environments, e.g. comets, and the surfaces and interiors of icy satellites, however the structural factors governing their formation and dissociation are poorly understood. We demonstrate the use of a high pressure gas cell, combined with variable temperature cooling and time-resolved data collection, to the in situ study of clathrate hydrates under conditions relevant to solar system environments. Clathrates formed and processed within the cell are monitored in situ using synchrotron X-ray powder diffraction and Raman spectroscopy. X-ray diffraction allows the formation of clathrate hydrates to be observed as CO2 gas is applied to ice formed within the cell. Complete conversion is obtained by annealing at temperatures just below the ice melting point. A subsequent rise in the quantity of clathrate is observed as the cell is thermally cycled. Four regions between 100-5000cm-1 are present in the Raman spectra that carry features characteristic of both ice and clathrate formation. This novel experimental arrangement is well suited to studying clathrate hydrates over a range of temperature (80-500K) and pressure (1-100bar) conditions and can be used with a variety of different gases and starting aqueous compositions. We propose the increase in clathrate formation observed during thermal cycling may be due to the formation of a quasi liquid-like phase that forms at temperatures below the ice melting point, but which allows easier formation of new clathrate cages, or the retention and delocalisation of previously formed clathrate structures, possibly as amorphous clathrate. The structural similarities between hexagonal ice, the quasi liquid-like phase, and crystalline CO2 hydrate mean that differences in the Raman spectrum are subtle; however, all features out to 5000cm-1 are diagnostic of clathrate structure.Comment: Astronomy & Astrophysics, in press. 6 page

Investigating the role of Hedgehog/GLI1 signaling in glioblastoma cell response to temozolomide.

Resistance to chemotherapy substantially hinders successful glioblastoma (GBM) treatment, contributing to an almost 100% mortality rate. Resistance to the frontline chemotherapy, temozolomide (TMZ), arises from numerous signaling pathways that are deregulated in GBM, including Hedgehog (Hh) signaling. Here, we investigate suppression of Hh signaling as an adjuvant to TMZ using U87-MG and T98G cell lines as in vitro models of GBM. We found that silencing GLI1 with siRNA reduces cell metabolic activity by up to 30% in combination with TMZ and reduces multidrug efflux activity by 2.5-fold. Additionally, pharmacological GLI inhibition modulates nuclear p53 levels and decreases MGMT expression in combination with TMZ. While we surprisingly found that silencing GLI1 does not induce apoptosis in the absence of TMZ co-treatment, we discovered silencing GLI1 without TMZ co-treatment induces senescence as evidenced by a significant 2.3-fold increase in senescence associated β-galactosidase staining, and this occurs in a loss of PTEN-dependent manner. Finally, we show that GLI inhibition increases apoptosis in glioma stem-like cells by up to 6.8-fold in combination with TMZ, and this reduces the size and number of neurospheres grown from glioma stem-like cells. In aggregate, our data warrant the continued investigation of Hh pathway inhibitors as adjuvants to TMZ chemotherapy and highlight the importance of identifying signaling pathways that determine whether co-treatment will be successful

Non-aqueous formation of the calcium carbonate polymorph vaterite: astrophysical implications

We study the formation of calcium carbonate, through the solid-gas interaction of amorphous Ca-silicate with gaseous CO2, at elevated pressures, and link this to the possible presence of calcium carbonate in a number of circumstellar and planetary environments. We use in-situ synchrotron X-Ray powder diffraction to obtain detailed structural data pertaining to the formation of the crystalline calcium carbonate phase vaterite and its evolution with temperature. We found that the metastable calcium carbonate phase vaterite was formed alongside calcite, at elevated CO2 pressure, at room temperature and subsequently remained stable over a large range of temperature and pressure. We report the formation of the calcium carbonate mineral vaterite whilst attempting to simulate carbonate dust grain formation in astrophysical environments. This suggests that vaterite could be a mineral component of carbonate dust and also presents a possible method of formation for vaterite and its polymorphs on planetary surfaces.Comment: Astronomy and Astrophysics, in pres

Phosphonopeptides Revisited, in an Era of Increasing Antimicrobial Resistance

Given the increase in resistance to antibacterial agents, there is an urgent need for the development of new agents with novel modes of action. As an interim solution, it is also prudent to reinvestigate old or abandoned antibacterial compounds to assess their efficacy in the context of widespread resistance to conventional agents. In the 1970s, much work was performed on the development of peptide mimetics, exemplified by the phosphonopeptide, alafosfalin. We investigated the activity of alafosfalin, di-alanyl fosfalin and β-chloro-L-alanyl-β-chloro-L-alanine against 297 bacterial isolates, including carbapenemase-producing Enterobacterales (CPE) (n = 128), methicillin-resistant Staphylococcus aureus (MRSA) (n = 37) and glycopeptide-resistant enterococci (GRE) (n = 43). The interaction of alafosfalin with meropenem was also examined against 20 isolates of CPE. The MIC50 and MIC90 of alafosfalin for CPE were 1 mg/L and 4 mg/L, respectively and alafosfalin acted synergistically when combined with meropenem against 16 of 20 isolates of CPE. Di-alanyl fosfalin showed potent activity against glycopeptide-resistant isolates of Enterococcus faecalis (MIC90; 0.5 mg/L) and Enterococcus faecium (MIC90; 2 mg/L). Alafosfalin was only moderately active against MRSA (MIC90; 8 mg/L), whereas β-chloro-L-alanyl-β-chloro-L-alanine was slightly more active (MIC90; 4 mg/L). This study shows that phosphonopeptides, including alafosfalin, may have a therapeutic role to play in an era of increasing antibacterial resistance

G1 Domain of Versican Regulates Hyaluronan Organization and the Phenotype of Cultured Human Dermal Fibroblasts

Variants of versican have wide-ranging effects on cell and tissue phenotype, impacting proliferation, adhesion, pericellular matrix composition, and elastogenesis. The G1 domain of versican, which contains two Link modules that bind to hyaluronan (HA), may be central to these effects. Recombinant human G1 (rhG1) with an N-terminal 8 amino acid histidine (His) tag, produced in Nicotiana benthamiana, was applied to cultures of dermal fibroblasts, and effects on proliferation and pericellular HA organization determined. rhG1 located to individual strands of cell surface HA which aggregated into structures resembling HA cables. On both individual and aggregated strands, the spacing of attached rhG1 was similar (~120 nm), suggesting interaction between rhG1 molecules. Endogenous V0/V1, present on HA between attached rhG1, did not prevent cable formation, while treatment with V0/V1 alone, which also bound to HA, did not induce cables. A single treatment with rhG1 suppressed cell proliferation for an extended period. Treating cells for 4 weeks with rhG1 resulted in condensed layers of elongated, differentiated α actin-positive fibroblasts, with rhG1 localized to cell surfaces, and a compact extracellular matrix including both collagen and elastin. These results demonstrate that the G1 domain of versican can regulate the organization of pericellular HA and affect phenotype

Light and electron microscopical observations of the effects of high-density lipoprotein on growth of Plasmodium falciparum in vitro

Author Posting. © Cambridge University Press, 2004. This article is posted here by permission of Cambridge University Press for personal use, not for redistribution. The definitive version was published in Parasitology 128 (2004): 577-584, doi:10.1017/S0031182004005025.Human serum high-density lipoprotein (HDL) is necessary and sufficient for the short-term maintenance of Plasmodium falciparum in in vitro culture. However, at high concentrations it is toxic to the parasite. A heat-labile component is apparently responsible for the stage-specific toxicity to parasites within infected erythrocytes 12–42 h after invasion, i.e. during trophozoite maturation. The effects of HDL on parasite metabolism (as determined by nucleic acid synthesis) are evident at about 30 h after invasion. Parasites treated with HDL show gross abnormalities by light and electron microscopy.Professor Hajduk was supported by NIH. Professor Day was supported by a Research Leave Fellowship from The Wellcome Trust. Dr Imrie and Ms Carter were supported by Programme Grant funding awarded to Professor Day from The Wellcome Trust. Dr Ferguson was supported by an equipment grant from The Wellcome Trust

MKID development for SuperSpec: an on-chip, mm-wave, filter-bank spectrometer

SuperSpec is an ultra-compact spectrometer-on-a-chip for millimeter and submillimeter wavelength astronomy. Its very small size, wide spectral bandwidth, and highly multiplexed readout will enable construction of powerful multibeam spectrometers for high-redshift observations. The spectrometer consists of a horn-coupled microstrip feedline, a bank of narrow-band superconducting resonator filters that provide spectral selectivity, and Kinetic Inductance Detectors (KIDs) that detect the power admitted by each filter resonator. The design is realized using thin-film lithographic structures on a silicon wafer. The mm-wave microstrip feedline and spectral filters of the first prototype are designed to operate in the band from 195-310 GHz and are fabricated from niobium with at Tc of 9.2K. The KIDs are designed to operate at hundreds of MHz and are fabricated from titanium nitride with a Tc of 2K. Radiation incident on the horn travels along the mm-wave microstrip, passes through the frequency-selective filter, and is finally absorbed by the corresponding KID where it causes a measurable shift in the resonant frequency. In this proceedings, we present the design of the KIDs employed in SuperSpec and the results of initial laboratory testing of a prototype device. We will also briefly describe the ongoing development of a demonstration instrument that will consist of two 500-channel, R=700 spectrometers, one operating in the 1-mm atmospheric window and the other covering the 650 and 850 micron bands.Comment: As submitted, except that "in prep" references have been update

Anthropometric and Physiological Characteristics of Elite Male Rugby Athletes

This is the first article to review the anthropometric and physiological characteristics required for elite rugby performance within both Rugby Union (RU) and Rugby League (RL). Anthropometric characteristics such as height and mass, and physiological characteristics such as speed and muscular strength, have previously been advocated as key discriminators of playing level within rugby. This review aimed to identify the key anthropometric and physiological properties required for elite performance in rugby, distinguishing between RU and RL, forwards and backs and competitive levels. There are differences between competitive standards such that, at the elite level, athletes are heaviest (RU forwards ~111 kg, backs ~93 kg; RL forwards ~103 kg, backs ~90 kg) with lowest % body fat (RU forwards ~15%, backs ~12%; RL forwards ~14%, backs ~11%), they have most fat-free mass and are strongest (Back squat: RU forwards ~176 kg, backs ~157 kg; RL forwards ~188 kg, backs ~ 168 kg; Bench press: RU forwards ~131 kg, backs ~118 kg; RL forwards ~122 kg, backs ~113 kg) and fastest (10 m: RU forwards ~1.87 s, backs ~1.77 s; 10 m RL forwards ~1.9 s, backs ~1.83 s). We also have unpublished data that indicate contemporary RU athletes have less body fat and are stronger and faster than the published data suggest. Regardless, well-developed speed, agility, lower-body power and strength characteristics are vital for elite performance, probably reflect both environmental (training, diet, etc.) and genetic factors, distinguish between competitive levels and are therefore important determinants of elite status in rugby.Published versio

Small Molecule Inhibitors of 15-PGDH Exploit a Physiologic Induced-Fit Closing System

15-prostaglandin dehydrogenase (15-PGDH) is a negative regulator of tissue stem cells that acts via enzymatic activity of oxidizing and degrading PGE2, and related eicosanoids, that support stem cells during tissue repair. Indeed, inhibiting 15-PGDH markedly accelerates tissue repair in multiple organs. Here we have used cryo-electron microscopy to solve the solution structure of native 15-PGDH and of 15-PGDH individually complexed with two distinct chemical inhibitors. These structures identify key 15-PGDH residues that mediate binding to both classes of inhibitors. Moreover, we identify a dynamic 15-PGDH lid domain that closes around the inhibitors, and that is likely fundamental to the physiologic 15-PGDH enzymatic mechanism. We furthermore identify two key residues, F185 and Y217, that act as hinges to regulate lid closing, and which both inhibitors exploit to capture the lid in the closed conformation, thus explaining their sub-nanomolar binding affinities. These findings provide the basis for further development of 15-PGDH targeted drugs as therapeutics for regenerative medicine