Isolation, purification and partial characterisation of cancer procoagulant from placental amnion-chorion membranes and its role in angiogenesis inflammation and metastasis

Cancer procoagulant (EC 3.4.22.26) is an enzyme that is derived from tumour and foetal tissue, but not normal tissue. It is a direct activator of factor X and has been isolated from amnion-chorion membranes as well as from extracts and cells from human melanoma. The presence of cancer procoagulant has been associated with the malignant phenotype, as well as having a particularly high activity in metastatic cells. Cancer procoagulant activity is elevated in the serum of early stage breast cancer patients and decreased to normal in the advanced stages of the disease. In this study, cancer procoagulant was successfully isolated from amnion-chorion membranes and purified to homogeneity. The molecular weight of cancer procoagulant was determined using SDS-PAGE and was found to be 68 kDa. Cancer procoagulant was delipidated and it was shown that its activity was increased by the presence of lipids in a dose-dependent manner. Recovery of cancer procoagulant after delipidation is poor, consequently, a larger mass of sample is required to obtain sufficient amounts of delipidated material for N-terminal amino acid analysis. The optimum pH of cancer procoagulant was determined to be pH 8 and its optimal temperature was found to be 50°C. Novel synthetic substrates were designed to assay for cancer procoagulant activity. Currently, 2 potential candidates have been identified, namely, PQVR-AMC and AVSQSKP-AMC. Cancer procoagulant-induced expression of cytokines is differently modulated in the less aggressive MCF-7 cell line as compared to the metastatic and more aggressive MDA-MB-231 cell line. There are marked similarities in the inflammatory response produced by cancer procoagulant in hTERT-HDLEC and MDA-MB-231 cells, which are both associated with migratory capacity. Furthermore, cancer procoagulant-induced PDGF-β expression in hTERT-HDLEC and MDA-MB-231 cells could point to involvement of cancer procoagulant in wound healing and metastatic spread, respectively. Cancer procoagulant induced the motility of MDA-MB-231, MCF-7 and hTERT- cells in vitro in a time- and dose-dependent manner. Together, these results suggest that cancer procoagulant plays a role in the migration of breast cancer cells as well as the migration of endothelial cells

Purification and partial characterisation of cathepsin D from ostrich skeletal muscle, and its activity during meat maturation

Cathepsin D, a muscle proteinase, participates in lysosomally mediated protein degradation in vivo. This enzyme has been proposed to play a significant role in the postmortem proteolysis process apparently associated with tenderisation. The lack of data on the postmortem characteristics of ostrich meat, especially on the ageing process and its influence on meat tenderness, called for an investigation into this process. There is no data available for purified ostrich cathepsin D, and the aim of this study was, therefore, to isolate, purify and characterise cathepsin D from ostrich skeletal muscle and subsequently investigate the possible role that it may have in the tenderisation process of meat. Cathepsin D was successfully isolated and purified from ostrich skeletal muscle using pepstatin A-agarose chromatography. The purified enzyme was composed of two subunits (14 and 29kDa). The amino acid composition as well as the N-terminal amino acid sequence of both subunits were determined. Kinetic parameters (Km and Vm), thermodynamic parameters (Ea, ∆H, ∆S and ∆G) and functional characteristics (effect of pH, temperature and various inhibitors on cathepsin D activity) were determined and are reported in this study. Ostrich muscle cathepsin D showed a pH optimum of 4 and a temperature optimum of 45°C. The activity of cathepsin D was strongly inhibited by pepstatin A and DTT. Purified ostrich cathepsin D displayed kinetic and functional properties similar to previously reported values from various species. The effect of storage on the activity of cathepsin D was investigated over a 30 day period. It was established that substantial postmortem cathepsin D activity remained throughout the storage period, to implicate cathepsin D, fulfilling a possible role in meat maturation

The experiential salience of music in identity for singing teachers

Professional musicians with strong identities in music may also have a high degree of music in their identities. Accordingly, a rigid identification with work may be problematic for musicians, particularly when forces beyond their control change their work circumstances. In this study, we conducted semi-structured interviews with 10 singing teachers, representing a subset of professional musicians, and used interpretative phenomenological analysis (IPA) to explore the ways in which they enacted music in their identities. The framework of musical identities in action was used to interpret the findings, revealing the dynamic, embodied, and situated complexity of music in participants’ identities. Music had existential salience in the accounts of nine participants. Its salience resulted from the dynamic and situated presence of music across the lifespan, the literal embodiment of the singing voice, and the metaphorical embodiment of the presence of music in participants’ experiences. While a strong sense of music in people’s identities can promote lifelong engagement with music, if the emphasis on music is existential, the identity of a professional musician may be at risk when they are faced with an external threat to their livelihood. This is important knowledge for music educators and professional musicians’ career thinking

The prime spectrum of algebras of quadratic growth

We study prime algebras of quadratic growth. Our first result is that if $A$ is a prime monomial algebra of quadratic growth then $A$ has finitely many prime ideals $P$ such that $A/P$ has GK dimension one. This shows that prime monomial algebras of quadratic growth have bounded matrix images. We next show that a prime graded algebra of quadratic growth has the property that the intersection of the nonzero prime ideals $P$ such that $A/P$ has GK dimension 2 is non-empty, provided there is at least one such ideal. From this we conclude that a prime monomial algebra of quadratic growth is either primitive or has nonzero locally nilpotent Jacobson radical. Finally, we show that there exists a prime monomial algebra $A$ of GK dimension two with unbounded matrix images and thus the quadratic growth hypothesis is necessary to conclude that there are only finitely many prime ideals such that $A/P$ has GK dimension 1.Comment: 23 page

Looking through the same lens: Shear calibration for LSST, Euclid, and WFIRST with stage 4 CMB lensing

The next-generation weak lensing surveys (i.e., LSST, Euclid, and WFIRST) will require exquisite control over systematic effects. In this paper, we address shear calibration and present the most realistic forecast to date for LSST/Euclid/WFIRST and CMB lensing from a stage 4 CMB experiment (“CMB S4”). We use the cosmolike code to simulate a joint analysis of all the two-point functions of galaxy density, galaxy shear, and CMB lensing convergence. We include the full Gaussian and non-Gaussian covariances and explore the resulting joint likelihood with Monte Carlo Markov chains. We constrain shear calibration biases while simultaneously varying cosmological parameters, galaxy biases, and photometric redshift uncertainties. We find that CMB lensing from CMB S4 enables the calibration of the shear biases down to 0.2%–3% in ten tomographic bins for LSST (below the ∼ 0.5 % requirements in most tomographic bins), down to 0.4%–2.4% in ten bins for Euclid, and 0.6%–3.2% in ten bins for WFIRST. For a given lensing survey, the method works best at high redshift where shear calibration is otherwise most challenging. This self-calibration is robust to Gaussian photometric redshift uncertainties and to a reasonable level of intrinsic alignment. It is also robust to changes in the beam and the effectiveness of the component separation of the CMB experiment, and slowly dependent on its depth, making it possible with third-generation CMB experiments such as AdvACT and SPT-3G, as well as the Simons Observatory

High-Frequency Density Oscillations from a Plasma Source Used for Simulating Low-Earth Orbit Plasma Environment

We present data from ground-based, vacuum-chamber tests demonstrating the ability to modulate the output of a plasma source capable of producing a low-Earth orbit (LEO) type plasma. We obtained plasma oscillations up to 2.5 kHz impingent on stationary test equipment, which corresponds to meter-level ionospheric structures in LEO. This plasma source is, therefore, suitable for developing scientific instruments that measure the LEO plasma environment, in situ, with meter-level spatial resolution. Measurements were made using a fixed-bias collector and an electrometer sampling at 40 kHz. A mechanical aperture was established at the output of the plasma source via two concentric grids. The outer grid was free to rotate in the azimuthal direction with respect to the fixed inner grid. An identical, alternating hole pattern in the two grids resulted in a variable aperture that cycles through 90 open/close cycles per revolution. The frequency of the plasma oscillations is limited by the mechanism used to spin the grids and the bearing assembly on which the grids rotate. Higher frequencies are obtainable by upgrading the drive mechanism, allowing the possibility of centimeter-level spatial resolution

The Development of Test Facilities for Induced, High-Frequency Plasma Instabilities

We present results from modifications to test facilities equipped with a plasma source capable of producing a LEO-type environment. The modifications impose an oscillation to the output, thus simulating ionospheric disturbance. The frequency of the oscillations is adjustable as well as the base-line output of the source. Test results indicate that the density of the plasma can be varied with minimal impact on other plasma properties such as electron temperature. It is, therefore, possible to simulate realistic plasma environments such as day/night transitions or localized turbulence. The modified source is an effective tool for testing space instruments in a relevant environment increasing the instruments technology readiness level

Purification and biochemical characterisation of a putative sodium channel agonist secreted from the South African Knobbly sea anemone Bunodosoma capense

Voltage gated ion channels have become a subject of investigation as possible pharmaceutical targets. Research has linked the activity of ion channels directly to anti-inflammatory pathways, energy homeostasis, cancer proliferation and painful diabetic neuropathy. Sea anemones secrete a diverse array of bioactive compounds including potassium and sodium channel toxins. A putative novel sodium channel agonist (molecular mass of 4619.7 Da) with a predicted sequence: CLCNSDGPSV RGNTLSGILW LAGCPSGWHN CKKHKPTIGW CCK was isolated from Bunodosoma capense using a modified stimulation technique to induce the secretion of the neurotoxin rich mucus confirmed by an Artemia nauplii bio-assay. The peptide purification combined size-exclusion and reverse-phase high performance liquid chromatography. A thallium-based ion flux assay confirmed the presence of a sodium channel agonist/inhibitor and purity was determined using a modified tricine SDS-PAGE system. The peptide isolated indicated the presence of multiple disulfide bonds in a tight β-defensin cystine conformation. An IC50 value of 26 nM was determined for total channel inhibition on MCF-7 cells. The unique putative sodium channel agonist initiating with a cystine bond indicates a divergent evolution to those previously isolated from Bunodosoma species