Expression of a tick toxin for the development of a canine vaccine

University of Technology, Sydney. Faculty of Science.Acute, ascending, flaccid motor paralysis of forelimbs and death due to respiratory failure is the dominant characteristic of tick toxicosis by the Australian paralysis tick, Ixodes holocyclus. A tick toxin vaccine has the potential to be an effective preventative measure against tick toxicosis that affects thousands of domestic and companion animals each year. In previous research for the development of an anti-tick vaccine, Masina (1999) used a maltose-binding protein (MBP) holocyclus toxin (HT-1) fusion protein as immunogen. It was partially protective against challenges with crude tick extract in neonatal mice but unable to protect dogs against paralysis caused by direct tick attachment. Subsequently, four monoclonal antibodies generated in this study against the fusion protein were found to be incapable of binding to the native toxin in crude tick extract in Western blots. These results indicated that the HT-1 component of the MBP fusion protein was incorrectly folded compared to the native toxin. An expression system using ubiquitin fusion protein was investigated. Ubiquitin is a 10 kDa carrier protein which is smaller than the 43 kDa MBP. A smaller fusion partner was considered as it is less likely to interfere with HT-1 folding. The ubiquitin-HT-1 fusion protein was expressed in both soluble and insoluble forms, corresponding to the cytoplasmic fraction and inclusion bodies. The soluble form could be purified under non-denaturing conditions utilising the incorporated His-6-tag and a Ni affinity column. The insoluble form, like the periplasmically expressed MBP-HT-1 fusion protein, could only be purified using denaturing conditions. The purified soluble ubiquitin-HT-1 fusion protein appeared to have the correct conformational folding as it was recognized by commercial dog anti-tick serum in Western blots. The purified soluble ubiquitin-HT-1 fusion protein was used to immunise rabbits and mice for protection experiments, but was found to be unprotective. However, serum from immunized animals was able to detect a 5 kDa protein from crude tick extract in a Western blot. This 5 kDa protein was also recognized by the commercial dog anti-tick serum and has the molecular weight corresponding to the HT-1 neurotoxin. Creation of monoclonal antibodies to this ubiquitin-HT-1 fusion protein may therefore aid future development towards a tick vaccine. These antibodies could in turn be used to isolate native HT-1 from the crude tick extract instead of a combination of conventional chromatography methods. This approach would allow isolation of HT-1 in more significant quantities than is currently possible to enable confirmation of the HT-1 sequence. Suspected problems associated with incorrect sequence or interference by the carrier protein to cause a non-protective response could also be resolved

Hyperoxia Elevates Adrenic Acid Peroxidation in Marine Fish and Is Associated with Reproductive Pheromone Mediators

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Common Genetic Variant Association with Altered HLA Expression, Synergy with Pyrethroid Exposure, and Risk for Parkinson's Disease: An Observational and Case-Control Study.

Background/objectivesThe common non-coding single nucleotide polymorphism (SNP) rs3129882 in HLA-DRA is associated with risk for idiopathic Parkinson's disease (PD). The location of the SNP in the major histocompatibility complex class II (MHC-II) locus implicates regulation of antigen presentation as a potential mechanism by which immune responses link genetic susceptibility to environmental factors in conferring lifetime risk for PD.MethodsFor immunophenotyping, blood cells from 81 subjects were analyzed by qRT-PCR and flow cytometry. A case-control study was performed on a separate cohort of 962 subjects to determine association of pesticide exposure and the SNP with risk of PD.ResultsHomozygosity for G at this SNP was associated with heightened baseline expression and inducibility of MHC class II molecules in B cells and monocytes from peripheral blood of healthy controls and PD patients. In addition, exposure to a commonly used class of insecticide, pyrethroids, synergized with the risk conferred by this SNP (OR = 2.48, p = 0.007), thereby identifying a novel gene-environment interaction that promotes risk for PD via alterations in immune responses.ConclusionsIn sum, these novel findings suggest that the MHC-II locus may increase susceptibility to PD through presentation of pathogenic, immunodominant antigens and/or a shift toward a more pro-inflammatory CD4+ T cell response in response to specific environmental exposures, such as pyrethroid exposure through genetic or epigenetic mechanisms that modulate MHC-II gene expression

Gene Expression Profiling Identified High-mobility Group AT-hook (HMGA2) as Being Frequently Upregulated in Esophageal Squamous Cell Carcinoma

Background: Esophageal cancer is one of the most deadly malignancies worldwide and esophageal squamous cell carcinoma (ESCC) is the most frequent type. Methods: We identified up-regulated genes from gene expression profiles of HKESC-4 cell line, its parental tumor tissues, non-tumoral esophageal epithelia and lymph nodes with metastatic carcinoma using Human Genome U133 Plus 2.0 microarray. Results: Four genes [High-mobility group AT-hook 2 (HMGA2), paternally expressed 10 (PEG10), SH3 and multiple ankyrin repeat domains 2 (SHANK2) and WNT1 inducible signaling pathway protein 3 (WISP3)] were selected for further validation with real-time quantitative polymerase chain reaction (qPCR) in a panel of ESCC cell lines and clinical specimens. HMGA2 was found to be overexpressed in the panel of ESCC cell lines tested. By using immunohistochemistry, HMGA2 was found to be up-regulated in 70% of ESCC tissues (21 out of 30 cases). Conclusion: This study demonstrates successful use of gene microarray to identify and reveal HMGA2 as a novel and consistently overexpressed gene in ESCC cell lines and clinical samples.published_or_final_versio

Requirements modelling and formal analysis using graph operations

The increasing complexity of enterprise systems requires a more advanced analysis of the representation of services expected than is currently possible. Consequently, the specification stage, which could be facilitated by formal verification, becomes very important to the system life-cycle. This paper presents a formal modelling approach, which may be used in order to better represent the reality of the system and to verify the awaited or existing system’s properties, taking into account the environmental characteristics. For that, we firstly propose a formalization process based upon properties specification, and secondly we use Conceptual Graphs operations to develop reasoning mechanisms of verifying requirements statements. The graphic visualization of these reasoning enables us to correctly capture the system specifications by making it easier to determine if desired properties hold. It is applied to the field of Enterprise modelling

Chemical ordering in PtNi nanocrystals

We investigated the chemical ordering in PtNi nanocrystals fabricated on sapphire substrate using in-situ synchrotron X-ray scattering. Nanocrystals with composition close to 1:1 were ordered in the tetragonal L1(0) structure at low temperatures. The transition to disordered FCC structure occurred at around 640 degrees C and substantial hysteresis of about 50 K was observed. Nanocrystals of smaller sizes fabricated under the same conditions were Ni rich and ordered into Cu3Au type L1(2) structure. Significantly higher degree of chemical ordering was observed in L1(2) structure than in L1(0) structure. (C) 2016 Elsevier B.V. All rights reserved.1144Ysciescopu

Clinical correlation of nuclear survivin in esophageal squamous cell carcinoma

To examine the correlation of survivin (both total and nuclear survivin) with clinicopathological parameters of esophageal squamous cell carcinoma (ESCC) patients. Tumors and non-tumor tissues near the proximal resection margins were resected from ESCC patients undergone esophagectomy. Quantitative polymerase chain reaction (qPCR) was performed to detect survivin mRNA expression level in the 10 paired tumor and adjacent non-tumor tissues. To confirm with the clinical situation, survivin mRNA and protein expression were measured by qPCR and immunoblot, respectively, in 5 ESCC cell lines and a non-neoplastic esophageal epithelial cell line. Immunohistochemistry was employed to reveal the cellular localization of survivin in tumor tissues isolated from the 64 ESCC patients undergone surgery alone. Up-regulation of survivin mRNA and protein was found in 5 ESCC lines (HKESC-1, HKESC-2, HKESC-3, HKESC-4, and SLMT-1) when compared to a non-neoplastic esophageal epithelial cell line NE-1. In particular, HKESC-3, HKESC-4, and SLMT-1 cells demonstrated ~50-fold increase in survivin mRNA. High level of survivin mRNA in tumor tissues when compared to non-tumor tissues was found in 70 % (7 of 10) of clinical cases. The increase in expression ranged from ~twofold to ~16-fold. Immunohistochemistry results showed that survivin was found at the cell nuclei in all specimens examined. Nuclear expression of survivin was inversely associated with the likelihood of developing nodal metastasis (p = 0.021) and significantly associated with early-stage ESCC (p = 0.039). Nuclear survivin could serve as a marker for indicating disease status in ESCC patients. © 2012 The Author(s).published_or_final_versio

MSSM Baryogenesis and Electric Dipole Moments: An Update on the Phenomenology

We explore the implications of electroweak baryogenesis for future searches for permanent electric dipole moments in the context of the minimal supersymmetric extension of the Standard Model (MSSM). From a cosmological standpoint, we point out that regions of parameter space that over-produce relic lightest supersymmetric particles can be salvaged only by assuming a dilution of the particle relic density that makes it compatible with the dark matter density: this dilution must occur after dark matter freeze-out, which ordinarily takes place after electroweak baryogenesis, implying the same degree of dilution for the generated baryon number density as well. We expand on previous studies on the viable MSSM regions for baryogenesis, exploring for the first time an orthogonal slice of the relevant parameter space, namely the (tan\beta, m_A) plane, and the case of non-universal relative gaugino-higgsino CP violating phases. The main result of our study is that in all cases lower limits on the size of the electric dipole moments exist, and are typically on the same order, or above, the expected sensitivity of the next generation of experimental searches, implying that MSSM electroweak baryogenesis will be soon conclusively tested.Comment: 23 pages, 10 figures, matches version published in JHE

Rewritable nanoscale oxide photodetector

Nanophotonic devices seek to generate, guide, and/or detect light using structures whose nanoscale dimensions are closely tied to their functionality. Semiconducting nanowires, grown with tailored optoelectronic properties, have been successfully placed into devices for a variety of applications. However, the integration of photonic nanostructures with electronic circuitry has always been one of the most challenging aspects of device development. Here we report the development of rewritable nanoscale photodetectors created at the interface between LaAlO3 and SrTiO3. Nanowire junctions with characteristic dimensions 2-3 nm are created using a reversible AFM writing technique. These nanoscale devices exhibit a remarkably high gain for their size, in part because of the large electric fields produced in the gap region. The photoconductive response is gate-tunable and spans the visible-to-near-infrared regime. The ability to integrate rewritable nanoscale photodetectors with nanowires and transistors in a single materials platform foreshadows new families of integrated optoelectronic devices and applications.Comment: 5 pages, 5 figures. Supplementary Information 7 pages, 9 figure