Biosynthesis and functions of eicosanoids generated by the coelomocytes of the starfish, Asterias rubens

Eicosanoids are a group of oxygenated fatty acid derivatives formed from C20 polyunsaturated fatty acids, including arachidonic and eicosapentaenoic acids. The potential of the coelomocytes of the starfish, Asterias rubens, to generate eicosanoids through the cyclooxygenase (COX) and lipoxygenase (LOX) pathways was investigated using reverse-phase high performance liquid chromatography, enzyme immunoassay and gas chromatography–mass spectrometry. The principal LOX product was identified as 8-hydroxyeicosatetraenoic acid (8-HETE) with 8-hydroxyeicosapentaenoic acid (8-HEPE) synthesised at significantly lower levels. No classical prostaglandins (PG), such as PGE₂ or PGD₂, were found to be generated by ionophore-challenged coelomocytes. Incubation of coelomocytes with lipopolysaccharides from either Escherichia coli or Salmonella abortus failed to induce an increase in generation of LOX products and the presence of 8-HETE (0–25 μM) had no significant effect on the in vitro phagocytic activity of Asterias coelomocytes. Neither indomethacin (a COX inhibitor) or esculetin (a LOX inhibitor) had any effect on the clearance of the bacterium, Vibrio splendidus, from the coelomic cavity of starfish suggesting that products of these enzymes are not involved in such coelomocyte responses to foreign particles.     

The development of a quick-running prediction tool for the assessment of human injury owing to terrorist attack within crowded metropolitan environments

In the aftermath of the London '7/7' attacks in 2005, UK government agencies required the development of a quick-running tool to predict the weapon and injury effects caused by the initiation of a person borne improvised explosive device (PBIED) within crowded metropolitan environments. This prediction tool, termed the HIP (human injury predictor) code, was intended to:--assist the security services to encourage favourable crowd distributions and densities within scenarios of 'sensitivity'; --provide guidance to security engineers concerning the most effective location for protection systems; --inform rescue services as to where, in the case of such an event, individuals with particular injuries will be located; --assist in training medical personnel concerning the scope and types of injuries that would be sustained as a consequence of a particular attack; --assist response planners in determining the types of medical specialists (burns, traumatic amputations, lungs, etc.) required and thus identify the appropriate hospitals to receive the various casualty types. This document describes the algorithms used in the development of this tool, together with the pertinent underpinning physical processes. From its rudimentary beginnings as a simple spreadsheet, the HIP code now has a graphical user interface (GUI) that allows three-dimensional visualization of results and intuitive scenario set-up. The code is underpinned by algorithms that predict the pressure and momentum outputs produced by PBIEDs within open and confined environments, as well as the trajectories of shrapnel deliberately placed within the device to increase injurious effects. Further logic has been implemented to transpose these weapon effects into forms of human injury depending on where individuals are located relative to the PBIED. Each crowd member is subdivided into representative body parts, each of which is assigned an abbreviated injury score after a particular calculation cycle. The injury levels of each affected body part are then summated and a triage state assigned for each individual crowd member based on the criteria specified within the 'injury scoring system'. To attain a comprehensive picture of a particular event, it is important that a number of simulations, using what is substantively the same scenario, are undertaken with natural variation being applied to the crowd distributions and the PBIED output. Accurate mathematical representation of such complex phenomena is challenging, particularly as the code must be quick-running to be of use to the stakeholder community. In addition to discussing the background and motivation for the algorithm and GUI development, this document also discusses the steps taken to validate the tool and the plans for further functionality implementation.     

Evaluation of large scale quantitative proteomic assay development using peptide affinity-based mass spectrometry

Stable isotope standards and capture by antipeptide antibodies (SISCAPA) couples affinity enrichment of peptides with stable isotope dilution and detection by multiple reaction monitoring mass spectrometry to provide quantitative measurement of peptides as surrogates for their respective proteins. In this report, we describe a feasibility study to determine the success rate for production of suitable antibodies for SISCAPA assays in order to inform strategies for large-scale assay development. A workflow was designed that included a multiplex immunization strategy in which up to five proteotypic peptides from a single protein target were used to immunize individual rabbits. A total of 403 proteotypic tryptic peptides representing 89 protein targets were used as immunogens. Antipeptide antibody titers were measured by ELISA and 220 antipeptide antibodies representing 89 proteins were chosen for affinity purification. These antibodies were characterized with respect to their performance in SISCAPA-multiple reaction monitoring assays using trypsin-digested human plasma matrix. More than half of the assays generated were capable of detecting the target peptide at concentrations of less than 0.5 fmol/μl in human plasma, corresponding to protein concentrations of less than 100 ng/ml. The strategy of multiplexing five peptide immunogens was successful in generating a working assay for 100% of the targeted proteins in this evaluation study. These results indicate it is feasible for a single laboratory to develop hundreds of assays per year and allow planning for cost-effective generation of SISCAPA assays.     

Identification of major clonal complexes and toxin producing strains among Staphylococcus aureus associated with atopic dermatitis

Staphylococcus aureus promotes the onset and severity of atopic dermatitis (AD), which is exacerbated by superantigen toxins SEB and SEC. The genetic identity of these isolates, and their relationship to common hospital- or community-associated methicillin resistant S. aureus (HA-MRSA and CA-MRSA) has not been defined. We conducted spa typing, partial multi-locus sequence typing (MLST), and toxin profiling (seb, sec, lukS-PV) of S. aureus from 119 pediatric and 40 adult AD patients. MLST clonal complexes CC45, CC5, CC15, CC1, CC8 and CC30 accounted for 79% of isolates, representing the same major groups reported for nosocomial S. aureus in hospital intensive care units. The highest disease severity was associated with CC1, which was significantly greater relative to CC15 (p?=?0.017) or CC30 (p?=?0.040), but with no significant difference relative to CC45, CC5 or CC8. Although there were two few lukS-PV, seb or sec isolates to infer a role in disease severity, CC45 was identified as a source of SEC producing strains, and lukS-PVL was associated with a small number of CC5 pediatric isolates. CC1 harbored the only CA-MRSA that was identified, and was a source of isolates that expressed both seb and sec, and closely resembled the USA400 strain of CA-MRSA.     

Uncovering genomic causes of co-morbidity in epilepsy: gene-driven phenotypic characterization of rare microdeletions

Background

Patients with epilepsy often suffer from other important conditions. The existence of such co-morbidities is frequently not recognized and their relationship with epilepsy usually remains unexplained.

Methodology/Principal Findings

We describe three patients with common, sporadic, non-syndromic epilepsies in whom large genomic microdeletions were found during a study of genetic susceptibility to epilepsy. We performed detailed gene-driven clinical investigations in each patient. Disruption of the function of genes in the deleted regions can explain co-morbidities in these patients.

Conclusions/Significance

Co-morbidities in patients with epilepsy can be part of a genomic abnormality even in the absence of (known) congenital malformations or intellectual disabilities. Gene-driven phenotype examination can also reveal clinically significant unsuspected condition.     

EVOLUTIONARY STUDIES OF ATRIPLEX: ADAPTIVE PRODUCTS FROM THE NATURAL HYBRID, 6N A. TRIDENTATA x 4N A. CANESCENS

Much of the variation in hexaploid Atriplex tridentata appears to have come from tetraploid A. canescens by introgression. Among the recombinations, three types appear to have become established as new, adaptive, hexaploid derivatives. One of these is a robust, woody form near Knolls, Utah, another is a widespread, low-growing, shrubby form in Lander County, Nevada, the other is an upright bushy, canescens-like form near Grantsville, Utah.