The protective effects of plasma gelsolin on stroke outcome in rats

<p>Abstract</p> <p>Background</p> <p>To date, recombinant tissue plasminogen activator (rtPA) is the only approved drug for ischemic stroke. It is intravenously administered functioning as a thrombolytic agent and is used to obtain reperfusion of the affected area of the brain. Excitotoxicity, inflammation and apoptosis are all involved in delayed neuronal death following stroke and offer multiple opportunities to intervene with neuroprotective agents. Gelsolin (GSN) is an actin- and calcium-binding protein mediating the disassembly of actin filaments and activity of calcium channels. It also functions as a regulator of apoptosis and inflammatory responses. This study tests the hypothesis that increasing the concentration of the form of GSN known as plasma GSN (pGSN) near an infarct will provide neuroprotection following ischemic stroke.</p> <p>Methods</p> <p>We induced middle cerebral artery occlusion (MCAO) in male rats via intracranial injection of endothelin-1 (ET-1), a potent vasoconstrictor, and then treated with local delivery of pGSN. Whole brain laser Doppler perfusion imaging was performed through the skull to assess MCAO effectiveness. Cylinder and vibrissae tests evaluated sensorimotor function before and 72 h after MCAO. Infarct volumes were examined 72 h after MCAO via 2, 3, 5-triphenyltetrazolium chloride (TTC) assay.</p> <p>Results</p> <p>Estimates of relative cerebral perfusion were significantly decreased in all groups receiving MCAO with no differences detected between treatments. Despite equivalent initial strokes, the infarct volume of the pGSN treatment group was significantly reduced compared with the untreated MCAO rats at 72 h. ET-1 induced significant deficits in both cylinder and vibrissae tests while pGSN significantly limited these deficits.</p> <p>Conclusion</p> <p>Gelsolin could be a promising drug for protection against neurodegeneration following ischemic stroke.</p

The Falklands Wargame

(U) The Falklands Wargame was conducted for the purposes of developing a maritime capability in the Contingency Force Analysis Wargame (CFAW) model and comparing wargame results with history. During the conduct of wargameexcursions, logistic constraint .was used to increase realism and to provide an additional control mechanism in the wargame. Two excursions using the historical scenario and one additional free play excursion were conducted as part of the wargame effort. (U) As a result of these wargames, a maritime capability was developed for the CFAW model which is consistent in detail with the rest of the model. Although casualty assessments did not statistically match h1storical results, they appeared to be reasonable, based on subjective military judgement. The CFAW model offers flexibility and robustness, which when combined with gamer imagination and innovation, provides a powerful wargaming tool. (U) The CFAW model is capable of portraying joint operations

Boresight Alignment Station

A boresight alignment system facilitates aligning a plurality of cameras and/or images of the cameras with respect to one another. The system may have a mount configured to facilitate attachment of a bezel containing a plurality of cameras to the mount. The system may have a plurality of targets and each target may be configured to provide light of at least two different wavelengths and/or ranges of wavelengths. One or more baffles may be disposed optically between the mount and the target assembly to inhibit stray light from being incident upon the cameras

A framework for understanding sources of bias in medication adherence research

The sources of bias in medication adherence research have not been comprehensively explored. We aimed to identify biases expected to affect adherence research and to develop a framework for mapping these onto the phases of adherence (initiation, implementation and discontinuation). A literature search was conducted, key papers were reviewed and a Catalogue of Bias was consulted. The specific biases related to adherence measurement and metrics were mapped onto the phases of adherence using a tabular matrix. Twenty-three biases were identified, of which 11 were specifically relevant to adherence measures and metrics. The mapping framework showed differences in the numbers and types of biases associated with each measure and metric while highlighting those common to many adherence study designs (e.g., unacceptability bias and apprehension bias). The framework will inform the design of adherence studies and the development of risk of bias tools for adherence research.<br/

Design of the HIV Prevention Trials Network (HPTN) Protocol 054: A cluster randomized crossover trial to evaluate combined access to Nevirapine in developing countries

HPTN054 is a cluster randomized trial designed to compare two approaches to providing single dose nevirapine to HIV-seropositive mothers and their infants to prevent mother-to-child transmission of HIV in resource limited settings. A number of challenging issues arose during the design of this trial. Most importantly, the need to achieve high participation rates among pregnant, HIV-seropositive women in selected prenatal care clinics led us to develop a method of collecting anonymous and unlinked information on a key surrogate endpoint instead of pursuing linked and identified information on a clinical endpoint. In addition, since group counseling is the standard model for prenatal care in sub-Saharan Africa, the prenatal care clinic serves as the unit of randomization. However, constraints on the number of suitable clinics and other logistical difficulties necessitated a unique type of hybrid parallel/stepped wedge cluster randomized design in which some clinics cross over between the two treatment modalities and some do not. We describe the design for the HPTN054 trial with an emphasis on the logistic and statistical features that allowed us to address these issues. We also provide some general statistical results that are useful for computing power in parallel, crossover, stepped wedge or mixed designs of cluster randomized trials

High-throughput screening of cellulase F mutants from multiplexed plasmid sets using an automated plate assay on a functional proteomic robotic workcell

<p>Abstract</p> <p>Background</p> <p>The field of plasmid-based functional proteomics requires the rapid assay of proteins expressed from plasmid libraries. Automation is essential since large sets of mutant open reading frames are being cloned for evaluation. To date no integrated automated platform is available to carry out the entire process including production of plasmid libraries, expression of cloned genes, and functional testing of expressed proteins.</p> <p>Results</p> <p>We used a functional proteomic assay in a multiplexed setting on an integrated plasmid-based robotic workcell for high-throughput screening of mutants of cellulase F, an endoglucanase from the anaerobic fungus <it>Orpinomyces </it>PC-2. This allowed us to identify plasmids containing optimized clones expressing mutants with improved activity at lower pH. A plasmid library of mutagenized clones of the <it>celF </it>gene with targeted variations in the last four codons was constructed by site-directed PCR mutagenesis and transformed into <it>Escherichia coli</it>. A robotic picker integrated into the workcell was used to inoculate medium in a 96-well deep well plate, combining the transformants into a multiplexed set in each well, and the plate was incubated on the workcell. Plasmids were prepared from the multiplexed culture on the liquid handler component of the workcell and used for <it>in vitro </it>transcription/translation. The multiplexed expressed recombinant proteins were screened for improved activity and stability in an azo-carboxymethylcellulose plate assay. The multiplexed wells containing mutants with improved activity were identified and linked back to the corresponding multiplexed cultures stored in glycerol. Spread plates were prepared from the glycerol stocks and the workcell was used to pick single colonies from the spread plates, prepare plasmid, produce recombinant protein, and assay for activity. The screening assay and subsequent deconvolution of the multiplexed wells resulted in identification of improved CelF mutants and corresponding optimized clones in expression-ready plasmids.</p> <p>Conclusion</p> <p>The multiplex method using an integrated automated platform for high-throughput screening in a functional proteomic assay allows rapid identification of plasmids containing optimized clones ready for use in subsequent applications including transformations to produce improved strains or cell lines.</p