Design, analysis and test verification of advanced encapsulation systems, phase 2 program results

Optical, electrical isolation, thermal structural, structural deflection, and thermal tests are reported. The utility of the optical, series capacitance, and structural deflection models was verified

Supersymmetry, quark confinement and the harmonic oscillator

We study some quantum systems described by noncanonical commutation relations formally expressed as [q,p]=ihbar(I + chi H), where H is the associated (harmonic oscillator-like) Hamiltonian of the system, and chi is a Hermitian (constant) operator, i.e. [H,chi]=0 . In passing, we also consider a simple (chi=0 canonical) model, in the framework of a relativistic Klein-Gordon-like wave equation.Comment: To be published in Journal of Physics A: Mathematical and Theoretical (2007

A Semantic Problem Solving Environment for Integrative Parasite Research: Identification of Intervention Targets for Trypanosoma cruzi

Effective research in parasite biology requires analyzing experimental lab data in the context of constantly expanding public data resources. Integrating lab data with public resources is particularly difficult for biologists who may not possess significant computational skills to acquire and process heterogeneous data stored at different locations. Therefore, we develop a semantic problem solving environment (SPSE) that allows parasitologists to query their lab data integrated with public resources using ontologies. An ontology specifies a common vocabulary and formal relationships among the terms that describe an organism, and experimental data and processes in this case. SPSE supports capturing and querying provenance information, which is metadata on the experimental processes and data recorded for reproducibility, and includes a visual query-processing tool to formulate complex queries without learning the query language syntax. We demonstrate the significance of SPSE in identifying gene knockout targets for T. cruzi. The overall goal of SPSE is to help researchers discover new or existing knowledge that is implicitly present in the data but not always easily detected. Results demonstrate improved usefulness of SPSE over existing lab systems and approaches, and support for complex query design that is otherwise difficult to achieve without the knowledge of query language syntax

Genome of the Avirulent Human-Infective Trypanosome—Trypanosoma rangeli

Background: Trypanosoma rangeli is a hemoflagellate protozoan parasite infecting humans and other wild and domestic mammals across Central and South America. It does not cause human disease, but it can be mistaken for the etiologic agent of Chagas disease, Trypanosoma cruzi. We have sequenced the T. rangeli genome to provide new tools for elucidating the distinct and intriguing biology of this species and the key pathways related to interaction with its arthropod and mammalian hosts.  Methodology/Principal Findings: The T. rangeli haploid genome is ,24 Mb in length, and is the smallest and least repetitive trypanosomatid genome sequenced thus far. This parasite genome has shorter subtelomeric sequences compared to those of T. cruzi and T. brucei; displays intraspecific karyotype variability and lacks minichromosomes. Of the predicted 7,613 protein coding sequences, functional annotations could be determined for 2,415, while 5,043 are hypothetical proteins, some with evidence of protein expression. 7,101 genes (93%) are shared with other trypanosomatids that infect humans. An ortholog of the dcl2 gene involved in the T. brucei RNAi pathway was found in T. rangeli, but the RNAi machinery is non-functional since the other genes in this pathway are pseudogenized. T. rangeli is highly susceptible to oxidative stress, a phenotype that may be explained by a smaller number of anti-oxidant defense enzymes and heatshock proteins.  Conclusions/Significance: Phylogenetic comparison of nuclear and mitochondrial genes indicates that T. rangeli and T. cruzi are equidistant from T. brucei. In addition to revealing new aspects of trypanosome co-evolution within the vertebrate and invertebrate hosts, comparative genomic analysis with pathogenic trypanosomatids provides valuable new information that can be further explored with the aim of developing better diagnostic tools and/or therapeutic targets

The steady-state transcriptome of the four major life-cycle stages of Trypanosoma cruzi

<p>Abstract</p> <p>Background</p> <p>Chronic chagasic cardiomyopathy is a debilitating and frequently fatal outcome of human infection with the protozoan parasite, <it>Trypanosoma cruzi</it>. Microarray analysis of gene expression during the <it>T. cruzi </it>life-cycle could be a valuable means of identifying drug and vaccine targets based on their appropriate expression patterns, but results from previous microarray studies in <it>T. cruzi </it>and related kinetoplastid parasites have suggested that the transcript abundances of most genes in these organisms do not vary significantly between life-cycle stages.</p> <p>Results</p> <p>In this study, we used whole genome, oligonucleotide microarrays to globally determine the extent to which <it>T. cruzi </it>regulates mRNA relative abundances over the course of its complete life-cycle. In contrast to previous microarray studies in kinetoplastids, we observed that relative transcript abundances for over 50% of the genes detected on the <it>T. cruzi </it>microarrays were significantly regulated during the <it>T. cruzi </it>life-cycle. The significant regulation of 25 of these genes was confirmed by quantitative reverse-transcriptase PCR (qRT-PCR). The <it>T. cruzi </it>transcriptome also mirrored published protein expression data for several functional groups. Among the differentially regulated genes were members of paralog clusters, nearly 10% of which showed divergent expression patterns between cluster members.</p> <p>Conclusion</p> <p>Taken together, these data support the conclusion that transcript abundance is an important level of gene expression regulation in <it>T. cruzi</it>. Thus, microarray analysis is a valuable screening tool for identifying stage-regulated <it>T. cruzi </it>genes and metabolic pathways.</p

Shoulder muscle endurance: the development of a standardized and reliable protocol

<p>Abstract</p> <p>Background</p> <p>Shoulder muscle fatigue has been proposed as a possible link to explain the association between repetitive arm use and the development of rotator cuff disorders. To our knowledge, no standardized clinical endurance protocol has been developed to evaluate the effects of muscle fatigue on shoulder function. Such a test could improve clinical examination of individuals with shoulder disorders. Therefore, the purpose of this study was to establish a reliable protocol for objective assessment of shoulder muscle endurance.</p> <p>Methods</p> <p>An endurance protocol was developed on a stationary dynamometer (Biodex System 3). The endurance protocol was performed in isotonic mode with the resistance set at 50% of each subject's peak torque as measured for shoulder external (ER) and internal rotation (IR). Each subject performed 60 continuous repetitions of IR/ER rotation. The endurance protocol was performed by 36 healthy individuals on two separate occasions at least two days apart. Maximal isometric shoulder strength tests were performed before and after the fatigue protocol to evaluate the effects of the endurance protocol and its reliability. Paired <it>t</it>-tests were used to evaluate the reduction in shoulder strength due to the protocol, while intraclass correlation coefficients (ICC) and minimal detectable change (MDC) were used to evaluate its reliability.</p> <p>Results</p> <p>Maximal isometric strength was significantly decreased after the endurance protocol (<it>P </it>< 0.001). The total work performed during the last third of the protocol was significantly less than the first third of the protocol (P < 0.05). The test-retest reliability of the post-fatigue strength measures was excellent (ICC >0.84).</p> <p>Conclusions</p> <p>Changes in muscular performance observed during and after the muscular endurance protocol suggests that the protocol did result in muscular fatigue. Furthermore, this study established that the resultant effects of fatigue of the proposed isotonic protocol were reproducible over time. The protocol was performed without difficulty by all volunteers and took less than 10 minutes to perform, suggesting that it might be feasible for clinical practice. This protocol could be used to induce local muscular fatigue in order to evaluate the effects of fatigue on shoulder kinematics or to evaluate changes in shoulder muscle endurance following rehabilitation.</p

Functional recombinant protein is present in the pre-induction phases of Pichia pastoris cultures when grown in bioreactors, but not shake-flasks

Background - Pichia pastoris is a widely-used host for recombinant protein production; expression is typically driven by methanol-inducible alcohol oxidase (AOX) promoters. Recently this system has become an important source of recombinant G protein-coupled receptors (GPCRs) for structural biology and drug discovery. The influence of diverse culture parameters (such as pH, dissolved oxygen concentration, medium composition, antifoam concentration and culture temperature) on productivity has been investigated for a wide range of recombinant proteins in P. pastoris. In contrast, the impact of the pre-induction phases on yield has not been as closely studied. In this study, we examined the pre-induction phases of P. pastoris bioreactor cultivations producing three different recombinant proteins: the GPCR, human A2a adenosine receptor (hA2aR), green fluorescent protein (GFP) and human calcitonin gene-related peptide receptor component protein (as a GFP fusion protein; hCGRP-RCP-GFP). Results - Functional hA2aR was detected in the pre-induction phases of a 1 L bioreactor cultivation of glycerol-grown P. pastoris. In a separate experiment, a glycerol-grown P. pastoris strain secreted soluble GFP prior to methanol addition. When glucose, which has been shown to repress AOX expression, was the pre-induction carbon source, hA2aR and GFP were still produced in the pre-induction phases. Both hA2aR and GFP were also produced in methanol-free cultivations; functional protein yields were maintained or increased after depletion of the carbon source. Analysis of the pre-induction phases of 10 L pilot scale cultivations also demonstrated that pre-induction yields were at least maintained after methanol induction, even in the presence of cytotoxic concentrations of methanol. Additional bioreactor data for hCGRP-RCP-GFP and shake-flask data for GFP, horseradish peroxidase (HRP), the human tetraspanins hCD81 and CD82, and the tight-junction protein human claudin-1, demonstrated that bioreactor but not shake flask cultivations exhibit recombinant protein production in the pre-induction phases of P. pastoris cultures. Conclusions - The production of recombinant hA2aR, GFP and hCGRP-RCP-GFP can be detected in bioreactor cultivations prior to methanol induction, while this is not the case for shake-flask cultivations of GFP, HRP, hCD81, hCD82 and human claudin-1. This confirms earlier suggestions of leaky expression from AOX promoters, which we report here for both glycerol- and glucose-grown cells in bioreactor cultivations. These findings suggest that the productivity of AOX-dependent bioprocesses is not solely dependent on induction by methanol. We conclude that in order to maximize total yields, pre-induction phase cultivation conditions should be optimized, and that increased specific productivity may result in decreased biomass yields

Trypanosoma cruzi Gene Expression in Response to Gamma Radiation

Trypanosoma cruzi is an organism highly resistant to ionizing radiation. Following a dose of 500 Gy of gamma radiation, the fragmented genomic DNA is gradually reconstructed and the pattern of chromosomal bands is restored in less than 48 hours. Cell growth arrests after irradiation but, while DNA is completely fragmented, RNA maintains its integrity. In this work we compared the transcriptional profiles of irradiated and non-irradiated epimastigotes at different time points after irradiation using microarray. In total, 273 genes were differentially expressed; from these, 160 were up-regulated and 113 down-regulated. We found that genes with predicted functions are the most prevalent in the down-regulated gene category. Translation and protein metabolic processes, as well as generation of precursor of metabolites and energy pathways were affected. In contrast, the up-regulated category was mainly composed of obsolete sequences (which included some genes of the kinetoplast DNA), genes coding for hypothetical proteins, and Retrotransposon Hot Spot genes. Finally, the tyrosyl-DNA phosphodiesterase 1, a gene involved in double-strand DNA break repair process, was up-regulated. Our study demonstrated the peculiar response to ionizing radiation, raising questions about how this organism changes its gene expression to manage such a harmful stress

A unified framework for managing provenance information in translational research

<p>Abstract</p> <p>Background</p> <p>A critical aspect of the NIH <it>Translational Research </it>roadmap, which seeks to accelerate the delivery of "bench-side" discoveries to patient's "bedside," is the management of the <it>provenance </it>metadata that keeps track of the origin and history of data resources as they traverse the path from the bench to the bedside and back. A comprehensive provenance framework is essential for researchers to verify the quality of data, reproduce scientific results published in peer-reviewed literature, validate scientific process, and associate trust value with data and results. Traditional approaches to provenance management have focused on only partial sections of the translational research life cycle and they do not incorporate "domain semantics", which is essential to support domain-specific querying and analysis by scientists.</p> <p>Results</p> <p>We identify a common set of challenges in managing provenance information across the <it>pre-publication </it>and <it>post-publication </it>phases of data in the translational research lifecycle. We define the semantic provenance framework (SPF), underpinned by the Provenir upper-level provenance ontology, to address these challenges in the four stages of provenance metadata:</p> <p>(a) Provenance <b>collection </b>- during data generation</p> <p>(b) Provenance <b>representation </b>- to support interoperability, reasoning, and incorporate domain semantics</p> <p>(c) Provenance <b>storage </b>and <b>propagation </b>- to allow efficient storage and seamless propagation of provenance as the data is transferred across applications</p> <p>(d) Provenance <b>query </b>- to support queries with increasing complexity over large data size and also support knowledge discovery applications</p> <p>We apply the SPF to two exemplar translational research projects, namely the Semantic Problem Solving Environment for <it>Trypanosoma cruzi </it>(<it>T.cruzi </it>SPSE) and the Biomedical Knowledge Repository (BKR) project, to demonstrate its effectiveness.</p> <p>Conclusions</p> <p>The SPF provides a unified framework to effectively manage provenance of translational research data during pre and post-publication phases. This framework is underpinned by an upper-level provenance ontology called Provenir that is extended to create domain-specific provenance ontologies to facilitate provenance interoperability, seamless propagation of provenance, automated querying, and analysis.</p