Reducing overweight through a multidisciplinary school-based intervention

Objective: This study sought to evaluate the effectiveness of a multidisciplinary elementary school—based intervention entitled Wellness, Academics & You. The primary areas of impact examined were BMI, consumption of fruits and vegetables, and physical activity. Research Methods and Procedures: The sample consisted of 1013 students in fourth and fifth grades from 69 classes in four states. Intervention and comparison classes were randomly selected at each school. Intervention teachers participated in workshops on the intervention and received program materials. BMI was calculated for baseline and post‐data points based on measured height and weight collected by trained research teams. Baseline surveys were administered by the students’ teacher in the presence of one of the research team members and collected. Results: SPSS (SPSS, Inc., Chicago, IL) analysis of post‐data shows significant positive shifts (p = 0.01) in BMI in the intervention group compared with the comparison group. Notable increases in the consumption of fruits and vegetables and increased physical activity levels were reported in the intervention group. Discussion: Data from this study indicate that this school‐based program may contribute to the reduction of overweight and obesity. The intervention was well received by the classroom teachers and shows promise for a school‐based obesity prevention initiative. This program has shown the capacity to effect positive changes in BMI through consumption of fruits and vegetables and physical activity. The evaluation covered a limited time period and, therefore, warrants additional studies to determine whether long‐term program fidelity will result in continued improvement.     

Application of the accurate mass and time tag approach to the proteome analysis of sub-cellular fractions obtained from Rhodobacter sphaeroides 2.4.1. Aerobic and photosynthetic cell cultures

The high-throughput accurate mass and time (AMT) tag proteomic approach was utilized to characterize the proteomes for cytoplasm, cytoplasmic membrane, periplasm, and outer membrane fractions from aerobic and photosynthetic cultures of the gram-nagtive bacterium Rhodobacter sphaeroides 2.4.1. In addition, we analyzed the proteins within purified chromatophore fractions that house the photosynthetic apparatus from photosynthetically grown cells. In total, 8,300 peptides were identified with high confidence from at least one subcellular fraction from either cell culture. These peptides were derived from 1,514 genes or 35% percent of proteins predicted to be encoded by the genome. A significant number of these proteins were detected within a single subcellular fraction and their localization was compared to in silico predictions. However, the majority of proteins were observed in multiple subcellular fractions, and the most likely subcellular localization for these proteins was investigated using a Z-score analysis of estimated protein abundance along with clustering techniques. Good (81%) agreement was observed between the experimental results and in silico predictions. The AMT tag approach provides localization evidence for those proteins that have no predicted localization information, those annotated as putative proteins, and/or for those proteins annotated as hypothetical and conserved hypothetical.     

High dynamic range characterization of the trauma patient plasma proteome

Although human plasma represents an attractive sample for disease biomarker discovery, the extreme complexity and large dynamic range in protein concentrations present significant challenges for characterization, candidate biomarker discovery, and validation. Herein we describe a strategy that combines immunoaffinity subtraction and subsequent chemical fractionation based on cysteinyl peptide and N-glycopeptide captures with two-dimensional LC-MS/MS to increase the dynamic range of analysis for plasma. Application of this "divide-and-conquer" strategy to trauma patient plasma significantly improved the overall dynamic range of detection and resulted in confident identification of 22,267 unique peptides from four different peptide populations (cysteinyl peptides, non-cysteinyl peptides, N-glycopeptides, and non-glycopeptides) that covered 3,654 different proteins with 1,494 proteins identified by multiple peptides. Numerous low abundance proteins were identified, exemplified by 78 "classic" cytokines and cytokine receptors and by 136 human cell differentiation molecules. Additionally a total of 2,910 different N-glycopeptides that correspond to 662 N-glycoproteins and 1,553 N-glycosylation sites were identified. A panel of the proteins identified in this study is known to be involved in inflammation and immune responses. This study established an extensive reference protein database for trauma patients that provides a foundation for future high throughput quantitative plasma proteomic studies designed to elucidate the mechanisms that underlie systemic inflammatory responses.     

Convergent evolution of clamp-like binding sites in diverse chaperones

Molecular chaperones have evolved diverse tertiary and quaternary structures to stabilize non-native polypeptides and facilitate their transition to the native state. Indeed, different families of chaperones lack sequence similarity, and few are represented ubiquitously in all three domains of life. Despite their discrete evolutionary paths, recent crystal structures reveal that many chaperones use seemingly convergent strategies to bind non-native proteins. This crystallographic evidence shows, or strongly suggests, that chaperones including prefoldin, Skp, trigger factor, Hsp40 and Hsp90 have clamp-like structural features used to grip substrate proteins. We explore the notion that clamp-like structures are evolutionarily favored by both ATP-dependent and ATP-independent molecular chaperones. Presumably, clamps present a multivalent binding surface ideal for protecting unstable protein conformers until they reach the native state or are transferred to another component of the folding machinery.     

Reticulate evolution in diploid and tetraploid species of Polystachya (Orchidaceae) as shown by plastid DNA sequences and low-copy nuclear genes

Background and Aims

Here evidence for reticulation in the pantropical orchid genus Polystachya is presented, using gene trees from five nuclear and plastid DNA data sets, first among only diploid samples (homoploid hybridization) and then with the inclusion of cloned tetraploid sequences (allopolyploids). Two groups of tetraploids are compared with respect to their origins and phylogenetic relationships.

Methods

Sequences from plastid regions, three low-copy nuclear genes and ITS nuclear ribosomal DNA were analysed for 56 diploid and 17 tetraploid accessions using maximum parsimony and Bayesian inference. Reticulation was inferred from incongruence between gene trees using supernetwork and consensus network analyses and from cloning and sequencing duplicated loci in tetraploids.

Key Results

Diploid trees from individual loci showed considerable incongruity but little reticulation signal when support from more than one gene tree was required to infer reticulation. This was coupled with generally low support in the individual gene trees. Sequencing the duplicated gene copies in tetraploids showed clearer evidence of hybrid evolution, including multiple origins of one group of tetraploids included in the study.

Conclusions

A combination of cloning duplicate gene copies in allotetraploids and consensus network comparison of gene trees allowed a phylogenetic framework for reticulation in Polystachya to be built. There was little evidence for homoploid hybridization, but our knowledge of the origins and relationships of three groups of allotetraploids are greatly improved by this study. One group showed evidence of multiple long-distance dispersals to achieve a pantropical distribution; another showed no evidence of multiple origins or long-distance dispersal but had greater morphological variation, consistent with hybridization between more distantly related parents.     

Optimization of square-wave electroporation for transfection of porcine fetal fibroblasts

Development of a transgenic porcine biomedical research model requires effective delivery of DNA into the donor cell followed by selection of genetically modified somatic cell lines to be used for nuclear transfer. The objective of the current study was 2-fold: (1) to compare the effectiveness of a single 1 ms pulse of different voltages (V; 100, 150, 200, 250, 300, 350) and multiple 1 ms pulses (1, 2, 3, 4 or 5) at 300 V for delivery and expression of super-coiled GFP vector in surviving cells of three fetal fibroblast cell lines, and (2) to determine the ability of these electroporation parameters to produce stably transfected fibroblast colonies following G418 selection. Cell line (P < 0.001) and voltage (P < 0.001) affected DNA delivery into the cell as assessed by GFP expression while survival at 24 h was affected by voltage (P < 0.001) and not by cell line (P = 0.797). Using a single pulse while increasing voltage resulted in the percentage of GFP expressing cells increasing from 3.2 ± 0.8% to 43.0 ± 3.4% while survival decreased from 90.5 ± 8.0% to 44.8 ± 2.0%. The number of pulses at 300 V significantly affected survival (P < 0.001) and GFP expression (P < 0.001). Survival steadily decreased following 1–5 pulses from 63.2 ± 6.3% to 3.0 ± 0.3% with GFP expression of surviving cells increasing from 35.6 ± 2.67% to 71.4 ± 6.1%. Electroporation of a selectable marker at a 1:1 copy number ratio to a co-electroporated transgene resulted in 83% of G418 resistant colonies also being PCR positive for the secondary transgene. These electroporation conditions, specifically, three 1 ms pulses of 300 V to 200 μL of 1 × 10⁶ cells/mL in the presence of 12.5 μg DNA/mL effectively introduced DNA into somatic cells. The utilization of these conditions produced numerous transgenic fibroblast colonies following G418 selection that when used for somatic cell nuclear transfer resulted in the production of live offspring.     

Synthesis,vasorelaxant activity and antihypertensive effect of benzo[d]imidazole derivatives

A series of 1H-benzo[d]imidazole analogues of Pimobendan, substituted at position 5 with either –CF₃ or –NO₂, were synthesized using a short synthetic route. All the nitro derivatives were potent, and exhibited a concentration- and partial endothelium-dependent vasorelaxant effects, with EC₅₀s <5 μM. 2-Methoxy-4-[5-nitro-1H-benzo[d]imidazol-2-yl]phenol (compound 13) was the most potent derivative of the series, showing an EC₅₀ value of 1.81 μM and E_max of 91.7% for ex vivo relaxant response in intact aortic rings, resulting in a 2.5-fold higher activity compared to Pimobendan. The closely related 5-CF₃ analogue (compound 8), was 19 times less potent than 13. The antihypertensive activity of compound 13 was evaluated at doses of 25, 50 and 100 mg kg^?1, using spontaneously hypertensive rats (SHR), showing a statistically significant dose-dependent effect.     

Spatial and temporal extinction dynamics in a freshwater cetacean

Geographical range contraction is a fundamental ecological characteristic of species population decline, but relatively little investigation has been conducted into general trends in the dynamic properties of range collapse. The Yangtze River dolphin or baiji (Lipotes vexillifer), probably the first large mammal species to have become extinct in over 50 years, was believed to have experienced major range collapse during its decline through progressive large-scale range contraction and fragmentation. This range-collapse model is challenged by a new dataset of 406 baiji last-sighting records collected from across the baiji''s historical range during an interview survey of Yangtze fishing communities. Although baiji regional abundance may have varied across its range, analyses of the extensive new sighting series provide comprehensive evidence that baiji population decline was not associated with any major contraction in geographical range across the middle–lower Yangtze drainage, even in the decade immediately before probable global extinction of the species. Extinction risk in baiji was therefore seemingly not related to evidence of range collapse. Baiji apparently underwent large-scale periodic and seasonal movements across their range, and we propose that range contraction and fragmentation may not be general biogeographic characteristics for declining populations of mobile species in connected landscapes.     

Upside-down spiders build upside-down orb webs: web asymmetry,spider orientation and running speed in Cyclosa

Almost all spiders building vertical orb webs face downwards when sitting on the hubs of their webs, and their webs exhibit an up–down size asymmetry, with the lower part of the capture area being larger than the upper. However, spiders of the genus Cyclosa, which all build vertical orb webs, exhibit inter- and intraspecific variation in orientation. In particular, Cyclosa ginnaga and C. argenteoalba always face upwards, and C. octotuberculata always face downwards, whereas some C. confusa face upwards and others face downwards or even sideways. These spiders provide a unique opportunity to examine why most spiders face downwards and have asymmetrical webs. We found that upward-facing spiders had upside-down webs with larger upper parts, downward-facing spiders had normal webs with larger lower parts and sideways-facing spiders had more symmetrical webs. Downward-facing C. confusa spiders were larger than upward- and sideways-facing individuals. We also found that during prey attacks, downward-facing spiders ran significantly faster downwards than upwards, which was not the case in upward-facing spiders. These results suggest that the spider''s orientation at the hub and web asymmetry enhance its foraging efficiency by minimizing the time to reach prey trapped in the web.