Transgenic production from in vivo-derived embryos: effect on calf birth weight and sex ratio

We examined transgenic-cattle production by DNA microinjection into 1-, 2-, and 4-cell embryos, analyzing the impact on calf size and subsequent viability. Embryos were either collected at an abattoir by flushing oviducts from superovulated and artificially inseminated cows (in vivo-derived) or obtained by in vitro maturation and in vitro fertilization of oocytes aspirated from excised ovaries (in vitro-derived). A human serum albumin (hSA) milk-expression DNA construct was microinjected, either in one of the visible pronuclei of in vitro- and in vivo-derived 1-cell embryos or in the nuclei of two blastomeres of 2- and 4-cell in vivo-derived embryos. Microinjection-induced mortality (lysis and developmental block) was equivalent ( approximately 40%) for all microinjected embryos. Embryos were co-cultured with BRL cells in B-2 medium containing 10% fetal calf serum (FSC). Overall, embryo development to morulae/blastocysts was significantly greater for in vivo-derived ova (15.5%) than for in vitro-derived oocytes (9.3%). All morulae and blastocysts were transferred to synchronized recipient females on Days 6-8 post-fertilization. A total of 189 calves were delivered. Birth weights were significantly greater for calves generated from in vitro-derived oocytes compared with those generated from in vivo-derived oocytes. One transgenic bull calf was obtained from the microinjection of a 2-cell embryo. Fluorescence in situ hybridization (FISH) analysis of lymphocytes detected one transgenic integration site in all cells. Transmission frequency of the hSA transgene in embryos obtained through IVM/IVF/IVC utilizing the semen of the transgenic calf confirmed that it was not mosaic.     

Quality of life and disease-specific functional status following microvascular reconstruction for advanced (T3 and T4) oropharyngeal cancers

In an effort to evaluate quality-of-life benefits of ablative head and neck cancer surgery and microvascular reconstruction, a longitudinal study was undertaken in which patients with T3 or T4 oropharyngeal cancers without systemic metastases at presentation were administered both general and disease-specific quality-of-life instruments preoperatively and postoperatively. In an initial prospective pilot study, 17 cancer patients were evaluated both preoperatively and postoperatively using the Medical Outcomes Short-Form Health Survey questionnaire (SF-36) and the Performance Status Scale for Head and Neck Cancer Patients. In the second part of the study, the need was recognized for a different disease-specific measure, for more frequent intervals of longitudinal follow-up (rather than be limited by a single data collection point), and for a noncancer control group. Since then, 17 more cancer patients were evaluated in the second part of the study and were compared with patients who had similar reconstructions after suffering head and neck trauma and also with age-matched controls. Instead of the performance status scale, the University of Washington Head and Neck Quality of Life questionnaire was substituted. Interval assessments were done at 1, 3, 6, and 12 months and preoperatively. Whereas many of the general and disease-specific quality of life subclasses initially worsened following extensive surgery and radiation therapy, most returned to the preoperative baseline by 6 months following conclusion of treatment and surpassed pretreatment values at 1 year. It can be concluded, based on this study, that large resections and reconstructions for head and neck cancer patients are justified in terms of outcome; the resection controls the local disease, and the microvascular reconstruction restores quality of life and functional status.     

Phosphorylation of ganciclovir phosphonate by cellular GMP kinase determines the stereoselectivity of anti-human cytomegalovirus activity

A racemic mixture of ganciclovir phosphonate was resolved by stereoselective phosphorylation using GMP kinase. The R-enantiomer of ganciclovir phosphonate was active against human cytomegalovirus but the S-enantiomer was less active. We show that enantiomeric selectivity of antiviral for ganciclovir phosphonate was conferred by stereoselective phosphorylations by mammalian enzymes, not by stereoselective inhibition of DNA polymerase from human cytomegalovirus.     

Determination of who may derive most benefit from aspirin in primary prevention: subgroup results from a randomised controlled trial

ObjectiveTo determine which groups of patients may derive particular benefit or experience harm from the use of low dose aspirin for the primary prevention of coronary heart disease.DesignRandomised controlled trial.Setting108 group practices in the Medical Research Council''s general practice research framework who were taking part in the thrombosis prevention trial.Participants5499 men aged between 45 and 69 years at entry who were at increased risk of coronary heart disease.ResultsAspirin reduced coronary events by 20%. This benefit, mainly for non-fatal events, was significantly greater the lower the systolic blood pressure at entry (interaction P=0.0015), the relative risk at pressures 130 mm Hg being 0.55 compared with 0.94 at pressures >145 mm Hg. Aspirin also reduced strokes at low but not high pressures, the relative risks being 0.41 and 1.42 (P=0.006) respectively. The relative risk of all major cardiovascular events—that is, the sum of coronary heart disease and stroke—was 0.59 at pressures <130 mm Hg compared with 1.08 at pressures >145 mm Hg (P=0.0001).ConclusionEven with the limitations of subgroup analyses the evidence suggests that the benefit of low dose aspirin in primary prevention may occur mainly in those with lower systolic blood pressures, although it is not clear even in these men that the benefit outweighs the potential hazards. Men with higher pressures may be exposed to the risks of bleeding while deriving no benefit through reductions in coronary heart disease and stroke.     

Proteus mirabilis interkingdom swarming signals attract blow flies

Flies transport specific bacteria with their larvae that provide a wider range of nutrients for those bacteria. Our hypothesis was that this symbiotic interaction may depend on interkingdom signaling. We obtained Proteus mirabilis from the salivary glands of the blow fly Lucilia sericata; this strain swarmed significantly and produced a strong odor that attracts blow flies. To identify the putative interkingdom signals for the bacterium and flies, we reasoned that as swarming is used by this bacterium to cover the food resource and requires bacterial signaling, the same bacterial signals used for swarming may be used to communicate with blow flies. Using transposon mutagenesis, we identified six novel genes for swarming (ureR, fis, hybG, zapB, fadE and PROSTU_03490), then, confirming our hypothesis, we discovered that fly attractants, lactic acid, phenol, NaOH, KOH and ammonia, restore swarming for cells with the swarming mutations. Hence, compounds produced by the bacterium that attract flies also are utilized for swarming. In addition, bacteria with the swarming mutation rfaL attracted fewer blow flies and reduced the number of eggs laid by the flies. Therefore, we have identified several interkingdom signals between P. mirabilis and blow flies.     

Amyloid fibrils as functionalizable components of nanocomposite materials

Amyloid fibrils are a form of protein nanofiber that show promise as components of multifunctional bionanomaterials. In this work, native bovine insulin and bovine insulin that had been previously converted into amyloid fibrils were combined with poly(vinyl alcohol) (PVOH) via solution casting to determine the effect of fibrillization on the thermomechanical properties of the resulting composite. The synthesis method was found to preserve the amyloid fibril structure and properties of the resulting fibril-PVOH composite were investigated. At a filling level of 0.6 wt %, the fibril-reinforced PVOH was 15% stiffer than the PVOH control. Various properties of the films, including the glass transition temperature, degradation temperature, microstructure, and film morphology were characterized. Although more work is required to optimize the properties of the composites, this study provides proof of principle that incorporation of amyloid fibrils into a polymeric material can impart useful changes to the mechanical and morphological properties of the films.     

Inhibition of platelet adhesion by peptidomimetics mimicking the interactive β-hairpin of glycoprotein Ibα

β-Hairpin peptidomimetics mimicking the interaction sites of the platelet receptor glycoprotein (GP)Ibα with von Willebrand factor (vWF) were synthesised and evaluated for their ability to increase platelet velocity under high shear conditions and to inhibit shear-induced platelet aggregation. A cyclic and bridged dodecapeptide 2e containing a heterochiral diproline motif was identified as a lead compound for the generation of a novel class of potential antiplatelet agents.     

The slow road to the eukaryotic genome

The eukaryotic genome is a mosaic of eubacterial and archaeal genes in addition to those unique to itself. The mosaic may have arisen as the result of two prokaryotes merging their genomes, or from genes acquired from an endosymbiont of eubacterial origin. A third possibility is that the eukaryotic genome arose from successive events of lateral gene transfer over long periods of time. This theory does not exclude the endosymbiont, but questions whether it is necessary to explain the peculiar set of eukaryotic genes. We use phylogenetic studies and reconstructions of ancestral first appearances of genes on the prokaryotic phylogeny to assess evidence for the lateral gene transfer scenario. We find that phylogenies advanced to support fusion can also arise from a succession of lateral gene transfer events. Our reconstructions of ancestral first appearances of genes reveal that the various genes that make up the eukaryotic mosaic arose at different times and in diverse lineages on the prokaryotic tree, and were not available in a single lineage. Successive events of lateral gene transfer can explain the unusual mosaic structure of the eukaryotic genome, with its content linked to the immediate adaptive value of the genes its acquired. Progress in understanding eukaryotes may come from identifying ancestral features such as the eukaryotic splicesome that could explain why this lineage invaded, or created, the eukaryotic niche.