Survey of egg farmers regarding the ban on conventional cages in the EU and their opinion of alternative layer housing systems in Flanders, Belgium.

On 1 January 2012, conventional cages for laying hens were banned in the European Union (EU); all egg farmers must now use alternative hen housing systems. In total, 218 Flemish egg farmers were surveyed in 2013 to 2014 regarding which housing systems they currently use, their degree of satisfaction with the system, and how they experienced the transition from conventional cages to an alternative system. The response rate was 58.3% (127 respondents). Of these, 43 (33.9%) were no longer active as an egg farmer, mainly due to the ban on conventional cages. The respondents who were active as egg farmers both before and after the transition (84, 66.1%) mainly judged the ban as negative for their own finances and for the competitive position of the Belgian egg industry, but were neutral or positive regarding the general consequences for their own business. Most respondents' hens were housed in either aviary systems (47.7%) or in alternative cage systems (38.2%). When choosing a new system, the fit into the farm and consumer demand were the most important factors. Consumer demand was the main reason for choosing a system with free-range access. In general, egg farmers were satisfied with the system they chose, although this differs between systems. When asked to compare the alternative systems to conventional cages, alternatives were judged to be better for hen welfare and consumer demand, but similar or worse for all other aspects, especially labor. Egg farmers previously using conventional cages judged alternative systems more negatively than those who had no prior experience with conventional cages. Farmers who had experience with free-range systems judged these more positively than those without this experience, e.g., for egg consumer demand, profitability, and hen welfare. These results can possibly be extrapolated to other EU countries in which conventional cages were the most common housing system until 2012, and lessons can be drawn from the farmers' experiences when implementing other animal welfare legislation that may require similar far-reaching adaptations for primary production.

Ideal versus corrected body weight for dosage of sugammadex in morbidly obese patients.

To date, the dosing of sugammadex is based on real body weight without taking fat content into account. We compared the reversal of profound rocuronium-induced neuromuscular blockade in morbidly obese patients using doses of sugammadex based on four different weight corrections. One hundred morbidly obese patients, scheduled for laparoscopic bariatric surgery under propofol-sufentanil anaesthesia, were randomly assigned four groups: ideal body weight; ideal body weight + 20%; ideal body weight + 40%; and real body weight. Patients received sugammadex 2 mg.kg(-1), when adductor pollicis monitoring showed two responses. The primary endpoint was full decurarisation. Secondary endpoints were the ability to get into bed independently on arrival to the post-anaesthetic care unit and clinical signs of residual paralysis. There was no residual paralysis in any patient. Morbidly obese patients can safely be decurarised from rocuronium-induced neuromuscular blockade T1-T2 with sugammadex dosed at 2 mg.kg(-1) ideal body weight + 40% (p < 0.0001).

Survey of egg producers on the introduction of alternative housing systems for laying hens in Flanders, Belgium.

In the context of the European Union ban on battery cages by 2012, a survey was conducted among Flemish egg producers (60% response rate, 140 completed questionnaires) about the introduction and opinion of alternative housing systems. Belgium appears to be among the countries in the European Union that are slower to adopt alternative housing. Belgium's egg industry is thus likely to undergo drastic changes to comply with the 2012 deadline. As of 2010, the battery cage was the dominant housing system (56% housing units, 67% hens), followed by floor housing (33% housing units, 15% hens) and aviary (10% housing units, 15% hens), whereas colony cages and furnished cages were extremely rare. Future- and market-oriented production was the most important reason for choosing a certain type of alternative system, although the importance of hen performance and amount of labor seemed to increase. A quarter of the producers with battery cages had detailed plans to convert to an alternative system (most planned to install aviaries, followed by colony cages, furnished cages, and floor housing) by 2012. Many older farmers indicated that they would stop farming, whereas others found it more profitable to delay the conversion as long as possible. Apart from hen welfare, producers expressed a negative opinion (relative to battery cages) about noncage systems and, to a lesser extent, furnished cages. However, users of alternative systems reported being quite satisfied, except for the amount of labor and hen health. The housing system had several effects on user satisfaction: positive effect of flock size, negative effect of experience with battery cages, and negative effect of outdoor area on hen health. Although not all opinions were supported by evidence, such surveys provide feedback about the success of alternative systems in practice. This information is valuable to further improve these systems and to producers who have yet to convert. Moreover, producer attitude may determine the extent to which legally imposed changes in husbandry environment result in the desired improvement of hen welfare in practice.

Digital hearing aids and future directions for hearing aids.

In recent years many technical evolutions have been applied in hearing aids. In this paper differences between analog, programmable and fully digital hearing aids, the basic and supplementary functions of a hearing aid, and some important issues and future directions for digital hearing aids will be mentioned.

A systematic gene expression screen of Caenorhabditis elegans cytochrome P450 genes reveals CYP35 as strongly xenobiotic inducible.

The soil nematode Caenorhabditis elegans is one of the simplest animals having the status of a laboratory model. Its genome contains 80 cytochrome P450 genes (CYP). In order to study CYP gene expression in C. elegans mixed stages and synchronized hermaphrodites were exposed to 18 known xenobiotic cytochrome P450 inducers. Messenger RNA expression was detected by DNA arrays and semiquantitative RT-PCR. Using subfamily-specific primers, a pooled set of exon-rich CYP fragments could be amplified. In this way it was possible to systematically check the influence of different inducers on CYP expression at the same time. The well-known CYP1A inducers beta-naphthoflavone, PCB52, and lansoprazol were the most active and in particular they strongly induced almost all CYP35 isoforms. A few number of further CYP forms were found to be inducible by other xenobiotics like phenobarbital, atrazine, and clofibrate. In addition, a transgenic C. elegans line expressing GFP under control of the CYP35A2 promoter showed a strong induction of the fusion by beta-naphthoflavone in the intestine.

The sarco-endoplasmic reticulum Ca2+ ATPase is required for development and muscle function in Caenorhabditis elegans.

The sarco-endoplasmic reticulum Ca(2+)-transport ATPase (SERCA) loads intracellular releasable Ca(2+) stores by transporting cytosolic Ca(2+) into the endoplasmic (ER) or sarcoplasmic reticulum (SR). We characterized the only SERCA homologue of the nematode Caenorhabditis elegans, which is encoded by the sca-1 gene. The sca-1 transcript is alternatively spliced in a similar mode as the vertebrate SERCA2 transcript, giving rise to two protein variants: CeSERCAa and CeSERCAb. These proteins showed structural and functional conservation to the vertebrate SERCA2a/b proteins. The CeSERCAs were primarily expressed in contractile tissues. Loss of CeSERCA through gene ablation or RNA interference resulted in contractile dysfunctioning and in early larval or embryonic lethality, respectively. Similar defects could be induced pharmacologically using the SERCA-specific inhibitor thapsigargin, which bound CeSERCA at a conserved site. The conservation of SERCA2 homologues in C. elegans will allow genetic and chemical suppressor analyses to identify promising drug targets and lead molecules for treatment of SERCA-related diseases such as heart disease.

Identification and characterization of a dimerization domain in CED-6, an adapter protein involved in engulfment of apoptotic cells.

Phagocytosis of apoptotic cells is a key step in the completion of programmed cell death that occurs throughout life in multicellular organisms. The molecular events involved in clearance of apoptotic cells are just beginning to be elucidated. Recently, CED-6, an adapter protein involved in engulfment has been cloned in Caenorhabditis elegans and in humans. CED-6 is composed of a phosphotyrosine-binding (PTB) domain and a proline-rich C-terminal domain with no apparent catalytic domain. Since PTB domains, originally identified in Shc, mediate intracellular signaling downstream of cell surface receptors, CED-6 has also been proposed to mediate intracellular signals leading to engulfment. In this report, we demonstrate that CED-6 dimerizes through a leucine zipper domain that is immediately adjacent to the PTB domain. Several lines of evidence based on co-immunoprecipitation studies, yeast two-hybrid assays, and gel filtration studies suggest that CED-6 exists as a dimer in vivo. Through mutational analyses, we show that the leucine zipper is necessary and sufficient for CED-6 dimerization and that this dimerization is conserved among C. elegans, rodent, and human CED-6 proteins. We propose that dimerization may have unique implications for ligand binding via CED-6 and its function during the phagocytosis of apoptotic cells.

The human homologue of Caenorhabditis elegans CED-6 specifically promotes phagocytosis of apoptotic cells.

A key feature of the process of programmed cell death (apoptosis) is the efficiency with which the dying cells are recognized and engulfed by phagocytes [1]. Apoptotic cells are rapidly cleared either by neighbouring cells acting as semi-professional phagocytes or by experts of the macrophage line, so that an inflammatory response is avoided [2]. The Caenorhabditis elegans gene ced-6 is required for efficient engulfment of apoptotic cells [3] and is one of a group of genes that define two partially redundant parallel pathways for the engulfment process [4] [5]. These pathways may be conserved across evolution, as two other engulfment genes have human homologues. A CED-5 homologue is part of a human CrkII-DOCK180-Rac signaling pathway proposed to mediate cytoskeletal reorganization [6] [7] [8] and a CED-7 homologue is similar to the ABC transporters [9] [10]. Here, we report the cloning and characterization of human CED-6, a human homologue of C. elegans CED-6. The 34 kDa hCED-6 protein is expressed in most tissues, some human cancer cells, and in primary human macrophages. We developed an assay that quantitates the phagocytic activity of mammalian macrophages: the number of apoptotic cells that have been internalized is measured by the uptake of lacZ-positive apoptotic cells by adherent transgenic macrophages. The results of this assay demonstrate that overexpression of hCED-6 promotes phagocytosis only of apoptotic cells and suggest that hCED-6 is the mammalian orthologue of C. elegans CED-6 and is a part of a highly conserved pathway that specifically mediates the phagocytosis of apoptotic cells.

Developmental genetic analysis of troponin T mutations in striated and nonstriated muscle cells of Caenorhabditis elegans.

We have been investigating a set of genes, collectively called mups, that are essential to striated body wall muscle cell positioning in Caenorhabditis elegans. Here we report our detailed characterization of the mup-2 locus, which encodes troponin T (TnT). Mutants for a heat-sensitive allele, called mup-2(e2346ts), and for a putative null, called mup-2(up1), are defective for embryonic body wall muscle cell contraction, sarcomere organization, and cell positioning. Characterizations of the heat-sensitive allele demonstrate that mutants are also defective for regulated muscle contraction in larval and adult body wall muscle, defective for function of the nonstriated oviduct myoepithelial sheath, and defective for epidermal morphogenesis. We cloned the mup-2 locus and its corresponding cDNA. The cDNA encodes a predicted 405-amino acid protein homologous to vertebrate and invertebrate TnT and includes an invertebrate-specific COOH-terminal tail. The mup-2 mutations lie within these cDNA sequences: mup-2(up1) is a termination codon near NH2 terminus (Glu94) and mup-2(e2346ts) is a termination codon in the COOH-terminal invertebrate-specific tail (Trp342). TnT is a muscle contractile protein that, in association with the thin filament proteins tropomyosin, troponin I and troponin C, regulates myosin-actin interaction in response to a rise in intracellular Ca2+. Our findings demonstrate multiple essential functions for TnT and provide a basis to investigate the in vivo functions and protein interactions of TnT in striated and nonstriated muscles.

Generation of structural and functional diversity in furin-like proteins in Drosophila melanogaster by alternative splicing of the Dfur1 gene.

To investigate whether or not alternative splicing might be a mechanism by which in Drosophila melanogaster diversity is generated in endoproteases of the novel eukaryotic family of subtilisin-like proprotein processing enzymes, we determined structural and functional characteristics of the Dfur1 gene. Northern blot analysis revealed Dfur1 transcripts of 7.6, 6.5, 4.5 and 4.0 kb. By comparative nucleotide sequence analysis of Dfur1 genomic and cDNA clones, 10 coding exons were identified and, together with Northern blot analysis using exon-specific probes, evidence was obtained that the four transcripts are generated by alternative splicing and polyadenylation. The apparently complete open reading frames of three Dfur1 cDNAs revealed that these coded for three furin-like proteins, Dfurin1 (892 residues), Dfurin1-CRR (1101 residues) and Dfurin1-X (1269 residues), which possessed common but also unique structural domains. These various isoforms of furin in Drosophila were characterized in gene transfer studies using immunoprecipitation analysis. Differential expression of Dfur1 transcripts was found in Northern blot analysis of RNA from various developmental stages of Drosophila. RNA in situ hybridization experiments revealed that the Dfurin1-X and Dfurin1-CRR isoforms are expressed in non-overlapping sets of tissues during Drosophila embryogenesis. In gene transfer experiments in which the Dfurin1, Dfurin1-CRR and Dfurin1-X proteins were expressed at high levels together with the precursor of the beta A-chain of activin-A, a member of the transforming growth factor beta (TGF beta) superfamily, or the precursor of von Willebrand factor, all three proteins appeared capable of processing these substrates. Our studies indicate that the Dfur1 gene encodes structurally different subtilisin-like proprotein processing enzymes with distinct physiological functions in Drosophila.

Positioning and maintenance of embryonic body wall muscle attachments in C. elegans requires the mup-1 gene.

As part of a general study of genes specifying a pattern of muscle attachments, we identified and genetically characterised mutants in the mup-1 gene. The body wall muscles of early stage mup-1 embryos have a wild-type myofilament pattern but may extend ectopic processes. Later in embryogenesis, some body wall muscles detach from the hypodermis. Genetic analysis suggests that mup-1 has both a maternal and a zygotic component and is not required for postembryonic muscle growth and attachment. mup-1 mutants are suppressed by mutations in several genes that encode extracellular matrix components. We propose that mup-1 may encode a cell surface/extracellular matrix molecule required both for the positioning of body wall muscle attachments in early embryogenesis and the subsequent maintenance of these attachments to the hypodermis until after cuticle synthesis.

The function of PS integrins during Drosophila embryogenesis.

The Drosophila position-specific (PS) antigens are homologous to the vertebrate fibronectin receptor family, or integrins. A Drosophila gene required for embryonic morphogenesis, l(1)myospheroid, codes for a product homologous to the beta subunit of the vertebrate integrins. l(1)myospheroid mutants die during embryogenesis. We show here that they lack the beta subunit of the PS antigens. In the absence of the beta subunit in mutant embryos, the PS alpha subunits are not expressed on the cell surface. We conclude that the l(1)myospheroid phenotype represents the lack-of-function phenotype for these Drosophila integrins. In wild-type embryos, PS antigens are found at the interface between mesoderm and ectoderm, and later mainly at the attachment sites of muscles to the epidermis and gut. Together these results indicate that during embryogenesis, Drosophila integrins are used to attach mesoderm to ectoderm, and are required for the proper assembly of the extracellular matrix and for muscle attachment.

The Drosophila PS2 antigen is an invertebrate integrin that, like the fibronectin receptor, becomes localized to muscle attachments.

We establish that the position-specific antigen 2 (PS2), a Drosophila cell surface glycoprotein complex, is an invertebrate member of the vertebrate fibronectin receptor (integrin) family. New monoclonal antibodies show that in Drosophila embryos and larvae PS2 alpha subunits have a size of ca. 140 kd. Analysis of cDNA and genomic clones revealed that the canonical PS2 alpha subunit contains 1394 amino acids and has extensive homology to the heavy and light chains of integrin alpha subunits. The distribution of the PS2 antigen is regulated at the level of PS2 alpha subunit mRNA. In early Drosophila development the protein is restricted to mesoderm and appears to be involved in muscle attachment. We suggest that PS2, like vertebrate fibronectin receptors, mediates changes in cell shape and cell-extracellular matrix adhesion by binding to a basement membrane protein.