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791.
James G. Burchfield Jinling Lu Daniel J. Fazakerley Shi‐Xiong Tan Yvonne Ng Katarina Mele Michael J. Buckley William E. Hughes David E. James 《Traffic (Copenhagen, Denmark)》2013,14(3):259-273
Regulated GLUT4 trafficking is a key action of insulin. Quantitative stepwise analysis of this process provides a powerful tool for pinpointing regulatory nodes that contribute to insulin regulation and insulin resistance. We describe a novel GLUT4 construct and workflow for the streamlined dissection of GLUT4 trafficking; from simple high throughput screens to high resolution analyses of individual vesicles. We reveal single cell heterogeneity in insulin action highlighting the utility of this approach – each cell displayed a unique and highly reproducible insulin response, implying that each cell is hard‐wired to produce a specific output in response to a given stimulus. These data highlight that the response of a cell population to insulin is underpinned by extensive heterogeneity at the single cell level. This heterogeneity is pre‐programmed within each cell and is not the result of intracellular stochastic events. 相似文献
792.
Maria Christina Dzul Stephen James Dinsmore Michael Carl Quist Daniel Bailey Gaines Kevin Patrick Wilson Michael Roy Bower Philip Michael Dixon 《Population Ecology》2013,55(2):325-341
The Devils Hole pupfish, Cyprinodon diabolis, is a federally-endangered fish that is endemic to Devils Hole, a discontiguous part of Death Valley National Park in Nye County, Nevada. Due to its status, Devils Hole pupfish monitoring must be non-obtrusive and thereby exclude techniques that require handling fish. Due to a recent decline in pupfish abundance, Devils Hole pupfish managers have expressed a need for a model that describes population dynamics. This population model would be used to identify vulnerable life history stage(s) and inform management actions. We constructed a set of individual-based simulation models designed to explore effects of population processes and evaluate assumptions. We developed a baseline model, whose output best resembled both observed length-frequency data and predicted intra-annual abundance patterns. We then ran simulations with 5 % increases in egg-larval, juvenile, and adult survival rates to better understand Devils Hole pupfish life history, thereby helping identify vulnerable life history stages that should become the target of management actions. Simulation models with temporally constant adult, juvenile, and egg-larval survival rates were able to reproduce observed length-frequency distributions and predicted intra-annual population patterns. In particular, models with monthly adult and juvenile survival rates of 80 % and an egg-larval survival rate of 4.7 % replicated patterns in observed data. Population growth was most affected by 5 % increases in egg-larval survival, whereas adult and juvenile survival rates had similar but lesser effects on population growth. Outputs from the model were used to assess factors suspected of influencing Devils Hole pupfish population decline. 相似文献
793.
Esther García‐Fernández Daniel J. Frank Beatriz Galán Petrea M. Kells Larissa M. Podust José L. García Paul R. Ortiz de Montellano 《Environmental microbiology》2013,15(8):2342-2359
Degradation of the cholesterol side‐chain in Mycobacterium tuberculosis is initiated by two cytochromes P450, CYP125A1 and CYP142A1, that sequentially oxidize C26 to the alcohol, aldehyde and acid metabolites. Here we report characterization of the homologous enzymes CYP125A3 and CYP142A2 from Mycobacterium smegmatis mc2 155. Heterologously expressed, purified CYP125A3 and CYP142A2 bound cholesterol, 4‐cholesten‐3‐one, and antifungal azole drugs. CYP125A3 or CYP142A2 reconstituted with spinach ferredoxin and ferredoxin reductase efficiently hydroxylated 4‐cholesten‐3‐one to the C‐26 alcohol and subsequently to the acid. The X‐ray structures of both substrate‐free CYP125A3 and CYP142A2 and of cholest‐4‐en‐3‐one‐bound CYP142A2 reveal significant differences in the substrate binding sites compared with the homologous M. tuberculosis proteins. Deletion only of cyp125A3 causes a reduction of both the alcohol and acid metabolites and a strong induction of cyp142 at the mRNA and protein levels, indicating that CYP142A2 serves as a functionally redundant back up enzyme for CYP125A3. In contrast to M. tuberculosis, the M. smegmatis Δcyp125Δcyp142 double mutant retains its ability to grow on cholesterol albeit with a diminished capacity, indicating an additional level of redundancy within its genome. 相似文献
794.
Javier Megías Victoria Maneu Pedro Salvador Daniel Gozalbo M. Luisa Gil 《Cellular microbiology》2013,15(7):1143-1153
Toll‐like receptors (TLRs) are expressed by haematopoietic stem and progenitor cells (HSPCs), and may play a role in haematopoiesis in response to pathogens during infection. We have previously demonstrated that (i) inactivated yeasts of Candida albicans induce in vitro differentiation of HSPCs towards the myeloid lineage, and (ii) soluble TLR agonists induce in vivo their differentiation towards macrophages. In this work, using an in vivo model of HSPCs transplantation, we report for the first time that HSPCs sense C. albicans in vivo and subsequently are directed to produce macrophages by a TLR2‐dependent signalling. Purified lineage‐negative cells (Lin?) from bone marrow of C57BL/6 mice (CD45.2 alloantigen) were transplanted into B6Ly5.1 mice (CD45.1 alloantigen), which were then injected with viable or inactivated C. albicans yeasts. Transplanted cells were detected in the spleen and in the bone marrow of recipient mice, and they differentiate preferentially to macrophages, both in response to infection or in response to inactivated yeasts. The generation of macrophages was dependent on TLR2 but independent of TLR4, as transplanted Lin? cells from TLR2?/? mice did not give rise to macrophages, whereas Lin? cells from TLR4?/? mice generated macrophages similarly to control cells. Interestingly, the absence of TLR2, or in a minor extent TLR4, gives Lin? cells an advantage in transplantation assays, as increases the percentage of transplanted recovered cells. Our results indicatethat TLR‐mediated recognition of C. albicans by HSPCs may help replace and/or increase cells that constitute the first line of defence against the fungus, and suggest that TLR‐mediated signalling may lead to reprogramming early progenitors to rapidly replenishing the innate immune system and generate the most necessary mature cells to deal with the pathogen. 相似文献
795.
Todd J Treangen Sergey Koren Daniel D Sommer Bo Liu Irina Astrovskaya Brian Ondov Aaron E Darling Adam M Phillippy Mihai Pop 《Genome biology》2013,14(1):R2
We describe MetAMOS, an open source and modular metagenomic assembly and analysis pipeline. MetAMOS represents an important step towards fully automated metagenomic analysis, starting with next-generation sequencing reads and producing genomic scaffolds, open-reading frames and taxonomic or functional annotations. MetAMOS can aid in reducing assembly errors, commonly encountered when assembling metagenomic samples, and improves taxonomic assignment accuracy while also reducing computational cost. MetAMOS can be downloaded from: https://github.com/treangen/MetAMOS. 相似文献
796.
Ants are powerful model systems for the study of cooperation and sociality. In this review, we discuss how recent advances in ant genomics have contributed to our understanding of the evolution and organization of insect societies at the molecular level. 相似文献
797.
Mohamed Abou-El-Enein Andy Römhild Daniel Kaiser Carola Beier Gerhard Bauer Hans-Dieter Volk Petra Reinke 《Cytotherapy》2013,15(3):362-383
Background aimsAdvanced therapy medicinal products (ATMP) have gained considerable attention in academia due to their therapeutic potential. Good Manufacturing Practice (GMP) principles ensure the quality and sterility of manufacturing these products. We developed a model for estimating the manufacturing costs of cell therapy products and optimizing the performance of academic GMP-facilities.MethodsThe “Clean-Room Technology Assessment Technique” (CTAT) was tested prospectively in the GMP facility of BCRT, Berlin, Germany, then retrospectively in the GMP facility of the University of California-Davis, California, USA. CTAT is a two-level model: level one identifies operational (core) processes and measures their fixed costs; level two identifies production (supporting) processes and measures their variable costs. The model comprises several tools to measure and optimize performance of these processes. Manufacturing costs were itemized using adjusted micro-costing system.ResultsCTAT identified GMP activities with strong correlation to the manufacturing process of cell-based products. Building best practice standards allowed for performance improvement and elimination of human errors. The model also demonstrated the unidirectional dependencies that may exist among the core GMP activities. When compared to traditional business models, the CTAT assessment resulted in a more accurate allocation of annual expenses. The estimated expenses were used to set a fee structure for both GMP facilities. A mathematical equation was also developed to provide the final product cost.ConclusionsCTAT can be a useful tool in estimating accurate costs for the ATMPs manufactured in an optimized GMP process. These estimates are useful when analyzing the cost-effectiveness of these novel interventions. 相似文献
798.
799.
Tremendous efforts have been made over the past few decades to discover novel cancer biomarkers for use in clinical practice. However, a striking discrepancy exists between the effort directed toward biomarker discovery and the number of markers that make it into clinical practice. One of the confounding issues in translating a novel discovery into clinical practice is that quite often the scientists working on biomarker discovery have limited knowledge of the analytical, diagnostic, and regulatory requirements for a clinical assay. This review provides an introduction to such considerations with the aim of generating more extensive discussion for study design, assay performance, and regulatory approval in the process of translating new proteomic biomarkers from discovery into cancer diagnostics. We first describe the analytical requirements for a robust clinical biomarker assay, including concepts of precision, trueness, specificity and analytical interference, and carryover. We next introduce the clinical considerations of diagnostic accuracy, receiver operating characteristic analysis, positive and negative predictive values, and clinical utility. We finish the review by describing components of the FDA approval process for protein-based biomarkers, including classification of biomarker assays as medical devices, analytical and clinical performance requirements, and the approval process workflow. While we recognize that the road from biomarker discovery, validation, and regulatory approval to the translation into the clinical setting could be long and difficult, the reward for patients, clinicians and scientists could be rather significant. 相似文献
800.
Nicolas Rochereau Daniel Drocourt Eric Perouzel Vincent Pavot Pierre Redelinghuys Gordon D. Brown Gerard Tiraby Xavier Roblin Bernard Verrier Christian Genin Blaise Corthésy Stéphane Paul 《PLoS biology》2013,11(9)
Intestinal microfold (M) cells possess a high transcytosis capacity and are able to transport a broad range of materials including particulate antigens, soluble macromolecules, and pathogens from the intestinal lumen to inductive sites of the mucosal immune system. M cells are also the primary pathway for delivery of secretory IgA (SIgA) to the gut-associated lymphoid tissue. However, although the consequences of SIgA uptake by M cells are now well known and described, the mechanisms whereby SIgA is selectively bound and taken up remain poorly understood. Here we first demonstrate that both the Cα1 region and glycosylation, more particularly sialic acid residues, are involved in M cell–mediated reverse transcytosis. Second, we found that SIgA is taken up by M cells via the Dectin-1 receptor, with the possible involvement of Siglec-5 acting as a co-receptor. Third, we establish that transcytosed SIgA is taken up by mucosal CX3CR1+ dendritic cells (DCs) via the DC-SIGN receptor. Fourth, we show that mucosal and systemic antibody responses against the HIV p24-SIgA complexes administered orally is strictly dependent on the expression of Dectin-1. Having deciphered the mechanisms leading to specific targeting of SIgA-based Ag complexes paves the way to the use of such a vehicle for mucosal vaccination against various infectious diseases. 相似文献