Production and impaired regulation of neutrophil extracellular traps following severe thermal injury, implications for sepsis and multiple organ failure

Advancements in burn care have improved immediate outcome, however, the prevalence of sepsis and multiple organ failure (MOF) remain significant. Although well characterised the mechanisms responsible for the pathogenesis of MOF and increased propensity to infection are poorly understood. Neutrophil extracellular traps (NETs) provide protection against invading pathogens but also contribute to thrombosis. Sepsis is required for NET generation following severe thermal injury. Quantification of circulating NET biomarkers shows good discriminatory power for diagnosis of sepsis. Interestingly, neutrophils isolated from 24 patients with severe thermal injuries, ≥ 15% total body surface area, had a significantly reduced ability to form NETs ex vivo, potentially mediated by phenotypical changes of neutrophils and inhibitory effects of formyl peptides. This thesis identified a major biological mechanism driving MOF after severe thermal injury, namely the compromise to the actin scavenging system which leads to reduced DNAse activity and a build-up of circulating DNA. Preliminary analysis suggests that DNAse activity can be restored by prehospital use of fresh frozen plasma following major trauma. Thus, administration of blood products or manipulation of the actin scavenging system is a potential therapeutic target. This thesis has identified a number of novel mechanisms responsible for the regulation of NETs following severe thermal injuries and their implications for sepsis and MOF

Traumatic injury and exposure to mitochondrial-derived damage associated molecular patterns suppresses neutrophil extracellular trap formation

Major traumatic injury induces significant remodeling of the circulating neutrophil pool and loss of bactericidal function. Although a well-described phenomenon, research to date has only analyzed blood samples acquired post-hospital admission, and the mechanisms that initiate compromised neutrophil function post-injury are therefore poorly understood. Here, we analyzed pre-hospital blood samples acquired from 62 adult trauma patients (mean age 44 years, range 19–95 years) within 1 h of injury (mean time to sample 39 min, range 13–59 min). We found an immediate impairment in neutrophil extracellular trap (NET) generation in response to phorbol 12-myristate 13-acetate (PMA) stimulation, which persisted into the acute post-injury phase (4–72 h). Reduced NET generation was accompanied by reduced reactive oxygen species production, impaired activation of mitogen-activated protein kinases, and a reduction in neutrophil glucose uptake and metabolism to lactate. Pre-treating neutrophils from healthy subjects with mitochondrial-derived damage-associated molecular patterns (mtDAMPs), whose circulating levels were significantly increased in our trauma patients, reduced NET generation. This mtDAMP-induced impairment in NET formation was associated with an N-formyl peptide mediated activation of AMP-activated protein kinase (AMPK), a negative regulator of aerobic glycolysis and NET formation. Indeed, activation of AMPK via treatment with the AMP-mimetic AICAR significantly reduced neutrophil lactate production in response to PMA stimulation, a phenomenon that we also observed for neutrophils pre-treated with mtDAMPs. Furthermore, the impairment in NET generation induced by mtDAMPs was partially ameliorated by pre-treating neutrophils with the AMPK inhibitor compound C. Taken together, our data demonstrate an immediate trauma-induced impairment in neutrophil anti-microbial function and identify mtDAMP release as a potential initiator of acute post-injury neutrophil dysfunction.</p

Traumatic Injury and Exposure to Mitochondrial-Derived Damage Associated Molecular Patterns Suppresses Neutrophil Extracellular Trap Formation

Major traumatic injury induces significant remodeling of the circulating neutrophil pool and loss of bactericidal function. Although a well-described phenomenon, research to date has only analyzed blood samples acquired post-hospital admission, and the mechanisms that initiate compromised neutrophil function post-injury are therefore poorly understood. Here, we analyzed pre-hospital blood samples acquired from 62 adult trauma patients (mean age 44 years, range 19–95 years) within 1 h of injury (mean time to sample 39 min, range 13–59 min). We found an immediate impairment in neutrophil extracellular trap (NET) generation in response to phorbol 12-myristate 13-acetate (PMA) stimulation, which persisted into the acute post-injury phase (4–72 h). Reduced NET generation was accompanied by reduced reactive oxygen species production, impaired activation of mitogen-activated protein kinases, and a reduction in neutrophil glucose uptake and metabolism to lactate. Pre-treating neutrophils from healthy subjects with mitochondrial-derived damage-associated molecular patterns (mtDAMPs), whose circulating levels were significantly increased in our trauma patients, reduced NET generation. This mtDAMP-induced impairment in NET formation was associated with an N-formyl peptide mediated activation of AMP-activated protein kinase (AMPK), a negative regulator of aerobic glycolysis and NET formation. Indeed, activation of AMPK via treatment with the AMP-mimetic AICAR significantly reduced neutrophil lactate production in response to PMA stimulation, a phenomenon that we also observed for neutrophils pre-treated with mtDAMPs. Furthermore, the impairment in NET generation induced by mtDAMPs was partially ameliorated by pre-treating neutrophils with the AMPK inhibitor compound C. Taken together, our data demonstrate an immediate trauma-induced impairment in neutrophil anti-microbial function and identify mtDAMP release as a potential initiator of acute post-injury neutrophil dysfunction

DEDD regulates degradation of intermediate filaments during apoptosis

Apoptosis depends critically on regulated cytoskeletal reorganization events in a cell. We demonstrate that death effector domain containing DNA binding protein (DEDD), a highly conserved and ubiquitous death effector domain containing protein, exists predominantly as mono- or diubiquitinated, and that diubiquitinated DEDD interacts with both the K8/18 intermediate filament network and pro–caspase-3. Early in apoptosis, both cytosolic DEDD and its close homologue DEDD2 formed filaments that colocalized with and depended on K8/18 and active caspase-3. Subsequently, these filamentous structures collapsed into intracellular inclusions that migrated into cytoplasmic blebs and contained DEDD, DEDD2, active caspase-3, and caspase-3–cleaved K18 late in apoptosis. Biochemical studies further confirmed that DEDD coimmunoprecipitated with both K18 and pro–caspase-3, and kinetic analyses placed apoptotic DEDD staining prior to caspase-3 activation and K18 cleavage. In addition, both caspase-3 activation and K18 cleavage was inhibited by expression of DEDDΔNLS1-3, a cytosolic form of DEDD that cannot be ubiquitinated. Finally, siRNA mediated DEDD knockdown cells exhibited inhibition of staurosporine-induced DNA degradation. Our data suggest that DEDD represents a novel scaffold protein that directs the effector caspase-3 to certain substrates facilitating their ordered degradation during apoptosis

Aroma delivery from spray dried coffee containing pressurised internalised gas

A non-chemical, ingredient free method of enhancing aroma delivery during the preparation of instant coffee is presented. The approach detailed introduces a method for entrapping high pressure internalised gas within the pores of spray dried soluble coffee by cycling of both pressure and temperature. The headspace delivery of volatile aroma compounds during powder hydration was tracked by an atmospheric pressure chemical ionisationquadrupole mass spectrometer, with a modified ion source. The release of pressurised gas on hydration caused a burst of volatile aroma compounds into the headspace, which was both faster (77%) and more intense (60%) than could be achieved using standard instant coffee or through the use of chemical effervescent agents alone (sodium hydrogen carbonate and citric acid). In addition, the process of natural gasification selectively accelerated the delivery of individual aroma compounds, with the time to maximum headspace intensity being reduced for both 2,3 butanedione and acetaldehyde. © 2012 Elsevier Ltd

Условия формирования и проблемы функционирования крупных диверсифицированных производственно-корпоративных структур в Украине

У статті розглянуто умови формування та функціонування, а також історія розвитку великих диверсифікованих виробничо-корпоративних структур в Україні. Пропонуються підходи оцінки результативності процесу диверсифікації з використанням різних методик. Визначено, що в даний час оцінка результативності процесу диверсифікації можлива лише непрямими математичними методами.В статье рассмотрены условия формирования и функционирования, а также история развития крупных диверсифицированных производственно-корпоративных структур в Украине. Предлагаются подходы оценки результативности процесса диверсификации с использованием разных методик. Определено, что в настоящее время оценка результативности процесса диверсификации возможна лишь косвенными математическими методами.In the article address the formation and functioning of the conditions, as well as story development of large industrial and corporate structures, becoming diversification in Ukraine. Proposes approaches assessing impact of the process of diversification, using of different methods. Proved that the current performance assessment process of diversification can only be indirect mathematical methods

Hecatomb: An integrated software platform for viral metagenomics

BACKGROUND: Modern sequencing technologies offer extraordinary opportunities for virus discovery and virome analysis. Annotation of viral sequences from metagenomic data requires a complex series of steps to ensure accurate annotation of individual reads and assembled contigs. In addition, varying study designs will require project-specific statistical analyses. FINDINGS: Here we introduce Hecatomb, a bioinformatic platform coordinating commonly used tasks required for virome analysis. Hecatomb means a great sacrifice. In this setting, Hecatomb is sacrificing false-positive viral annotations using extensive quality control and tiered-database searches. Hecatomb processes metagenomic data obtained from both short- and long-read sequencing technologies, providing annotations to individual sequences and assembled contigs. Results are provided in commonly used data formats useful for downstream analysis. Here we demonstrate the functionality of Hecatomb through the reanalysis of a primate enteric and a novel coral reef virome. CONCLUSION: Hecatomb provides an integrated platform to manage many commonly used steps for virome characterization, including rigorous quality control, host removal, and both read- and contig-based analysis. Each step is managed using the Snakemake workflow manager with dependency management using Conda. Hecatomb outputs several tables properly formatted for immediate use within popular data analysis and visualization tools, enabling effective data interpretation for a variety of study designs. Hecatomb is hosted on GitHub (github.com/shandley/hecatomb) and is available for installation from Bioconda and PyPI

Draft genomes of 12 Bifidobacterium isolates from human IBD fecal samples

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