The effects of sleep hypoxia on coagulant factors and hepatic inflammation in emphysematous rats

Objectives

To develop a sleep hypoxia (SH) in emphysema (SHE) rat model and to explore whether SHE results in more severe hepatic inflammation than emphysema alone and whether the inflammation changes levels of coagulant/anticoagulant factors synthesized in the liver.

Methods

Seventy-five rats were put into 5 groups: SH control (SHCtrl), treated with sham smoke exposure (16 weeks) and SH exposure (12.5% O₂, 3 h/d, latter 8 weeks); emphysema control (ECtrl), smoke exposure and sham SH exposure (21% O₂); short SHE (SHEShort), smoke exposure and short SH exposure (1.5 h/d); mild SHE (SHEMild), smoke exposure and mild SH exposure (15% O₂); standard SHE (SHEStand), smoke exposure and SH exposure. Therefore, ECtrl, SHEShort, SHEMild and SHEStand group were among emphysematous groups. Arterial blood gas (ABG) data was obtained during preliminary tests. After exposure, hepatic inflammation (interleukin -6 [IL-6] mRNA and protein, tumor necrosis factor α [TNFα] mRNA and protein) and liver coagulant/anticoagulant factors (antithrombin [AT], fibrinogen [FIB] and Factor VIII [F VIII]) were evaluated. SPSS 11.5 software was used for statistical analysis.

Results

Characteristics of emphysema were obvious in emphysematous groups and ABGs reached SH criteria on hypoxia exposure. Hepatic inflammation parameters and coagulant factors are the lowest in SHCtrl and the highest in SHEStand while AT is the highest in SHCtrl and the lowest in SHEStand. Inflammatory cytokines of liver correlate well with coagulant factors positively and with AT negatively.

Conclusions

When SH is combined with emphysema, hepatic inflammation and coagulability enhance each other synergistically and produce a more significant liver-derivative inflammatory and prothrombotic status.     

Pharmacodynamics in the study of drug resistance and establishing in vitro susceptibility breakpoints: ready for prime time

Considerable advancements have been made in providing informative, relevant interpretations of the results of antimicrobial susceptibility tests to clinicians, clinical microbiologists, epidemiologists, and researchers. At the same time, the science of pharmacokinetics has flourished, and the importance of drug exposure in vivo on outcome is now recognized by researchers and clinicians alike. More recently, pharmacokinetic and quantitative measures of antimicrobial susceptibility have been integrated using pharmacokinetic-pharmacodynamic (PK-PD) models to forecast clinical and microbiological outcomes. Stochastic methods utilizing patient population pharmacokinetics, target organism minimum inhibitory concentration (MIC) distributions, and PK-PD targets from non-clinical models of infection or clinical data have established a new paradigm for determining in vitro susceptibility breakpoints and selection of empirical therapy in clinical practice. Given the increasing problem of antimicrobial resistance, these new tools are valuable additions for clinicians, researchers, and regulatory authorities.     

A conserved peptide in West Nile virus NS4A protein contributes to proteolytic processing and is essential for replication

The West Nile virus strain Kunjin virus (WNV(KUN)) NS4A protein is a multifunctional protein involved in membrane proliferation, stimulation of cellular pathways, and evasion of host defense and is a major component of the WNV(KUN) RNA replication complex. We identified a highly conserved region ((120)P-E-P-E(123)) upstream of the viral protease dibasic cleavage site and investigated whether this motif was required for WNV(KUN) replication. Single point mutations to alanine and a PEPE deletion mutation were created in a full-length infectious WNV(KUN) molecular clone. All mutations drastically impaired viral replication and virion production, except that of the P122A mutant, which was slightly attenuated. These mutations were subsequently transferred to a WNV(KUN) replicon to specifically assess effects on RNA replication alone. Again, all mutants, except P122A, showed severely reduced negative-sense RNA production as well as decreased viral protein production. Correspondingly, immunofluorescence analyses showed a lack of double-stranded RNA (dsRNA) labeling and a dispersed localization of the WNV(KUN) proteins, suggesting that replication complex formation was additionally impaired. Attempts to rescue replication via conservative mutants largely failed except for substitution of Asp at E121, suggesting that a negative charge at this residue is equally important. Analysis of viral protein processing suggested that cleavage of the 2K peptide from NS4A did not occur with the mutant constructs. These observations imply that the combined effects of proline and negatively charged residues within the PEPE peptide are essential to promote the cleavage of 2K from NS4A, which is a prerequisite for efficient WNV replication.     

Broadening horizons: holistic viewpoints from the Biology of Genomes

A report on the Cold Spring Harbor Laboratory 27th annual meeting on the Biology of Genomes, held in Cold Spring Harbor, New York, USA, 6-10 May 2014.     

Dietary and environmental reconstruction with stable isotope analyses of herbivore tooth enamel from the Miocene locality of Fort Ternan, Kenya

Tooth enamel of nine Middle Miocene mammalian herbivores from Fort Ternan, Kenya, was analyzed for δ¹³C and δ¹⁸O. The δ¹⁸O values of the tooth enamel compared with pedogenic and diagenetic carbonate confirm the use of stable isotope analysis of fossil tooth enamel as a paleoenvironmental indicator. Furthermore, the δ¹⁸O of tooth enamel indicates differences in water sources between some of the mammals. The δ¹³C values of tooth enamel ranged from −8·6–−13·0‰ which is compatible with a pure C₃diet, though the possibility of a small C₄fraction in the diet of a few of the specimens sampled is not precluded. The carbon isotopic data do not support environmental reconstructions of a Serengeti-typed wooded grassland with a significant proportion of C₄grasses. This study does not preclude the presence of C₃grasses at Fort Ternan; it is possible that C₃grasses could have had a wider geographic range if atmospheric CO₂levels were higher than the present values.     

Terminal Regions Confer Plasticity to the Tetrameric Assembly of Human HspB2 and HspB3

Heterogeneity in small heat shock proteins (sHsps) spans multiple spatiotemporal regimes—from fast fluctuations of part of the protein, to conformational variability of tertiary structure, plasticity of the interfaces, and polydispersity of the inter-converting, and co-assembling oligomers. This heterogeneity and dynamic nature of sHsps has significantly hindered their structural characterization. Atomic coordinates are particularly lacking for vertebrate sHsps, where most available structures are of extensively truncated homomers. sHsps play important roles in maintaining protein levels in the cell and therefore in organismal health and disease. HspB2 and HspB3 are vertebrate sHsps that are found co-assembled in neuromuscular cells, and variants thereof are associated with disease. Here, we present the structure of human HspB2/B3, which crystallized as a hetero-tetramer in a 3:1 ratio. In the HspB2/B3 tetramer, the four α-crystallin domains (ACDs) assemble into a flattened tetrahedron which is pierced by two non-intersecting approximate dyads. Assembly is mediated by flexible “nuts and bolts” involving IXI/V motifs from terminal regions filling ACD pockets. Parts of the N-terminal region bind in an unfolded conformation into the anti-parallel shared ACD dimer grooves. Tracts of the terminal regions are not resolved, most likely due to their disorder in the crystal lattice. This first structure of a full-length human sHsp heteromer reveals the heterogeneous interactions of the terminal regions and suggests a plasticity that is important for the cytoprotective functions of sHsps.     

amontillado,the Drosophila homolog of the prohormone processing protease PC2, is required during embryogenesis and early larval development

Biosynthesis of most peptide hormones and neuropeptides requires proteolytic excision of the active peptide from inactive proprotein precursors, an activity carried out by subtilisin-like proprotein convertases (SPCs) in constitutive or regulated secretory pathways. The Drosophila amontillado (amon) gene encodes a homolog of the mammalian PC2 protein, an SPC that functions in the regulated secretory pathway in neuroendocrine tissues. We have identified amon mutants by isolating ethylmethanesulfonate (EMS)-induced lethal and visible mutations that define two complementation groups in the amon interval at 97D1 of the third chromosome. DNA sequencing identified the amon complementation group and the DNA sequence change for each of the nine amon alleles isolated. amon mutants display partial embryonic lethality, are defective in larval growth, and arrest during the first to second instar larval molt. Mutant larvae can be rescued by heat-shock-induced expression of the amon protein. Rescued larvae arrest at the subsequent larval molt, suggesting that amon is also required for the second to third instar larval molt. Our data indicate that the amon proprotein convertase is required during embryogenesis and larval development in Drosophila and support the hypothesis that AMON acts to proteolytically process peptide hormones that regulate hatching, larval growth, and larval ecdysis.