docid: kzyxzlrm

title: Identification of Novel Compounds Inhibiting Chikungunya Virus-Induced Cell Death by High Throughput Screening of a Kinase Inhibitor Library

text: Chikungunya virus (CHIKV) is a mosquito-borne arthrogenic alphavirus that causes acute febrile illness in humans accompanied by joint pains and in many cases, persistent arthralgia lasting weeks to years. The re-emergence of CHIKV has resulted in numerous outbreaks in the eastern hemisphere, and threatens to expand in the foreseeable future. Unfortunately, no effective treatment is currently available. The present study reports the use of resazurin in a cell-based high-throughput assay, and an image-based high-content assay to identify and characterize inhibitors of CHIKV-infection in vitro. CHIKV is a highly cytopathic virus that rapidly kills infected cells. Thus, cell viability of HuH-7 cells infected with CHIKV in the presence of compounds was determined by measuring metabolic reduction of resazurin to identify inhibitors of CHIKV-associated cell death. A kinase inhibitor library of 4,000 compounds was screened against CHIKV infection of HuH-7 cells using the resazurin reduction assay, and the cell toxicity was also measured in non-infected cells. Seventy-two compounds showing ≥50% inhibition property against CHIKV at 10 µM were selected as primary hits. Four compounds having a benzofuran core scaffold (CND0335, CND0364, CND0366 and CND0415), one pyrrolopyridine (CND0545) and one thiazol-carboxamide (CND3514) inhibited CHIKV-associated cell death in a dose-dependent manner, with EC(50) values between 2.2 µM and 7.1 µM. Based on image analysis, these 6 hit compounds did not inhibit CHIKV replication in the host cell. However, CHIKV-infected cells manifested less prominent apoptotic blebs typical of CHIKV cytopathic effect compared with the control infection. Moreover, treatment with these compounds reduced viral titers in the medium of CHIKV-infected cells by up to 100-fold. In conclusion, this cell-based high-throughput screening assay using resazurin, combined with the image-based high content assay approach identified compounds against CHIKV having a novel antiviral activity - inhibition of virus-induced CPE - likely by targeting kinases involved in apoptosis.


title: Strategies for Designing Transgenic DNA Constructs

text: Generation and characterization of transgenic mice are important elements of biomedical research. In recent years, transgenic technology has become more versatile and sophisticated, mainly because of the incorporation of recombinase-mediated conditional expression and targeted insertion, site-specific endonuclease-mediated genome editing, siRNA-mediated gene knockdown, various inducible gene expression systems, and fluorescent protein marking and tracking techniques. Site-specific recombinases (such as PhiC31) and engineered endonucleases (such as ZFN and Talen) have significantly enhanced our ability to target transgenes into specific genomic loci, but currently a great majority of transgenic mouse lines are continuingly being created using the conventional random insertion method. A major challenge for using this conventional method is that the genomic environment at the integration site has a substantial influence on the expression of the transgene. Although our understanding of such chromosomal position effects and our means to combat them are still primitive, adhering to some general guidelines can significantly increase the odds of successful transgene expression. This chapter first discusses the major problems associated with transgene expression, and then describes some of the principles for using plasmid and bacterial artificial chromosomes (BACs) for generating transgenic constructs. Finally, the strategies for conducting each of the major types of transgenic research are discussed, including gene overexpression, promoter characterization, cell-lineage tracing, mutant complementation, expression of double or multiple transgenes, siRNA knockdown, and conditional and inducible systems.


title: Vaccines for the future: learning from human immunology

text: Conventional vaccines have been extremely successful in preventing infections by pathogens expressing relatively conserved antigens through antibody‐mediated effector mechanisms. Thanks to vaccination some diseases have been eradicated and mortality due to infectious diseases has been significantly reduced. However, there are still many infections that are not preventable with vaccination, which represent a major cause of mortality worldwide. Some of these infections are caused by pathogens with a high degree of antigen variability that cannot be controlled only by antibodies, but require a mix of humoral and cellular immune responses. Novel technologies for antigen discovery, expression and formulation allow now for the development of vaccines that can better cope with pathogen diversity and trigger multifunctional immune responses. In addition, the application of new genomic assays and systems biology approaches in human immunology can help to better identify vaccine correlates of protection. The availability of novel vaccine technologies, together with the knowledge of the distinct human immune responses that are required to prevent different types of infection, should help to rationally design effective vaccines where conventional approaches have failed.


title: Docking, virtual high throughput screening and in silico fragment-based drug design

text: The drug discovery process has been profoundly changed recently by the adoption of computational methods helping the design of new drug candidates more rapidly and at lower costs. In silico drug design consists of a collection of tools helping to make rational decisions at the different steps of the drug discovery process, such as the identification of a biomolecular target of therapeutical interest, the selection or the design of new lead compounds and their modification to obtain better affinities, as well as pharmacokinetic and pharmacodynamic properties. Among the different tools available, a particular emphasis is placed in this review on molecular docking, virtual high-throughput screening and fragment-based ligand design.


title: Ubiquitin-Specific Proteases 25 Negatively Regulates Virus-Induced Type I Interferon Signaling

text: Ubiquitination and deubiquitination have emerged as critical regulatory processes in the virus-triggered type I interferon (IFN) induction pathway. In this study, we carried out a targeted siRNA screen of 54 ubiquitin-specific proteases (USPs) and identified USP25 as a negative regulator of the virus-triggered type I IFN signaling pathway. Overexpression of USP25 inhibited virus-induced activation of IFN-β, interferon regulation factor 3 (IRF3) and nuclear factor-kappa B (NF-κB), as well as the phosphorylation of IRF3 and NF-κB subunit p65. Furthermore, Knockdown of USP25 potentiated virus-induced induction of the IFN-β. In addition, detailed analysis demonstrated that USP25 cleaved lysine 48- and lysine 63-linked polyubiquitin chains in vitro and in vivo, and its deubiquitinating enzyme (DUB) activity, were dependent on a cysteine residue (Cys178) and a histidine residue (His607). USP25 mutants lacking DUB activity lost the ability to block virus-induced type I IFN to some degree. Mechanistically, USP25 deubiquitinated retinoic acid-inducible gene I (RIG-I), tumornecrosis factor (TNF) receptor-associated factor 2 (TRAF2), and TRAF6 to inhibit RIG-I-like receptor-mediated IFN signaling. Our findings suggest that USP25 is a novel DUB negatively regulating virus-induced type I IFN production.


title: Novel Virus Discovery and Genome Reconstruction from Field RNA Samples Reveals Highly Divergent Viruses in Dipteran Hosts

text: We investigated whether small RNA (sRNA) sequenced from field-collected mosquitoes and chironomids (Diptera) can be used as a proxy signature of viral prevalence within a range of species and viral groups, using sRNAs sequenced from wild-caught specimens, to inform total RNA deep sequencing of samples of particular interest. Using this strategy, we sequenced from adult Anopheles maculipennis s.l. mosquitoes the apparently nearly complete genome of one previously undescribed virus related to chronic bee paralysis virus, and, from a pool of Ochlerotatus caspius and Oc. detritus mosquitoes, a nearly complete entomobirnavirus genome. We also reconstructed long sequences (1503-6557 nt) related to at least nine other viruses. Crucially, several of the sequences detected were reconstructed from host organisms highly divergent from those in which related viruses have been previously isolated or discovered. It is clear that viral transmission and maintenance cycles in nature are likely to be significantly more complex and taxonomically diverse than previously expected.


title: Inflammation and Immune System Activation in Aging: A Mathematical Approach

text: Memory and learning declines are consequences of normal aging. Since those functions are associated with the hippocampus, I analyzed the global gene expression data from post-mortem hippocampal tissue of 25 old (age ≥ 60 yrs) and 15 young (age ≤ 45 yrs) cognitively intact human subjects. By employing a rigorous, multi-method bioinformatic approach, I identified 36 genes that were the most significant in terms of differential expression; and by employing mathematical modeling, I demonstrated that 7 of the 36 genes were able to discriminate between the old and young subjects with high accuracy. Remarkably, 90% of the known genes from those 36 most significant genes are associated with either inflammation or immune system activation. This suggests that chronic inflammation and immune system over-activity may underlie the aging process of the human brain, and that potential anti-inflammatory treatments targeting those genes may slow down this process and alleviate its symptoms.


title: Spironolactone Attenuates Bleomycin-Induced Pulmonary Injury Partially via Modulating Mononuclear Phagocyte Phenotype Switching in Circulating and Alveolar Compartments

text: BACKGROUND: Recent experimental studies provide evidence indicating that manipulation of the mononuclear phagocyte phenotype could be a feasible approach to alter the severity and persistence of pulmonary injury and fibrosis. Mineralocorticoid receptor (MR) has been reported as a target to regulate macrophage polarization. The present work was designed to investigate the therapeutic potential of MR antagonism in bleomycin-induced acute lung injury and fibrosis. METHODOLOGY/PRINCIPAL FINDINGS: We first demonstrated the expression of MR in magnetic bead-purified Ly6G-/CD11b+ circulating monocytes and in alveolar macrophages harvested in bronchoalveolar lavage fluid (BALF) from C57BL/6 mice. Then, a pharmacological intervention study using spironolactone (20mg/kg/day by oral gavage) revealed that MR antagonism led to decreased inflammatory cell infiltration, cytokine production (downregulated monocyte chemoattractant protein-1, transforming growth factor β1, and interleukin-1β at mRNA and protein levels) and collagen deposition (decreased lung total hydroxyproline content and collagen positive area by Masson’ trichrome staining) in bleomycin treated (2.5mg/kg, via oropharyngeal instillation) male C57BL/6 mice. Moreover, serial flow cytometry analysis in blood, BALF and enzymatically digested lung tissue, revealed that spironolactone could partially inhibit bleomycin-induced circulating Ly6C(hi) monocyte expansion, and reduce alternative activation (F4/80+CD11c+CD206+) of mononuclear phagocyte in alveoli, whereas the phenotype of interstitial macrophage (F4/80+CD11c-) remained unaffected by spironolactone during investigation. CONCLUSIONS/SIGNIFICANCE: The present work provides the experimental evidence that spironolactone could attenuate bleomycin-induced acute pulmonary injury and fibrosis, partially via inhibition of MR-mediated circulating monocyte and alveolar macrophage phenotype switching.


title: Ultra-Deep Sequencing of Intra-host Rabies Virus Populations during Cross-species Transmission

text: One of the hurdles to understanding the role of viral quasispecies in RNA virus cross-species transmission (CST) events is the need to analyze a densely sampled outbreak using deep sequencing in order to measure the amount of mutation occurring on a small time scale. In 2009, the California Department of Public Health reported a dramatic increase (350) in the number of gray foxes infected with a rabies virus variant for which striped skunks serve as a reservoir host in Humboldt County. To better understand the evolution of rabies, deep-sequencing was applied to 40 unpassaged rabies virus samples from the Humboldt outbreak. For each sample, approximately 11 kb of the 12 kb genome was amplified and sequenced using the Illumina platform. Average coverage was 17,448 and this allowed characterization of the rabies virus population present in each sample at unprecedented depths. Phylogenetic analysis of the consensus sequence data demonstrated that samples clustered according to date (1995 vs. 2009) and geographic location (northern vs. southern). A single amino acid change in the G protein distinguished a subset of northern foxes from a haplotype present in both foxes and skunks, suggesting this mutation may have played a role in the observed increased transmission among foxes in this region. Deep-sequencing data indicated that many genetic changes associated with the CST event occurred prior to 2009 since several nonsynonymous mutations that were present in the consensus sequences of skunk and fox rabies samples obtained from 20032010 were present at the sub-consensus level (as rare variants in the viral population) in skunk and fox samples from 1995. These results suggest that analysis of rare variants within a viral population may yield clues to ancestral genomes and identify rare variants that have the potential to be selected for if environment conditions change.


docid: sxohecoi

title: The Neonatal Fc Receptor (FcRn) Enhances Human Immunodeficiency Virus Type 1 (HIV-1) Transcytosis across Epithelial Cells

text: The mechanisms by which human immunodeficiency virus type 1 (HIV-1) crosses mucosal surfaces to establish infection are unknown. Acidic genital secretions of HIV-1-infected women contain HIV-1 likely coated by antibody. We found that the combination of acidic pH and Env-specific IgG, including that from cervicovaginal and seminal fluids of HIV-1-infected individuals, augmented transcytosis across epithelial cells as much as 20-fold compared with Env-specific IgG at neutral pH or non-specific IgG at either pH. Enhanced transcytosis was observed with clinical HIV-1 isolates, including transmitted/founder strains, and was eliminated in Fc neonatal receptor (FcRn)-knockdown epithelial cells. Non-neutralizing antibodies allowed similar or less transcytosis than neutralizing antibodies. However, the ratio of total:infectious virus was higher for neutralizing antibodies, indicating that they allowed transcytosis while blocking infectivity of transcytosed virus. Immunocytochemistry revealed abundant FcRn expression in columnar epithelia lining the human endocervix and penile urethra. Acidity and Env-specific IgG enhance transcytosis of virus across epithelial cells via FcRn and could facilitate translocation of virus to susceptible target cells following sexual exposure.


title: Defining the Range of Pathogens Susceptible to Ifitm3 Restriction Using a Knockout Mouse Model

text: The interferon-inducible transmembrane (IFITM) family of proteins has been shown to restrict a broad range of viruses in vitro and in vivo by halting progress through the late endosomal pathway. Further, single nucleotide polymorphisms (SNPs) in its sequence have been linked with risk of developing severe influenza virus infections in humans. The number of viruses restricted by this host protein has continued to grow since it was first demonstrated as playing an antiviral role; all of which enter cells via the endosomal pathway. We therefore sought to test the limits of antimicrobial restriction by Ifitm3 using a knockout mouse model. We showed that Ifitm3 does not impact on the restriction or pathogenesis of bacterial (Salmonella typhimurium, Citrobacter rodentium, Mycobacterium tuberculosis) or protozoan (Plasmodium berghei) pathogens, despite in vitro evidence. However, Ifitm3 is capable of restricting respiratory syncytial virus (RSV) in vivo either through directly restricting RSV cell infection, or by exerting a previously uncharacterised function controlling disease pathogenesis. This represents the first demonstration of a virus that enters directly through the plasma membrane, without the need for the endosomal pathway, being restricted by the IFITM family; therefore further defining the role of these antiviral proteins.


title: Quantification of Hantaan Virus with a SYBR Green Ⅰ-Based One-Step qRT-PCR Assay

text: Hantaan virus (HTNV) is a major zoonotic pathogen that causes hemorrhagic fever with renal syndrome (HFRS) in Asia, especially in China. Shaanxi province, which is located in northwest of China, is one of the areas in China most severely afflicted with HFRS epidemics annually. This study aims to establish a quantitative RT-PCR (qRT-PCR) assay to detect HTNV both in cell culture and clinical serum samples. We established a SYBR Green Ⅰ-based one-step qRT-PCR assay that targets the S segment of the HTNV genome for rapid detection and quantification. The HTNV cRNA standards were constructed by in vitro transcription, and the copy numbers of the HTNV cRNA were quantified. Standard curve was generated by determining the mean cycle threshold (Ct) values versus 10-fold serial dilutions of the HTNV cRNA over a range of 1×10(8) to 1×10(3) copies/μl. The standard curve had a reaction efficiency of 102.1%, a correlation coefficient (R(2)) of 0.998, and a slope of -3.273. The coefficient of variation (CV) of the intra- and inter-assays ranged from 0.68% to 3.00% and from 0.86% to 3.21%, respectively. The cycle intervals of the qRT-PCR assay between each dilution ranged from 2.9 to 3.8 cycles, and the lowest detection limit of the qRT-PCR assay was 10 copies/μl. The assay exhibited high specificity that was confirmed by melting curve analysis, and no cross reaction with the Seoul virus (SEOV) and other viruses (HBV, HCV and HIV) was observed. HTNV RNA was also detected in the 27 serum samples of clinical HFRS patients using the assay, and the HTNV RNA viral load ranged from 2.06×10(1) to 1.95×10(5) copies/μl. The SYBR Green Ⅰ-based one-step qRT-PCR assay is a sensitive, specific, reproducible, and simple method for detecting and quantifying HTNV in cell culture and clinical samples.


title: Machine learning approach identifies new pathways associated with demyelination in a viral model of multiple sclerosis

text: Theiler’s murine encephalomyelitis is an experimentally virus-induced inflammatory demyelinating disease of the spinal cord, displaying clinical and pathological similarities to chronic progressive multiple sclerosis. The aim of this study was to identify pathways associated with chronic demyelination using an assumption-free combined microarray and immunohistology approach. Movement control as determined by rotarod assay significantly worsened in Theiler’s murine encephalomyelitis -virus-infected SJL/J mice from 42 to 196 days after infection (dpi). In the spinal cords, inflammatory changes were detected 14 to 196 dpi, and demyelination progressively increased from 42 to 196 dpi. Microarray analysis revealed 1001 differentially expressed genes over the study period. The dominating changes as revealed by k-means and functional annotation clustering included up-regulations related to intrathecal antibody production and antigen processing and presentation via major histocompatibility class II molecules. A random forest machine learning algorithm revealed that down-regulated lipid and cholesterol biosynthesis, differentially expressed neurite morphogenesis and up-regulated toll-like receptor-4-induced pathways were intimately associated with demyelination as measured by immunohistology. Conclusively, although transcriptional changes were dominated by the adaptive immune response, the main pathways associated with demyelination included up-regulation of toll-like receptor 4 and down-regulation of cholesterol biosynthesis. Cholesterol biosynthesis is a rate limiting step of myelination and its down-regulation is suggested to be involved in chronic demyelination by an inhibition of remyelination.


title: The Soluble Form of the EIAV Receptor Encoded by an Alternative Splicing Variant Inhibits EIAV Infection of Target Cells

text: Equine lentivirus receptor 1 (ELR1) has been identified as the sole receptor for equine infectious anemia virus (EIAV) and is a member of the tumor necrosis factor receptor (TNFR) superfamily. In addition to the previously described membrane-associated form of ELR1, two other major alternative splicing variant mRNAs were identified in equine monocyte-derived macrophages (eMDMs). One major spliced species (ELR1-IN) contained an insertion of 153 nt, which resulted in a premature stop codon situated 561 nt upstream of the predicted membrane spanning domain. The other major species (ELR1-DE) has a deletion of 109 nt that causes a shift of the open reading frame and generates a stop codon 312 nt downstream. Because ELR1-DE presumably encodes a peptide of a mere 23 residues, only ELR1-IN was further analyzed. The expression of a soluble form of ELR1 (sELR1) by ELR1-IN was confirmed by Western blot and immunofluorescence analyses. Similar to ELR1, the transcription level of ELR1-IN varied among individual horses and at different time points in the same individuals. The ratio of ELR1-IN mRNA species to ELR1 mRNA was approximately 1∶2.5. Pre-incubation of the recombinant sELR1 with EIAV significantly inhibited EIAV infection in equine macrophages, the primary in vivo target cell of the virus. Fetal equine dermal (FED) cells are susceptible to EIAV in vitro, and the replication of EIAV in FED cells transiently transfected with ELR1-IN was markedly reduced when compared with replication in cells transfected with the empty vector. Finally, the expression levels of both forms of the EIAV receptor were significantly regulated by infection with this virus. Taken together, our data indicate that sELR1 acts as a secreted cellular factor that inhibits EIAV infection in host cells.


docid: qcjbtflt

title: Confounding roles for type I interferons during bacterial and viral pathogenesis

text: Although type I interferons (IFN-I) were initially defined as potent antiviral agents, they can also cause decreased host resistance to some bacterial and viral infections. The many antiviral functions of the IFN-I include direct suppression of viral replication and activation of the immune response against viruses. In addition to their antiviral effects, IFN-I are also protective against several extracellular bacterial infections, in part, by promoting the induction of TNF-α and nitric oxide. In contrast, there is a negative effect of IFN-I on host resistance during chronic infection with lymphocytic choriomeningitis virus (LCMV) and acute infections with intracellular bacteria. In the case of LCMV, chronic IFN-I signaling induces adaptive immune system suppression. Blockade of IFN-I signaling removes the suppression and allows CD4 T-cell- and IFN-γ-mediated resolution of the infection. During acute intracellular bacterial infection, IFN-I suppress innate immunity by at least two defined mechanisms. During Francisella infection, IFN-I prevent IL-17 upregulation on γδ T cells and neutrophil recruitment. Following Listeria infection, IFN-I promote the cell death of macrophages and lymphocytes, which leads to innate immune suppression. These divergent findings for the role of IFN-I on pathogen control emphasize the complexity of the interferons system and force more mechanistic evaluation of its role in pathogenesis. This review evaluates IFN-I during infection with an emphasis on work carried out IFN-I-receptor-deficient mice.


docid: qtbrelfv

title: Porcine Epidemic Diarrhea Virus Variants with High Pathogenicity, China


title: Synonymous Codon Usage in TTSuV2: Analysis and Comparison with TTSuV1

text: Two species of the DNA virus Torque teno sus virus (TTSuV), TTSuV1 and TTSuV2, have become widely distributed in pig-farming countries in recent years. In this study, we performed a comprehensive analysis of synonymous codon usage bias in 41 available TTSuV2 coding sequences (CDS), and compared the codon usage patterns of TTSuV2 and TTSuV1. TTSuV codon usage patterns were found to be phylogenetically conserved. Values for the effective number of codons (ENC) indicated that the overall extent of codon usage bias in both TTSuV2 and TTSuV1 was not significant, the most frequently occurring codons had an A or C at the third codon position. Correspondence analysis (COA) was performed and TTSuV2 and TTSuV1 sequences were located in different quadrants of the first two major axes. A plot of the ENC revealed that compositional constraint was the major factor determining the codon usage bias for TTSuV2. In addition, hierarchical cluster analysis of 41 TTSuV2 isolates based on relative synonymous codon usage (RSCU) values suggested that there was no association between geographic distribution and codon bias of TTSuV2 sequences. Finally, the comparison of RSCU for TTSuV2, TTSuV1 and the corresponding host sequence indicated that the codon usage pattern of TTSuV2 was similar to that of TTSuV1. However the similarity was low for each virus and its host. These conclusions provide important insight into the synonymous codon usage pattern of TTSuV2, as well as better understangding of the molecular evolution of TTSuV2 genomes.


title: Bovine respiratory syncytial virus: infection dynamics within and between herds

text: The infection dynamics of bovine respiratory syncytial virus (BRSV) were studied in randomly selected Norwegian dairy herds. A total of 134 herds were tested twice, six months apart. The herds were classified as positive for BRSV if at least one animal between 150 and 365 days old tested positive for antibodies against BRSV, thereby representing herds that had most likely had the virus present during the previous year. The prevalence of positive herds at the first and second sampling was 34 per cent and at 41 per cent, respectively, but varied greatly between regions. Negative herds were found in close proximity to positive herds. Some of these herds remained negative despite several new infections nearby. Of the herds initially being negative, 42 per cent changed status to positive during the six months. This occurred at the same rate during summer as winter, but a higher rate of animals in the herds was positive if it took place during winter. Of the herds initially being positive, 33 per cent changed to negative. This indicates that an effective strategy to lower the prevalence and the impact of BRSV could be to employ close surveillance and place a high biosecurity focus on the negative herds.


title: Review on Sphaeranthus indicus Linn. (Koṭṭaikkarantai)

text: Sphaeranthus indicus Linn. is from the aroma family Asteraceae. It is also known with other synonyms such as Munditika, Mundi, Shravana, Bhikshu, Tapodhana, Mahashravani, Shravanahva, Shravanashirshaka. It is abundantly distributed in damp areas in plains and also as a weed in the rice fields. In the Indian system of medicine, the plant as a whole plant or its different anatomical parts viz., leaf, stem, bark, root, flower and seed are widely used for curing many diseases. The plant is bitter, stomachic, restorative, alterative, pectoral, demulcent and externally soothing. The whole plant and its anatomical parts have been reported with different types of secondary metabolites which include eudesmanolides, sesquiterpenoids, sesquiterpene lactones, sesquiterpene acids, flavone glycosides, flavonoid C-glycosides, isoflavone glycoside, sterols, sterol glycoside, alkaloid, peptide alkaloids, amino acids and sugars. The essential oils obtained from the flowers and whole plants were analyzed by different authors and reported the presence of many monoterpene hydrocarbons, oxygenated monoterpenes, sesquiterpene hydrocarbons and oxygenated sesquiterpenes. The whole plants, its isolated secondary metabolites and different anatomical parts have been reported for ovicidal, antifeedant, anthelmintic, antimicrobial, antiviral, macrofilaricidal, larvicidal, analgesic, antipyretic, hepatoprotective, antitussive, wound healing, bronchodilatory, mast cell stabilizing activity, anxiolytic, neuroleptic, immunomodulatory, anti-diabetic, antihyperlipidemic and antioxidant, antioxidant, central nervous system depressant, anti-arthritic, nephroprotective, anticonvulsant activities and many other activities. It is also effective on psoriasis. In the present paper, the plant is reviewed for its phytochemical and pharmacological reports in detail.


title: Positive Evolutionary Selection On the RIG-I-Like Receptor Genes in Mammals

text: The mammalian RIG-I-like receptors, RIG-I, MDA5 and LGP2, are a family of DExD/H box RNA helicases responsible for the cytoplasmic detection of viral RNA. These receptors detect a variety of RNA viruses, or DNA viruses that express unusual RNA species, many of which are responsible for a great number of severe and lethal diseases. Host innate sentinel proteins involved in pathogen recognition must rapidly evolve in a dynamic arms race with pathogens, and thus are subjected to long-term positive selection pressures to avoid potential infections. Using six codon-based Maximum Likelihood methods, we were able to identify specific codons under positive selection in each of these three genes. The highest number of positively selected codons was detected in MDA5, but a great percentage of these codons were located outside of the currently defined protein domains for MDA5, which likely reflects the imposition of both functional and structural constraints. Additionally, our results support LGP2 as being the least prone to evolutionary change, since the lowest number of codons under selection was observed for this gene. On the other hand, the preponderance of positively selected codons for RIG-I were detected in known protein functional domains, suggesting that pressure has been imposed by the vast number of viruses that are recognized by this RNA helicase. Furthermore, the RIG-I repressor domain, the region responsible for recognizing and binding to its RNA substrates, exhibited the strongest evidence of selective pressures. Branch-site analyses were performed and several species branches on the three receptor gene trees showed evidence of episodic positive selection. In conclusion, by looking for evidence of positive evolutionary selection on mammalian RIG-I-like receptor genes, we propose that a multitude of viruses have crafted the receptors biological function in host defense, specifically for the RIG-I gene, contributing to the innate species-specific resistance/susceptibility to diverse viral pathogens.


title: Investigation of serum protein profiles in scrapie infected sheep by means of SELDI-TOF-MS and multivariate data analysis

text: BACKGROUND: Classical scrapie in sheep is a fatal neurodegenerative disease associated with the conversion PrP(C) to PrP(Sc). Much is known about genetic susceptibility, uptake and dissemination of PrP(Sc) in the body, but many aspects of prion diseases are still unknown. Different proteomic techniques have been used during the last decade to investigate differences in protein profiles between affected animals and healthy controls. We have investigated the protein profiles in serum of sheep with scrapie and healthy controls by SELDI-TOF-MS and LC-MS/MS. Latent Variable methods such as Principal Component Analysis, Partial Least Squares-Discriminant Analysis and Target Projection methods were used to describe the MS data. RESULTS: The serum proteomic profiles showed variable differences between the groups both throughout the incubation period and at the clinical end stage of scrapie. At the end stage, the target projection model separated the two groups with a sensitivity of 97.8%, and serum amyloid A was identified as one of the protein peaks that differed significantly between the groups. CONCLUSIONS: At the clinical end stage of classical scrapie, ten SELDI peaks significantly discriminated the scrapie group from the healthy controls. During the non-clinical incubation period, individual SELDI peaks were differently expressed between the groups at different time points. Investigations of differences in -omic profiles can contribute to new insights into the underlying disease processes and pathways, and advance our understanding of prion diseases, but comparison and validation across laboratories is difficult and challenging.


title: The Effect of Postoperative Corticosteroid Administration on Free Vascularized Fibular Grafting for Treating Osteonecrosis of the Femoral Head

text: Free vascularized fibular grafting (FVFG) has been reported to be an effective method of treating osteonecrosis of the femoral head (ONFH). This study evaluated whether postoperative maintenance doses of corticosteroids had an adverse effect on FVFG outcomes in patients with corticosteroid-induced ONFH. We retrospectively reviewed the records of 39 patients (67 hips) who had received maintenance doses of corticosteroids following FVFG. This group was matched to a group of patients who had not received corticosteroids treatment after operation. The mean follow-up duration was 5.4 years for the postoperative corticosteroid administration group (PCA group) and 5.0 years for the control group. At the latest follow-up, the average increase in Harris hip score was 11.1 ± 8.7 points for all hips in the PCA group and 12.6 ± 7.4 points for all hips in the control group (P > 0.05). In the PCA group, through radiographic evaluation, 49 hips were improved, 10 hips appeared unchanged, and 8 hips appeared worse. In the control group, 47 hips were improved, 13 hips appeared unchanged, and 7 hips appeared worse. The results suggested that postoperative maintenance doses of corticosteroids do not have an adverse effect on FVFG outcomes in patients with corticosteroid-induced ONFH.


title: Short Message Service (SMS) Applications for Disease Prevention in Developing Countries

text: BACKGROUND: The last decade has witnessed unprecedented growth in the number of mobile phones in the developing world, thus linking millions of previously unconnected people. The ubiquity of mobile phones, which allow for short message service (SMS), provides new and innovative opportunities for disease prevention efforts. OBJECTIVE: The aim of this review was to describe the characteristics and outcomes of SMS interventions for disease prevention in developing countries and provide recommendations for future work. METHODS: A systematic search of peer-reviewed and gray literature was performed for papers published in English, French, and German before May 2011 that describe SMS applications for disease prevention in developing countries. RESULTS: A total of 34 SMS applications were described, among which 5 had findings of an evaluation reported. The majority of SMS applications were pilot projects in various levels of sophistication; nearly all came from gray literature sources. Many applications were initiated by the project with modes of intervention varying between one-way or two-way communication, with or without incentives, and with educative games. Evaluated interventions were well accepted by the beneficiaries. The primary barriers identified were language, timing of messages, mobile network fluctuations, lack of financial incentives, data privacy, and mobile phone turnover. CONCLUSION: This review illustrates that while many SMS applications for disease prevention exist, few have been evaluated. The dearth of peer-reviewed studies and the limited evidence found in this systematic review highlight the need for high-quality efficacy studies examining behavioral, social, and economic outcomes of SMS applications and mobile phone interventions aimed to promote health in developing country contexts.


title: DC-SIGN gene promoter variants and IVIG treatment response in Kawasaki disease

text: BACKGROUND: Genetic variants in the inhibiting FcγRIIB mediate anti-inflammatory responses and influence IVIG refractoriness (IVIG-R). However, these variants are rare in Asian and Hispanic populations so other genes in the pathway could be potentially involved. IVIG is ineffective in mice lacking SIGN-R1, a related molecule to human DC-SIGN. Further, DC-SIGN is a known receptor for sialylated Fc, the component responsible for the anti-inflammatory action of IVIG. Thus, we hypothesized that DC-SIGN would also be involved in the pathway of IVIG response in Kawasaki Disease (KD) patients. FINDINGS: A case-control approach was performed to examine the differential distribution of five single nucleotide polymorphisms (SNPs) in DC-SIGN promoter with IVIG-R among White (158 vs. 62), Asian (64 vs. 12) and Hispanic (55 vs. 20) KD patients. Distinct differences in allele frequency distributions of several variants in the DC-SIGN promoter were observed in the three ethnic groups. Further, Asians with the major allele “A” in rs2287886 were more likely (OR = 1.76, p = 0.04) to be IVIG non-responder, but this allele is a minor allele in other two ethnic groups, where the association was not apparent. CONCLUSIONS: DC-SIGN can potentially complement the role of FcγRIIB in the anti-inflammatory cascade involved in the IVIG response mechanism.


title: Comparison of viremia of type II porcine reproductive and respiratory syndrome virus in naturally infected pigs by zip nucleic acid probe-based real-time PCR

text: BACKGROUND: Porcine reproductive and respiratory syndrome virus (PRRSV) is a RNA virus with high genetic variation. This virus causes significant economic losses in most pig-producing countries. The clinical presentation of PRRSV ranges from asymptomatic to devastating. In this study, we developed a sensitive and specific zip nucleic acid probe-based real-time PCR assay to evaluate the viremia of natural PRRSV-infected pigs in Taiwan. Serum samples were collected from 577 pigs aged 5–12 weeks. These include 444 clinically healthy pigs and 133 symptomatic pigs were confirmed to have porcine respiratory disease complex (PRDC). RESULTS: Viremia was quantified in 79 of the 444 (17.8%) clinically healthy pigs and in 112 of the 133 (84.2%) PRDC cases. Viremias were significantly more common in pigs with PRDC compared with the clinically healthy pigs (P <0.0001). These results suggest that a high viral load is a major feature of PRRSV-affected pigs. CONCLUSIONS: ZNA probe-based real-time PCR can be a useful tool to diagnose symptomatic and asymptomatic PRRSV-infected pigs. The presence of this marker in a sample of animals with high PRRSV loads (>10(4.2) PRRSV genomes/μl of serum) seems to indicate that it correlates with the presence of PRDC in pigs.


title: Epigenetic analysis leads to identification of HNF1B as a subtype-specific susceptibility gene for ovarian cancer

text: HNF1B is overexpressed in clear cell epithelial ovarian cancer, and we observed epigenetic silencing in serous epithelial ovarian cancer, leading us to hypothesize that variation in this gene differentially associates with epithelial ovarian cancer risk according to histological subtype. Here we comprehensively map variation in HNF1B with respect to epithelial ovarian cancer risk and analyse DNA methylation and expression profiles across histological subtypes. Different single-nucleotide polymorphisms associate with invasive serous (rs7405776 odds ratio (OR)=1.13, P=3.1 × 10(−10)) and clear cell (rs11651755 OR=0.77, P=1.6 × 10(−8)) epithelial ovarian cancer. Risk alleles for the serous subtype associate with higher HNF1B-promoter methylation in these tumours. Unmethylated, expressed HNF1B, primarily present in clear cell tumours, coincides with a CpG island methylator phenotype affecting numerous other promoters throughout the genome. Different variants in HNF1B associate with risk of serous and clear cell epithelial ovarian cancer; DNA methylation and expression patterns are also notably distinct between these subtypes. These findings underscore distinct mechanisms driving different epithelial ovarian cancer histological subtypes. SUPPLEMENTARY INFORMATION: The online version of this article (doi:10.1038/ncomms2629) contains supplementary material, which is available to authorized users.


docid: jquxicdr

title: Calculation of Evolutionary Correlation between Individual Genes and Full-Length Genome: A Method Useful for Choosing Phylogenetic Markers for Molecular Epidemiology

text: Individual genes or regions are still commonly used to estimate the phylogenetic relationships among viral isolates. The genomic regions that can faithfully provide assessments consistent with those predicted with full-length genome sequences would be preferable to serve as good candidates of the phylogenetic markers for molecular epidemiological studies of many viruses. Here we employed a statistical method to evaluate the evolutionary relationships between individual viral genes and full-length genomes without tree construction as a way to determine which gene can match the genome well in phylogenetic analyses. This method was performed by calculation of linear correlations between the genetic distance matrices of aligned individual gene sequences and aligned genome sequences. We applied this method to the phylogenetic analyses of porcine circovirus 2 (PCV2), measles virus (MV), hepatitis E virus (HEV) and Japanese encephalitis virus (JEV). Phylogenetic trees were constructed for comparisons and the possible factors affecting the method accuracy were also discussed in the calculations. The results revealed that this method could produce results consistent with those of previous studies about the proper consensus sequences that could be successfully used as phylogenetic markers. And our results also suggested that these evolutionary correlations could provide useful information for identifying genes that could be used effectively to infer the genetic relationships.


title: Incubation periods of viral gastroenteritis: a systematic review

text: BACKGROUND: Accurate knowledge of incubation period is important to investigate and to control infectious diseases and their transmission, however statements of incubation period in the literature are often uncited, inconsistent, and/or not evidence based. METHODS: In a systematic review of the literature on five enteric viruses of public health importance, we found 256 articles with incubation period estimates, including 33 with data for pooled analysis. RESULTS: We fit a log-normal distribution to pooled data and found the median incubation period to be 4.5 days (95% CI 3.9-5.2 days) for astrovirus, 1.2 days (95% CI 1.1-1.2 days) for norovirus genogroups I and II, 1.7 days (95% CI 1.5-1.8 days) for sapovirus, and 2.0 days (95% CI 1.4-2.4 days) for rotavirus. CONCLUSIONS: Our estimates combine published data and provide sufficient quantitative detail to allow for these estimates to be used in a wide range of clinical and modeling applications. This can translate into improved prevention and control efforts in settings with transmission or the risk of transmission.


title: Multi-criteria decision analysis as an innovative approach to managing zoonoses: results from a study on Lyme disease in Canada

text: BACKGROUND: Zoonoses are a growing international threat interacting at the human-animal-environment interface and call for transdisciplinary and multi-sectoral approaches in order to achieve effective disease management. The recent emergence of Lyme disease in Quebec, Canada is a good example of a complex health issue for which the public health sector must find protective interventions. Traditional preventive and control interventions can have important environmental, social and economic impacts and as a result, decision-making requires a systems approach capable of integrating these multiple aspects of interventions. This paper presents the results from a study of a multi-criteria decision analysis (MCDA) approach for the management of Lyme disease in Quebec, Canada. MCDA methods allow a comparison of interventions or alternatives based on multiple criteria. METHODS: MCDA models were developed to assess various prevention and control decision criteria pertinent to a comprehensive management of Lyme disease: a first model was developed for surveillance interventions and a second was developed for control interventions. Multi-criteria analyses were conducted under two epidemiological scenarios: a disease emergence scenario and an epidemic scenario. RESULTS: In general, we observed a good level of agreement between stakeholders. For the surveillance model, the three preferred interventions were: active surveillance of vectors by flagging or dragging, active surveillance of vectors by trapping of small rodents and passive surveillance of vectors of human origin. For the control interventions model, basic preventive communications, human vaccination and small scale landscaping were the three preferred interventions. Scenarios were found to only have a small effect on the group ranking of interventions in the control model. CONCLUSIONS: MCDA was used to structure key decision criteria and capture the complexity of Lyme disease management. This facilitated the identification of gaps in the scientific literature and enabled a clear identification of complementary interventions that could be used to improve the relevance and acceptability of proposed prevention and control strategy. Overall, MCDA presents itself as an interesting systematic approach for public health planning and zoonoses management with a “One Health” perspective.


title: Immunosuppression of the Trimellitic Anhydride-Induced Th2 Response by Novel Nonanatural Products Mixture in Mice

text: Many natural dietary products prevent or cure allergic inflammation; however, the ability of mixtures of these natural medicinals to suppress allergic skin inflammation is unknown. We examined the inhibitory effects of nonanatural products mixture (NPM-9), which provides immunoregulatory activation, on Th2-mediated skin allergic inflammation. Oral administration of NPM-9 in mice reduced ear thickness and specific IgE production in trimellitic anhydride- (TMA-)induced contact hypersensitivity (CHS). NPM-9 also suppressed IL-4 and IL-1β production in splenocytes but prevented only TMA-induced IL-1β production in inflamed ears. To characterize the mechanism of this effect, we examined NPM-9 immunosuppression on an OVA-induced Th2 allergic state. Oral administration of NPM-9 inhibited Th2-mediated serum IgE overproduction. NPM-9 also downregulated the polarized Th2 response, whereas it upregulated Th1 response in splenocytes. These data suggest that NPM-9 may be a useful therapeutic agent for allergic inflammatory diseases through its suppression of the Th2-mediated allergic response.


title: Effect of hepatocyte growth factor and angiotensin II on rat cardiomyocyte hypertrophy

text: Angiotensin II (Ang II) plays an important role in cardiomyocyte hypertrophy. The combined effect of hepatocyte growth factor (HGF) and Ang II on cardiomyocytes is unknown. The present study was designed to determine the effect of HGF on cardiomyocyte hypertrophy and to explore the combined effect of HGF and Ang II on cardiomyocyte hypertrophy. Primary cardiomyocytes were isolated from neonatal rat hearts and cultured in vitro. Cells were treated with Ang II (1 µM) alone, HGF (10 ng/mL) alone, and Ang II (1 µM) plus HGF (10 ng/mL) for 24, 48, and 72 h. The amount of [(3)H]-leucine incorporation was then measured to evaluate protein synthesis. The mRNA levels of β-myosin heavy chain and atrial natriuretic factor were determined by real-time PCR to evaluate the presence of fetal phenotypes of gene expression. The cell size of cardiomyocytes was also studied. Ang II (1 µM) increased cardiomyocyte hypertrophy. Similar to Ang II, treatment with 1 µM HGF promoted cardiomyocyte hypertrophy. Moreover, the combination of 1 µM Ang II and 10 ng/mL HGF clearly induced a combined pro-hypertrophy effect on cardiomyocytes. The present study demonstrates for the first time a novel, combined effect of HGF and Ang II in promoting cardiomyocyte hypertrophy.


title: Identification of Three Antiviral Inhibitors against Japanese Encephalitis Virus from Library of Pharmacologically Active Compounds 1280

text: Japanese encephalitis virus (JEV) can cause severe central nervous disease with a high mortality rate. There is no antiviral drug available for JEV-specific treatment. In this study, a cytopathic-effect-based, high-throughput screening assay was developed and applied to screen JEV inhibitors from Library of Pharmacologically Active Compounds 1280. The antiviral effects of three hit compounds including FGIN-1-27, cilnidipine, and niclosamide were evaluated in cells by western blotting, indirect immunofluorescence assay, and plaque reduction assay. A time-of-addition assay proved that all three compounds inhibited JEV at the stage of replication. The EC50s of FGIN-1-27, cilnidipine, and niclosamide were 3.21, 6.52, and 5.80 µM, respectively, while the selectivity indexes were 38.79, 30.67, and 7.49. FGIN-1-27 and cilnidipine have high efficiency and selectivity against JEV. This study provided two JEV antiviral inhibitors as candidates for treatment of JEV infection.


title: The Panhandle Formed by Influenza A and C Virus NS Non-Coding Regions Determines NS Segment Expression

text: Exchange of the extremities of the NS segment of type A and C influenza viruses in reverse genetics systems was used to assess their putative role in type specificity. Restoration of each specific proximal panhandle was mandatory to allow the rescue of viruses with heterotypic extremities. Moreover, the transcription level of the modified segment seemed to be directly affected by the distal panhandle strength.


title: Research Resource: Identification of Novel Growth Hormone-Regulated Phosphorylation Sites by Quantitative Phosphoproteomics

text: GH and GH receptors are expressed throughout life, and GH elicits a diverse range of responses, including growth and altered metabolism. It is therefore important to understand the full spectrum of GH signaling pathways and cellular responses. We applied mass spectrometry-based phosphoproteomics combined with stable isotope labeling with amino acids in cell culture to identify proteins rapidly phosphorylated in response to GH in 3T3-F442A preadipocytes. We identified 132 phosphosites in 95 proteins that exhibited rapid (5 or 15 min) GH-dependent statistically significant increases in phosphorylation by more than or equal to 50% and 96 phosphosites in 46 proteins that were down-regulated by GH by more than or equal to 30%. Several of the GH-stimulated phosphorylation sites were known (e.g. regulatory Thr/Tyr in Erks 1 and 2, Tyr in signal transducers and activators of transcription (Stat) 5a and 5b, Ser939 in tuberous sclerosis protein (TSC) 2 or tuberin). The remaining 126 GH-stimulated sites were not previously associated with GH. Kyoto Encyclopedia of Genes and Genomes pathway analysis of GH-stimulated sites indicated enrichment in proteins associated with the insulin and mammalian target of rapamycin (mTOR) pathways, regulation of the actin cytoskeleton, and focal adhesions. Akt/protein kinase A consensus sites (RXRXXS/T) were the most commonly phosphorylated consensus sites. Immunoblotting confirmed GH-stimulated phosphorylation of all seven novel GH-dependent sites tested [regulatory sites in proline-rich Akt substrate, 40 kDA (PRAS40), regulatory associated protein of mTOR, ATP-citrate lyase, Na(+)/H(+) exchanger-1, N-myc downstream regulated gene 1, and Shc]). The immunoblot results suggest that many, if not most, of the GH-stimulated phosphosites identified in this large-scale quantitative phosphoproteomics analysis, including sites in multiple proteins in the Akt/ mTOR complex 1 pathway, are phosphorylated in response to GH. Their identification significantly broadens our thinking of GH-regulated cell functions.


title: Cellular microRNA miR-181b Inhibits Replication of Mink Enteritis Virus by Repression of Non-Structural Protein 1 Translation

text: Mink enteritis virus (MEV) is one of the most important viral pathogens in the mink industry. Recent studies have showed that microRNAs (miRNAs), small noncoding RNAs of length ranging from 18–23 nucleotides (nt) participate in host-pathogen interaction networks; however, whether or not miRNAs are involved in MEV infection has not been reported. Our study revealed that miRNA miR-181b inhibited replication of MEV in the feline kidney (F81) cell line by targeting the MEV non-structural protein 1 (NS1) messenger RNA (mRNA) coding region, resulting in NS1 translational repression, while MEV infection reduced miR-181b expression. This is the first description of cellular miRNAs modulating MEV infection in F81 cells, providing further insight into the mechanisms of viral infection, and may be useful in development of naturally-occurring miRNAs antiviral strategies.


title: Endoplasmic Reticulum Stress Contributes to Helicobacter Pylori VacA-Induced Apoptosis

text: Vacuolating cytotoxin A (VacA) is one of the important virulence factors produced by H. pylori. VacA induces apoptotic cell death, which is potentiated by ammonia. VacA also causes cell death by mitochondrial damage, via signaling pathways that are not fully defined. Our aim was to determine whether endoplasmic reticulum (ER) stress is associated with VacA-induced mitochondrial dysfunction and apoptosis. We found that C/EBP homologous protein (CHOP), a key signaling protein of ER stress-induced apoptosis, was transcriptionally up-regulated following incubation of gastric epithelial cells with VacA. The effect of VacA on CHOP induction was significantly enhanced by co-incubation with ammonium chloride. Phosphorylation of eukaryotic initiation factor 2 (eIF2)-alpha, which is known to occur downstream of the ER stress sensor PKR-like ER-localized eIF2-alpha kinase (PERK) and to regulate CHOP expression, was also observed following incubation with VacA in the presence of ammonium chloride. Knockdown of CHOP by siRNA resulted in inhibition of VacA-induced apoptosis. Further studies showed that silencing of the PERK gene with siRNA attenuated VacA-mediated phosphorylation of eIF2-alpha, CHOP induction, expression of BH3-only protein Bim and Bax activation, and cell death induced by VacA with ammonium chloride, indicating that ER stress may lead to mitochondrial dysfunction during VacA-induced toxicity. Activation of ER stress and up-regulation of BH3-only proteins were also observed in human H. pylori-infected gastric mucosa. Collectively, this study reveals a possible association between VacA-induced apoptosis in gastric epithelial cells, and activation of ER stress in H. pylori-positive gastric mucosa.


title: miRNomes of haematopoietic stem cells and dendritic cells identify miR-30b as a regulator of Notch1

text: Dendritic cells (DCs) are critical to initiate the immune response and maintain tolerance, depending on different status and subsets. The expression profiles of microRNAs (miRNAs) in various DC subsets and haematopoietic stem cells (HSCs), which generate DCs, remain to be fully identified. Here we examine miRNomes of mouse bone marrow HSCs, immature DCs, mature DCs and IL-10/NO-producing regulatory DCs by deep sequencing. We identify numerous stage-specific miRNAs and histone modification in HSCs and DCs at different differentiation stages. miR-30b, significantly upregulated via a TGF-beta/Smad3-mediated epigenetic pathway in regulatory DCs, can target Notch1 to promote IL-10 and NO production, suggesting that miR-30b is a negative regulator of immune response. We also identify miRNomes of in vivo counterparts of mature DCs and regulatory DCs and systematically compare them with DCs cultured in vitro. These results provide a resource for studying roles of miRNAs in stem cell biology, development and functional regulation of DC subsets.


title: Tat Peptide-Mediated Soluble Expression of the Membrane Protein LSECtin-CRD in Escherichia coli

text: The human liver and lymph node sinusoidal endothelial cell C-type lectin (hLSECtin), a type II integral membrane protein, containing a Ca(2+)-dependent carbohydrate recognition domain (CRD), has a well-established biological activity, yet its three-dimensional structure is unknown due to low expression yields and aggregation into inclusion bodies. Previous study has demonstrated that the HIV-1 virus-encoded Tat peptide (‘YGRKKRRQRRR’) can increase the yields and the solubility of heterologous proteins. However, whether the Tat peptide could promote the high-yield and soluble expression of membrane proteins in Escherichia coli is not known. Therefore, the prokaryotic expression vector pET28b-Tat-hLSECtin-CRD (using pET28b and pET28b-hLSECtin-CRD as controls) was constructed, and transformed into E. coli BL21 (DE3) cells and induced with isopropyl-β-d-thiogalactoside (IPTG) followed with identifying by SDS-PAGE and Western blot. Subsequently, the bacterial subcellular structure, in which overexpressed the heterologous proteins Tat-hLSECtin-CRD and Tat-free hLSECtin-CRD, was analyzed by transmission electron microscope (TEM) respectively, and the mannose-binding activity of Tat-hLSECtin-CRD was also determined. Expectedly, the solubility of Tat-LSECtin-CRD significantly increased compared to Tat-free LSECtin-CRD (**p < 0.01) with prolonged time, and the Tat-LSECtin-CRD had a significant mannose-binding activity. The subcellular structure analysis indicated that the bacterial cells overexpressed Tat-hLSECtin-CRD exhibited denser region compared with controls, while dot denser region aggregated in the two ends of bacterial cells overexpressed Tat-free hLSECtin-CRD. This study provided a novel method for improving the soluble expression of membrane proteins in prokaryotic systems by fusion with the Tat peptide, which may be potentially expanded to the expression of other membrane proteins.


title: Antisense suppression of donor splice site mutations in the dystrophin gene transcript

text: We describe two donor splice site mutations, affecting dystrophin exons 16 and 45 that led to Duchenne muscular dystrophy (DMD), through catastrophic inactivation of the mRNA. These gene lesions unexpectedly resulted in the retention of the downstream introns, thereby increasing the length of the dystrophin mRNA by 20.2 and 36 kb, respectively. Splice-switching antisense oligomers targeted to exon 16 excised this in-frame exon and the following intron from the patient dystrophin transcript very efficiently in vitro, thereby restoring the reading frame and allowing synthesis of near-normal levels of a putatively functional dystrophin isoform. In contrast, targeting splice-switching oligomers to exon 45 in patient cells promoted only modest levels of an out-of-frame dystrophin transcript after transfection at high oligomer concentrations, whereas dual targeting of exons 44 and 45 or 45 and 46 resulted in more efficient exon skipping, with concomitant removal of intron 45. The splice site mutations reported here appear highly amenable to antisense oligomer intervention. We suggest that other splice site mutations may need to be evaluated for oligomer interventions on a case-by-case basis.


title: Acetylation regulates Cyclophilin A catalysis, immunosuppression and HIV isomerisation

text: Cyclophilin A (CypA) is a ubiquitous cis-trans-prolyl isomerase with key roles in immunity and viral infection. CypA suppresses T-cell activation through cyclosporine (Cs) complexation and is required for effective HIV-1 replication in host cells. We show that CypA is acetylated in diverse human cell lines and use a synthetically evolved acetyl-lysyl-tRNA synthetase/tRNA(CUA) pair to produce recombinant acetylated CypA in E. coli. We determine atomic resolution structures of acetylated CypA and its complexes with Cs and HIV-1 capsid. Acetylation dramatically inhibits CypA catalysis of cis to trans isomerisation and stabilises cis rather than trans forms of the HIV-1 capsid. Furthermore, CypA acetylation antagonizes the immunosuppressive effects of Cs, by inhibiting the sequential steps of Cs binding and calcineurin inhibition. Our results reveal that acetylation regulates key functions of CypA in immunity and viral infection and provide a general set of mechanisms by which acetylation modulates interactions to regulate cell function.


title: Application of an Amine Functionalized Biopolymer in the Colonic Delivery of Glycyrrhizin: A Design and In Vivo Efficacy Study

text: In our current study, a newer amine functionalized guar gum derivative was studied for its efficacy in colonic drug delivery. Glycyrrhizic acid mono-ammonium salt was used as the model drug. Drug-loaded microparticles were formulated by ionic crosslinking using sodium tripolyphosphate. The Scanning Electron Microscopic study revealed spherical particles of sizes from 4.9 ± 3.8 μm to 6.9 ± 3.9 μm. The FT-IR studies presented a possible interaction between the drug and the polymer. The drug was encapsulated in amorphous form as observed from the powder X-Ray Diffraction studies. A cumulative drug release study was carried out in simulated gastric, intestinal, and colonic fluids. The cumulative drug release studies presented a burst release followed by a sustained release of the drug in simulated colonic fluid containing rat cecal contents. The drug-polymer ratio was optimised using a 3(2) factorial design by taking the amounts of glycyrrhizic acid (X(1)) and guar gum alkyl amine (X(2)) as the independant variables. The percent cumulative drug release at 240 mins (Q(240)), 720 mins (Q(720)), and at 1,440 mins (Q(1440)) were considered as the dependant variables. The efficacy of the optimized formulation was studied in a 2,4,6-trinitrobenzene sulfonic acid-induced rat colitis model. The tissue’s nitric oxide, malondialdehyde, and myeloperoxidase activities were found to be much lower in the microparticle-treated group compared to free drug-treated group. The histology of the colonic tissue from the treated group of animals revealed almost no infiltration of inflammatory cells in the tissue for the microparticle-treated group of animals. The synthesized amine derivative of guar gum was found to be better in vitro with a better in vivo efficacy in the colonic delivery of glycyrrhizic acid monoammonium salt and can be considered as a newer modified biopolymer for colonic drug delivery.


title: Identification of RNase L-Dependent, 3′-End-Modified, Viral Small RNAs in Sindbis Virus-Infected Mammalian Cells

text: Small RNAs play a critical role in host-pathogen interaction. Indeed, small RNA-mediated silencing or RNA interference (RNAi) is one of the earliest forms of antiviral immunity. Although it represents the main defense system against viruses in many organisms, the antiviral role of RNAi has not been clearly proven in higher vertebrates. However, it is well established that their response to viral infection relies on the recognition of viral RNAs by host pattern recognition receptors (PRRs) to trigger activation of the interferon pathway. In the present work, we report the existence of a novel small noncoding RNA population produced in mammalian cells upon RNA virus infection. Using Sindbis virus (SINV) as a prototypic arbovirus model, we profiled the small RNA population of infected cells in both human and African green monkey cell lines. Here, we provide evidence for the presence of discrete small RNAs of viral origin that are not associated with the RNA-induced silencing complex (RISC), that are highly expressed and detected by Northern blot analysis, and that accumulate as 21- to 28-nucleotide (nt) species during infection. We report that the cellular antiviral endoribonuclease RNase L cleaves the viral genome, producing in turn the small RNAs. Surprisingly, we uncovered the presence of a modification on the 3′-end nucleotide of SINV-derived viral small RNAs (SvsRNAs) that might be at the origin of their stability. Altogether, our findings show that stable modified small viral RNAs could represent a novel way to modulate host-virus interaction upon SINV infection.


title: Functional Limitations of Plasmacytoid Dendritic Cells Limit Type I Interferon, T Cell Responses and Virus Control in Early Life

text: Infant mortality from viral infection remains a major global health concern: viruses causing acute infections in immunologically mature hosts often follow a more severe course in early life, with prolonged or persistent viral replication. Similarly, the WE strain of lymphocytic choriomeningitis virus (LCMV-WE) causes acute self-limiting infection in adult mice but follows a protracted course in infant animals, in which LCMV-specific CD8(+) T cells fail to expand and control infection. By disrupting type I IFNs signaling in adult mice or providing IFN-α supplementation to infant mice, we show here that the impaired early life T cell responses and viral control result from limited early type I IFN responses. We postulated that plasmacytoid dendritic cells (pDC), which have been identified as one major source of immediate-early IFN-I, may not exert adult-like function in vivo in the early life microenvironment. We tested this hypothesis by studying pDC functions in vivo during LCMV infection and identified a coordinated downregulation of infant pDC maturation, activation and function: despite an adult-like in vitro activation capacity of infant pDCs, the expression of the E2-2 pDC master regulator (and of critical downstream antiviral genes such as MyD88, TLR7/TLR9, NF-κB, IRF7 and IRF8) is downregulated in vivo at baseline and during LCMV infection. A similar pattern was observed in response to ssRNA polyU, a model ligand of the TLR7 viral sensor. This suggests that the limited T cell-mediated defense against early life viral infections is largely attributable to / regulated by infant pDC responses and provides incentives for novel strategies to supplement or stimulate immediate-early IFN-α responses.


title: The Natural History of Influenza Infection in the Severely Immunocompromised vs Nonimmunocompromised Hosts

text: Introduction. Medical advances have led to an increase in the world's population of immunosuppressed individuals. The most severely immunocompromised patients are those who have been diagnosed with a hematologic malignancy, solid organ tumor, or who have other conditions that require immunosuppressive therapies and/or solid organ or stem cell transplants. Materials and methods. Medically attended patients with a positive clinical diagnosis of influenza were recruited prospectively and clinically evaluated. Nasal washes and serum were collected. Evaluation of viral shedding, nasal and serum cytokines, clinical illness, and clinical outcomes were performed to compare severely immunocompromised individuals to nonimmunocompromised individuals with influenza infection. Results. Immunocompromised patients with influenza had more severe disease/complications, longer viral shedding, and more antiviral resistance while demonstrating less clinical symptoms and signs on clinical assessment. Conclusions. Immunocompromised patients are at risk for more severe or complicated influenza induced disease, which may be difficult to prevent with existing vaccines and antiviral treatments. Specific issues to consider when managing a severely immunocompromised host include the development of asymptomatic shedding, multi-drug resistance during prolonged antiviral therapy, and the potential high risk of pulmonary involvement. Clinical trials registration, ClinicalTrials.gov identifier NCT00533182.


title: Exome Sequencing Reveals RAG1 Mutations in a Child with Autoimmunity and Sterile Chronic Multifocal Osteomyelitis Evolving into Disseminated Granulomatous Disease

text: We describe a boy who developed autoinflammatory (chronic sterile multifocal osteomyelitis) and autoimmune (autoimmune cytopenias; vitiligo) phenotypes who subsequently developed disseminated granulomatous disease. Whole exome sequencing revealed homozygous RAG1 mutations thus expanding the spectrum of combined immunodeficiency with autoimmunity and granuloma that can occur with RAG deficiency.


title: Minimizing the threat of pandemic emergence from avian influenza in poultry systems

text: BACKGROUND: Live-animal markets are a culturally important feature of meat distribution chains in many populations, yet they provide an opportunity for the maintenance and transmission of potentially emergent zoonotic pathogens. The ongoing human outbreak of avian H7N9 in China highlights the need for increased surveillance and control in these live-bird markets (LBMs). DISCUSSION: Closure of retail markets in affected areas rapidly decreased human cases to rare, sporadic occurrence, but little attention has been paid thus far to the role of upstream elements of the poultry distribution chain such as wholesale markets. This could partly explain why transmission in poultry populations has not been eliminated more broadly. We present surveillance data from both wholesale live-bird markets (wLBMs) and rLBMs in Shantou, China (from 2004–2006), and call on disease-dynamic theory to illustrate why closing rLBMs has only minor effects on the overall volume of transmission. We show that the length of time birds stay in rLBMs can severely limit transmission there, but that the system-wide effect may be reduced substantially by high levels of transmission upstream of retail markets. SUMMARY: Management plans that minimize transmission throughout the entire poultry supply chain are essential for minimizing exposure to the public. These include reducing stay-time of birds in markets to 1 day, standardizing poultry supply chains to limit transmission in pre-retail settings, and monitoring strains with epidemiological traits that pose a high risk of emergence. These actions will further limit human exposure to extant viruses and reduce the likelihood of the emergence of novel strains by decreasing the overall volume of transmission.


title: The Gene Expression Profile of Peripheral Blood Mononuclear Cells from EV71-Infected Rhesus Infants and the Significance in Viral Pathogenesis

text: Enterovirus 71 (EV71) is the major pathogen responsible for fatal hand, foot and mouth disease (HFMD). Our previous work reported on an EV71-infected rhesus monkey infant model that presented with histo-pathologic changes of the central nervous system (CNS) and lungs. This study is focused on the correlated modulation of gene expression in the peripheral blood mononuclear cells (PBMCs) from EV71-infected rhesus monkey infants. The expression of more than 500 functional genes associated with multiple pathways was modulated. The expression of genes associated with immune inflammatory responses was up-regulated during the period from days 4 to 10 post-infection. The expression of two genes (TAC1 and IL17A), which play major roles in inflammatory reactions, was remarkably up-regulated during the infection period. Furthermore, a higher expression level of the TAC1 gene was identified in the CNS compared to the lungs, but a high expression level of the IL-17A gene was observed in the lungs and not in the CNS. The results of this study suggest at least two facts about EV71 infection, which are that: the TAC1 gene that encodes substance P and neurokinin-A is present in both PBMCs and the hypothalamus; and the up-regulation of IL-17A is sustained in the peripheral blood.


62(ND) ASMS Conference on Mass Spectrometry and Allied Topics


title: Design and Experimental Approach to the Construction of a Human Signal-Molecule-Profiling Database

text: The human signal-molecule-profiling database (HSMPD) is designed as a prospective medical database for translational bioinformatics (TBI). To explore the feasibility of low-cost database construction, we studied the roadmap of HSMPD. A HSMPD-oriented tool, called “signal-molecule-profiling (SMP) chip” was developed for data acquisition, which can be employed in the routine blood tests in hospitals; the results will be stored in the HSMPD system automatically. HSMPD system can provide data services for the TBI community, which generates a stable income to support the data acquisition. The small-scale experimental test was performed in the hospital to verify SMP chips and the demo HSMPD software. One hundred and eighty nine complete SMP records were collected, and the demo HSMPD system was also evaluated in the survey study on patients and doctors. The function of SMP chip was verified, whereas the demo HSMPD software needed to be improved. The survey study showed that patients would only accept free tests of SMP chips when they originally needed blood examinations. The study indicated that the construction of HSMPD relies on the self-motivated cooperation of the TBI community and the traditional healthcare system. The proposed roadmap potentially provides an executable solution to build the HSMPD without high costs.


title: A Neutralization Epitope in the Hepatitis C Virus E2 Glycoprotein Interacts with Host Entry Factor CD81

text: The identification of a specific immunogenic candidate that will effectively activate the appropriate pathway for neutralizing antibody production is fundamental for vaccine design. By using a monoclonal antibody (1H8) that neutralizes HCV in vitro, we have demonstrated here that 1H8 recognized an epitope mapped between residues A524 and W529 of the E2 protein. We also found that the epitope residues A524, P525, Y527 and W529 were crucial for antibody binding, while the residues T526, Y527 and W529 within the same epitope engaged in the interaction with the host entry factor CD81. Furthermore, we detected “1H8-like” antibodies, defined as those with amino acid-specificity similar to 1H8, in the plasma of patients with chronic HCV infection. The time course study of plasma samples from Patient H, a well-characterized case of post-transfusion hepatitis C, showed that “1H8-like” antibodies could be detected in a sample collected almost two years after the initial infection, thus confirming the immunogenicity of this epitope in vivo. The characterization of this neutralization epitope with a function in host entry factor CD81 interaction should enhance our understanding of antibody-mediated neutralization of HCV infections.


title: Normal Thoracic Radiographic Appearance of the Cynomolgus Monkey (Macaca fascicularis)

text: BACKGROUND: The cynomolgus monkey (Macaca fascicularis) has been increasingly used as a non-human primate model in biomedical research. As establishing baseline thoracic radiography for the cynomolgus monkey is essential, we tested the hypothesis that age and sex may affect the thoracic radiography parameters of this species. METHODS: Here, 697 healthy cynomolgus monkeys were segregated by sex and age (three age groups: 25–36 months, 37–48 months, 49–60 months). The lung length (LL), maximal interior thoracic depth (TD), maximal interior thoracic breadth (TBr), cardiac silhouette breadth (CBr), cardiothoracic ratio (CR), right and left costophrenic angles (RCA and LCA), and right hilar height ratio (R-HHR) were assessed by chest film. Statistical analysis was applied to examine the effect of age, sex, and age × sex interactions. RESULTS: Significant effects by age were shown for LL, TD, TBr, CBr, and CR. Significant effects by sex were found for TD, TBr, CBr, CR, and R-HHR. Significant effects by age × sex were observed for TD, TBr, CBr, and CR. Both TD and TBr increased with age in both sexes, and both were significantly higher in males than in females in the group aged 49–60 months. CBr increased with age and was significantly higher in males than in females across all age groups. CR declined with age and was significantly higher in males than females across all age groups, and CR was similar or slightly higher relative to those previously found in other non-human primate species. As to the other parameters with no significant sex nor age-related differences, the R-HHR was greater than 1.00, and the angulation of bilateral costophrenic angles were sharp. CONCLUSIONS: The thoracic radiographic parameters for the healthy cynomolgus monkey presented here should prove useful in veterinary practice, research involving non-human primate models of respiratory or cardiovascular disorders, and morphological studies on cynomolgus monkeys.


title: Pandemic Influenza Virus 2009 H1N1 and Adenovirus in a High Risk Population of Young Adults: Epidemiology, Comparison of Clinical Presentations, and Coinfection

text: BACKGROUND: In 2009, pandemic H1N1 influenza virus (2009 H1N1) emerged worldwide, causing morbidity and mortality that disproportionately affected young adults. Upper respiratory infection (URI), largely due to adenovirus, is an endemic cause of morbidity in military training. Whether clinical presentations differ or excess morbidity results from coinfection is unclear. METHODS: The Center for Advanced Molecular Detection evaluates epidemiology and rapid diagnostics of respiratory pathogens in trainees with URI. From May 1, 2009, to November 30, 2009, demographic, clinical, and PCR data from throat and nasal specimens for adenovirus and 2009 H1N1 were prospectively collected. RESULTS: 375 trainees with URI enrolled and were tested for both adenovirus and 2009 H1N1 by PCR (median age 20; 89% male). Adenovirus PCR was positive in 72% (96% serotype E-4) and 2009 H1N1 in 20%. Males were more likely to have adenovirus and females more likely to have 2009 H1N1 (p = 0.047). Subjects with 2009 H1N1 presented an average of 1 week earlier in training, had shorter illness duration before enrollment, less sore throat, diarrhea, and fewer abnormal findings on throat exam. Coryza and cough were more common with 2009 H1N1 compared to adenovirus. Subjects with 2009 H1N1 were less likely to have adenovirus than those without, despite persistently high frequencies of adenovirus detections during peak 2009 H1N1 weeks (15% vs. 83%, p < 0.01). Coinfection with adenovirus and 2009 H1N1 was rare (4%). Rates of hospitalization and pneumonia did not differ between the adenovirus, 2009 H1N1, or coinfected groups. CONCLUSION: Military trainees with 2009 H1N1 vs. adenovirus have differing clinical presentations, and males are more likely to have adenovirus. Despite high frequencies of adenovirus infection, coinfection with adenovirus and 2009 H1N1 is rare and apparently does not result in increased morbidity.


title: CD13 promotes mesenchymal stem cell-mediated regeneration of ischemic muscle

text: Mesenchymal stem cells (MSCs) are multipotent, tissue-resident cells that can facilitate tissue regeneration and thus, show great promise as potential therapeutic agents. Functional MSCs have been isolated and characterized from a wide array of adult tissues and are universally identified by the shared expression of a core panel of MSCs markers. One of these markers is the multifunctional cell surface peptidase CD13 that has been shown to be expressed on human and murine MSCs from many tissues. To investigate whether this universal expression indicates a functional role for CD13 in MSC biology we isolated, expanded and characterized MSCs from bone marrow of wild type (WT) and CD13(KO) mice. Characterization of these cells demonstrated that both WT and CD13(KO) MSCs expressed the full complement of MSC markers (CD29, CD44, CD49e, CD105, Sca1), showed comparable proliferation rates and were capable of differentiating toward the adipogenic and osteogenic lineages. However, MSCs lacking CD13 were unable to differentiate into vascular cells, consistent with our previous characterization of CD13 as an angiogenic regulator. Compared to WT MSCs, adhesion and migration on various extracellular matrices of CD13(KO) MSCs were significantly impaired, which correlated with decreased phospho-FAK levels and cytoskeletal alterations. Crosslinking human MSCs with activating CD13 antibodies increased cell adhesion to endothelial monolayers and induced FAK activation in a time dependent manner. In agreement with these in vitro data, intramuscular injection of CD13(KO) MSCs in a model of severe ischemic limb injury resulted in significantly poorer perfusion, decreased ambulation, increased necrosis and impaired vascularization compared to those receiving WT MSCs. This study suggests that CD13 regulates FAK activation to promote MSC adhesion and migration, thus, contributing to MSC-mediated tissue repair. CD13 may present a viable target to enhance the efficacy of mesenchymal stem cell therapies.


title: Update on the Angiotensin Converting Enzyme 2-Angiotensin (1–7)-Mas Receptor Axis: Fetal Programing, Sex Differences, and Intracellular Pathways

text: The renin-angiotensin-system (RAS) constitutes an important hormonal system in the physiological regulation of blood pressure. Indeed, dysregulation of the RAS may lead to the development of cardiovascular pathologies including kidney injury. Moreover, the blockade of this system by the inhibition of angiotensin converting enzyme (ACE) or antagonism of the angiotensin type 1 receptor (AT(1)R) constitutes an effective therapeutic regimen. It is now apparent with the identification of multiple components of the RAS that the system is comprised of different angiotensin peptides with diverse biological actions mediated by distinct receptor subtypes. The classic RAS can be defined as the ACE-Ang II-AT(1)R axis that promotes vasoconstriction, sodium retention, and other mechanisms to maintain blood pressure, as well as increased oxidative stress, fibrosis, cellular growth, and inflammation in pathological conditions. In contrast, the non-classical RAS composed of the ACE2-Ang-(1–7)-Mas receptor axis generally opposes the actions of a stimulated Ang II-AT(1)R axis through an increase in nitric oxide and prostaglandins and mediates vasodilation, natriuresis, diuresis, and oxidative stress. Thus, a reduced tone of the Ang-(1–7) system may contribute to these pathologies as well. Moreover, the non-classical RAS components may contribute to the effects of therapeutic blockade of the classical system to reduce blood pressure and attenuate various indices of renal injury. The review considers recent studies on the ACE2-Ang-(1–7)-Mas receptor axis regarding the precursor for Ang-(1–7), the intracellular expression and sex differences of this system, as well as an emerging role of the Ang1-(1–7) pathway in fetal programing events and cardiovascular dysfunction.


title: Immunomorphologic Manifestations in Mice Liver Infected with Influenza A/H5N1, A/Goose/Krasnoozerskoye/627/05 Strain

text: Highly pathogenic avian influenza H5N1 (HPAI H5N1) viruses can infect mammals, including humans, causing severe systemic disease with the inhibition of the immune system and a high mortality rate. In conditions of lymphoid tissue depletion, the liver plays an important role in host defence against viruses. The changes in mice liver infected with HPAI H5N1 virus A/goose/Krasnoozerskoye/627/05 have been studied. It has been shown that the virus persistence in the liver leads to the expression of proinflammatory cytokines (TNF-α, IL-6) and intracellular proteases (lysozyme, cathepsin D, and myeloperoxidase) by Kupffer cells. Defective antiviral response exacerbates destructive processes in the liver accelerating the development of liver failure.


title: Relative antioxidant activities of quercetin and its structurally related substances and their effects on NF-κB/CRE/AP-1 signaling in murine macrophages

text: Reactive oxygen species (ROS) and reactive nitrogen species (RNS) produced by the oxidative burst in activated macrophages and neutrophils cause oxidative stressimplicated diseases. Quercetin is flavonoid that occurs naturally in plants and is widely used as a nutritional supplement due to its antioxidant and anti-inflammatory properties. In this study, we investigated antioxidant activities and mechanisms of action in zymosan-induced macrophages of quercetin and quercetin-related flavonoids such as quercitrin, isoquercitrin, quercetin 3-O-β-(2″-galloyl)-rhamnopyranoside (QGR) and quercetin 3-O-β-(2″-galloyl)-glucopyranoside (QGG) as well as gallic acid, a building moiety of QGR and QGG. QGR and QGG exhibited stronger antioxidant activities compared with quercetin, whereas quercitrin, isoquercitrin and gallic acid exhibited weak-tono antioxidant activities, assessed by 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging, superoxide production, superoxide scavenging, nitric oxide (NO) production, peroxynitrite (ONOO(−)) scavenging and myeloperoxidase (MPO) activity. Regarding mechanisms, the quercetincontaining flavonoids QGR and QGG differentially targeted compared with quercetin in the NF-κB signaling pathway that inhibited the DNA binding activity of the NF-κB complex without affecting the degradation and phosphorylation of IκBα and NF-κB phosphorylation. In addition, QGR and QGG inhibited CRE and activator protein (AP-1) transcriptional activity and JNK phosphorylation by inhibiting the cAMP/protein kinase A (PKA) and protein kinase C (PKC) signaling in a different manner than quercetin. Our results showed that although QGR and QGG exhibited stronger antioxidant activities than querce-tin in macrophages, their mechanisms of action in terms of the NF-κB, PKA and PKC signaling pathways were different.


title: ESCRT regulates surface expression of the Kir2.1 potassium channel

text: Protein quality control (PQC) is required to ensure cellular health. PQC is recognized for targeting the destruction of defective polypeptides, whereas regulated protein degradation mechanisms modulate the concentration of specific proteins in concert with physiological demands. For example, ion channel levels are physiologically regulated within tight limits, but a system-wide approach to define which degradative systems are involved is lacking. We focus on the Kir2.1 potassium channel because altered Kir2.1 levels lead to human disease and Kir2.1 restores growth on low-potassium medium in yeast mutated for endogenous potassium channels. Using this system, first we find that Kir2.1 is targeted for endoplasmic reticulum–associated degradation (ERAD). Next a synthetic gene array identifies nonessential genes that negatively regulate Kir2.1. The most prominent gene family that emerges from this effort encodes members of endosomal sorting complex required for transport (ESCRT). ERAD and ESCRT also mediate Kir2.1 degradation in human cells, with ESCRT playing a more prominent role. Thus multiple proteolytic pathways control Kir2.1 levels at the plasma membrane.


docid: iogmaulh

title: Activity of sputum p38 MAPK is correlated with airway inflammation and reduced FEV(1) in COPD patients

text: BACKGROUND: Inflammation and remodeling of the small airways are major determinants of the progression and severity of COPD. The present study explored the correlation between sputum p38 mitogen-activated protein kinase (MAPK) activity and airway inflammation and reduction of lung function in the patients with chronic obstructive pulmonary disease (COPD). MATERIAL/METHODS: Sputum samples were collected from 48 COPD patients and 12 healthy persons. Sputum p38 MAPK activity was measured by Western blotting and sputum levels of CXCL8 and neutrophil, and lung function was measured. The correlation between p38MAPK activity and airway inflammation and reduction of lung function was analyzed. RESULT: Our results showed the significantly increased expression of phospho-p38 MAPK and CXCL8 in the sputum samples of the COPD patients. The p38 MAPK activity was remarkably correlated with the CXCL8 level and neutrophils infiltration in the airway, and the decline of lung function in the COPD patients. CONCLUSIONS: These findings suggest the pivotal role of p38 MAPK in the airway inflammation of COPD patients. We propose p38 MAPK as a potential target for the treatment of COPD.


title: Spatio-temporal analysis on enterovirus cases through integrated surveillance in Taiwan

text: BACKGROUND: Severe epidemics of enterovirus have occurred frequently in Malaysia, Singapore, Taiwan, Cambodia, and China, involving cases of pulmonary edema, hemorrhage and encephalitis, and an effective vaccine has not been available. The specific aim of this study was to understand the epidemiological characteristics of mild and severe enterovirus cases through integrated surveillance data. METHODS: All enterovirus cases in Taiwan over almost ten years from three main databases, including national notifiable diseases surveillance, sentinel physician surveillance and laboratory surveillance programs from July 1, 1999 to December 31, 2008 were analyzed. The Pearson’s correlation coefficient was applied for measuring the consistency of the trends in the cases between different surveillance systems. Cross correlation analysis in a time series model was applied for examining the capability to predict severe enterovirus infections. Poisson temporal, spatial and space-time scan statistics were used for identifying the most likely clusters of severe enterovirus outbreaks. The directional distribution method with two standard deviations of ellipse was applied to measure the size and the movement of the epidemic. RESULTS: The secular trend showed that the number of severe EV cases peaked in 2008, and the number of mild EV cases was significantly correlated with that of severe ones occurring in the same week [r = 0.553, p < 0.01]. These severe EV cases showed significantly higher association with the weekly positive isolation rates of EV-71 than the mild cases [severe: 0.498, p < 0.01 vs. mild: 0.278, p < 0.01]. In a time series model, the increase of mild EV cases was the significant predictor for the occurrence of severe EV cases. The directional distribution showed that both the mild and severe EV cases spread extensively during the peak. Before the detected spatio-temporal clusters in June 2008, the mild cases had begun to rise since May 2008, and the outbreak spread from south to north. CONCLUSIONS: Local public health professionals can monitor the temporal and spatial trends plus spatio-temporal clusters and isolation rate of EV-71 in mild and severe EV cases in a community when virus transmission is high, to provide early warning signals and to prevent subsequent severe epidemics.


title: Tuberculosis mortality: patient characteristics and causes

text: BACKGROUND: In the antibiotic era, tuberculosis (TB) still causes a substantial number of mortalities. We aimed to identify the causes and risks of death among TB patients. METHODS: Medical records of mortality cases of culture-proven TB diagnosed during 2003–2007 were reviewed. All TB deaths were classified into 2 groups (TB-related and non-TB-related), based on the underlying cause of death. RESULTS: During the study period, 2016 cases (male: 71.1%) of culture-proven TB were identified. The mean age was 59.3 (range: 0.3–96) years. The overall mortality rate was 12.3% (249 cases) and the mean age at death was 74 years; 17.3% (43 cases) of all TB deaths were TB-related. Most of the TB-related deaths occurred early (median survival: 20 days), and the patient died of septic shock. Malignancy, liver cirrhosis, renal failure, and miliary and pneumonic radiographic patterns were all independent predictors for all TB deaths. Cavitary, miliary and pneumonic radiographic patterns were all significant predictive factors for TB-related death. Extrapulmonary involvement and liver cirrhosis were also factors contributing to TB-related death. CONCLUSIONS: The majority of TB deaths were ascribed to non-TB-related causes. Managing TB as well as underlying comorbidities in a multidisciplinary approach is essential to improve the outcome of patients in an aging population. However, the clinical manifestations of patients with TB-related death vary; many progressed to fulminant septic shock requiring timely recognition with prompt treatment to prevent early death.


title: Early responses to H7N9 in southern Mainland China

text: BACKGROUND: H7N9 posed potentially serious health challenges for Chinese society. The previous SARS outbreak in this country was accompanied by contradictory information, while worries about wide-spread influenza led to discrimination worldwide. Early understanding of public threat perceptions is therefore important for effective public health communication and intervention. METHODS: We interviewed 1011 respondents by phone two weeks after the first case. Questions examined risk awareness and media use, beliefs about the emergence of the threat and those most at risk, anxiety about infection and preventive and avoidant behaviours. RESULTS: Results demonstrate moderate levels of anxiety but relatively high levels of trust towards government officials. Threat emergence was associated with hygiene levels, temperature change, floating pigs in the Huangpu River and migration to the city. Anxiety predicted both recommended and non-recommended behavioural changes. CONCLUSIONS: Comparatively high levels of trust in Chinese government advice about H7N9 contrast positively with previous pandemic communications in China. Anxiety helped drive both recommended and non-recommended behaviours, with potentially important economic and social implications. This included evidence of 'othering’ of those associated with the threat (e.g. migrants). Findings emphasise the need to manage public communications early during new influenza outbreaks.


title: Overexpression of PTEN suppresses lipopolysaccharide-induced lung fibroblast proliferation, differentiation and collagen secretion through inhibition of the PI3-K-Akt-GSK3beta pathway

text: BACKGROUND: Abnormal and uncontrolled proliferation of lung fibroblasts may contribute to pulmonary fibrosis. Lipopolysaccharide (LPS) can induce fibroblast proliferation and differentiation through activation of phosphoinositide3-Kinase (PI3-K) pathway. However, the detail mechanism by which LPS contributes to the development of lung fibrosis is not clearly understood. To investigate the role of phosphatase and tensin homolog (PTEN), a PI3-K pathway suppressor, on LPS-induced lung fibroblast proliferation, differentiation, collagen secretion and activation of PI3-K, we transfected PTEN overexpression lentivirus into cultured mouse lung fibroblasts with or without LPS treatment to evaluate proliferation by MTT and Flow cytometry assays. Expression of PTEN, alpha-smooth muscle actin (alpha-SMA), glycogen synthase kinase 3 beta (GSK3beta) and phosphorylation of Akt were determined by Western-blot or real-time RT-PCR assays. The PTEN phosphorylation activity was measured by a malachite green-based assay. The content of C-terminal propeptide of type I procollagen (PICP) in cell culture supernatants was examined by ELISA. RESULTS: We found that overexpression of PTEN effectively increased expression and phosphatase activity of PTEN, and concomitantly inhibited LPS-induced fibroblast proliferation, differentiation and collagen secretion. Phosphorylation of Akt and GSK3beta protein expression levels in the LPS-induced PTEN overexpression transfected cells were significantly lower than those in the LPS-induced non-transfected cells, which can be reversed by the PTEN inhibitor, bpV(phen). CONCLUSIONS: Collectively, our results show that overexpression and induced phosphatase activity of PTEN inhibits LPS-induced lung fibroblast proliferation, differentiation and collagen secretion through inactivation of PI3-K-Akt-GSK3beta signaling pathways, which can be abrogated by a selective PTEN inhibitor. Thus, expression and phosphatase activity of PTEN could be a potential therapeutic target for LPS-induced pulmonary fibrosis. Compared with PTEN expression level, phosphatase activity of PTEN is more crucial in affecting lung fibroblast proliferation, differentiation and collagen secretion.


title: Isolation, Characterization, and Molecular Modeling of a Rheumatoid Factor from a Hepatitis C Virus Infected Patient with Sjögren's Syndrome

text: We have previously isolated several IgG rheumatoid factors (RFs) from patients with both rheumatoid arthritis and idiopathic thrombocytopenia purpura using phage display system. To study IgG RFs in patients with other autoimmune diseases, phage display antibody libraries from a hepatitis C virus infected patient with Sjögren's syndrome were constructed. After panning, a specific clone RFL11 was isolated for characterization in advance. The binding activity and specificity of RFL11 to IgG Fc fragment were comparable to those of RFs previously isolated. The analysis with existed RF-Fc complex structures indicated the homology model of RFL11 is similar to IgM RF61 complex with high binding affinity of about 6 × 10(−8) M. This effect resulted from longer complementarity-determining region (CDR) combining key somatic mutations. In the RFL11-Fc interfaces, the CDR-H3 loop forms a finger-like structure extending into the bottom of Fc pocket and resulting in strong ion and cation-pi interactions. Moreover, a process of antigen-driven maturation was proven by somatically mutated VH residues on H2 and H3 CDR loops in the interfaces. Taken together, these results suggested that high affinity IgG RFs can be generated in patients with Sjögren's syndrome and may play an important role in the pathogenesis of this autoimmune disease.