An unbiased genetic screen reveals the polygenic nature of the influenza virus anti-interferon response.

Influenza A viruses counteract the cellular innate immune response at several steps, including blocking RIG I-dependent activation of interferon (IFN) transcription, interferon (IFN)-dependent upregulation of IFN-stimulated genes (ISGs), and the activity of various ISG products; the multifunctional NS1 protein is responsible for most of these activities. To determine the importance of other viral genes in the interplay between the virus and the host IFN response, we characterized populations and selected mutants of wild-type viruses selected by passage through non-IFN-responsive cells. We reasoned that, by allowing replication to occur in the absence of the selection pressure exerted by IFN, the virus could mutate at positions that would normally be restricted and could thus find new optimal sequence solutions. Deep sequencing of selected virus populations and individual virus mutants indicated that nonsynonymous mutations occurred at many phylogenetically conserved positions in nearly all virus genes. Most individual mutants selected for further characterization induced IFN and ISGs and were unable to counteract the effects of exogenous IFN, yet only one contained a mutation in NS1. The relevance of these mutations for the virus phenotype was verified by reverse genetics. Of note, several virus mutants expressing intact NS1 proteins exhibited alterations in the M1/M2 proteins and accumulated large amounts of deleted genomic RNAs but nonetheless replicated to high titers. This suggests that the overproduction of IFN inducers by these viruses can override NS1-mediated IFN modulation. Altogether, the results suggest that influenza viruses replicating in IFN-competent cells have tuned their complete genomes to evade the cellular innate immune system and that serial replication in non-IFN-responsive cells allows the virus to relax from these constraints and find a new genome consensus within its sequence space. IMPORTANCE In natural virus infections, the production of interferons leads to an antiviral state in cells that effectively limits virus replication. The interferon response places considerable selection pressure on viruses, and they have evolved a variety of ways to evade it. Although the influenza virus NS1 protein is a powerful interferon antagonist, the contributions of other viral genes to interferon evasion have not been well characterized. Here, we examined the effects of alleviating the selection pressure exerted by interferon by serially passaging influenza viruses in cells unable to respond to interferon. Viruses that grew to high titers had mutations at many normally conserved positions in nearly all genes and were not restricted to the NS1 gene. Our results demonstrate that influenza viruses have fine-tuned their entire genomes to evade the interferon response, and by removing interferon-mediated constraints, viruses can mutate at genome positions normally restricted by the interferon response

The use of fluoroproline in MUC1 antigen enables efficient detection of antibodies in patients with prostate cancer

A structure-based design of a new gene22ration tumor-associated glycopeptides with improved affinity against two anti-MUC1 antibodies is described. These unique antigens feature a fluorinated proline residue, such as a (4S)-4-fluoro-L-proline or 4,4-difluoroproline, at the most immunogenic domain. Binding assays using bio-layer interferometry reveal 3-fold to 10-fold affinity improvement with respect to the natural (glyco)peptides. According to X-ray crystallography and MD simulations, the fluorinated residues stabilize the antigen-antibody complex by enhancing key CH/ interactions. Interestingly, a notable improvement in detection of cancer-associated anti-MUC1 antibodies from serum of patients with prostate cancer is achieved with the non-natural antigens, which proves that these derivatives can be considered better diagnostic tools than the natural antigen for this type of cancer.We thank the Ministerio de Economía y Competitividad (projects CTQ2015-67727-R, UNLR13-4E-1931, CTQ2013-44367-C2-2-P, CTQ2015-64597-C2-1P, and BFU2016-75633-P). I. A. B. thanks the Asociación Española Contra el Cancer en La Rioja for a grant. I. S. A. and G. J. L. B. thank FCT Portugal (PhD studentship and FCT Investigator, respectively) and the EPSRC for funding. G. J. L. B. holds a Royal Society URF and an ERC StG (TagIt). F.C. and G. J. L. B thank the EU (Marie-Sklodowska Curie ITN, Protein Conjugates). R.H-G. thanks Agencia Aragonesa para la Investigación y Desarrollo (ARAID) and the Diputación General de Aragón (DGA, B89) for financial support. The research leading to these results has also received funding from the FP7 (2007-2013) under BioStruct-X (grant agreement N°283570 and BIOSTRUCTX_5186). We thank synchrotron radiation source DIAMOND (Oxford) and beamline I04 (number of experiment mx10121-19). Hokkaido University group acknowledges to JSPS KAKENHI Grant Number 25220206 and JSPS Wakate B KAKENHI Grant Number 24710242. We also thank CESGA (Santiago de Compostela) for computer support

Immediate thoracotomy for penetrating injuries: Ten years' experience at a Dutch level I trauma center

Background: An emergency department thoracotomy (EDT) or an emergency thoracotomy (ET) in the operating theater are both beneficial in selected patients following thoracic penetrating injuries. Since outcome-descriptive European studies are lacking, the aim of this retrospective study was to evaluate ten years of experience at a Dutch level I trauma center. Method: Data on patients who underwent an immediate thoracotomy after sustaining a penetrating thoracic injury between October 2000 and January 2011 were collected from the trauma registry and hospital files. Descriptive and univariate analyses were performed. Results: Among 56 patients, 12 underwent an EDT and 44 an ET. Forty-six patients sustained one or multiple stab wounds, versus ten with one or multiple gunshot wounds. Patients who had undergone an EDT had a lower GCS (p < 0. 001), lower pre-hospital RTS and hospital triage RTS (p < 0. 001 and p = 0. 009, respectively), and a lower SBP (p = 0. 038). A witnessed loss of signs of life generally occurred in EDT patients and was accompanied by 100 % mortality. Survival following EDT was 25 %, which was significantly lower than in the ET group (75 %; p = 0. 002). Survivors had lower ISS (p = 0. 011), lower rates of pre-hospital (p = 0. 031) and hospital (p = 0. 003) hemodynamic instability, and a lower prevalence of concomitant abdominal injury (p = 0. 002). Conclusion: The overall survival rate in our study was 64 %. The outcome of immediate thoracotomy performed in this level I trauma center was similar to those obtained in high-incidence regions like the US and South Africa. This suggests that trauma units where immediate thoracotomies are not part of the daily routine can achieve similar results, if properly trained

Chemokines in cerebrospinal fluid correlate with cerebral metabolite patterns in HIV-infected individuals

Chemokines influence HIV neuropathogenesis by affecting the HIV life cycle, trafficking of macrophages into the nervous system, glial activation, and neuronal signaling and repair processes; however, knowledge of their relationship to in vivo measures of cerebral injury is limited. The primary objective of this study was to determine the relationship between a panel of chemokines in cerebrospinal fluid (CSF) and cerebral metabolites measured by proton magnetic resonance spectroscopy (MRS) in a cohort of HIV-infected individuals. One hundred seventy-one stored CSF specimens were assayed from HIV-infected individuals who were enrolled in two ACTG studies that evaluated the relationship between neuropsychological performance and cerebral metabolites. Concentrations of six chemokines (fractalkine, IL-8, IP-10, MCP-1, MIP-1β, and SDF-1) were measured and compared with cerebral metabolites individually and as composite neuronal, basal ganglia, and inflammatory patterns. IP-10 and MCP-1 were the chemokines most strongly associated with individual cerebral metabolites. Specifically, (1) higher IP-10 levels correlated with lower N-acetyl aspartate (NAA)/creatine (Cr) ratios in the frontal white matter and higher MI/Cr ratios in all three brain regions considered and (2) higher MCP-1 levels correlated with lower NAA/Cr ratios in frontal white matter and the parietal cortex. IP-10, MCP-1, and IL-8 had the strongest associations with patterns of cerebral metabolites. In particular, higher levels of IP-10 correlated with lower neuronal pattern scores and higher basal ganglia and inflammatory pattern scores, the same pattern which has been associated with HIV-associated neurocognitive disorders (HAND). Subgroup analysis indicated that the effects of IP-10 and IL-8 were influenced by effective antiretroviral therapy and that memantine treatment may mitigate the neuronal effects of IP-10. This study supports the role of chemokines in HAND and the validity of MRS as an assessment tool. In particular, the findings identify relationships between the immune response—particularly an interferon-inducible chemokine, IP-10—and cerebral metabolites and suggest that antiretroviral therapy and memantine modify the impact of the immune response on neurons

HIV-1 Tat Co-Operates with IFN-γ and TNF-α to Increase CXCL10 in Human Astrocytes

HIV-associated neurological disorders (HAND) are estimated to affect 60% of the HIV infected population. HIV-encephalitis (HIVE), the pathological correlate of the most severe form of HAND is often characterized by glial activation, cytokine/chemokine dysregulation, and neuronal damage and loss. However, the severity of HIVE correlates better with glial activation rather than viral load. One of the characteristic features of HIVE is the increased amount of the neurotoxic chemokine, CXCL10. This chemokine can be released from astroglia activated with the pro-inflammatory cytokines IFN-γ and TNF-α, in conjunction with HIV-1 Tat, all of which are elevated in HIVE. In an effort to understand the pathogenesis of HAND, this study was aimed at exploring the regulation of CXCL10 by cellular and viral factors during astrocyte activation. Specifically, the data herein demonstrate that the combined actions of HIV-1 Tat and the pro-inflammatory cytokines, IFN-γ and TNF-α, result in the induction of CXCL10 at both the RNA and protein level. Furthermore, CXCL10 induction was found to be regulated transcriptionally by the activation of the p38, Jnk, and Akt signaling pathways and their downstream transcription factors, NF-κB and STAT-1α. Since CXCL10 levels are linked to disease severity, understanding its regulation could aid in the development of therapeutic intervention strategies for HAND

Trichomonas vaginalis: Clinical relevance, pathogenicity and diagnosis

Trichomonas vaginalis is the etiological agent of trichomoniasis, the most prevalent non-viral sexually transmitted disease worldwide. Trichomoniasis is a widespread, global health concern and occurring at an increasing rate. Infections of the female genital tract can cause a range of symptoms, including vaginitis and cervicitis, while infections in males are generally asymptomatic. The relatively mild symptoms, and lack of evidence for any serious sequelae, have historically led to this disease being under diagnosed, and under researched. However, growing evidence that T. vaginalis infection is associated with other disease states with high morbidity in both men and women has increased the efforts to diagnose and treat patients harboring this parasite. The pathology of trichomoniasis results from damage to the host epithelia, caused by a variety of processes during infection and recent work has highlighted the complex interactions between the parasite and host, commensal microbiome and accompanying symbionts. The commercial release of a number of nucleic acid amplification tests (NAATs) has added to the available diagnostic options. Immunoassay based Point of Care testing is currently available, and a recent initial evaluation of a NAAT Point of Care system has given promising results, which would enable testing and treatment in a single visit

Common controversies in total knee replacement surgery

Total knee replacement (TKR) is an operation that has radically improved the quality of life of millions of people during the last decades. However, some technical details, concerning the surgical procedure, are still a matter of a strong debate. In this review of the literature, we have included the best evidence available of the last decade, related to TKR surgery like the use of tourniquet during operation, patellar resufacing, all-polyethilene tibial component, cemented or cementless fixation, mobile bearings, or posterior-stabilized versus cruciate-retaining prosthesis.YesLa artroplastia total de rodilla (ATR) es una operación que ha mejorado radicalmente la calidad de vida de millones de personas en las últimas décadas. Sin embargo, algunos detalles técnicos, relativos al procedimiento quirúrgico, son todavía un tema de fuerte debate. En esta revisión de la literatura se ha incluido la mejor evidencia al respecto de la última década, sobre algunos de los temas más controvertidos de este procedimiento como el uso de torniquete, gestos quirúrgicos sobre la rótula, bandeja tibial toda de polietileno (all-poly) o con respaldo metálico, la fijación cementada o sin cementar, el uso de platillos móviles, ó la conservación / sustitución del ligamento cruzado posterior

Deciphering the Non-Equivalence of Serine and Threonine O-glycosylation Points: Implications for Molecular Recognition of the Tn Antigen by an anti-MUC1 Antibody

© 2015 The Authors. The structural features of MUC1-like glycopeptides bearing the Tn antigen (α-O-GalNAc-Ser/Thr) in complex with an anti MUC-1 antibody are reported at atomic resolution. For the α-O-GalNAc-Ser derivative, the glycosidic linkage adopts a high-energy conformation, barely populated in the free state. This unusual structure (also observed in an α-S-GalNAc-Cys mimic) is stabilized by hydrogen bonds between the peptidic fragment and the sugar. The selection of a particular peptide structure by the antibody is thus propagated to the carbohydrate through carbohydrate/peptide contacts, which force a change in the orientation of the sugar moiety. This seems to be unfeasible in the α-O-GalNAc-Thr glycopeptide owing to the more limited flexibility of the side chain imposed by the methyl group. Our data demonstrate the non-equivalence of Ser and Thr O-glycosylation points in molecular recognition processes. These features provide insight into the occurrence in nature of the APDTRP epitope for anti-MUC1 antibodies