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The innate immune system provides a primary line of defense against pathogens. Stimulator of IFN genes (STING), encoded by the TMEM173 gene, is a critical protein involved in IFN-b induction in response to infection by different pathogens. In this study, we describe the expression of three different alternative-spliced human (h) TMEM173 mRNAs producing STING truncated isoforms 1, 2, and 3 in addition to the full-length wild-type (wt) hSTING. All of the truncated isoforms lack exon 7 and share the N-terminal transmembrane region with wt hSTING. Overexpression of the three STING truncated isoforms failed to induce IFN-b, and they acted as selective pathway inhibitors of wt hSTING even in combination with upstream inducer cyclic-di-GMP-AMP synthase. Truncated isoforms alter the stability of wt hSTING, reducing protein t1/2 to some extent by the induction of proteasome-dependent degradation. Knocking down expression of truncated isoforms increased production of IFN-b by THP1 monocytes in response to intracellular cytosolic DNA or HSV-1 infection. At early stages of infection, viruses like HSV-1 or vesicular stomatitis virus reduced the ratio of full-length wt hSTING/truncated STING isoforms, suggesting the skewing of alternative splicing of STING toward truncated forms as a tactic to evade antiviral responses. Finally, in silico analysis revealed that the human intron–exon gene architecture of TMEM173 (splice sites included) is preserved in other mammal species, predominantly primates, stressing the relevance of alternative splicing in regulating STING antiviral biology.

Durmart le Galois no es una novela artúrica convencional. Los caballeros de la Tabla Redonda son relegados a un segundo plano frente al protagonismo que cobra el héroe epónimo. Su estructura carece de una etapa de crisis personal o social del héroe, de forma que la aventura constituye un itinerario cuyas etapas se despliegan a través de una vasta geografía. Los castillos y ciudades se imponen sobre otros espacios salvajes, como el bosque, más frecuentes en otras obras caballerescas. La descripción de estos lugares construidos parte de los cánones establecidos por la tradición literaria, pero se aleja después y muestra un espacio amable, abierto a la sociabilidad y al entretenimiento. El ambiente festivo inunda la novela y encuentra su culminación en el banquete, con sus rituales y su protocolo. En torno a la mesa, los comensales intercambian conversaciones que marcarán el progreso de la intriga. Durmart le Galois is not a conventional Arthurian novel. The knights of the Round Table are pushed into the background, while relevance is given to the eponymous hero. Its structure lacks a stage of personal or social crisis of the hero, so, adventure covers an itinerary whose stages are displayed throughout a vast geography. Castles and cities replace other wild spaces, such as the forest, which are more frequent in other chivalric works. The description of these built-up spaces starts from the pre-established literary canons, but later it moves away from conventions, and it depicts a gentle space, which is open to sociability and entertainment. A merry atmosphere floods the novel, and it reaches its climax in the banquet, with its rituals and protocol. Around the table, guests exchange conversations, which will influence the unfolding of the plot.

Decisions on transport policy measures proposed by the European Union (EU) have long-term and important impacts on economy, environment and society. Transport policy measures can lock up capital for decades and cause manifold external effects – thus, policy measures may have a tremendous scope, especially if proposed on a European level. In order to allow European policy-makers to evaluate transport polices, a strategic assessment tool has been developed to compute economic, environmental and social impacts of transport policies.

The frequency, distribution and impact of small ruminant Sarcocystis infection in the European Union is largely unknown; this study reports the prevalence of macroscopic Sarcocystis sp. cysts and associated carcass condemnation, in 6065 adult, cull, small ruminants from 145 farms in Spain. Macrocysts were detected in 12% of sheep from 60% flocks, and in none of the 345 goats examined. Most affected sheep had cysts in more than one body part and as a result, 79% of sheep carcasses with cysts were totally condemned. Consequently, it is estimated that Sarcocystis spp. infection could be costing the Spanish sheep industry € 20 million yr-1. Three types of cysts were identified according to size, shape and location: narrow, filament-shaped measuring 2-10 × ≤1 mm, present striated muscles only, and two wider types measuring 2-20 × 2-6 mm, including oval-shaped oesophageal cysts and more elongated cysts in striated muscles. Narrow and wide macrocysts were found in the same sheep and are compatible with Sarcocystis gigantea and Sarcocystis medusiformis, respectively, as described in New Zealand in the 1970s. However, cyst size and morphology varies with age and location. Moreover, S. medusiformis has not been reported in Europe and species-specific diagnosis is necessary to ascertain the ethiology of macrocysts in this study., Apenas existe información sobre la frecuencia, distribución e impacto actual de la sarcocistiosis en la Unión Europea; este trabajo describe la prevalencia de quistes macroscópicos de Sarcocystis spp. y el porcentaje de decomisos asociados a los quistes, en 6.065 pequeños rumiantes adultos de desvieje, de 145 granjas de España. Se observaron quistes en 12% de ovejas de 60% de rebaños, y en ninguna de las 345 cabras examinadas. La mayoría de las ovejas afectadas presentó quistes en varias zonas corporales decomisándose el 79% de las canales afectadas. Según esto, las pérdidas por la infección por Sarcocystis spp. para la industria ovina española se estimaron en 20 millones de euros año–1. Se observaron tres tipos de quistes según el tamaño, forma y localización: estrechos, filiformes, de 2-10 × ≤ 1 mm, en músculo estriado solamente y dos tipos de quistes anchos, de 2-20 × 2-6 mm, ovales en esófago y los mas elongados en musculatura estriada, que podrían corresponderse con las especies Sarcocystis gigantea y Sarcocystis medusiformis, descritas en Nueva Zelanda en la década de 1970. Sin embargo, sería necesario un diagnóstico específico para confirmar la etiología de los quistes de este estudio, ya que el tamaño y morfología de éstos varía según la edad y la localización, y S. medusiformis no está descrita en Europa.

The high levels of interleukin 10 (IL-10) present in chronic hepatitis C virus (HCV) infection have been suggested as responsible for the poor antiviral cellular immune responses found in these patients. To overcome the immunosuppressive effect of IL-10 on antigen-presenting cells such as dendritic cells (DCs), we developed peptide inhibitors of IL-10 to restore DC functions and concomitantly induce efficient antiviral immune responses. Two IL-10-binding peptides (p9 and p13) were selected using a phage-displayed library and their capacity to inhibit IL-10 was assessed in a bioassay and in STAT-3 (signal transducer and activator of transcription 3) phosphorylation experiments in vitro. In cultures of human leukocytes where HCV core protein induces the production of IL-10, p13 restored the ability of plasmacytoid DC to produce interferon alpha (IFN-α) after Toll-like receptor 9 (TLR9) stimulation. Similarly, when myeloid DCs were stimulated with CD40L in the presence of HCV core, p9 enhanced IL-12 production by inhibiting HCV core-induced as well as CD40L-induced IL-10. Moreover, in vitro, p13 potentiated the effect of maturation stimuli on human and murine DC, increasing their IL-12 production and stimulatory activity, which resulted in enhanced proliferation and IFN-γ production by responding T-cells. Finally, immunization with p13-treated murine DC induced stronger anti-HCV T-cell responses not only in wildtype mice but also in HCV transgenic mice and in mice transiently expressing HCV core in the liver. CONCLUSION: These results suggest that IL-10 inhibiting peptides may have important applications to enhance anti-HCV immune responses by restoring the immunostimulatory capabilities of DC.

Broad T cell and B cell responses to multiple HCV antigens are observed early in individuals who control or clear HCV infection. The prevailing hypothesis has been that similar immune responses induced by prophylactic immunization would reduce acute virus replication and protect exposed individuals from chronic infection. Here, we demonstrate that immunization of naïve chimpanzees with a multicomponent HCV vaccine induced robust HCV-specific immune responses, and that all vaccinees exposed to heterologous chimpanzee-adapted HCV 1b J4 significantly reduced viral RNA in serum by 84%, and in liver by 99% as compared to controls (P=0.024 and 0.028, respectively). However, despite control of HCV in plasma and liver in the acute period, in the chronic phase, 3 of 4 vaccinated animals developed persistent infection. Analysis of expression levels of proinflammatory cytokines in serial hepatic biopsies failed to reveal an association with vaccine outcome. However, expression of IDO, CTLA-4 [corrected] and PD-1 levels in liver correlated with clearance or chronicity. CONCLUSION: Despite early control of virus load, a virus-associated tolerogenic-like state can develop in certain individuals independent of vaccination history.

Indoleamine 2,3-dioxygenase (IDO) is induced by proinflammatory cytokines and by CTLA-4-expressing T cells and constitutes an important mediator of peripheral immune tolerance. In chronic hepatitis C, we found upregulation of IDO expression in the liver and an increased serum kynurenine/tryptophan ratio (a reflection of IDO activity). Huh7 cells supporting hepatitis C virus (HCV) replication expressed higher levels of IDO mRNA than noninfected cells when stimulated with gamma interferon or when cocultured with activated T cells. In infected chimpanzees, hepatic IDO expression decreased in animals that cured the infection, while it remained high in those that progressed to chronicity. For both patients and chimpanzees, hepatic expression of IDO and CTLA-4 correlated directly. Induction of IDO may dampen T-cell reactivity to viral antigens in chronic HCV infection

During a viral infection, binding of viral double-stranded RNAs (dsRNAs) to the cytosolic RNA helicase RIG-1 leads to recruitment of the mitochondria-associated Cardif protein, involved in activation of the IRF3-phosphorylating IKKepsilon/TBK1 kinases, interferon (IFN) induction, and development of the innate immune response. The hepatitis C virus (HCV) NS3/4A protease cleaves Cardif and abrogates both IKKepsilon/TBK1 activation and IFN induction. By using an HCV replicon model, we previously showed that ectopic overexpression of IKKepsilon can inhibit HCV expression. Here, analysis of the IKKepsilon transcriptome profile in these HCV replicon cells showed induction of several genes associated with the antiviral action of IFN. Interestingly, IKKepsilon still inhibits HCV expression in the presence of neutralizing antibodies to IFN receptors or in the presence of a dominant negative STAT1alpha mutant. This suggests that good IKKepsilon expression levels are important for rapid activation of the cellular antiviral response in HCV-infected cells, in addition to provoking IFN induction. To determine the physiological importance of IKKepsilon in HCV infection, we then analyzed its expression levels in liver biopsy specimens from HCV-infected patients. This analysis also included genes of the IFN induction pathway (RIG-I, MDA5, LGP2, Cardif, TBK1), and three IKKepsilon-induced genes (IFN-beta, CCL3, and ISG15). The results show significant inhibition of expression of IKKepsilon and of the RNA helicases RIG-I/MDA5/LGP2 in the HCV-infected patients, whereas expression of TBK1 and Cardif was not significantly altered. In conclusion, given the antiviral potential of IKKepsilon and of the RNA helicases, these in vivo data strongly support an important role for these genes in the control of HCV infection.

Systems for in vitro culture of Hepatitis C virus (HCV) are essential tools to analyse virus-cell interactions and to investigate relevant pathophysiological aspects of HCV infection. Although the HCV replicon methodology has increased our understanding of HCV biology, this system does not reproduce the natural infection. Recently, tupaia (Tupaia belangeri chinensis) hepatocytes have been utilized for in vitro culture of HCV. In the present work, primary tupaia hepatocytes infected in vitro with HCV were used to analyse the evolution of HCV quasispecies in infected cells and the ability of the virus to influence antiviral and proinflammatory responses in cells sustaining virus replication. The results confirmed the potential of tupaia hepatocytes as a model for HCV infection, although this system is limited by rapid loss of differentiated cell phenotype in culture. These findings revealed an extraordinary plasticity of HCV quasispecies, which underwent rapid evolution to tupaia-tropic variants as early as 24 h after infection. It was also shown that HCV could activate interferon-sensitive genes, albeit modestly in comparison with other viruses such as Semliki Forest virus. Importantly, HCV activated NF-kappaB in primary hepatocytes and upregulated NF-kappaB-responsive genes including the chemokines MCP-1 and CXCL2 (MIP-2). This effect may play a role in induction of the hepatic inflammatory reaction in vivo. In summary, HCV quasispecies adapt rapidly to the specific biology of the host and HCV stimulates a blunted interferon response while inducing a proinflammatory phenotype in the infected cell.

Se presenta la experiencia de la Maternidad del Hospital Clínico de la Universidad de Chile de un programa diseñado especialmente con el objetivo de disminuir las cesáreas evitables. Tomando el porcentaje de cesáreas del trimestre enero-marzo de 2003 como base. Se efectuó un programa de intervención entre los meses de abril-septiembre de 2003, que considera: entrega diaria del turno de residencia, segunda opinión para operación cesárea, diagnóstico pormenorizado de pelvis en ficha clínica, auditoría mensual de cesáreas "evitables" y estandarización de un protocolo de presentaciones distócicas. El propósito del estudio fue evaluar el efecto de la introducción de un sistema de auditoría en la reducción de la tasa de cesáreas, según propuesta de Robson y cols. El porcentaje global de cesáreas disminuyó de 44,9% a 37,1%, en nulíparas de 39,0% a 29,5% y en multíparas de 48,1% a 42,2%, diferencias estadísticamente significativas SUMMARY The experience at the Maternity Ward of Hospital Clínico de la Universidad de Chile in reducing avoidable cesarean sections is presented. The purpose of this study is to analyze the effects of an audit cycle in the cesarean section rate, using the proposal of Robson et al. A specifically designed program is used. Using the cesarean section rate of january to march trimester as a baseline, we designed an intervention protocol for april to september period which involves: daily resident report, second opinion for cesarean section, detailed pelvis diagnosis in the medical file, monthly "avoidable" cesarean section audit cycle and abnormal lie protocol standardization. The overall cesarean section rate was reduced from 44.9% to 37.1%, in nulliparous from 39.0% to 29.5% and in multiparous from 48.1% to 42.2%, all these differences were statistically significant

Background: Ultrasonography is useful in trauma patients to detect pleural effusions or peritoneal fluid. Aim: To assess the value of ultrasonography performed by surgeons in the assessment of trauma patients. Material and methods: A retrospective review of ultrasonography reports and clinical history of 284 trauma patients. Results: One hundred fifty six patients had blunt trauma and 128 had penetrating trauma. Ultrasonography detected peritoneal fluid in 20%, pericardial effusion in 1% and pleural effusion in 1%. Eight percent had visceral damage or hematomas, without peritoneal fluid. None of the patients with a normal ultrasonography required surgery for hemoperitoneum; however, four patients had intestinal perforations and required surgery. Conclusions: Ultrasonography had a 100% sensitivity and specificity for the detection of clinically significant hemoperitoneum. Emergency ultrasonography performed by surgeons is useful and accurate (Rev Med Chile 2002; 130: 892-6)

Interferon (IFN)-alpha is a family of antiviral proteins encoded by different genes. The biological significance of the existence of various IFN-alpha subtypes is not clear. We have investigated the interferon system in chronic hepatitis C virus (HCV) infection, a disease that responds to interferon-alpha2 therapy in only a limited proportion of cases. We analysed the expression of interferon regulatory factor (IRF)-1, IRF-2, and IFN-alpha subtypes in nonstimulated and Sendai virus-stimulated peripheral blood mononuclear cells (PBMC) from HCV infected patients and healthy controls. We observed that the IRF-1 mRNA and IRF-1/IRF-2 ratios were increased in PBMC from hepatitis C patients with respect to normal subjects. Sendai virus stimulation of PBMC led to a significant increase in the levels of IRF-1, IRF-2 and IFN-alpha mRNAs and in the production of IFN-alpha protein with respect to basal values in healthy controls as well as in patients with HCV infection. In addition, we found that while natural HCV infection induced increased IFN-alpha5 expression in PBMC, in vitro infection of these cells with Sendai virus caused a raise in the expression of IFN-alpha8 in both patients and normal controls. In summary, our results indicate that virus-induced activation of the IFN system in human PBMC is associated with selective expression of individual IFN-alpha subtypes, IFN-alpha5 being the specific subtype induced in PBMC from patients with chronic HCV infection.

The mechanisms of liver damage in chronic hepatitis C virus (HCV) infection are poorly understood. The transcription factor, nuclear factor-kappa B (NF-kappa B), regulates the expression of genes involved in apoptosis, inflammation, and antiviral response. It plays a protective role in several forms of liver damage. In this study, we analyzed NF-kappa B by gel mobility shift assay and immunohistochemistry in liver biopsies from HCV-infected patients, and we have determined the hepatic levels of the components of the NF-kappa B system by semiquantitative polymerase chain reaction (PCR). We found that NF-kappa B was activated in the liver of patients with chronic hepatitis C. Neither NF-kappa B activity nor the RNA levels of NF-kappa B subunits showed correlation with liver inflammatory activity, viral load, or HCV genotype. By contrast, hepatic mRNA values of RelA, the main element of active NF-kappa B, correlated inversely with apoptosis (r = -.68; P

Exposure of human peripheral blood mononuclear cells (PBMC) to the stilbene derivative DIDS (4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid) (60 microM and above) significantly increased the release of tumor necrosis factor-alpha (TNF-alpha), as determined by TNF-alpha activity in the incubation media. When the TNF-alpha message was analyzed in PBMC by a reverse transcription/polymerase chain reaction (RT/PCR)-based procedure, it was found that incubation with DIDS (60 microM) was followed by a time-dependent accumulation of TNF-alpha mRNA. Measurements of intracellular pH showed that the presence of increasing concentrations of DIDS resulted in a progressive intracellular alkalinization of PBMC. It is suggested that the known DIDS effect of inhibiting transmembrane anion exchange, i.e., chloride/bicarbonate exchange, might play a role in the stimulation of TNF-alpha production by PBMC exposed to DIDS.