Pyloric atresia and Down's syndrome: prenatal double bubble false sign. / Atresia pilórica y síndrome de Down: falso signo de la doble burbuja prenatal.

Pyloric atresia is a rare malformation, with an incidence of 1:100,000 live newborns. Male to female ratio is 1/1. Typically, it is an isolated malformation, with a good prognosis, but 20-40% of cases present epidermolysis bullosa, and to a lesser extent, multiple intestinal atresias. We present the case of a pre-term newborn prenatally diagnosed with polyhydramnios, duodenal atresia with "double bubble" sign, and suspected Down's syndrome, who eventually had pyloric atresia.

La atresia pilórica es una malformación rara, presenta una incidencia de 1:100.000 recién nacidos vivos y la ratio hombre/mujer es de 1/1. Generalmente es una malformación aislada, con buen pronóstico, pero entre el 20-40% de los casos se asocia a epidermólisis bullosa y en menor frecuencia a otras atresias intestinales múltiples. Presentamos un caso de recién nacido pretérmino con atresia pilórica con el diagnóstico prenatal de polihidramnios, atresia duodenal con signo de 'doble burbuja' y sospecha de síndrome de Down.

Chemokines cooperate with TNF to provide protective anti-viral immunity and to enhance inflammation.

The role of cytokines and chemokines in anti-viral defense has been demonstrated, but their relative contribution to protective anti-viral responses in vivo is not fully understood. Cytokine response modifier D (CrmD) is a secreted receptor for TNF and lymphotoxin containing the smallpox virus-encoded chemokine receptor (SECRET) domain and is expressed by ectromelia virus, the causative agent of the smallpox-like disease mousepox. Here we show that CrmD is an essential virulence factor that controls natural killer cell activation and allows progression of fatal mousepox, and demonstrate that both SECRET and TNF binding domains are required for full CrmD activity. Vaccination with recombinant CrmD protects animals from lethal mousepox. These results indicate that a specific set of chemokines enhance the inflammatory and protective anti-viral responses mediated by TNF and lymphotoxin, and illustrate how viruses optimize anti-TNF strategies with the addition of a chemokine binding domain as soluble decoy receptors.

Full Interchangeability in Regard to Immunogenicity Between the Infliximab Reference Biologic and Biosimilars CT-P13 and SB2 in Inflammatory Bowel Disease.

Background: Infliximab (IFX) biosimilars CT-P13 and SB2 have comparable efficacy, safety, and immunogenicity to the originator Remicade (RMC). However, concerns about cross-switching patients between the 3 brands were raised in the absence of cross reactivity data between them. We aimed to determine whether antibodies to infliximab (ATI) in inflammatory bowel disease (IBD) patients cross-react with RMC, CT-P13, and SB2. Methods: Based on previous ATI status, samples from 34 patients participating in the BIOSIM01 study (13 RMC, 9 CT-P13, and 12 switchers) were selected. Patients were treated with either RMC only, or CT-P13 only, or with RMC switched to CT-P13. Additionally, 28 IFX-naïve patients were assayed as controls. In total, 180 samples were analyzed. ATI trough levels were measured in parallel with 3 different bridging Enzyme Linked Immunosorbent Assays constructed using the 3 drugs. Spearman's coefficient and percentages of agreement were used to study the correlation between each assay. Results: In total, 76 samples out of 152 IFX-treated patient samples were ATI-positive (30 RMC, 14 CT-P13, and 32 switchers). All resulted ATI-positive when either CT-P13 or SB2 bridging assays were used. The overall percentage of agreement was 100% when compared either with CT-P13 or SB2 assays. No significant differences were found among ATI levels and coefficients (Spearman's 0.98 to 1.0, P < 0.0001). Conclusions: ATI of RMC-treated, CT-P13-treated or RMC to CT-P13 switched patients show full cross-reactivity with CT-P13 and SB2. Findings suggest that immunodominant epitopes in the reference and CT-P13 drugs are equally present in SB2. Data support full interchangeability between biosimilars in regard to immunogenicity.

Antibodies to infliximab in Remicade-treated rheumatic patients show identical reactivity towards biosimilars.

OBJECTIVES: The aim of this study was to determine whether antibodies to infliximab (IFX) in Remicade-treated patients cross-react with the biosimilar CT-P13. METHODS: 250 consecutive patients with rheumatic diseases under Remicade and 77 controls were retrospectively selected for the study. Anti-IFX antibodies at drug through levels were measured in parallel with three different bridging ELISA assays: Promonitor-ANTI-IFX kit, which uses Remicade to detect antibodies, and two more assays that use either Inflectra or Remsima with the same format. Correlation and association between each assay was studied. RESULTS: 50.4% of patients were tested positive with Promonitor-ANTI-IFX. All were antibodies to IFX (ATI)-positive when either Inflectra or Remsima assays were used. In all comparisons positive and negative percentage agreements were 100%, and correlation coefficients were ≥0.995. No differences between rheumatoid arthritis and spondyloarthritis, or between concomitant immunosuppressives, were observed. CONCLUSIONS: Anti-IFX antibodies of Remicade-treated patients cross-react with either Inflectra or Remsima. Although additional epitopes may be present in the biosimilar, results suggest that epitopes influencing the immune response to IFX are also present in the biosimilar. Antibody-positive patients treated with Remicade should not be switched to the biosimilar, since antibodies will interact with the new drug and potentially lead to loss of response. This finding supports the utility for therapeutic drug monitoring before a switching strategy is considered.

[Postcholecystectomy syndrome in children, fact or fiction?] / Síndrome postcolecistectomía en niños ¿realidad o mito?

INTRODUCTION: The postcholecystectomy syndrome (SPC) is broadly defined and published in adults, whereas in the pediatric population are hardly any articles about it. Up to a third of adults have dyspeptic symptoms without organic cause the first year after cholecystectomy. Our goal is to determine the incidence of SPC in our population. METHODS: An observational study was performed, collecting data from patients who had been done laparoscopic cholecystectomy in our hospital since 2005. Patients diagnosed choledochal cyst and biliary atresia were excluded. The following data were collected: type of dyspeptic symptoms, scheduled office visits and emergency units in the first postoperative year and in the following. Children who did not make any visits, a telephone survey was conducted. RESULTS: Data from 36 patients, including 3 patients who were excluded for presenting organic cause, were collected. The most frequent diagnosis was idiopathic cholelithiasis (64,7%). Sixteen children (48,5%) had postoperative symptoms in the first year, of which 14 went to scheduled office visit and 6 emergent (2 required hospitalization). The main symptoms were abdominal postoperative pain (100%), nausea (62,5%) and vomiting (50%). After the first year (6 patients were excluded for less follow-up), only 5 patients (18,5%) continued to symptoms (p= 0,015), 2 required visit to programmatically consultation and no one emergent. CONCLUSION: In our sample, SPC in children exists and improves after the first year. So postoperative follow-up is an important fact, and only further tests must be done if signs of organic cause.

INTRODUCCION: El síndrome postcolecistectomía (SPC) está ámpliamente definido y publicado en adultos, en cambio en la población pediátrica apenas hay artículos al respecto. Hasta un tercio de los adultos presentan síntomas dispépticos sin causa orgánica el primer año después de una colecistectomía. Nuestro objetivo es conocer la incidencia del SPC en nuestro medio. MATERIAL Y METODOS: Se realizó un estudio observacional, recogiendo datos de los pacientes colecistectomizados por laparoscopia en nuestro hospital desde 2005. Se excluyeron pacientes diagnosticados de quiste de colédoco y atresia de vías biliares. Se recogieron los siguientes datos: tipo de síntomas dispépticos, visitas a consulta de forma programada y urgente en el primer año postquirúrgico y en los años sucesivos. Se realizó encuesta telefónica a los pacientes que no efecturaron ninguna visita. RESULTADOS: Se recogieron datos de 36 pacientes, de los cuales se excluyeron 3 pacientes por presentar causa orgánica. El diagnóstico más frecuente fue la colelitiasis idiopática (64,7%). Dieciséis pacientes (48,5%) presentaron síntomas en el primer año postquirúrgico, de los cuales 14 acudieron a consultas de forma programada y 6 urgente (2 precisaron ingreso). Los síntomas principales postquirúrgicos fueron el dolor abdominal (100%), náuseas (62,5%) y vómitos (50%). Tras el primer año (6 pacientes excluidos por seguimiento menor), solo 5 (18,5%) continuaron con los síntomas (p= 0,015), 2 requirieron visita a consultas de forma programada y ninguna urgente. CONCLUSION: Según nuestra muestra, el SPC en niños existe y mejora tras el primer año, por lo que es importarte el seguimiento postquirúrgico de los mismos y solo realizar pruebas complementarias ante signos de causa orgánica.

[Ileostomy closure after necrotizing enterocolitis. When is the best moment?] / Cierre de ileostomía tras enterocolitis necrotizante. ¿Cuándo es el mejor momento?

INTRODUCTION: In more than 50% of the necrotizing enterocolitis that underwent surgery will require an ileostomy. The optimal time to reestablish intestinal transit still is a controversial subject. Many times ileostomies cause medical issues that require early intestinal reconstruction. Our objective is to compare the early closure against late close, being the shift point 35 days according to other published research. MATERIAL AND METHODS: Retrospective study off all patients that in the last 10 years have had an episode of necrotizing enterocolitis which required an intestinal derivation like ileostomy. RESULTS: We studied 39 patients, 22 had an early closure (EC) and 17 in had a late closure (LC). There were statistically significant differences in age and weight between both groups, being younger in the EC group (p<0,05). All the morbidity factors were greater in the EC group (days of parenteral nutrition, days of central venous catheter, inotropic use, surgical wound infection and intestinal occlusions). The days of mechanical ventilation were greater in the EC group (2,33 vs p=0,017). The rate of reoperation was higher in the EC group (31%) against the LE group (17%). CONCLUSIONS: It is necessary to perform prospective studies with larger number of patients to be able to recommend a late closure ileostomy. In our experience the early closure has more morbidity and a higher rate of surgical reoperations.

INTRODUCCION: En más del 50% de las enterocolitis necrotizantes intervenidas es necesario realizar una ileostomía. El tiempo óptimo para restablecer el tránsito intestinal continúa siendo un tema controvertido. En muchas ocasiones las ileostomías dan problemas, requiriendo una reconstrucción precoz. El objetivo es comparar el cierre precoz con el cierre diferido, estableciendo el punto de corte en 35 días, desde el momento de realización del estoma, de acuerdo con otros trabajos publicados así como con la práctica realizada en nuestro hospital.. MATERIAL Y METODOS: Revisión retrospectiva de todos los pacientes que en los últimos diez años han presentado un episodio de enterocolitis necrotizante en nuestro hospital, precisando una derivación intestinal tipo ileostomía y en los que, además, se realizó el cierre de la misma. RESULTADOS: Se han estudiado 39 pacientes, en 22 se realizó un cierre precoz (CP) y en 17 un cierre diferido (CD). En ambos grupos, la edad y el peso presentaron diferencias estadísticamente significativas, siendo menores en el grupo de CP (p<0,05). Todas las variables de morbilidad estudiadas fueron mayores en el grupo de CP (días de nutrición parenteral total, días de catéter venoso central, uso de inotrópicos, infección de herida quirúrgica y oclusiones intestinales). Los días de ventilación mecánica fueron mayores en el grupo CP (2,33 vs 0 p=0,017). La tasa de reintervención quirúrgica fue mayor en el grupo CP (31%) frente al grupo CD (17%). CONCLUSIONES: Es necesario realizar estudios prospectivos y con mayor número de pacientes para poder recomendar un cierre diferido. En nuestra experiencia el cierre precoz presenta mayor morbilidad, así como mayor tasa de reintervenciones.

A new biomarker panel in bronchoalveolar lavage for an improved lung cancer diagnosis.

INTRODUCTION: The enormous biological complexity and high mortality rate of lung cancer highlights the need for new global approaches for the discovery of reliable early diagnostic biomarkers. The study of bronchoalveolar lavage samples by proteomic techniques could identify new lung cancer biomarkers and may provide promising noninvasive diagnostic tools able to enhance the sensitivity of current methods. METHODS: First, an observational prospective study was designed to assess protein expression differences in bronchoalveolar lavages from patients with (n = 139) and without (n = 49) lung cancer, using two-dimensional gel electrophoresis and subsequent protein identification by mass spectrometry. Second, validation of candidate biomarkers was performed by bead-based immunoassays with a different patient cohort (204 patients, 48 controls). RESULTS: Thirty-two differentially expressed proteins were identified in bronchoalveolar lavages, 10 of which were confirmed by immunoassays. The expression levels of APOA1, CO4A, CRP, GSTP1, and SAMP led to a lung cancer diagnostic panel that reached 95% sensitivity and 81% specificity, and the quantification of STMN1 and GSTP1 proteins allowed the two main lung cancer subtypes to be discriminated with 90% sensitivity and 57% specificity. CONCLUSIONS: Bronchoalveolar lavage represents a promising noninvasive source of lung cancer specific protein biomarkers with high diagnostic accuracy. Measurement of APOA1, CO4A, CRP, GSTP1, SAMP, and STMN1 in this fluid may be a useful tool for lung cancer diagnosis, although a further validation in a larger clinical set is required for early stages.

Impact of the endoscopic teaching process on colonic adenoma detection.

BACKGROUND: There has been little reported experience in the Latin American hospital setting in relation to the impact of the endoscopic training process on colonoscopy quality. AIMS: To determine the effect that training in the technique of colonoscopy has on adenoma detection in an Argentinian teaching hospital. MATERIAL AND METHOD: Within the time frame of July 2012 and July 2013, 3 physicians received training in colonoscopy from 4 experienced endoscopists. The colonoscopies performed by the supervised trainees were compared with those carried out by the experienced endoscopists. RESULTS: A total of 318 colonoscopies performed by any one of the 3 supervised trainees and 367 carried out by any one of the experienced endoscopists were included. The univariate analysis showed a non-significant difference in the detection rate of adenomas (30.4 vs. 24.7%, P=.09). In the multivariate analysis, the detection rate of adenomas was significantly higher in the colonoscopies performed by one of the 3 trainees (odds ratio = 1.72 [1.19-2.48]). CONCLUSIONS: The supervised involvement of endoscopic trainees has a positive effect on adenoma detection.

The eukaryotic translation initiation factor eIF4E is a direct transcriptional target of NF-κB and is aberrantly regulated in acute myeloid leukemia.

The eukaryotic translation initiation factor eIF4E is a potent oncogene elevated in many cancers, including the M4 and M5 subtypes of acute myeloid leukemia (AML). Although eIF4E RNA levels are elevated 3- to 10-fold in M4/M5 AML, the molecular underpinnings of this dysregulation were unknown. Here, we demonstrate that EIF4E is a direct transcriptional target of nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) that is dysregulated preferentially in M4 and M5 AML. In primary hematopoietic cells and in cell lines, eIF4E levels are induced by NF-κB activating stimuli. Pharmacological or genetic inhibition of NF-κB represses this activation. The endogenous human EIF4E promoter recruits p65 and cRel to evolutionarily conserved κB sites in vitro and in vivo following NF-κB activation. Transcriptional activation is demonstrated by recruitment of p300 to the κB sites and phosphorylated Pol II to the coding region. In primary AML specimens, generally we observe that substantially more NF-κB complexes associate with eIF4E promoter elements in M4 and M5 AML specimens examined than in other subtypes or unstimulated normal primary hematopoietic cells. Consistently, genetic inhibition of NF-κB abrogates eIF4E RNA levels in this same population. These findings provide novel insights into the transcriptional control of eIF4E and a novel molecular basis for its dysregulation in at least a subset of M4/M5 AML specimens.

The experience in nicaragua: childhood leukemia in low income countries-the main cause of late diagnosis may be "medical delay".

Background. The event-free survival for pediatric leukemia in low-income Countries is much lower than in high-income countries. Late diagnosis, which is regarded as a contributing factor, may be due to "parental" or "medical" delay. Procedures. The present study analyses determinants of lag time from first symptoms to diagnosis of leukemia, comparing pediatric (0-16 years old) patients in two referral centers, one in Nicaragua and one in Italy. An observational retrospective study was conducted to assess factors influencing the time to diagnosis. Results. 81 charts of children diagnosed with acute myeloid leukemia or lymphoblastic leukemia were analyzed from each centre. Median lag time to diagnosis was higher in Nicaragua than in Italy (29 versus 14 days, P < 0.001) and it was mainly due to "physician delay" (16.5 versus 7 days, P < 0.001), whereas "patient delay" from symptoms to first medical assessment was similar in the two centers (7 versus 5 days, P = 0.27). Moreover, median lag time from symptoms to diagnosis was decreased in Nicaraguan districts were a specific training program upon childhood oncological diseases was carried out (20.5 versus 40 days, P = 0.0019). Conclusions. Our study shows that delay in diagnosis of childhood leukemia is mainly associated with "physician delay" and it may be overcome by programs of continuous medical education.

Perfiles de personalidad de estudiantes de psicología de universidad privada comparados em similitud y congruencia / Personality profiles of psychology students from a privateuniversity compared by similarity and congruence

En continuidad con los estudios del área de Perfiles de la Cátedra de Psicología Experimental I y II, se llevó a cabo un análisis comparativo de Perfiles de Personalidad en estudiantes de psicología. La muestra fue intencional y autoseleccionada, quedó conformada por 153 estudiantes de Psicología. Se ha utilizado un diseño Descriptivo y Comparado. El instrumento utilizado fue el Inventario Multifásico de la Personalidad Minnesota-2 (MMPI-2). Los resultados reportan que existe un patrón de personalidad con similitudes muy acentuadas en los estudiantes de la carrera de psicología, independiente del curso al que pertenecen, y en comparación a los perfiles de años anteriores (n = 152, muestra 2001: n=168, muestra 2007: y n= 101 muestra 2008).

In continuity with the studies of the area of Profiles of the Professorship of Experimental Psychology I and II, a comparative analysis of Profiles of Personality in students of psychology was carried out. The sample was intentional and autoseleccionada, remained conformed by 153 (166) students of Psychology. A Descriptive design has been utilized and Compared. The instrument utilized was the Polyphase Inventory of the Personality Minnesota-2 (MMPI-2). The results report that a boss of personality with similarities exists very accentuated in the students of the career of psychology, independent of the course to which they belong, and in comparison to the profiles of previous years (n = 152 sample 2001; n = 168, sample 2007 and n = 101 sample 2008).

[Incorporation of a psychologist into a nephrology service: criteria and process]. / Incorporación de un psicólogo en un servicio de nefrología: criterios y proceso.

Chronic kidney disease is associated with a wide range of stressful situations causing important physical and psychological repercussions. It is not usual that psychology professionals are active members of the nephrology teams. In consequence, these alterations are not properly assisted. Our aim is to present the introduction process of a psychologist in a nephrology department and its preliminary results. We designed a clearly defined introduction process, starting with a therapeutic communication training program for all the staff. In the model we have priorized pre-emptive interventions in order to promote the adaptation process, far from simple psychological symptom control. It is assumed the binomial patient-family as the major objective for care, choosing an interdisciplinary approach. We worked more from a health psychology perspective than from a mental health perspective. Over the year 2008 the number of patients assisted by the psychologist were 571 (mean 48 patients/month). The total number of interventions was 1,022. Majority of cases (45.2%) were derived from the advanced chronic kidney disease program, mostly related to demands about emotional impact of renal replacement therapy commencement. Others were: suspect of depression episode, adherence, primary caregiver emotional overwhelming, bereavement, anxiety and support in decision making process. This experience is a stimulus for the integral approach of the renal patient.

An ectromelia virus protein that interacts with chemokines through their glycosaminoglycan binding domain.

Poxviruses encode a number of secreted virulence factors that modulate the host immune response. The vaccinia virus A41 protein is an immunomodulatory protein with amino acid sequence similarity to the 35-kDa chemokine binding protein, but the host immune molecules targeted by A41 have not been identified. We report here that the vaccinia virus A41 ortholog encoded by ectromelia virus, a poxvirus pathogen of mice, named E163 in the ectromelia virus Naval strain, is a secreted 31-kDa glycoprotein that selectively binds a limited number of CC and CXC chemokines with high affinity. A detailed characterization of the interaction of ectromelia virus E163 with mutant forms of the chemokines CXCL10 and CXCL12alpha indicated that E163 binds to the glycosaminoglycan binding site of the chemokines. This suggests that E163 inhibits the interaction of chemokines with glycosaminoglycans and provides a mechanism by which E163 prevents chemokine-induced leukocyte migration to the sites of infection. In addition to interacting with chemokines, E163 can interact with high affinity with glycosaminoglycan molecules, enabling E163 to attach to cell surfaces and to remain in the vicinity of the sites of viral infection. These findings identify E163 as a new chemokine binding protein in poxviruses and provide a molecular mechanism for the immunomodulatory activity previously reported for the vaccinia virus A41 ortholog. The results reported here also suggest that the cell surface and extracellular matrix are important targeting sites for secreted poxvirus immune modulators.

Manipulation of the nuclear factor-kappaB pathway and the innate immune response by viruses.

Viral and microbial constituents contain specific motifs or pathogen-associated molecular patterns (PAMPs) that are recognized by cell surface- and endosome-associated Toll-like receptors (TLRs). In addition, intracellular viral double-stranded RNA is detected by two recently characterized DExD/H box RNA helicases, RIG-I and Mda-5. Both TLR-dependent and -independent pathways engage the IkappaB kinase (IKK) complex and related kinases TBK-1 and IKKvarepsilon. Activation of the nuclear factor kappaB (NF-kappaB) and interferon regulatory factor (IRF) transcription factor pathways are essential immediate early steps of immune activation; as a result, both pathways represent prime candidates for viral interference. Many viruses have developed strategies to manipulate NF-kappaB signaling through the use of multifunctional viral proteins that target the host innate immune response pathways. This review discusses three rapidly evolving areas of research on viral pathogenesis: the recognition and signaling in response to virus infection through TLR-dependent and -independent mechanisms, the involvement of NF-kappaB in the host innate immune response and the multitude of strategies used by different viruses to short circuit the NF-kappaB pathway.

Magnetic resonance urography: a pictorial overview.

Magnetic resonance urography (MRU) can be performed on the basis of two different imaging strategies: static-fluid MRU, based on heavily T2 weighted turbo spin echo (TSE) sequences, and gadolinium-enhanced excretory MRU. Both MR urographic techniques in combination with standard MRI permit a comprehensive examination of the entire urinary tract. This pictorial review illustrates the MRU features of the a wide spectrum of pathological conditions affecting the urinary tract.

A chemokine-binding domain in the tumor necrosis factor receptor from variola (smallpox) virus.

Variola virus (VaV) is the causative agent of smallpox, one of the most devastating diseases encountered by man, that was eradicated in 1980. The deliberate release of VaV would have catastrophic consequences on global public health. However, the mechanisms that contribute to smallpox pathogenesis are poorly understood at the molecular level. The ability of viruses to evade the host defense mechanisms is an important determinant of viral pathogenesis. Here we show that the tumor necrosis factor receptor (TNFR) homologue CrmB encoded by VaV functions not only as a soluble decoy TNFR but also as a highly specific binding protein for several chemokines that mediate recruitment of immune cells to mucosal surfaces and the skin, sites of virus entry and viral replication at late stages of smallpox. CrmB binds chemokines through its C-terminal domain, which is unrelated to TNFRs, was named smallpox virus-encoded chemokine receptor (SECRET) domain and uncovers a family of poxvirus chemokine inhibitors. An active SECRET domain was found in another viral TNFR (CrmD) and three secreted proteins encoded by orthopoxviruses. These findings identify a previously undescribed chemokine-binding and inhibitory domain unrelated to host chemokine receptors and a mechanism of immune modulation in VaV that may influence smallpox pathogenesis.

Both soluble and membrane-anchored forms of Felid herpesvirus 1 glycoprotein G function as a broad-spectrum chemokine-binding protein.

Recently, glycoprotein G (gG) of several alphaherpesviruses infecting large herbivores was shown to belong to a new family of chemokine-binding proteins (vCKBPs). In the present study, the function of Felid herpesvirus 1 (FeHV-1) gG as a vCKBP was investigated and the following conclusions were reached: (i) FeHV-1 secreted gG is a high-affinity broad-spectrum vCKBP that binds CC, CXC and C chemokines; (ii) gG is the only vCKBP expressed by FeHV-1 that binds CCL3 and CXCL1; (iii) secreted gG blocks chemokine activity by preventing their interaction with high-affinity cellular receptors; (iv) the membrane-anchored form of gG expressed on the surface of infected cells is also able to bind chemokines; and (v) the vCKBP activity is conserved among different field isolates of FeHV-1. Altogether, these data demonstrate that FeHV-1 gG is a new member of the vCKBP-4 family. Moreover, this study is the first to demonstrate that gG expressed at the surface of FeHV-1-infected cells can also bind chemokines.

Thermostability of the human respiratory syncytial virus fusion protein before and after activation: implications for the membrane-fusion mechanism.

Anchorless fusion (F) proteins () of human respiratory syncytial virus (RSV) are seen by electron microscopy as unaggregated cones when the proteolytic cleavage at two furin sites required for membrane-fusion activity is incomplete, but aggregate into rosettes of lollipop-shaped spikes following cleavage. To show that this aggregation occurred by interactions of the fusion peptide, a deletion mutant of lacking the first half of the fusion peptide was generated. This mutant remained unaggregated even after completion of cleavage, supporting the notion that aggregation of involved the fusion peptide. As exposure of the fusion peptide is a key event that occurs after activation of F proteins, the uncleaved and cleaved forms of may represent the pre- and post-active forms of RSV F protein. In an analysis of the structural differences between the two forms, their thermostability before and after proteolytic cleavage was examined. In contrast to other viral proteins involved in membrane fusion (e.g. influenza haemagglutinin), the pre-active (uncleaved) and post-active (cleaved) forms of were equally resistant to heat denaturation, assessed by spectrofluorimetry, circular dichroism or antibody binding. These results are interpreted in terms of the proposed structural changes associated with the process of membrane fusion mediated by RSV F protein.