Characterization of a new human monoclonal antibody directed against the Vel antigen.

BACKGROUND AND OBJECTIVES: The Vel blood group antigen is a poorly characterized high-prevalence antigen. Until now, anti-Vel antibodies have been observed in only alloimmunized Vel-negative individuals. In this study, we aimed to establish a human hybridoma cell line secreting the first anti-Vel monoclonal antibody (mAb), clone SpG213Dc. MATERIALS AND METHODS: Peripheral blood lymphocytes from a French Vel-negative woman with anti-Vel in her plasma were transformed with Epstein-Barr virus and then hybridized with the myeloma cell line Sp2/O-Ag14 using the polyethylene glycol (PEG) method. A specific anti-Vel mAb was successfully produced and was extensively characterized by serological, flow cytometry and Western blot analyses. RESULTS: One human anti-Vel-secreting clone was produced and the secreted anti-Vel mAb (SpG213Dc) was examined. The specificity of the SpG213Dc mAb was assessed by its reactivity against a panel of nine genotyped RBCs including, respectively, three Vel-negative and six Vel-positive (three wild-type homozygous and three heterozygous) samples using flow cytometry method. Vel-positive RBCs were specifically stained and were subsequently used to perform Western blot and immunoprecipitation analysis of the Vel antigen. CONCLUSION: Serological characterization of the new monoclonal anti-Vel SpG213Dc showed a heterogeneous level of expression of the Vel antigen on the different RBCs. Our results suggest that the mAb SpG213Dc can be reliably used as a blood grouping reagent, thus allowing the mass-scale phenotyping of blood donors to strengthen rare blood banks with Vel-negative RBC units.

The use of computed tomography in identifying radiologically occult hip fractures in the elderly.

INTRODUCTION: Fractured neck of femur (NOF) is a cause of significant morbidity and mortality. Approximately 4% of patients with an initial normal hip x-ray in the emergency department (ED) will in fact have an occult fracture. In cases where there is ongoing clinical suspicion of NOF fracture despite a normal hip x-ray, alternative imaging should be used. Although available evidence supports the use of magnetic resonance imaging (MRI) for this, it is often not readily accessible from the ED. In our department, it is common practice to request computed tomography (CT). METHODS: A historical review was undertaken of all patients who presented between October 2007 and January 2011 who had CT requested by ED staff. Patients included in the study were those who presented following low impact trauma in whom fractured NOF was suspected despite a normal x-ray. RESULTS: Of the 65 included patients, fractures (pelvic and hip) were identified in 38 patients on CT. Fractured NOFs were found in 13 patients. Acetabular fractures were found in nine patients, five of whom required further orthopaedic management. One patient went on to have MRI to confirm the diagnosis of an impacted NOF fracture, suspected both on x-ray and CT. Further review was undertaken of the medical notes of discharged patients to identify any who reattended or required further imaging. No such cases were found. CONCLUSIONS: This review has shown the use of CT to be a practical approach to improving the care of patients with occult hip fractures.

Propofol for adult procedural sedation in a UK emergency department: safety profile in 1008 cases.

BACKGROUND: Concerns exist regarding the safe use of propofol by Emergency Physicians for procedural sedation. The World SIVA International Sedation Task Force has recently created an adverse event tool, in an effort to standardize reporting. We present a safety analysis of our use of propofol using this tool. METHOD: Propofol was given according to a previously published guideline. We analysed our dedicated departmental sedation database between December 2006 and March 2012 and cross-examined the original sedation chart for each case recorded. We stratified the identified adverse events according to consensus agreement. RESULTS: Of the 1008 consecutive cases, we identified 11 sentinel (5 cases of hypoxia, 6 of hypotension), 34 moderate, 25 minor, and 3 minimal risk adverse events. There were no adverse outcomes. CONCLUSIONS: Our large series of propofol sedations performed by emergency physicians supports the safety of this practice. The sentinel adverse event rate of 1% that we identify prompts review: we will in future emphasize adherence to the reduced 0.5 mg kg(-1) propofol dose in the elderly, and reconsider our use of metaraminol. We believe that our application of the World SIVA adverse event tool sets a benchmark for further studies.

The impact of a temporary ice rink on a local emergency department service.

Ice skating is becoming more popular throughout the UK, with temporary ice rinks opening in many city centres during holiday periods, especially during Christmas. Data were collected from patients who presented to the local emergency department with injuries sustained on a nearby city-centre temporary ice rink. Injuries related to ice rinks accounted for 0.76% of all emergency department attendances and represented 0.29% of ice rink participants (2.9 per 1000). Women in the older age range sustained the most significant injuries. Our study has shown that the rate of injuries per 1000 ice rink participants is comparable with data recorded when a new ice rink is opened.

Is propofol a safe and effective sedative for relocating hip prostheses?

OBJECTIVE: To explore the safety and efficacy of propofol as a sedative for the relocation of hip prostheses in the emergency department. METHODS: A prospective observational study was performed in 100 patients aged 37-93 years who received sedation with propofol in the emergency department for hip prosthetic relocation. All patients received intravenous titrated morphine prior to radiography, followed by a 1 mg/kg bolus of propofol after adequate preoxygenation. At 60 s, joint relocation was attempted by an independent physician. Data for each patient were recorded, with particular attention given to adverse outcomes. RESULTS: Two patients were excluded because of protocol violation. The use of propofol achieved hip relocation in 94 of the remaining 98 patients (96%). The four unsuccessful cases required general anaesthesia with muscle relaxation. Eight patients experienced a fall in oxygen saturation, four responding to airway repositioning and four requiring brief supplemental ventilation. Four patients became hypotensive and required titrated intravenous boluses of a vasopressor (metaraminol) to restore normal blood pressure. 42 patients required additional doses of propofol, 36 for inadequate sedation and 6 for prolonged reduction attempts. CONCLUSIONS: Significant adverse effects of propofol in this case series were uncommon (12/98 patients) and readily countered. This case series suggests that propofol is a safe and effective sedative for relocating hip prostheses.

Microbial impact of small tributaries on water and shellfish quality in shallow coastal areas.

This study aimed to investigate the impact of small tributaries on seawater and shellfish quality in coastal area subjected to brief episodes leading to fecal contamination. Escherichia coli and F-RNA-specific bacteriophages were selected as fecal indicators and astroviruses were chosen as being representative of pathogens in the human population during winter viral epidemics. A two-dimensional hydrodynamic model was built to simulate the current and dispersion in the model domain, which includes areas uncovered at low tide. The model also includes decay rates to simulate microorganism behavior and assess the influence of fecal input on shellfish quality. The originality lies in the fact that specific features of the study area were considered. Modeling results indicate limited particle movements and long flushing times at the back of the bay, where shellfish are farmed. Computational results showed that under normal conditions, i.e. 94% of the time, when rainfall was less than 10 mm per day, the sector shows acceptable water quality. These results are in agreement with shellfish concentration measured in the field. Under high flow conditions, high concentrations of fecal indicators and astrovirus were measured in the river and tributaries. The corresponding fluxes were over 50 times higher than under normal weather conditions. The location of the shellfish beds near the coast makes them vulnerable and fecal indicators and viruses were detected in shellfish after short rainfall events. Our modeling approach makes a contribution to shellfish management and consumer protection, by indicating the "risk period" as defined by EU regulations. Molecular development such as viral quantification in conjunction with model developments will help to prevent shellfish contamination and thus provide safer products to consumers and an effective tool for shellfish producers.

High stability of yellow fever 17D-204 vaccine: a 12-year restrospective analysis of large-scale production.

We have retrospectively analyzed 12 bulk lots of yellow fever vaccine Stamaril, produced between 1990 and 2002 and prepared from the same seed lot that has been in continuous use since 1990. All vaccine batches displayed identical genome sequence. Only four nucleotide substitutions were observed, compared to previously published sequence, with no incidence at amino-acid level. Fine analysis of viral plaque size distribution was used as an additional marker for genetic stability and demonstrated a remarkable homogeneity of the viral population. The total virus load, measured by qRT-PCR, was also homogeneous pointing out reproducibility of the vaccine production process. Mice inoculated intracerebrally with the different bulks exhibited a similar average survival time, and ratio between in vitro potency and mouse LD(50) titers remained constant from batch-to-batch. Taken together, these data demonstrate the genetic stability of the strain at mass production level over a period of 12 years and reinforce the generally admitted idea of the safety of YF17D-based vaccines.

Fate and characterization of circulating tumor cells in a NOD/SCID mouse model of human hepatocellular carcinoma.

There is much debate about the way in which epithelial tumors metastasize. It has been proposed that the bone marrow (BM) acts as a tumor cell reservoir. We injected human hepatocellular carcinoma (HCC) cells (Mahlavu cell line) into the livers, circulation or BM of NOD/SCID mice and circulating tumor cells were quantified. When injected under the Glisson capsule, a primary tumor developed and continuously yielded circulating tumor cells. Liver tumor removal led to a very low level of Mahlavu cells both in blood and BM 30 days later. When Mahlavu cells (cultured or from BM of primary mice femurs) were intravenously injected into mice, the number of cells in the bloodstream (BS) steadily decreased, whereas the BM was not significantly colonized. When Mahlavu cells were directly injected into one femur, the controlateral femur was not colonized. Microscopic analysis and a sensitive PCR assay (<1 Mahlavu cell/nuclear cells) both failed to detect human tumor cells in other organs regardless of injection route. In conclusion, our model strongly supports the hypothesis that HCCs continuously release cells into the BS. However, in sharp contrast with the current hypothesis, the BM is not specifically colonized by tumor cells but could store them at a very low level.

Human T-cell leukemia virus type 1 tax protein inhibits the expression of the basic helix-loop-helix transcription factor MyoD in muscle cells: maintenance of proliferation and repression of differentiation.

Human T-cell leukemia virus type 1 Tax protein, a transcriptional activator of viral expression, promotes uncontrolled cellular proliferation. In this report, we show that Tax-expressing myoblasts do not exit the cell cycle and fail to differentiate into myotubes despite the deprivation of serum. In these cells, which displayed unchanged levels of the ubiquitous basic helix-loop-helix E2A factors and Id proteins, Tax was found to target the muscle-specific basic helix-loop-helix transcription factor MyoD. The Tax-induced increase in cyclin-dependent kinase 2 activity correlated with the phosphorylation of MyoD. However, the half-life of this hyperphosphorylated form of MyoD increased in Tax-expressing myoblasts, contrary to that in control cells. Furthermore, MyoD mRNA levels were reduced in Tax-expressing cells. Tax was found to repress MyoD expression at the transcriptional step by preventing MyoD from activating its own transcription. Interestingly, overexpression of the transcriptional coactivator p300 restored the capacity of Tax-expressing muscle cells to differentiate. These observations underscore the critical effect of the trans-repressing ability of Tax on the MyoD-controlled proliferation and differentiation processes of the myoblast lineage.

The human T cell leukemia/lymphotropic virus type 1 Tax protein represses MyoD-dependent transcription by inhibiting MyoD-binding to the KIX domain of p300. A potential mechanism for Tax-mediated repression of the transcriptional activity of basic helix-loop-helix factors.

The human T cell leukemia/lymphotropic virus type 1 (HTLV-1) Tax protein strongly activates viral and cellular gene transcription. It mainly functions by interacting with cellular transcription factors and the KIX domain of the p300/CBP coactivators. Tax can also repress the transcription of cellular genes through the basic helix-loop-helix (bHLH) protein family. To investigate the molecular mechanisms of this Tax-mediated inhibition, we analyzed its effect on the transcriptional activity of the myogenic MyoD protein, which was used as a paradigm of bHLH factors. In this study, we show that overexpression of the p300 coactivator in transient transfection assays was sufficient to rescue MyoD repression by Tax. Furthermore, an N-terminal domain of p300 (amino acids 379-654) containing the region of KIX serving as the Tax binding site was found, when overexpressed, to potentiate Tax-mediated transactivation of HTLV-1 proviral as well as MyoD-dependent transcription, and to antagonize the inhibition by Tax of the transcriptional activity of MyoD. These results revealing the presence of an N-terminal MyoD binding site were confirmed by in vitro protein-protein interaction assays that demonstrate that MyoD binds to the KIX domain of p300 and that Tax competes with MyoD binding in a nonreciprocal manner. These observations provide evidence that Tax binding to the KIX domain of p300 prevents bHLH proteins from contacting this N-terminal domain of the coactivator, thus resulting in their transcriptional repression. As bHLH proteins are implicated in many developmental fate decisions, especially during thymopoiesis, Tax-mediated inhibition of their transcriptional activity may contribute to the induction of HTLV-1-linked leukemogenesis.

Esophageal rupture in a patient with idiopathic eosinophilic esophagitis.

Idiopathic eosinophilic esophagitis is an extremely rare condition with fewer than 20 cases described in the literature. We present a case presenting as an emergency with esophageal perforation that eventually required subtotal esophagectomy.