Quartet analysis of putative horizontal gene transfer in Crenarchaeota.

Horizontal gene transfers (HGT) between four Crenarchaeota species (Metallosphaera cuprina Ar-4T, Acidianus hospitalis W1T, Vulcanisaeta moutnovskia 768-28T, and Pyrobaculum islandicum DSM 4184T) were investigated with quartet analysis. Strong support was found for individual genes that disagree with the phylogeny of the majority, implying genomic mosaicism. One such gene, a ferredoxin-related gene, was investigated further and incorporated into a larger phylogeny, which provided evidence for HGT of this gene from the Vulcanisaeta lineage to the Acidianus lineage. This is the first application of quartet analysis of HGT for the phylum Crenarchaeota. The results have shown that quartet analysis is a powerful technique to screen homologous sequences for putative HGTs and is useful in visually describing genomic mosaicism and HGT within four taxa.

Defining quality assessment in vascular surgery training: an expert Delphi process.

INTRODUCTION: A robust and reproducible way of assessing training should optimise and standardise vascular surgical training. This study describes the methodology supporting the Vascular Surgery Specialty Advisory Committee regional quality assurance reports for vascular surgery training programmes in the UK. METHODS: A Delphi consensus exercise was performed to establish the domains of training that most appropriately assess the quality of a vascular surgery training programme. A total of 54 stakeholders were invited to participate, including trainees, training programme directors and members of the vascular speciality advisory committee (SAC), vascular society executive and education committees. RESULTS: A total of 39 stakeholders successfully completed the three-stage Delphi process over 15 weeks. The domains identified as most appropriate to assess the quality of a vascular training programme were: Joint Committee on Surgical Training (JCST) survey results, clinical experience, regional education programmes, radiology support, timetable, regional support for trainees, trainer support for trainees, opportunities for professional development, trainee-rated quality of consultant teaching and training, and trainee recommendation of the post. CONCLUSIONS: This study describes a method to identify and prioritise domains that are appropriate to assess the quality of a vascular training programme. The domains that were identified as appropriate to assess quality are transferable internationally and the Delphi methodology could be used by other training schemes to 'fine-tune' their own domains to review and optimise the quality of their own training programmes.

Safety and efficacy of ocrelizumab in patients with rheumatoid arthritis and an inadequate response to methotrexate: results of a forty-eight-week randomized, double-blind, placebo-controlled, parallel-group phase III trial.

OBJECTIVE: To evaluate the efficacy and safety of treatment with ocrelizumab plus methotrexate (MTX) in patients with active rheumatoid arthritis (RA) and an inadequate response to MTX. METHODS: STAGE was a phase III randomized, double-blind, parallel-group international study to evaluate the safety and efficacy of ocrelizumab compared with placebo in patients with active RA continuing MTX treatment. Patients receiving stable doses of MTX were randomized to receive 2 infusions of placebo (n = 320), ocrelizumab 200 mg (n = 343), or ocrelizumab 500 mg (n = 343) on days 1 and 15 as well as weeks 24 and 26. Coprimary end points were the proportion of patients with an American College of Rheumatology 20% improvement criteria (ACR20) response at weeks 24 and 48. Secondary end points included the change from baseline in the modified Sharp/van der Heijde score (SHS) and the ACR50/70 responses. RESULTS: The ACR20 response rates were 35.7% in the placebo group, 56.9% in the ocrelizumab 200 mg group, and 54.5% in the ocrelizumab 500 mg group at 24 weeks, and 27.6%, 58.3%, and 62.1%, respectively, at 48 weeks (P < 0.0001 versus placebo for each dose at both time points). At week 48, both of the ocrelizumab doses improved the ACR50 and ACR70 response rates 3-fold as compared with placebo and showed a statistically significant (P < 0.0001) reduction in joint damage progression relative to placebo (mean change in SHS reduced by 85% and 100% for the 200-mg and 500-mg doses, respectively). Rates of serious infection were comparable in the placebo (3.48 per 100 patient-years) and ocrelizumab 200 mg (3.54 per 100 patient-years) groups but were elevated in the ocrelizumab 500 mg group (8.66 per 100 patient-years). CONCLUSION: With both ocrelizumab doses, the primary end point was met, and the signs and symptoms of RA were significantly improved at weeks 24 and 48. Ocrelizumab also significantly inhibited the progression of joint damage. A higher rate of serious infections was observed with 500 mg of ocrelizumab as compared with ocrelizumab 200 mg or placebo.

Safety and efficacy of ocrelizumab in patients with rheumatoid arthritis and an inadequate response to at least one tumor necrosis factor inhibitor: results of a forty-eight­week randomized, double-blind, placebo-controlled, parallel-group phase III trial.

OBJECTIVE: To evaluate the safety and efficacy of ocrelizumab plus methotrexate (MTX) or leflunomide (LEF) in patients with active rheumatoid arthritis (RA) and an inadequate response to tumor necrosis factor α inhibitors. METHODS: This was a multicenter randomized, double-blind, placebo-controlled, parallel-group study that continued over 48 weeks. Patients receiving stable doses of MTX or LEF were randomized to receive 2 infusions of placebo (n = 277), ocrelizumab 200 mg (n = 278), or ocrelizumab 500 mg (n = 285) on days 1 and 15 as well as at weeks 24 and 26. Coprimary end points were the proportion of patients with response according to the American College of Rheumatology 20% improvement criteria (ACR20) at weeks 24 and 48. Secondary end points included the change from baseline in the modified Sharp/van der Heijde score (SHS) and the ACR50/70 responses. RESULTS: ACR20 responses were 22.0% in the placebo group, 42.2% in the ocrelizumab 200 mg group, and 47.9% in the ocrelizumab 500 mg group at 24 weeks and 19.5%, 48.7%, and 50.7%, respectively, at 48 weeks (P < 0.0001 versus placebo for each comparison at each time point). At 48 weeks, patients receiving both doses of ocrelizumab showed significantly improved ACR50 and ACR70 responses of ~3-fold versus placebo. Only those in the ocrelizumab 500 mg group showed statistically significant (P = 0.0017) inhibition of joint damage progression (mean change in the SHS) relative to placebo (61% inhibition) at 48 weeks. Overall adverse events and infections during the 48 weeks of study were comparable in all treatment groups. Serious infections were observed more frequently in patients taking ocrelizumab (5.1% and 4.3%) than in those taking placebo (2.5%). CONCLUSION: Patients in both of the ocrelizumab groups met the clinical primary efficacy end points. Inhibition of change in the SHS was statistically significant at 48 weeks for those in the ocrelizumab 500 mg group. The rate of serious infections in this trial was higher for both ocrelizumab doses as compared with placebo.

Force amplification response of actin filaments under confined compression.

Actin protein is a major component of the cell cytoskeleton, and its ability to respond to external forces and generate propulsive forces through the polymerization of filaments is central to many cellular processes. The mechanisms governing actin's abilities are still not fully understood because of the difficulty in observing these processes at a molecular level. Here, we describe a technique for studying actin-surface interactions by using a surface forces apparatus that is able to directly visualize and quantify the collective forces generated when layers of noninterconnected, end-tethered actin filaments are confined between 2 (mica) surfaces. We also identify a force-response mechanism in which filaments not only stiffen under compression, which increases the bending modulus, but more importantly generates opposing forces that are larger than the compressive force. This elastic stiffening mechanism appears to require the presence of confining surfaces, enabling actin filaments to both sense and respond to compressive forces without additional mediating proteins, providing insight into the potential role compressive forces play in many actin and other motor protein-based phenomena.

Adhesive interactions between vesicles in the strong adhesion limit.

We consider the adhesive interaction energy between a pair of vesicles in the strong adhesion limit, in which bending forces play a negligible role in determining vesicle shape compared to forces due to membrane stretching. Although force−distance or energy−distance relationships characterizing adhesive interactions between fluid bilayers are routinely measured using the surface forces apparatus, the atomic force microscope, and the biomembrane force probe, the interacting bilayers in these methods are supported on surfaces (e.g., mica sheet) and cannot be deformed. However, it is known that, in a suspension, vesicles composed of the same bilayer can deform by stretching or bending, and can also undergo changes in volume. Adhesively interacting vesicles can thus form flat regions in the contact zone, which will result in an enhanced interaction energy as compared to rigid vesicles. The focus of this paper is to examine the magnitude of the interaction energy between adhesively interacting, deformed vesicles relative to free, undeformed vesicles as a function of the intervesicle separation. The modification of the intervesicle interaction energy due to vesicle deformability can be calculated knowing the undeformed radius of the vesicles, R0, the bending modulus, k(b), the area expansion modulus, k(a), and the adhesive minimum, W(P)(0), and separation, D(P)(0), in the energy of interaction between two flat bilayers, which can be obtained from the force−distance measurements made using the above supported-bilayer methods. For vesicles with constant volumes, we show that adhesive potentials between nondeforming bilayers such as |W(P)(0)| 5 × 10(−4) mJ/m2, which are ordinarily considered weak in the colloidal physics literature, can result in significantly deep (>10×) energy minima due to increase in vesicle area and flattening in the contact region. If the osmotic expulsion of water across the vesicles driven by the tense, stretched membrane in the presence of an osmotically active solute is also taken into account, the vesicles can undergo additional deformation (flattening), which further enhances the adhesive interaction between them. Finally, equilibration of ions and solutes due to the concentration differences created by the osmotic exchange of water can lead to further enhancement of the adhesion energy. Our result of the progressively increasing adhesive interaction energy between vesicles in the above regimes could explain why suspensions of very weakly attractive vesicles may undergo flocculation and eventual instability due to separation of vesicles from the suspending fluid by gravity. The possibility of such an instability is an extremely important issue for concentrated vesicle-based products and applications such as fabric softeners, hair therapeutics and drug delivery.

The Contribution of DOPA to Substrate-Peptide Adhesion and Internal Cohesion of Mussel-Inspired Synthetic Peptide Films.

Mussels use a variety of 3, 4-dihydroxyphenyl-l-alanine (DOPA) rich proteins specifically tailored to adhering to wet surfaces. Synthetic polypeptide analogues of adhesive mussel foot proteins (specifically mfp-3) are used to study the role of DOPA in adhesion. The mussel-inspired peptide is a random copolymer of DOPA and N(5) -(2-hydroxyethyl)-l-glutamine synthesized with DOPA concentrations of 0-27 mol% and molecular weights of 5.9-7.1 kDa. Thin films (3-5 nm thick) of the mussel-inspired peptide are used in the surface forces apparatus (SFA) to measure the force-distance profiles and adhesion and cohesion energies of the films in an acetate buffer. The adhesion energies of the mussel-inspired peptide films to mica and TiO(2) surfaces increase with DOPA concentration. The adhesion energy to mica is 0.09 µJ m(-2) mol(DOPA) (-1) and does not depend on contact time or load. The adhesion energy to TiO(2) is 0.29 µJ m(-2) mol(DOPA) (-1) for short contact times and increases to 0.51 µJ m(-2) mol(DOPA) (-1) for contact times >60 min in a way suggestive of a phase transition within the film. Oxidation of DOPA to the quinone form, either by addition of periodate or by increasing the pH, increases the thickness and reduces the cohesion of the films. Adding thiol containing polymers between the oxidized films recovers some of the cohesion strength. Comparison of the mussel-inspired peptide films to previous studies on mfp-3 thin films show that the strong adhesion and cohesion in mfp-3 films can be attributed to DOPA groups favorably oriented within or at the interface of these films.

Direct measurement of double-layer, van der Waals, and polymer depletion attraction forces between supported cationic bilayers.

The interactions of supported cationic surfactant bilayers and the effects of nonadsorbing cationic polyelectrolytes on these interactions were studied using the surface forces apparatus (SFA) technique. Bilayers of the cationic surfactant di(tallow ethyl ester) dimethyl ammonium chloride (DEEDMAC) were deposited on mica surfaces using the Langmuir-Blodgett technique, and the interactions between the bilayers were measured in various salt, nonionic polymer (PEG), and cationic polyelectrolyte solutions at different polymer molecular weights and concentrations. The forces between the bilayers in CaCl(2) solution are purely repulsive and follow the DLVO theory quantitatively down to bilayer separations of ∼2 nm. Addition of nonadsorbing polymer or polyelectrolyte has a number of effects on the interactions including the induction of a depletion-attraction between the bilayers and screening of the double-layer repulsion due to the added ions in the solution from the polyelectrolyte. The experimental results are shown to agree well with standard theories of depletion attraction and double-layer screening associated with dissolved polyelectrolyte. We also observed significant time and rate effects on measuring the equilibrium bilayer-bilayer interactions possibly due to the unexpectedly long times (>1 min) associated with the charge regulation of the bilayer surfaces. Implications for the interactions and stability of vesicle dispersions, i.e., of free rather than supported bilayers, in polymer solutions are discussed.

Transient filamentous network structure of a colloidal suspension excited by stepwise electric fields.

Jamming and force networks observed in electrorheological (ER) fluids bear many similarities to those observed in various granular and colloidal systems. We have measured the time evolution (transient stresses) of filamentous networks of colloidal particles in suspensions subjected to continuous tensile strain concomitant with the switching on and off of electric fields. The density of particle chains was found to increase exponentially with the applied tensile strain via a rapid formation of single chains followed by a slower coarsening (aggregation) of the chains. The two processes can be ascribed to the field-induced short-range and long-range interparticle forces, respectively, along with the tensile viscous force.

Diaphragmatic crural augmentation utilising cross-linked porcine dermal collagen biologic mesh (Permacol) in the repair of large and complex para-oesophageal herniation: a retrospective cohort study.

OBJECTIVE: To evaluate the safety, efficacy and durability of selective integration of porcine dermal collagen (Permacol) biologic mesh for crural re-construction in large or complex para-oesophageal hernia surgery. BACKGROUND: Surgical repair of para-oesophageal herniation has been associated with high rates of failure. The utilisation of prosthetic mesh is controversial with complications including erosion and fistulation. Long-term outcomes for biologic mesh crural augmentation are unclear. METHODS: A retrospective analysis of patients who underwent a biologic mesh (Permacol) augmented cruroplasty in the repair of large and/or complex para-oesophageal hernia was performed utilising the prospectively maintained oesophago-gastric database at the Royal Devon and Exeter Hospital between October 2004 and January 2013. This technique was selectively used for patients where the lateral extent of the diaphragmatic-crural defect prevented the fashioning of a sound, tension-free repair with sutures alone, or previous surgery had failed. Successful outcome was defined by resolution of symptoms and structural integrity of the repair. RESULTS: Fifty one procedures were performed on 49 patients (15 male), median age 75 (range 25-91). Post-operative morbidity included 2 (3.9%) oesophageal injuries managed conservatively, and 2 (3.9%) patients who suffered early repair breakdown requiring immediate surgical re-intervention. Four patients (8%) required endoscopic dilatation due to dysphagia, one (2%) in the early post-operative phase. The median follow-up was 36 months (range 6-105). All patients reported initial symptomatic resolution. Two patients (4%) were demonstrated to have breakdown of their repair during the follow-up period, both of whom underwent revision mesh-augmented surgery and are re-incorporated in this series. Late-onset dysphagia in two (4%) patients may be mesh-related, but no other complications were observed and a Kaplan-Meier analysis of this series predicts a symptom-free rate of approximately 94% at 5 years. CONCLUSIONS: The selective integration of biologic mesh to augment the crural repair in para-oesophageal hernia with extensive diaphragmatic defects appears to be safe, effective and infers the potential of long-term satisfactory outcomes.

Localized neuroblastoma treated by surgery: a Pediatric Oncology Group Study.

A prospective study was designed to evaluate the outcome of patients with localized resectable neuroblastoma without regional lymph node involvement when no therapy beyond surgical resection was administered. One hundred one patients observed for 3 to 60 months had a 2-year disease-free survival of 89% (SE = 5%). Of the nine patients experiencing relapse, only three have died. There were no apparent distinguishing characteristics of the nine failures. Due to the favorable prognosis of the subset of neuroblastoma patients, prognostic factor analysis had very limited power and lacked clinical importance. Complete gross removal of the localized tumors is adequate therapy to ensure the survival of the majority of these patients.

Huntington's disease in Venezuela: neurologic features and functional decline.

We studied 65 Huntington's disease patients and 225 at-risk individuals over the past 4 years. The rate of decline of these untreated patients from Venezuela was similar to that seen in US patients who had received neuroleptic drugs. Chorea, oculomotor dysfunction, and dysdiadochokinesis were early symptoms; parkinsonian features and dystonia came later. Juvenile patients declined nearly twice as fast as adult-onset patients. No distinctive neurologic phenotypes were seen in children of two affected parents.

Protocols for high efficiency, stage-specific retroviral transduction of murine fetal thymocytes and thymic epithelial cells.

Viral vectors have the potential to provide a fast and economic alternative to transgenic methods for manipulating gene expression in studies of immune system development and function. Although protocols exist for the infection of hematopoietic precursors and peripheral T cells in vitro, critical stages of T cell differentiation are strictly dependent upon a three-dimensional thymic architecture and their analysis poses unique technical challenges. Whole fetal thymic lobes have been used as targets for retroviral and adenoviral infection, both in situ and in vitro, but this approach does not allow for discrimination between lymphoid and stromal components. Isolated thymocytes have been infected by co-culture with viral producer cells, but under these conditions they rapidly lose their developmental potential. To overcome these problems we have combined a number of efficient techniques for retroviral production, concentration, and infection that allow us to rapidly achieve significant transduction rates of purified populations of double-negative (DN) and double-positive (DP) thymocytes, single-positive (SP) T lymphocytes, as well as fetal thymic MHC II(+) epithelial cells without the need for co-culture with viral producer cells. Reaggregate thymic organ culture (RTOC) techniques were used to assess the development and function of transduced cells in defined cellular environments. As a demonstration of the utility of these methods, CD80 (B7.1) was transduced into thymic epithelial cells and shown to allow them to mediate negative selection of DP thymocytes, and to act as antigen-presenting cells (APC) to mature T cells. The ability to genetically manipulate primary cells of a specified type and differentiation stage provides a powerful complement to RTOC techniques for the study of T cell development.

Papillary adenocarcinoma of rete testis. Autopsy findings, histochemistry, immunohistochemistry, ultrastructure, and clinical correlations.

Adenocarcinoma of the rete testis is a rare neoplasm that usually occurs in men after the age of 60 and carries a variable prognosis. We report an occurrence of this tumor in a 91-year-old man who had been treated for 2 years for an hydrocele. At the time of diagnosis, metastases were not evident; and the patient was treated with local radiotherapy. The diagnosis of papillary adenocarcinoma of the rete testis was made on the basis of: (a) a transition from normal rete testis to atypical and neoplastic rete epithelium; (b) exclusion of primary germinal and nongerminal testicular tumors and spread from distant sources; and (c) electron-microscopic findings, histochemical and immunological studies, and autopsy findings supporting the diagnosis. This is the first reported case of adenocarcinoma of the rete testis that includes documentation of the tumor's metastatic pattern.

Non-Hodgkin's lymphoma: identification of the monoclonal B lymphocyte component in the presence of polyclonal immunoglobulin.

A high proportion of non-Hodgkin's lymphomas are neoplastic proliferations of B lymphocytes, and, as such, express integral membrane and/or cytoplasmic immunoglobulin (Ig). Because these cellular proliferations are monoclonal, the Ig of all neoplastic lymphocytes will have identical light chain type and idiotype. These tumors sometimes contain significant amounts of polyclonal Ig. In this study we demonstrate that the polyclonal non-B-lymphocyte-associated Ig may be removed by washing tissue at low pH to reveal either neoplastic B lymphocytes or neoplastic "null" lymphocytes. These observations should facilitate the application of immunohistology to the routine diagnosis of lymphoma.

A large kindred with X-linked mental retardation, marker X and macroorchidism.

Thirty-eight members of a black kindred with mental retardation and the Marker X were studied. Ten of 14 affected males, 6 of 6 affected females, 18 carriers or possible carriers, and 7 normal males were examined. Eight of 9 affected males who could be measured had macroorchidism, but their ears and mandibles were not prominent. No distinct facies were evident, although minor anomalies, such as a slight pectus, were present in some. Clinical diagnosis in the absence of a strongly positive family history may be even more difficult among prepubertal black children than in whites. The risk of retardation among children of carriers was estimated at 20-40%.

Dominant inheritance of bifid nose.

To our knowledge, median cleft of nose without apparent hypertelorism (bifid nose) has been reported only twice [Esser, 1939; Boo-chai, 1965]. We report five individuals in 3 generations of a family who had a bifid nose without apparent hypertelorism or mental retardation. In this family bifid nose was most likely an autosomal dominant trait.

Congenital polycystic tumor of the atrioventricular node: possible familial occurrence and critical review of reported cases with special emphasis on histogenesis.

Congenital polycystic tumor of the atrioventricular node is a rare tumor of disputed histogenesis. It constant location in the atrioventricular node separates it from other cardiac cysts and tumors of indisputable endodermal origin. New studies of the embryogenesis of conduction tissues provide a reasonable explanation for the site of the lesion. Morphologic and histochemical features of the tumor fail to conclusively favor either a mesothelial or an endodermal genesis. A recently studied case provides some new morphologic observations (complex desmosomes, granule-containing epithelial cells) and suggests the possibility of of familial occurrence.

Chemical cholecystitis associated with hepatic arterial chemotherapy delivered by a permanently implanted pump.

The introduction of chemotherapeutic agents directly into the proper hepatic artery via an indwelling catheter results in perfusion of the gallbladder, because the cystic artery is usually a branch of the right hepatic artery. Five gallbladders, removed two to 16 months after insertion of permanently implanted Infusaid model 400 pumps, were examined. All of the gallbladders had significant arteritis, with narrowing or occlusion of lumina or necrosis of vessel walls. Fibrosis of the gallbladder wall was also a constant finding. Nuclear atypia of mucosal epithelium and connective tissue was common. Varying degrees of acute and chronic inflammation were present. These abnormalities may have a radiomimetic and direct irritant pathogenesis.

Intraplaque hemorrhage: its significance in cerebrovascular disease.

Recently, carotid plaque factors, specifically intraplaque hemorrhage, have been studied with respect to the production of cerebrovascular symptoms. Ninety-five carotid endarterectomies were performed and the plaques that were removed were examined for intraplaque hemorrhage. Patients were separated into three groups: those with specific neurologic symptoms, those with nonlateralizing symptoms, and those who were asymptomatic. In the group of patients who presented with specific neurologic symptoms, correlation was made between the age of the intraplaque hemorrhage and the timing of symptoms. The vast majority of patients with specific neurologic symptoms exhibited carotid plaque hemorrhage, but patients with nonlateralizing symptoms and those who were asymptomatic also demonstrated an unexpectedly high percentage of intraplaque hemorrhage. Moreover, our results show a poor relationship between the timing of symptoms and the age of the intraplaque hemorrhage. These data do not refute the concept that intraplaque hemorrhage may play a role in the production of cerebrovascular symptoms, but they do refute the notion that the mere presence of hemorrhage causes specific neurologic symptoms and they also refute the previous report that demonstrates a good correlation between the timing of symptoms and the age of the intraplaque hemorrhage.