An allosteric role for receptor activity-modifying proteins in defining GPCR pharmacology

G protein-coupled receptors are allosteric proteins that control transmission of external signals to regulate cellular response. Although agonist binding promotes canonical G protein signalling transmitted through conformational changes, G protein-coupled receptors also interact with other proteins. These include other G protein-coupled receptors, other receptors and channels, regulatory proteins and receptor-modifying proteins, notably receptor activity-modifying proteins (RAMPs). RAMPs have at least 11 G protein-coupled receptor partners, including many class B G protein-coupled receptors. Prototypic is the calcitonin receptor, with altered ligand specificity when co-expressed with RAMPs. To gain molecular insight into the consequences of this protein–protein interaction, we combined molecular modelling with mutagenesis of the calcitonin receptor extracellular domain, assessed in ligand binding and functional assays. Although some calcitonin receptor residues are universally important for peptide interactions (calcitonin, amylin and calcitonin gene-related peptide) in calcitonin receptor alone or with receptor activity-modifying protein, others have RAMP-dependent effects, whereby mutations decreased amylin/calcitonin gene-related peptide potency substantially only when RAMP was present. Remarkably, the key residues were completely conserved between calcitonin receptor and AMY receptors, and between subtypes of AMY receptor that have different ligand preferences. Mutations at the interface between calcitonin receptor and RAMP affected ligand pharmacology in a RAMP-dependent manner, suggesting that RAMP may allosterically influence the calcitonin receptor conformation. Supporting this, molecular dynamics simulations suggested that the calcitonin receptor extracellular N-terminal domain is more flexible in the presence of receptor activity-modifying protein 1. Thus, RAMPs may act in an allosteric manner to generate a spectrum of unique calcitonin receptor conformational states, explaining the pharmacological preferences of calcitonin receptor-RAMP complexes. This provides novel insight into our understanding of G protein-coupled receptor-protein interaction that is likely broadly applicable for this receptor class.

Relação Entre Motivação e Desempenho Escolar em Alunos do 1º Ciclo

O presente estudo teve como objectivo analisar a existência de uma possível relação entre motivação (intrínseca ou extrínseca) e o desempenho escolar em alunos do 1º ciclo que frequentam o 4º ano. A amostra foi constituída por 62 alunos, com idades compreendidas entre os 9 e 10 anos, 15 que frequentavam a Escola Básica do 1º Ciclo de Brancas e 47 a Escola Básica da Batalha (24 alunos da Turma A e 23 da Turma B). Considerando a possível existência de variáveis sociodemográficas associadas ao desempenho, no processo de recolha de dados foi aplicado um questionário sóciodemográfico, que pela sua potencial natureza descritiva, interpretativa e reflexiva, constituiu uma importante ferramenta na análise e avaliação dos resultados obtidos. Com o objectivo de avaliar a motivação (intrínseca e extrínseca) foi administrada uma escala de motivação escolar, aferida para a população portuguesa contendo 18 questões. Cada uma destas questões contém 4 possibilidades de resposta (duas referentes à motivação intrínseca e outras duas referentes à motivação extrínseca). Quanto ao desempenho escolar, este foi avaliado através das notas que os alunos obtiveram no final do ano lectivo e que foram facultadas pelas professoras dos mesmos. A análise dos resultados evidenciou que existe uma relação entre a motivação e o desempenho escolar, concluindo-se que os alunos intrinsecamente motivados apresentam melhor desempenho do que os alunos extrinsecamente motivados. Consideramos deste modo, que a implementação de estratégias na sala de aula que promovam a motivação intrÍnseca será mais benéfico para o sucesso escolar dos alunos. / The objective of this study, is to analyze the possible relation between intrinsic and extrinsic motivation and the educational achievement in students in the fourth grade of grade school. The sample was composed by 62 students, with a ages between 9 and 10 years of age. 15 students of this sample attended the grade school of Brancas, 47 attended the grade school of Batalha ( 24 of the students are from class A and 23 are from class B). Considering a possible existence of sociodemographic variables related to achievement, in the process of data collecting, a sociodemographic questionnaire was used, considering its great potential to describe, interpret analyze and evaluate the results obtained. To evaluate intrinsic and extrinsic motivation, a scale of school motivation adapted for Portuguese population with eighteen questions was applied. Each of these questions has 4 possibilities of response (two referring to intrinsic motivation and two referring to extrinsic motivation). In what concerns to school achievement, this was evaluated trough school grades obtained at the end of the school year given by the teachers. The analyses of the results demonstrated that there are differences between motivation and school achievement, concluding that the students with intrinsic motivation present better achievement then the students with extrinsic motivation. The implementation of strategies in the classroom, will promote intrinsic motivation and bring more success to students.

Performance en fluage des assemblages antiglissement avec des surfaces métallisées dans les ponts en acier

Les éléments des ponts en acier sont exposés à de sévères conditions environnementales, tel l’épandage de sels déglaçant sur les routes. Ces éléments ont besoin d’un niveau suffisant de protection contre la corrosion afin de préserver leur intégrité à long terme. Une solution efficace, devenue populaire au Canada, est la métallisation. La métallisation est un revêtement anticorrosion formé par projection thermique de métal, généralement du zinc, sur la surface à protéger. La protection fournie au substrat d’acier est assurée par une barrière physique et une protection galvanique. Pour le calcul des assemblages boulonnés antiglissement, les codes de conception, tel le code Canadien sur le calcul des ponts routiers CAN/CSA S6-14, spécifient, en fonction des conditions des surfaces de contact désirées, un coefficient de glissement à utiliser. Actuellement, ces codes ne fournissent aucun coefficient de glissement entre deux surfaces métallisées. Donc, il est pratique courante pour les fabricants de ponts en acier de masquer les surfaces de contact des joints boulonnés avant de métalliser, ce qui est très couteux pour l’industrie puisque ce travail doit se faire manuellement. Récemment, des études ont évalué la résistance au glissement à court terme d’assemblages antiglissement ayant des surfaces de contact métallisées. Les résultats ont révélé une résistance au glissement supérieure aux assemblages sur acier nu grenaillés. Dans la présente recherche, la performance en fluage des assemblages antiglissement métallisés a été caractérisée pour s’assurer qu’une résistance en glissement de Classe B soit toujours valide à long terme. L’effet de la relaxation de la force de serrage sur la charge de glissement a aussi été évalué. Les résultats ont démontré une bonne performance en fluage. Aussi, les résultats ont révélé que la relaxation de la force de serrage n’a pas d’impact significatif sur la résistance au glissement des assemblages antiglissement métallisés. Les conclusions de cette étude pourraient bien influencer de futures révisions des codes de conception et avoir un impact sur l’industrie de l’acier en Amérique du Nord.Mots clés : Assemblage boulonné antiglissement, métallisation, pont routier en acier, fluage, relaxation

Apprendre avec la technologie : l'impact sur l'enseignement et l'apprentissage de l’utilisation de la technologie numérique (SMARTBOARD) dans les classes

This research was devoted to gaining information on teachers? use of technology, specifically SMARTBOARD technology, for teaching and promoting learning in the classroom. Research has suggested that use of technology can enhance learning and classroom practices. This has resulted in administrators encouraging the use of SMARTBOARDS, installing them in classrooms and providing training and support for teachers to use this technology. Adoption of new technology, however, is not simple. It is even more challenging because making the best use of new technologies requires more than training; it requires a paradigm shift in teachers? pedagogical approach. Thus, while it may be reasonable to believe that all we need to do is show teachers the benefits of using the SMARTBOARD; research tells us that changing paradigms is difficult for a variety of reasons. This research had two main objectives. First, to discover what factors might positively or negatively affect teachers? decisions to take up this technology. Second, to investigate how the SMARTBOARD is used by teachers who have embraced it and how this impacts participation in classrooms. The project was divided into two parts; the first was a survey research (Part 1), and the second was an ethnographic study (Part 2). A thirty-nine item questionnaire was designed to obtain information on teachers? use of technology and the SMARTBOARD. The questionnaire was distributed to fifty teachers at two EMSB schools: James Lyng Adult Centre (JLAC) and the High School of Montreal (HSM). Part 2 was an ethnographic qualitative study of two classes (Class A, Class B) at JLAC. Class A was taught by a male teacher, an early-adopter of technology and a high-level user of the SMARTBOARD; Class B was taught by a female teacher who was more traditional and a low-level user. These teachers were selected because they had similar years of experience and general competence in their subject matter but differed in their use of the technology. The enrollment in Class A and Class B were twenty-three and twenty-four adult students, respectively. Each class was observed for 90 minutes on three consecutive days in April 2010. Data collection consisted of videotapes of the entire period, and observational field notes with a graphical recording of participatory actions. Information from the graphical recording was converted to sociograms, a graphic representation of social links among individuals involved in joint action. The sociogram data was tabulated as quantified data. The survey results suggest that although most teachers are interested in and use some form of technology in their teaching, there is a tendency for factors of gender and years of experience to influence the use of and opinions on using technology. A Chi Square analysis of the data revealed (a) a significant difference (2 = 6.031, p < .049) for gender in that male teachers are more likely to be interested in the latest pedagogic innovation compared to female teachers; and, (b) a significant difference for years of experience (2 = 10.945, p < .004), showing that teachers with ?6 years experience were more likely to use the SMARTBOARD, compared to those with more experience (>6 years). All other items from the survey data produced no statistical difference. General trends show that (a) male teachers are more willing to say yes to using the SMARTBOARD compared to female teachers, and (b) teachers with less teaching experience were more likely to have positive opinions about using the SMARTBOARD compared to teachers with more experience. The ethnographic study results showed differences in students? response patterns in the two classrooms. Even though both teachers are experienced and competent, Teacher A elicited more participation from his students than Teacher B. This was so partly because he used the SMARTBOARD to present visual materials that the students could easily respond to. By comparison, Teacher B used traditional media or methods to present most of her course material. While these methods also used visual materials, students were not able to easily relate to these smaller, static images and did not readily engage with the material. This research demonstrates a generally positive attitude by teachers towards use of the SMARTBOARD and a generally positive role of this technology in enhancing students? learning and engagement in the classroom. However, there are many issues related to the SMARTBOARD use that still need to be examined. A particular point is whether teachers feel adequately trained to integrate SMARTBOARD technology into their curricula. And, whether the gender difference revealed is related to other factors like a need for more support, other responsibilities, or a general sense of anxiety when it comes to technology. Greater opportunity for training and ongoing support may be one way to increase teacher use of the SMARTBOARD; particularly for teachers with more experience (>6 years) and possibly also for female teachers.

Vidéos contre lectures: Une évaluation de l'utilisation de vidéos pour accomplir la construction initiale de connaissances

Abstract : The goal of this study was to determine whether providing videos for students to watch before class would be more effective than assigning readings. The study took place within a flipped classroom: a methodology designed to engage students in the initial construction of knowledge before class, freeing up classroom time for active learning pedagogies. Preparing for class by watching videos should require less of the students’ time than doing readings, and they should respond better to videos than to readings because these more closely mimic this generation’s interactions with information and media. Consequently, flipped classroom students provided with videos should perceive a lower workload, which could translate into more positive learning outcomes. From an instructor’s perspective, however, developing and organizing videos is extremely time consuming. Thus, a teacher giving consideration to flipping their class would likely want to know whether videos would lead to positive outcomes for their students before actually committing to developing these. However, no research to date was identified which has examined the question of whether flipped class videos would be more effective than readings. The hypotheses for the study were that videos would result in measurable learning gains for the students, and would lead to lower time demands and perceived workloads, as well as more positive attitudes. These were tested using a quasi-experimental design involving a convenience sample of two small college General Biology 1 courses taught by the same teacher. One group had videos to watch before class for the first 1/3 of the course (treatment; Class A), during which the other was assigned readings (control; Class B). Following this, both groups were provided with videos. Student scores were compared on pre-instruction and in-class quizzes, activities, four unit tests, and a final exam. Further, students completed a 44 item survey as well as a demographic questionnaire. Results suggest greater learning gains for students provided with videos. Certainly, students from Class B improved significantly when provided with videos, especially as compared to Class A, whose improvement over the same time span was marginal. However, conclusions based on these results are somewhat tentative, as Class A performed rather poorly on all summative assessments, and this could have driven the patterns observed, at least partially. From the surveys, Class B students reported that they spent more time preparing for class, were less likely to do the necessary preparations, and generally perceived their effort and workload levels to be higher. These factors were significant enough that they contributed to four students from Class B (10% of the class) dropping the course over the semester. In addition, students from both groups also reported more positive attitudes towards videos than readings, although they did not necessarily feel that videos were (or would be) a more effective study and learning tool. The results of this study suggest that any effort on the part of an instructor to prepare or organize videos as pre-class instructional tools would likely be well spent. However, even outside of the context of a flipped classroom, this study provides an indication that assigned readings can place heavy workload requirements on students, which should give cause for any instructor employing mandatory readings to reassess their approach. Finally, since both groups were statistically equivalent across all measured demographic variables, it appears that some of the observed disparities in assessment scores may have been driven by differences in group dynamics. Consequently, the suggestion is made that an instrument to measure classroom climate should be incorporated into any research design comparing two or more interacting groups of students, as group dynamics have the potential to play a key role in any outcomes.||Résumé : L'objectif de cette étude était de déterminer si fournir des vidéos aux étudiants avant une classe serait plus efficace comme méthode d’apprentissage individuel que de leur donner une liste de lectures à compléter. L'étude a eu lieu dans une classe renversée – une stratégie plus reconnu sous le nom de «classroom flipping» – ce qui ce trouve à être une méthodologie qui vise à obliger les élèves d’accomplir le transfert de connaissances initial avant la classe, en vue de libérer du temps de classe pour des activités pédagogiques plus approfondie. En théorie, pour les élèves, se préparer pour une classe renversé en regardant des vidéos devrait exiger moins de temps que d’être obligé à apprendre le matériel en lisant. En plus, présenter le matériel d’un cours avec des vidéos imite de plus près les genres d’interactions qu’on les étudiants de cette génération avec de l'information et les médias, ce qui devrait faciliter leur tâche. Par conséquent, les élèves d’une classe renversée fournies avec des vidéos devraient percevoir une charge de travail moins élevée, et ceci pourrait se traduire en bilans d'apprentissage plus positifs. Cependant, du point de vue du professeur, avoir à développer et organiser des vidéos se présente comme un gros défi, surtout en vue du temps et de l’effort qui sont requis. Ainsi, un enseignant qui songe à renversée sa classe voudrait probablement savoir si le fait d’offrir des vidéos mène à des résultats positifs avant de réellement s'engager dans le développement de ces derniers. Par contre, lors de l’écriture de ce texte, aucune étude n’a été identifié qui répond à cette question, et aucune publication compare la performance des étudiants lorsqu’ils ont des vidéos à regarder avec lorsqu’ils sont obligés de faire de la lecture pour se préparer. Alors, les hypothèses de l'étude étaient que des étudiants ayant accès à des vidéos démontreraient des gains d’apprentissages évidents, qu’ils apercevraient des requêtes de temps moins encombrantes et une charge de travail plus faible, et, en vue de ces derniers, que les étudiants auraient des attitudes plus positives envers le cours et le matériel. Les hypothèses ont été testés à l'aide d'un modèle quasi-expérimental, avec comme échantillon deux classes collégiales de Biologie générale 1, tout les deux enseignés par le même professeur. Un de ces groupes avaient accès à des vidéos pour se préparer pour chaque classe lors du premier tiers du cours, alors que l’autre groupe étaient obligé de faire de la lecture. Après le premier tiers du cours, ce qui fut 10 des 30 classes du semestre, les deux groupes ont été fournis avec des vidéos pour le restant du cours. Des notes ont été ramassées et comparées sur des mini-examens préparatoires avant et durant les classes, sur des activités complétées en classe, sur quatre examens en classe, et un examen final. En outre, les étudiants ont rempli un questionnaire composé de 44 items, ainsi qu’un autre questionnaire démographique. Les résultats de l’étude suggèrent qu’une décision d’offrir des vidéos aux élèves peut mener à des gains d'apprentissage plus élevés comparativement à l’option lecture. En particulier, les élèves du groupe initial de lecture se sont considérablement améliorées lorsqu'ils ont été fournis avec des vidéos, surtout par rapport au groupe initial de vidéos, dont l’amélioration durant la même période était marginale. Cependant, ces conclusions sont un peu incertaines, parce que la performance du groupe initial de vidéos était globalement médiocre, ce qui aurait pu mener aux résultats observés. Par contre, les résultats obtenus des questionnaires étaient moins incertains, et indiquent que les élèves qui se sont vues attribués des lectures passaient plus de temps pour préparer leurs classes, étaient plus aptes à ne pas faire les préparatifs nécessaires, et ont aperçus leurs niveaux d'efforts et leurs charges de travails comme étant plus élevés. Ces facteurs étaient suffisamment importants qu'ils ont contribué au fait que quatre étudiants du groupe de lecture, soit 10% de la classe, ont abandonnés le cours durant le premier tiers du semestre. Finalement, les élèves ont démontrés des attitudes plus positives envers les vidéos, bien qu'ils ne trouvaient pas nécessairement que ceux-ci étaient plus efficaces que les lectures lorsqu’ils voulaient étudier pour les examens. Basé sur ces résultats, il est possible de conclure que tout effort de la part d'un instructeur pour préparer ou organiser des vidéos pédagogiques serait probablement un bon investissement. En plus, et même en dehors du contexte d'une classe renversée, cette étude offre une indication qu’une stratégie pédagogique qui oblige les étudiants à faire beaucoup de lecture peut imposer des exigences et une charge de travail très élevée pour les étudiants, ce qui devrait donner cause à tout instructeur qui attribue des lectures à réévaluer leur approche. Enfin, puisque les deux groupes étaient équivalents d’un point de vue statistique sur toutes les mesures démographiques, il semble que certaines disparités entre les deux groupes dans les notes d'évaluation et d’examens peuvent possiblement avoir été entraînées par des différentes dynamiques dans chaque group. Par conséquent, il est suggéré que des mesures de relations entre étudiants et de dynamiques de groupes devraient être incorporé dans n’importe qu’elle recherche comparant deux ou plusieurs groupes d’étudiants, et surtout quand ces étudiants sont en interaction, parce-que en toute apparence les dynamiques de groupes ont le potentiel de jouer un rôle clé dans les résultats obtenus.

Effect of light-sticks and electralunie attractors on surface-longline catches of swordfish (Xiphias gladius, Linnaeus, 1959) in the southwest equatorial Atlantic

Two experimental fishing trials were carried out off the coast of Pernambuco, Brazil, in 1999 and 2001, using a small artisanal longliner. In experiment 1, six-hook baskets with three chemical light-sticks on alternating hooks had significantly higher catch rates than those with zero or with a light-stick on every hook, with most swordfish accounted for by hooks with light-sticks. Analysis of the data from experiment 2 showed no significant difference between electralume attractors, consisting of AA lithium batteries protected by a solid cover and light-sticks that produce a fluorescent light when two chemical products are mixed. Significant differences were detected in mean CPUE by size class, with most swordfish belonging to class 'b' (125-170 cm lower jaw to fork length (LJFL)). No differences, however, were found for swordfish catches in classes 'a' (< 125 cm LJFL) and V, and no evidence was found of interaction between the two factors (attractor and size class). Although there was no significant difference between the total length-frequency distributions of swordfish caught with light-sticks and electralume attractors, signibcant differences were found for fish smaller than 125 cm LJFL, with electralume catches consisting of smaller swordfish than those of gear using light-sticks. (c) 2004 Elsevier B.V. All rights reserved.

Effect of light-sticks and electralume attractors on surface-longline catches of swordfish (Xiphias gladius, Linnaeus, 1959) in the southwest equatorial Atlantic

Two experimental fishing trials were carried out off the coast of Pernambuco, Brazil, in 1999 and 2001, using a small artisanal longliner. In experiment 1, six-hook baskets with three chemical light-sticks on alternating hooks had significantly higher catch rates than those with zero or with a light-stick on every hook, with most swordfish accounted for by hooks with light-sticks. Analysis of the data from experiment 2 showed no significant difference between electralume attractors, consisting of AA lithium batteries protected by a solid cover and light-sticks that produce a fluorescent light when two chemical products are mixed. Significant differences were detected in mean CPUE by size class, with most swordfish belonging to class ‘b’ (125–170 cm lower jaw to fork length (LJFL)). No differences, however, were found for swordfish catches in classes ‘a’ (<125 cm LJFL) and ‘b’, and no evidence was found of interaction between the two factors (attractor and size class). Although there was no significant difference between the total length-frequency distributions of swordfish caught with light-sticks and electralume attractors, significant differences were found for fish smaller than 125 cm LJFL, with electralume catches consisting of smaller swordfish than those of gear using light-sticks.

Evolutionary patterns of MHC class II B in owls and their implications for the understanding of avian MHC evolution

Owing to its special mode of evolution and central role in the adaptive immune system, the major histocompatibility complex (MHC) has become the focus of diverse disciplines such as immunology, evolutionary ecology, and molecular evolution. MHC evolution has been studied extensively in diverse vertebrate lineages over the last few decades, and it has been suggested that birds differ from the established mammalian norm. Mammalian MHC genes evolve independently, and duplication history (i.e., orthology) can usually be traced back within lineages. In birds, this has been observed in only 3 pairs of closely related species. Here we report strong evidence for the persistence of orthology of MHC genes throughout an entire avian order. Phylogenetic reconstructions of MHC class II B genes in 14 species of owls trace back orthology over tens of thousands of years in exon 3. Moreover, exon 2 sequences from several species show closer relationships than sequences within species, resembling transspecies evolution typically observed in mammals. Thus, although previous studies suggested that long-term evolutionary dynamics of the avian MHC was characterized by high rates of concerted evolution, resulting in rapid masking of orthology, our results question the generality of this conclusion. The owl MHC thus opens new perspectives for a more comprehensive understanding of avian MHC evolution.

Isolation and characterization of major histocompatibility complex (MHC) class II B genes in the Barn owl (Aves: Tyto alba)

We isolated major histocompatibility complex class II B (MHCIIB) genes in the Barn owl (Tyto alba). A PCR-based approach combined with primer walking on genomic and complementary DNA as well as Southern blot analyses revealed the presence of two MHCIIB genes, both being expressed in spleen, liver, and blood. Characteristic structural features of MHCIIB genes as well as their expression and high non-synonymous substitution rates in the region involved in antigen binding suggest that both genes are functional. MHC organization in the Barn owl is simple compared to passerine species that show multiple duplications, and resembles the minimal essential MHC of chicken.

Peptide-antibody conjugates for tumour therapy: a MHC-class-II-restricted tetanus toxin peptide coupled to an anti-Ig light chain antibody can induce cytotoxic lysis of a human B-cell lymphoma by specific CD4 T cells.

Anti-idiotype antibody therapy of B-cell lymphomas, despite numerous promising experimental and clinical studies, has so far met with limited success. Tailor-made monoclonal anti-idiotype antibodies have been injected into a large series of lymphoma patients, with a few impressive complete tumour remissions but a large majority of negative responses. The results presented here suggest that, by coupling to antilymphoma idiotype antibodies a few molecules of the tetanus toxin universal epitope peptide P2 (830-843), one could markedly increase the efficiency of this therapy. We show that after 2-hr incubation with conjugates consisting of the tetanus toxin peptide P2 coupled by an S-S bridge to monoclonal antibodies directed to the lambda light chain of human immunoglobulin, human B-lymphoma cells can be specifically lysed by a CD4 T-lymphocyte clone specific for the P2 peptide. Antibody without peptide did not induce B-cell killing by the CD4 T-lymphocyte clone. The free cysteine-peptide was also able to induce lysis of the B-lymphoma target by the T-lymphocyte clone, but at a molar concentration 500 to 1000 times higher than that of the coupled peptide. Proliferation assays confirmed that the antibody-peptide conjugate was antigenically active at a much lower concentration than the free peptide. They also showed that antibody-peptide conjugates required an intact processing function of the B cell for peptide presentation, which could be selectively inhibited by leupeptin and chloroquine.(ABSTRACT TRUNCATED AT 250 WORDS)

Regeneration of class III furcation defects with basic fibroblast growth factor (b-FGF) associated with GTR. A descriptive and histometric study in dogs

Background: the poor predictability of periodontal regenerative treatment of Class III furcation defects stimulates the study of alternatives to improve its results, such as the use of polypeptide growth factors. The objective of this study was to evaluate, both histologically and histometrically, the effects of topical application of basic fibroblast growth factor (b-FGF) associated with guided tissue regeneration (GTR) in the treatment of Class III defects surgically induced in dogs.Methods: All second and fourth premolars of 5 mongrel dogs were used and randomly assigned to one of three treatment groups: group 1 (control), treated with scaling and root planing, tetracycline hydrochloride (125 mg/ml) conditioning, and GTR with a collagen membrane; group 2, same treatment as group 1 plus 0.5 mg of b-FGF; group 3, same treatment as group 1 plus 1.0 mg of b-FGF. After a 90-day healing period, routine histologic processing and staining with hematoxylin and eosin and Masson trichrome were performed.Results: the descriptive analysis indicated better regenerative results in both groups treated with b-FGF while the histometric data, analyzed by means of analysis of variance (ANOVA), showed greater filling of the defects in group 2 in comparison to the defects in groups 3 and 1, respectively, which was represented by a smaller area of plaque-occupied space (P = 0.004) as well as a greater amount of newly formed cementum (P = 0.002).Conclusions: These results indicate that b-FGF, especially in smaller doses, may enhance the regenerative results in Class III furcation lesions, leading to greater filling of these defects with both mineralized and non-mineralized tissues.

Structural and functional characterization of Group B Streptococcus class C sortases

In Group B Streptococcus (GBS) three structurally distinct types of pili have been discovered as potential virulence factors and vaccine candidates. The pilus-forming proteins are assembled into high-molecular weight polymers via a transpeptidation mechanism mediated by specific class C sortases. Using a multidisciplinary approach including bioinformatics, structural and biochemical studies and in vivo mutagenesis we performed a broad characterization of GBS sortase C. The high resolution X-ray structure of the enzymes revealed that the active site, located into the β-barrel core of the enzyme, is made of the catalytic triad His157-Cys219-Arg228 and covered by a loop, known as the “lid”. We show that the catalytic triad and the predicted N- and C-terminal trans-membrane regions are required for the enzyme activity. Interestingly, by in vivo complementation mutagenesis studies we found that the deletion of the entire lid loop or mutations in specific lid key residues had no effect on catalytic activity of the enzyme. In addition, kinetic characterizations of recombinant enzymes indicate that the lid mutants can still recognize and cleave the substrate-mimicking peptide at least as well as the wild type protein.

Class-switzched B cells display response to therapeutic B-cell depletion in rheumatoid arthritis

Introduction Reconstitution of peripheral blood (PB) B cells after therapeutic depletion with the chimeric anti-CD20 antibody rituximab (RTX) mimics lymphatic ontogeny. In this situation, the repletion kinetics and migratory properties of distinct developmental B-cell stages and their correlation to disease activity might facilitate our understanding of innate and adaptive B-cell functions in rheumatoid arthritis (RA). Methods Thirty-five 'RTX-naïve' RA patients with active arthritis were treated after failure of tumour necrosis factor blockade in an open-label study with two infusions of 1,000 mg RTX. Prednisone dose was tapered according to clinical improvement from a median of 10 mg at baseline to 5 mg at 9 and 12 months. Conventional disease-modifying antirheumatic drugs were kept stable. Subsets of CD19+ B cells were assessed by flow cytometry according to their IgD and CD27 surface expression. Their absolute number and relative frequency in PB were followed every 3 months and were determined in parallel in synovial tissue (n = 3) or synovial fluid (n = 3) in the case of florid arthritis. Results Six of 35 patients fulfilled the European League Against Rheumatism criteria for moderate clinical response, and 19 others for good clinical response. All PB B-cell fractions decreased significantly in number (P < 0.001) after the first infusion. Disease activity developed independently of the total B-cell number. B-cell repopulation was dominated in quantity by CD27-IgD+ 'naïve' B cells. The low number of CD27+IgD- class-switched memory B cells (MemB) in the blood, together with sustained reduction of rheumatoid factor serum concentrations, correlated with good clinical response. Class-switched MemB were found accumulated in flaring joints. Conclusions The present data support the hypothesis that control of adaptive immune processes involving germinal centre-derived, antigen, and T-cell-dependently matured B cells is essential for successful RTX treatment.