Recombinant adeno associated viral (AAV) vector type 9 delivery of Ex1-Q138-mutant huntingtin in the rat striatum as a short-time model for in vivo studies in drug discovery

Huntington's disease (HD) is an inherited neurodegenerative disorder characterized by dyskinesia, cognitive impairment and emotional disturbances, presenting progressive neurodegeneration in the striatum and intracellular mutant Huntingtin (mHTT) aggregates in various areas of the brain. Recombinant Adeno Associated Viral (rAAV) vectors have been successfully used to transfer foreign genes to the brain of adult animals. In the present study we report a novel in vivo rat HD model obtained by stereotaxic injection of rAAV serotype2/9 containing Exon1-Q138 mHTT (Q138) and Exon1-Q17 wild type HTT (Q17; control), respectively in the right and in the left striatum, and expressed as C-terminal GFP fusions to facilitate detection of infected cells and aggregate production. Immunohistochemical analysis of brain slices from animals sacrificed twenty-one days after viral infection showed that Q138 injection resulted in robust formation of GFP-positive aggregates in the striatum, increased GFAP and microglial activation and neurodegeneration, with little evidence of any of these events in contralateral tissue infected with wild type (Q17) expressing construct. Differences in the relative metabolite concentrations (N-Acetyl Aspartate/Creatine and Myo-Inositol/Creatine) were observed by H1 MR Spectroscopy. By quantitative RT-PCR we also demonstrated that mHTT induced changes in the expression of genes previously shown to be altered in other rodent HD models. Importantly, administration of reference compounds previously shown to ameliorate the aggregation and neurodegeneration phenotypes in preclinical HD models was demonstrated to revert the mutant HTT-dependent effects in our model. In conclusion, the AAV2/9-Q138/Q17 exon 1 HTT stereotaxic injection represents a useful first-line in vivo preclinical model for studying the biology of mutant HTT exon 1 in the striatum and to provide early evidence of efficacy of therapeutic approaches

Elaboração de um instrumento de consulta de enfermagem para suspeita e casos de COVID-19 e/ou Influenza

The aim was to build a facilitating tool for nursing consultation, aimed at caring for patients with suspected or confirmed cases of COVID-19 and/or Influenza in Primary Health Care. This is an experience report of nursing students and professors during the construction of the nursing consultation support instrument. The elaboration took place in two stages: bibliographical research and elaboration of the instrument. The bibliographical research was carried out from scientific articles in a database with a focus on Primary Health Care. The construction of the instrument took place in eleven domains, namely: (1) Personal Data; (2) Environment; (3) Complaints; (4) Personal and Family Background; (5) Habits of Life; (6) Symptoms; (7) Epidemiology; (8) Vital Signs; (9) Physical Examination; (10) Vaccines; (11). Exams. The creation of an instrument becomes opportune since it acts as a facilitator for the professional during the nursing consultation. The study allowed to acquire knowledge about COVID-19 and Influenza. The instrument was designed to guide nurses in screening cases of flu syndrome in Basic Health Units.Objetivou-se construção de uma ferramenta facilitadora para consulta de enfermagem, voltada ao atendimento de pacientes com suspeitas ou casos confirmados de COVID-19 e/ou Influenza na Atenção Primária à Saúde. Trata-se de um relato de experiência das discentes e docentes de enfermagem durante a construção do instrumento de apoio à consulta de enfermagem. A elaboração ocorreu em duas etapas: pesquisa bibliográfica e elaboração do instrumento. A pesquisa bibliográfica foi realizada a partir de artigos científicos em base de dados com enfoque na Atenção Primária à Saúde. A construção do instrumento ocorreu em onze domínios, a saber: (1) Dados Pessoais; (2) Ambiente; (3) Queixas; (4) Antecedentes Pessoais e Familiares; (5) Hábitos de Vida; (6) Sintomas; (7) Epidemiologia; (8) Sinais Vitais; (9) Exame Físico; (10) Vacinas; (11). Exames. A criação de um instrumento se torna oportuna uma vez que atua como facilitador para o profissional durante a consulta de enfermagem. O estudo permitiu adquirir conhecimento sobre COVID-19 e Influenza. O instrumento foi elaborado para nortear o enfermeiro na triagem de casos de síndromes gripais nas Unidades Básicas de Saúde

Elaboração de um instrumento de consulta de enfermagem para suspeita e casos de COVID-19 e/ou Influenza

The aim was to build a facilitating tool for nursing consultation, aimed at caring for patients with suspected or confirmed cases of COVID-19 and/or Influenza in Primary Health Care. This is an experience report of nursing students and professors during the construction of the nursing consultation support instrument. The elaboration took place in two stages: bibliographical research and elaboration of the instrument. The bibliographical research was carried out from scientific articles in a database with a focus on Primary Health Care. The construction of the instrument took place in eleven domains, namely: (1) Personal Data; (2) Environment; (3) Complaints; (4) Personal and Family Background; (5) Habits of Life; (6) Symptoms; (7) Epidemiology; (8) Vital Signs; (9) Physical Examination; (10) Vaccines; (11). Exams. The creation of an instrument becomes opportune since it acts as a facilitator for the professional during the nursing consultation. The study allowed to acquire knowledge about COVID-19 and Influenza. The instrument was designed to guide nurses in screening cases of flu syndrome in Basic Health Units.Objetivou-se construção de uma ferramenta facilitadora para consulta de enfermagem, voltada ao atendimento de pacientes com suspeitas ou casos confirmados de COVID-19 e/ou Influenza na Atenção Primária à Saúde. Trata-se de um relato de experiência das discentes e docentes de enfermagem durante a construção do instrumento de apoio à consulta de enfermagem. A elaboração ocorreu em duas etapas: pesquisa bibliográfica e elaboração do instrumento. A pesquisa bibliográfica foi realizada a partir de artigos científicos em base de dados com enfoque na Atenção Primária à Saúde. A construção do instrumento ocorreu em onze domínios, a saber: (1) Dados Pessoais; (2) Ambiente; (3) Queixas; (4) Antecedentes Pessoais e Familiares; (5) Hábitos de Vida; (6) Sintomas; (7) Epidemiologia; (8) Sinais Vitais; (9) Exame Físico; (10) Vacinas; (11). Exames. A criação de um instrumento se torna oportuna uma vez que atua como facilitador para o profissional durante a consulta de enfermagem. O estudo permitiu adquirir conhecimento sobre COVID-19 e Influenza. O instrumento foi elaborado para nortear o enfermeiro na triagem de casos de síndromes gripais nas Unidades Básicas de Saúde

Morphometrical evaluation of neurodegeneration: an integrated multiparametric approach using whole slide imaging

Assessing degree of neurodegeneration and eventually identifying a compound related neuroprotection can be much more arduous than one might imagine: number of animals, number of samples/animal, experimental design and statistics, together with the choice of proper IHC markers and the technology whereby they are evaluated are prominent factors in order to obtain reliable results. In acute models, histopathological measurements are an essential part of the experimental procedure; here we describe an integrated (qualitative/quantitative) histological assessment to be applied for the evaluation of neuroprotection in acute models of Huntington’s disease. We used whole slide imaging to validate a rat model of Huntington disease obtained by intrastriatal viral vector delivery of mutant huntingtin (Htt). Recombinant Adeno Associated Viral (AAV) vectors have been used successfully to transfer genes in a variety of tissues, including the brain, in adult animals [1]. Here we used rAAV9, charged with Exon 1 Htt carrying 17 and 138 CAG repeats. AAV9-Ex1- GFP-Q138 injection induced the formation of GFP positive Htt aggregates in the entire striatal area, increased GFAP and microglial activation with respect to Q17 injected striatum. NeuN, ChAT, GFAP, OX42 immunohistochemistry and GFP epifluorescence were evaluated contemporaneously to qualitatively evaluate the degree of induced lesions; qualitative evaluation allowed to exclude animals that have not responded to rAAV9 infection. The remaining selected animals were used for a multivariate statistical analysis based on whole slide imaging of immunohistochemically stained sections. We believe this approach increases results reliability when evaluating animal models of neurodegeneration

Work addiction and its association with personality traits, general distress, and self-esteem among adult Italian workers

Background: Work addiction has become a topic of increasing research interest but has been little studied in Italy. Therefore, the aim of the present study was to investigate the associations between work addiction, assessed with a recently validated psychometric scale (i.e., Italian version of Bergen Work Addiction Scale, [BWAS]) and other psychological constructs. Methods: The sample comprised 367 Italian workers (Mean 16.11 years; SD±11.28) who completed a survey including the BWAS (Mean 19.422; SD±6.365), Depression Anxiety Stress Scales-21 (Mean 40.866; SD±29.865), Dutch Workaholism Scale (Mean 24.837; SD±6.488), Need for Recovery Scale (Mean 12.946; SD±7.340), Ten Item Personality Inventory (TIPI, Extraversion (Mean 4.253; SD±1.506); Agreeableness (Mean 5.431; SD±1.111), Conscientiousness (Mean 5.792; SD±1.067), Neuroticism (Mean 4.507; SD±1.480), Openness (Mean 4.801; SD±1.122), and Rosenberg’s Self-Esteem Scale (Mean 21.850; SD±6.796). Results: The results indicated that work addiction was positively associated with stress, anxiety, and depression, as well as with the number of hours worked and need for recovery. Moreover, BWAS scores explained 20.1% of an individual’s general psychological distress (i.e., depression, anxiety, and stress). Personality variables explained only a small amount of the variance in work addiction (15.4%). Conclusion: In the present study, a positive and significant association was found between the BWAS (assessing work addiction) and the DUWAS (assessing workaholism). Although work addiction and workaholism are different constructs, they have many characteristics in common. The study expands the work addiction literature base and demonstrates important associating factors in the Italian context

A potent and selective Sirtuin 1 inhibitor alleviates pathology in multiple animal and cell models of Huntington's disease

Protein acetylation, which is central to transcriptional control as well as other cellular processes, is disrupted in Huntington's disease (HD). Treatments that restore global acetylation levels, such as inhibiting histone deacetylases (HDACs), are effective in suppressing HD pathology in model organisms. However, agents that selectively target the disease-relevant HDACs have not been available. SirT1 (Sir2 in Drosophila melanogaster) deacetylates histones and other proteins including transcription factors. Genetically reducing, but not eliminating, Sir2 has been shown to suppress HD pathology in model organisms. To date, small molecule inhibitors of sirtuins have exhibited low potency and unattractive pharmacological and biopharmaceutical properties. Here, we show that highly selective pharmacological inhibition of Drosophila Sir2 and mammalian SirT1 using the novel inhibitor selisistat (selisistat; 6-chloro-2,3,4,9-tetrahydro-1H-carbazole-1-carboxamide) can suppress HD pathology caused by mutant huntingtin exon 1 fragments in Drosophila, mammalian cells and mice. We have validated Sir2 as the in vivo target of selisistat by showing that genetic elimination of Sir2 eradicates the effect of this inhibitor in Drosophila. The specificity of selisistat is shown by its effect on recombinant sirtuins in mammalian cells. Reduction of HD pathology by selisistat in Drosophila, mammalian cells and mouse models of HD suggests that this inhibitor has potential as an effective therapeutic treatment for human disease and may also serve as a tool to better understand the downstream pathways of SirT1/Sir2 that may be critical for H

siRNA screen identifies QPCT as a druggable target for Huntington's disease.

Huntington's disease (HD) is a currently incurable neurodegenerative condition caused by an abnormally expanded polyglutamine tract in huntingtin (HTT). We identified new modifiers of mutant HTT toxicity by performing a large-scale 'druggable genome' siRNA screen in human cultured cells, followed by hit validation in Drosophila. We focused on glutaminyl cyclase (QPCT), which had one of the strongest effects on mutant HTT-induced toxicity and aggregation in the cell-based siRNA screen and also rescued these phenotypes in Drosophila. We found that QPCT inhibition induced the levels of the molecular chaperone αB-crystallin and reduced the aggregation of diverse proteins. We generated new QPCT inhibitors using in silico methods followed by in vitro screening, which rescued the HD-related phenotypes in cell, Drosophila and zebrafish HD models. Our data reveal a new HD druggable target affecting mutant HTT aggregation and provide proof of principle for a discovery pipeline from druggable genome screen to drug development

A potent and selective Sirtuin 1 inhibitor alleviates pathology in multiple animal and cell models of Huntington's disease

Protein acetylation, which is central to transcriptional control as well as other cellular processes, is disrupted in Huntington's disease (HD). Treatments that restore global acetylation levels, such as inhibiting histone deacetylases (HDACs), are effective in suppressing HD pathology in model organisms. However, agents that selectively target the disease-relevant HDACs have not been available. SirT1 (Sir2 in Drosophila melanogaster) deacetylates histones and other proteins including transcription factors. Genetically reducing, but not eliminating, Sir2 has been shown to suppress HD pathology in model organisms. To date, small molecule inhibitors of sirtuins have exhibited low potency and unattractive pharmacological and biopharmaceutical properties. Here, we show that highly selective pharmacological inhibition of Drosophila Sir2 and mammalian SirT1 using the novel inhibitor selisistat (selisistat; 6-chloro-2,3,4,9-tetrahydro-1H-carbazole-1-carboxamide) can suppress HD pathology caused by mutant huntingtin exon 1 fragments in Drosophila, mammalian cells and mice. We have validated Sir2 as the in vivo target of selisistat by showing that genetic elimination of Sir2 eradicates the effect of this inhibitor in Drosophila. The specificity of selisistat is shown by its effect on recombinant sirtuins in mammalian cells. Reduction of HD pathology by selisistat in Drosophila, mammalian cells and mouse models of HD suggests that this inhibitor has potential as an effective therapeutic treatment for human disease and may also serve as a tool to better understand the downstream pathways of SirT1/Sir2 that may be critical for HD

BHPR research: qualitative1. Complex reasoning determines patients' perception of outcome following foot surgery in rheumatoid arhtritis

Background: Foot surgery is common in patients with RA but research into surgical outcomes is limited and conceptually flawed as current outcome measures lack face validity: to date no one has asked patients what is important to them. This study aimed to determine which factors are important to patients when evaluating the success of foot surgery in RA Methods: Semi structured interviews of RA patients who had undergone foot surgery were conducted and transcribed verbatim. Thematic analysis of interviews was conducted to explore issues that were important to patients. Results: 11 RA patients (9 ♂, mean age 59, dis dur = 22yrs, mean of 3 yrs post op) with mixed experiences of foot surgery were interviewed. Patients interpreted outcome in respect to a multitude of factors, frequently positive change in one aspect contrasted with negative opinions about another. Overall, four major themes emerged. Function: Functional ability & participation in valued activities were very important to patients. Walking ability was a key concern but patients interpreted levels of activity in light of other aspects of their disease, reflecting on change in functional ability more than overall level. Positive feelings of improved mobility were often moderated by negative self perception ("I mean, I still walk like a waddling duck”). Appearance: Appearance was important to almost all patients but perhaps the most complex theme of all. Physical appearance, foot shape, and footwear were closely interlinked, yet patients saw these as distinct separate concepts. Patients need to legitimize these feelings was clear and they frequently entered into a defensive repertoire ("it's not cosmetic surgery; it's something that's more important than that, you know?”). Clinician opinion: Surgeons' post operative evaluation of the procedure was very influential. The impact of this appraisal continued to affect patients' lasting impression irrespective of how the outcome compared to their initial goals ("when he'd done it ... he said that hasn't worked as good as he'd wanted to ... but the pain has gone”). Pain: Whilst pain was important to almost all patients, it appeared to be less important than the other themes. Pain was predominately raised when it influenced other themes, such as function; many still felt the need to legitimize their foot pain in order for health professionals to take it seriously ("in the end I went to my GP because it had happened a few times and I went to an orthopaedic surgeon who was quite dismissive of it, it was like what are you complaining about”). Conclusions: Patients interpret the outcome of foot surgery using a multitude of interrelated factors, particularly functional ability, appearance and surgeons' appraisal of the procedure. While pain was often noted, this appeared less important than other factors in the overall outcome of the surgery. Future research into foot surgery should incorporate the complexity of how patients determine their outcome Disclosure statement: All authors have declared no conflicts of interes

Studio dell'ipofunzione colinergica indotta nel ratto dall'invecchiamento, dall'etanolo e dalla (beta(-amiloide(

Dottorato di ricerca in farmacologia e tossicologia. 7. ciclo. A.a. 1994-95. Coordinatore P. F. MannaioniConsiglio Nazionale delle Ricerche - Biblioteca Centrale - P.le Aldo Moro, 7, Rome; Biblioteca Nazionale Centrale - P.za Cavalleggeri, 1, Florence / CNR - Consiglio Nazionale delle RichercheSIGLEITItal