Protein Quality Control Disruption by PKCβII in Heart Failure; Rescue by the Selective PKCβII Inhibitor, βIIV5-3

Myocardial remodeling and heart failure (HF) are common sequelae of many forms of cardiovascular disease and a leading cause of mortality worldwide. Accumulation of damaged cardiac proteins in heart failure has been described. However, how protein quality control (PQC) is regulated and its contribution to HF development are not known. Here, we describe a novel role for activated protein kinase C isoform βII (PKCβII) in disrupting PQC. We show that active PKCβII directly phosphorylated the proteasome and inhibited proteasomal activity in vitro and in cultured neonatal cardiomyocytes. Importantly, inhibition of PKCβII, using a selective PKCβII peptide inhibitor (βIIV5-3), improved proteasomal activity and conferred protection in cultured neonatal cardiomyocytes. We also show that sustained inhibition of PKCβII increased proteasomal activity, decreased accumulation of damaged and misfolded proteins and increased animal survival in two rat models of HF. Interestingly, βIIV5-3-mediated protection was blunted by sustained proteasomal inhibition in HF. Finally, increased cardiac PKCβII activity and accumulation of misfolded proteins associated with decreased proteasomal function were found also in remodeled and failing human hearts, indicating a potential clinical relevance of our findings. Together, our data highlights PKCβII as a novel inhibitor of proteasomal function. PQC disruption by increased PKCβII activity in vivo appears to contribute to the pathophysiology of heart failure, suggesting that PKCβII inhibition may benefit patients with heart failure. (218 words

Identification of New Genes Related to Virulence of Xanthomonas axonopodis Pv. Citri during Citrus Host Interactions

A mutant library of the bacterium Xanthomonas citri subsp. citri strain 306 pathotype A (Xac), the causative agent of most aggressive Asiatic type A citrus canker, was screened regarding altered canker symptoms after inoculations into Citrus sinensis and Citrus limonia host leaves. Twenty-six mutants have shown phenotypic virulence changes and have respectively knocked out gene identified by sequencing. In vivo growth curves were obtained for nine mutants to quantify how the mutations could affect pathogen's adaptability to growth inside and attack host plant infected tissue. Among identified genes in mutated strains, we could find those that until now had not been reported as being involved in Xac adaptation and/or virulence, such as predicted to encode for xylose repressor-like protein (XAC Delta xylR), Fe-S oxidoredutase (XAC Delta aslB), helicase IV (XAC Delta helD), ubiquinol cytochrome c oxidoreductase iron-sulfur subunit (XAC Delta petA), chromosome partitioning protein (XAC Delta parB) and cell division protein FtsB (XAC Delta ftsB), in addition to genes predicted to encode for hypothetical proteins. The new genes found in this study as being relevant to adaptation and virulence, improve the understanding of Xac fitness during citrus plant attack and canker symptoms development.Fundacao de Amparo a Pesquisa do Estado de Sao Paulo-FAPESPCoordenacao de Aperfeicoamento de Pessoal de Nivel Superior-CAPESCNPqFundo de Defesa da Citricultura (FUNDECITRUS)CAPESFUNDECITRUSFAPESPSao Paulo State Univ UNESP, Fac Agr Sci & Vet, Dept Technol, Jaboticabal, SP, BrazilCitriculture Res Ctr Sylvio Moreira IAC Campinas, Campinas, SP, BrazilUniv Fed Ouro Preto, Inst Exact & Biol Sci, Dept Biol Sci, Ouro Preto, MG, BrazilUniv Fed Ouro Preto, Res Ctr Biol Sci NUPEB, Inst Exact & Biol Sci, Dept Biol Sci, Ouro Preto, MG, BrazilSao Paulo State Univ UNESP, Fac Agr Sci & Vet, Plant Hlth Dept, Jaboticabal, SP, BrazilFed Univ Sao Paulo UNIFESP, Dept Biol Sci, Diadema, SP, BrazilFed Univ Sao Paulo UNIFESP, Dept Biol Sci, Diadema, SP, BrazilFAPESP: 04/02006-7Web of Scienc

Proteomics-based identification of differentially abundant proteins reveals adaptation mechanisms of Xanthomonas citri subsp citri during Citrus sinensis infection

Background: Xanthomonas citri subsp. citri (Xac) is the causal agent of citrus canker. A proteomic analysis under in planta infectious and non-infectious conditions was conducted in order to increase our knowledge about the adaptive process of Xac during infection. Results: For that, a 2D-based proteomic analysis of Xac at 1, 3 and 5 days after inoculation, in comparison to Xac growth in NB media was carried out and followed by MALDI-TOF-TOF identification of 124 unique differentially abundant proteins. Among them, 79 correspond to up-regulated proteins in at least one of the three stages of infection. Our results indicate an important role of proteins related to biofilm synthesis, lipopolysaccharides biosynthesis, and iron uptake and metabolism as possible modulators of plant innate immunity, and revealed an intricate network of proteins involved in reactive oxygen species adaptation during Plants'Oxidative Burst response. We also identified proteins previously unknown to be involved in Xac-Citrus interaction, including the hypothetical protein XAC3981. A mutant strain for this gene has proved to be non-pathogenic in respect to classical symptoms of citrus canker induced in compatible plants. Conclusions: This is the first time that a protein repertoire is shown to be active and working in an integrated manner during the infection process in a compatible host, pointing to an elaborate mechanism for adaptation of Xac once inside the plant.Fundacao de Amparo a Pesquisa do Estado de Sao PauloFundacao de Amparo a Pesquisa do Estado de Minas GeraisBIGA grant-CAPESUniv Fed Ouro Preto, Inst Ciencias Exatas & Biol, Dept Ciencias Biol DECBI, Ouro Preto, MG, BrazilUniv Fed Ouro Preto, Nucleo Pesquisas Ciencias Biol NUPEB, Ouro Preto, MG, BrazilUniv Fed Rio de Janeiro, Inst Quim, Dept Bioquim DBq, Rio De Janeiro, RJ, BrazilUniv Estadual Paulista, UNESP, Dept Tecnol, Fac Ciencias Agr & Vet Jaboticabal, Jaboticabal, SP, BrazilUniv Estadual Campinas, UNICAMP, Inst Quim, Campinas, SP, BrazilUniv Sao Paulo, Inst Quim, Dept Bioquim, Sao Paulo, SP, BrazilUniv Fed Sao Paulo UNIFESP, Dept Ciencias Biol, Diadema, SP, BrazilVirginia Tech, Biocomplex Inst, Blacksburg, VA USAUniv Sao Paulo, Inst Quim, Dept Bioquim, Sao Paulo, SP, BrazilUniv Fed Sao Paulo UNIFESP, Dept Ciencias Biol, Diadema, SP, BrazilCAPESFAPESP: 04/02006-7Fundacao de Amparo a Pesquisa do Estado de Minas Gerais: CBB-APQ-04425-10BIGA grant-CAPES: 3385/2013Web of Scienc

AVALIAÇÃO DA QUALIDADE DE CARCAÇA DA TILÁPIA ALIMENTADA COM LEVEDURAS

O desenvolvimento do setor aquícola tem expandido a intensidade dos sistemas produtivos,expondo os peixes a fatores estressantes que podem causar enfermidades nos animais.Aumentando a utilização indiscriminada de quimioterápicos para controle e profilaxia o queinstigou a busca por alternativas mais sustentáveis para o tratamento de doenças e promotoresde crescimento, como o uso de probióticos como agentes de controle biológico. A leveduraSaccharomyces cerevisiae é um dos produtos de origem microbiana que vêm sendoamplamente estudada por ser um fungo não patogênico, que por possuir rápida velocidade decrescimento, ocorre sempre excesso de produção, e pode ser usada para os mais diversos finsdepois de desidratada, como por exemplo, na alimentação animal. Já sendo utilizada emdiversas espécies, como suínos, bovinos, frangos de corte, entre outras. Diante do exposto opresente estudo objetivou avaliar a suplementação da levedura Saccharomyces cerevisiaecomo probiótico para tilápias-do-nilo (Oreochromis niloticus). No experimento foramutilizadas 60 tilápias-do-nilo com peso médio de 150 g, e cepas de S. cerevisae foram cedidase previamente incorporadas à ração pela Biosyn Nutrição Animal LTDA. Os peixes foramdistribuídos em dez unidades experimentais (6/EU), equipadas com sistema de recirculação deágua e aeração constante. As caixas foram divididas em dois tratamentos com cincorepetições, peixes alimentados com dieta: 1º) suplementada por Saccharomyces cerevisae(SAC); e 2º) controle (sem suplementação); Após o período experimental de 120 dias, todosos peixes foram pesados para determinação dos parâmetros zootécnicos, e quatro peixes decada tanque (20 por tratamento) foram utilizados para coleta sanguínea e realização deanálises hematológicas, e posteriormente eutanasiados e eviscerados para coleta dos dados afim de obter os valores de peso de carcaça, peso e comprimento de trato digestório e fígado etambém para a determinação da composição centesimal do filé. Ao avaliar os parâmetroshematológicos pôde-se observar aumento significativo no número de eritrócitos, trombócitos,linfócitos e monócitos no tratamento SAC. Os índices zootécnicos apresentaram melhora nosanimais suplementados com a levedura com aumento no ganho de peso semanal, eficiênciaalimentar, taxa de retenção proteica e produtividade e uma menor conversão alimentar, alémdo aumento tendencioso nos níveis de proteína observados nos filés em relação ao grupocontrole. Portanto pode-se concluir que a suplementação com S. cerevisiae demonstroupropriedades probióticas na fase de engorda e melhora na qualidade de carcaça de juvenis detilápia-do-nilo (Oreochromis niloticus)

Low medication adherence in patients with Short Bowel Syndrome dependent on parenteral nutrition

Study design: Cross-sectional. Aim: We aimed to investigate adherence to oral medications in patients with severe Short Bowel Syndrome (SBS). Casuistic and Methods: Ten patients with severe intestinal failure attending a specialized unit in a university hospital were enrolled in this study. All patients received intermittent parenteral nutrition for at least 4 months. The participants (six men and four women) had 56 ± 13 years and body mass index of 18 ± 3 kg/m2. Information related to the use of medications at home was recorded, and adherence behavior was assessed by using the Morisky Medication Adherence Scale. Results: Four patients (40%) had high medication adherence. Among patients with low medication adherence, unintentional behavior was observed in two patients, intentional behavior was observed in one patient, and an association of unintentional with intentional behavior was identified in three patients. Conclusion: Outside the hospital setting, most patients with SBS have low adherence to oral medications. These results suggest the development and implementation of targeted strategies aimed to promote awareness and adherence to medication treatmentModelo do estudo: Transversal. Objetivo do estudo: Avaliar a adesão de pacientes com Síndrome do Intestino Curto ao tratamento medicamentoso por via oral. Casuística e Métodos: O estudo inclui 10 pacientes com Síndrome do Intestino Curto (SIC) acompanhados em unidade especializada de um hospital universitário. Todos os pacientes recebiam nutrição parenteral por no mínimo quatro meses. A casuística consistia em seis homens e quatro mulheres, com idade de 56 ± 13 anos e Índice de Massa Corporal de 18 ± 3kg/m2. Foram registradas informações sobre o uso de medicamentos no domicílio. A adesão ao tratamento medicamentoso foi avaliada pelo teste de Morisky e foi identificada a natureza do comportamento em relação à não adesão. Resultados Apenas 40% dos pacientes foram classificados como “mais aderentes”. Dentre os “menos aderentes”, a baixa adesão por associação do comportamento intencional e não intencional foi observada em três casos; dois pacientes apresentavam apenas o comportamento intencional e o comportamento não intencional foi documentado em um paciente. Conclusões: A maioria dos pacientes tem baixa adesão ao tratamento medicamentoso por via oral durante o período em que eles permanecem em seus domicílios. Tais resultados sugerem o desenvolvimento e a implementação de estratégias que visem à percepção da relevância do uso correto dos medicamentos e a promoção da adesão ao tratamento medicamentos

Exercise training prevents oxidative stress and ubiquitin-proteasome system overactivity and reverse skeletal muscle atrophy in heart failure

Background: Heart failure (HF) is known to lead to skeletal muscle atrophy and dysfunction. However, intracellular mechanisms underlying HF-induced myopathy are not fully understood. We hypothesized that HF would increase oxidative stress and ubiquitin-proteasome system (UPS) activation in skeletal muscle of sympathetic hyperactivity mouse model. We also tested the hypothesis that aerobic exercise training (AET) would reestablish UPS activation in mice and human HF. Methods/Principal Findings: Time-course evaluation of plantaris muscle cross-sectional area, lipid hydroperoxidation, protein carbonylation and chymotrypsin-like proteasome activity was performed in a mouse model of sympathetic hyperactivity-induced HF. At the 7th month of age, HF mice displayed skeletal muscle atrophy, increased oxidative stress and UPS overactivation. Moderate-intensity AET restored lipid hydroperoxides and carbonylated protein levels paralleled by reduced E3 ligases mRNA levels, and reestablished chymotrypsin-like proteasome activity and plantaris trophicity. In human HF (patients randomized to sedentary or moderate-intensity AET protocol), skeletal muscle chymotrypsin-like proteasome activity was also increased and AET restored it to healthy control subjects' levels. Conclusions: Collectively, our data provide evidence that AET effectively counteracts redox imbalance and UPS overactivation, preventing skeletal myopathy and exercise intolerance in sympathetic hyperactivity-induced HF in mice. Of particular interest, AET attenuates skeletal muscle proteasome activity paralleled by improved aerobic capacity in HF patients, which is not achieved by drug treatment itself. Altogether these findings strengthen the clinical relevance of AET in the treatment of HF.Fundacao de Amparo a Pesquisa do Estado de Sao Paulo (FAPESP)Fundacao de Amparo a Pesquisa do Estado de Sao Paulo (FAPESP) [2006/61523-7, 2006/58460-4]Conselho Nacional de Pesquisa e Desenvolvimento (CNPq) [473251/2009-4, 301519/2008-0, 301867/2010-0]Conselho Nacional de Pesquisa e Desenvolvimento (CNPq

Distribution and biological role of the oligopeptide-binding protein (OppA) in Xanthomonas species

In this study we investigated the prevalence of the oppA gene, encoding the oligopeptide binding protein (OppA) of the major bacterial oligopeptide uptake system (Opp), in different species of the genus Xanthomonas. The oppA gene was detected in two Xanthomonas axonopodis strains among eight tested Xanthomonas species. The generation of an isogenic oppA-knockout derivative of the Xac 306 strain, showed that the OppA protein neither plays a relevant role in oligopeptide uptake nor contributes to the infectivity and multiplication of the bacterial strain in leaves of sweet orange (Citrus sinensis) and Rangpur lime (Citrus limonia). Taken together these results suggest that the oppA gene has a recent evolutionary history in the genus and does not contribute in the physiology or pathogenesis of X. axonopodis

Acidity and characterization of 12-Tungstophosphoric acid supported on Silica-alumina

This work deals with preparation and characterization of H3PW12O40 (H3PW) supported on silica-alumina. Impregnation of H3PW (15, 20, 30 and 40 wt.%) on commercial silica-alumina support in acidic aqueous solution is effective for preparing this catalyst keeping its Keggin structure, according to different methods of characterization. The catalysts were tested in a model reaction of acetic acid with ethanol and 30 wt.% H3PW/SiO2-Al2O3 had the highest activity under the conditions: catalyst calcination at 300 ºC, temperature of 100 ºC, acetic acid:ethanol molar ratio of 2:1 and catalyst:acetic acid mass ratio of 10 wt.%. The reaction yield was 79 and 100% selectivity for ethyl acetate over three reutilizations, for reaction time of 2 h. The calculated total acid site distribution was 0.299 mmol g-1 (97% of the theoretical probed by pyridine), and most of these (0.236 mmol g-1) were Brønsted weak-medium strength (pyridine desorption between 300 and 500 ºC)

The HIBEAM program: search for neutron oscillations at the ESS

With the construction of the European Spallation Source, a remarkable opportunity has emerged to conduct high sensitivity searches for neutron oscillations, including a first search for thirty years for free neutrons converting to antineutrons. Furthermore, searches can be made for transitions of neutrons and antineutrons to sterile neutron states. The HIBEAM program provides an increase in sensitivity of an order of magnitude compared to previous work. The HIBEAM program corresponds to baryon number violation by one and two units. The observation of a process satisfying a Sakharov condition addresses the open question of the origin of the matter-antimatter asymmetry in the Universe. Sterile neutron states would belong to a `dark' sector of particles which may explain dark matter. As electrically neutral, meta-stable objects that can be copiously produced and studied, neutrons represent an attractive portal to a `dark' sector. This paper describes the capability, design, infrastructure, and potential of the HIBEAM program. This includes a dedicated beamline, neutron optical system, magnetic shielding and control, and detectors for neutrons and antineutrons.Comment: 41 pages, 12 figure