Cathepsin K induces platelet dysfunction and affects cell signaling in breast cancer - molecularly distinct behavior of cathepsin K in breast cancer

Background: Breast cancer comprises clinically and molecularly distinct tumor subgroups that differ in cell histology and biology and show divergent clinical phenotypes that impede phase III trials, such as those utilizing cathepsin K inhibitors. Here we correlate the epithelial-mesenchymal-like transition breast cancer cells and cathepsin K secretion with activation and aggregation of platelets. Cathepsin K is up-regulated in cancer cells that proteolyze extracellular matrix and contributes to invasiveness. Although proteolytically activated receptors (PARs) are activated by proteases, the direct interaction of cysteine cathepsins with PARs is poorly understood. In human platelets, PAR-1 and -4 are highly expressed, but PAR-3 shows low expression and unclear functions. Methods: Platelet aggregation was monitored by measuring changes in turbidity. Platelets were immunoblotted with anti-phospho and total p38, Src-Tyr-416, FAK-Tyr-397, and TGF beta monoclonal antibody. Activation was measured in a flow cytometer and calcium mobilization in a confocal microscope. Mammary epithelial cells were prepared from the primary breast cancer samples of 15 women with Luminal-B subtype to produce primary cells. Results: We demonstrate that platelets are aggregated by cathepsin K in a dose-dependent manner, but not by other cysteine cathepsins. PARs-3 and -4 were confirmed as the cathepsin K target by immunodetection and specific antagonists using a fibroblast cell line derived from PARs deficient mice. Moreover, through co-culture experiments, we show that platelets activated by cathepsin K mediated the up-regulation of SHH, PTHrP, OPN, and TGF beta in epithelial-mesenchymal-like cells from patients with Luminal B breast cancer. Conclusions: Cathepsin K induces platelet dysfunction and affects signaling in breast cancer cells.Associacao Beneficente de Coleta de Sangue (Colsan)Fundacao de Amparo a Pesquisa do Estado de Sao Paulo (FAPESP)Coordenacao de Aperfeicoamento de Pessoal de Nivel Superior (CAPES)Conselho Nacional de Desenvolvimento Cientifico e Tecnologico (CNPq)Univ Fed Sao Paulo, Dept Gynecol, BR-04024002 Sao Paulo, SP, BrazilCOLSAN, Charitable Assoc Blood Collect, BR-04080006 Sao Paulo, SP, BrazilUniv Fed Sao Paulo, Dept Biophys, BR-04024002 Sao Paulo, SP, BrazilUniv Fed Sao Paulo, Dept Biochem, BR-04024002 Sao Paulo, SP, BrazilAntonio Prudente Fdn, AC Camargo Canc Ctr, AC Camargo Hosp Biobank, Dept Pathol, BR-01509010 Sao Paulo, SP, BrazilUniv Fed Sao Paulo, Cellular Gynecol Lab, Dept Gynecol, Rua Napoleao Barros 608, BR-04024002 Sao Paulo, BrazilUniv Fed Sao Paulo, Dept Gynecol, BR-04024002 Sao Paulo, SP, BrazilUniv Fed Sao Paulo, Dept Biophys, BR-04024002 Sao Paulo, SP, BrazilUniv Fed Sao Paulo, Dept Biochem, BR-04024002 Sao Paulo, SP, BrazilUniv Fed Sao Paulo, Cellular Gynecol Lab, Dept Gynecol, Rua Napoleao Barros 608, BR-04024002 Sao Paulo, BrazilFAPESP: 2012/19780-3FAPESP: 2012/19851-8FAPESP: 2009/53766-5Web of Scienc

Minimal Symptom Expression' in Patients With Acetylcholine Receptor Antibody-Positive Refractory Generalized Myasthenia Gravis Treated With Eculizumab

The efficacy and tolerability of eculizumab were assessed in REGAIN, a 26-week, phase 3, randomized, double-blind, placebo-controlled study in anti-acetylcholine receptor antibody-positive (AChR+) refractory generalized myasthenia gravis (gMG), and its open-label extension

Post-intervention Status in Patients With Refractory Myasthenia Gravis Treated With Eculizumab During REGAIN and Its Open-Label Extension

OBJECTIVE: To evaluate whether eculizumab helps patients with anti-acetylcholine receptor-positive (AChR+) refractory generalized myasthenia gravis (gMG) achieve the Myasthenia Gravis Foundation of America (MGFA) post-intervention status of minimal manifestations (MM), we assessed patients' status throughout REGAIN (Safety and Efficacy of Eculizumab in AChR+ Refractory Generalized Myasthenia Gravis) and its open-label extension. METHODS: Patients who completed the REGAIN randomized controlled trial and continued into the open-label extension were included in this tertiary endpoint analysis. Patients were assessed for the MGFA post-intervention status of improved, unchanged, worse, MM, and pharmacologic remission at defined time points during REGAIN and through week 130 of the open-label study. RESULTS: A total of 117 patients completed REGAIN and continued into the open-label study (eculizumab/eculizumab: 56; placebo/eculizumab: 61). At week 26 of REGAIN, more eculizumab-treated patients than placebo-treated patients achieved a status of improved (60.7% vs 41.7%) or MM (25.0% vs 13.3%; common OR: 2.3; 95% CI: 1.1-4.5). After 130 weeks of eculizumab treatment, 88.0% of patients achieved improved status and 57.3% of patients achieved MM status. The safety profile of eculizumab was consistent with its known profile and no new safety signals were detected. CONCLUSION: Eculizumab led to rapid and sustained achievement of MM in patients with AChR+ refractory gMG. These findings support the use of eculizumab in this previously difficult-to-treat patient population. CLINICALTRIALSGOV IDENTIFIER: REGAIN, NCT01997229; REGAIN open-label extension, NCT02301624. CLASSIFICATION OF EVIDENCE: This study provides Class II evidence that, after 26 weeks of eculizumab treatment, 25.0% of adults with AChR+ refractory gMG achieved MM, compared with 13.3% who received placebo

Mortality from gastrointestinal congenital anomalies at 264 hospitals in 74 low-income, middle-income, and high-income countries: a multicentre, international, prospective cohort study

Summary Background Congenital anomalies are the fifth leading cause of mortality in children younger than 5 years globally. Many gastrointestinal congenital anomalies are fatal without timely access to neonatal surgical care, but few studies have been done on these conditions in low-income and middle-income countries (LMICs). We compared outcomes of the seven most common gastrointestinal congenital anomalies in low-income, middle-income, and high-income countries globally, and identified factors associated with mortality. Methods We did a multicentre, international prospective cohort study of patients younger than 16 years, presenting to hospital for the first time with oesophageal atresia, congenital diaphragmatic hernia, intestinal atresia, gastroschisis, exomphalos, anorectal malformation, and Hirschsprung’s disease. Recruitment was of consecutive patients for a minimum of 1 month between October, 2018, and April, 2019. We collected data on patient demographics, clinical status, interventions, and outcomes using the REDCap platform. Patients were followed up for 30 days after primary intervention, or 30 days after admission if they did not receive an intervention. The primary outcome was all-cause, in-hospital mortality for all conditions combined and each condition individually, stratified by country income status. We did a complete case analysis. Findings We included 3849 patients with 3975 study conditions (560 with oesophageal atresia, 448 with congenital diaphragmatic hernia, 681 with intestinal atresia, 453 with gastroschisis, 325 with exomphalos, 991 with anorectal malformation, and 517 with Hirschsprung’s disease) from 264 hospitals (89 in high-income countries, 166 in middleincome countries, and nine in low-income countries) in 74 countries. Of the 3849 patients, 2231 (58·0%) were male. Median gestational age at birth was 38 weeks (IQR 36–39) and median bodyweight at presentation was 2·8 kg (2·3–3·3). Mortality among all patients was 37 (39·8%) of 93 in low-income countries, 583 (20·4%) of 2860 in middle-income countries, and 50 (5·6%) of 896 in high-income countries (p<0·0001 between all country income groups). Gastroschisis had the greatest difference in mortality between country income strata (nine [90·0%] of ten in lowincome countries, 97 [31·9%] of 304 in middle-income countries, and two [1·4%] of 139 in high-income countries; p≤0·0001 between all country income groups). Factors significantly associated with higher mortality for all patients combined included country income status (low-income vs high-income countries, risk ratio 2·78 [95% CI 1·88–4·11], p<0·0001; middle-income vs high-income countries, 2·11 [1·59–2·79], p<0·0001), sepsis at presentation (1·20 [1·04–1·40], p=0·016), higher American Society of Anesthesiologists (ASA) score at primary intervention (ASA 4–5 vs ASA 1–2, 1·82 [1·40–2·35], p<0·0001; ASA 3 vs ASA 1–2, 1·58, [1·30–1·92], p<0·0001]), surgical safety checklist not used (1·39 [1·02–1·90], p=0·035), and ventilation or parenteral nutrition unavailable when needed (ventilation 1·96, [1·41–2·71], p=0·0001; parenteral nutrition 1·35, [1·05–1·74], p=0·018). Administration of parenteral nutrition (0·61, [0·47–0·79], p=0·0002) and use of a peripherally inserted central catheter (0·65 [0·50–0·86], p=0·0024) or percutaneous central line (0·69 [0·48–1·00], p=0·049) were associated with lower mortality. Interpretation Unacceptable differences in mortality exist for gastrointestinal congenital anomalies between lowincome, middle-income, and high-income countries. Improving access to quality neonatal surgical care in LMICs will be vital to achieve Sustainable Development Goal 3.2 of ending preventable deaths in neonates and children younger than 5 years by 2030

Percepção de risco no uso de agrotóxicos em cinco comunidades rurais no município de Pombal, PB

The indiscriminate use of pesticides in Brazil has been growing over time and the consequences of food contamination, environmental pollution and health problems of rural workers. For the foregoing held lectures in five rural communities in the municipality of Pombal-PB, with goals to inform farmers about correct ways of applying pesticides, the health risks by excessive exposure of these substances and the proper way of disposal of empty containers of pesticides. The knowledge about the subject in question was conducted through a questionnaire administered before the lectures and found that 93% of respondents reported using pesticides on their crops, 83% responded that they use pesticides to more than five years, 40% reported agricultural pesticides applied once a week, was diagnosed also that 54% of respondents did not receive technical information on application and handling of pesticides, and 51% reported that they do not know the personal protective equipment (PPE) and 96% of respondents not properly designed empty containers of pesticides.O uso indiscriminado de agrotóxicos no Brasil vem crescendo ao longo do tempo e cujas consequências são a contaminação de alimentos, a poluição ambiental e problemas de saúde dos trabalhadores rurais. Diante do exposto realizou-se palestras em cinco comunidades rurais do município de Pombal-PB, com objetivos de informar aos agricultores as formas corretas de aplicação dos defensivos agrícolas, os riscos à saúde pela exposição excessiva dessas substâncias e a correta maneira da destinação final das embalagens vazias dos agrotóxicos. O conhecimento sobre o assunto em questão foi realizado por meio de um questionário aplicado antes das palestras e verificou-se que 93% dos entrevistados informaram que utilizam agrotóxicos em suas lavouras, 83% responderam que utilizam produtos fitossanitários a mais de cinco anos, 40% reportaram aplicar defensivos agrícolas uma vez por semana, diagnosticou-se também que 54% dos entrevistados não receberam informação técnica sobre aplicação e manejo de agrotóxico, bem como 51% relataram que não conhecem os equipamentos de proteção individual (EPI) e 96 % dos entrevistados não destinam corretamente as embalagens vazias dos agrotóxicos

A TRANSMISSÃO MARTENO-INFANTIL DO HIV:: UMA REVISÃO DE LITERATURA NA PERSPECTIVA DA ENFERMAGEM

O objetivo deste estudo é realizar uma revisão de literatura sobre osfatores envolvidos na transmissão vertical do HIV. Metodologia: Trata-se de umarevisão de literatura, realizada nos últimos dez anos (2001 a 2010), no qualrealizou-se uma consulta a periódicos presentes na Biblioteca Virtual de Saúde,Periódicos do Portal Capes e documentos oficiais do Ministério da Saúde. Foramencontrados 4.000 artigos no período mencionado, os mesmos foram selecionadospelo idioma português, e exclusão de artigos coincidentes relacionados ao tema.Resultados e Discussão: No Brasil, de 1980 a junho de 2007 foram notificados 474.273casos da doença. As possibilidades de controle dessa pandemia ainda parecemremotas, apesar do desenvolvimento de novas terapias e do esforço mundial nabusca de uma vacina eficaz contra a infecção. Cerca de 40% das transmissões doHIV da mãe para o bebê ocorrem durante a gestação e 65%, no parto. Há, ainda,um risco acrescido de transmissão por meio da amamentação natural, com índicesentre 7% e 22% por exposição durante a mamada. Conclusão: Sendo assim, osserviços de saúde devem estar preparados para esse atendimento e a equipemultiprofissional precisa estar envolvida na busca de solução para astransformações que a infecção pelo HIV trouxe para a assistência da gestante,do feto e da família

Reformulação da política estadual de custeio da assistência farmacêutica com foco nos serviços farmacêuticos

Introdução: O custeio da Assistência Farmacêutica (AF) ambulatorial no Brasil foi historicamente voltado para a disponibilidade de medicamentos. Recentemente, a Política Nacional de Atenção Básica(1) expõe a necessidade de execução de outros serviços farmacêuticos que não a mera disponibilidade deste insumo. Somado a isso, percebe-se a sistematização de instrumentos(2,3) que trazem elementos clínicos e de educação em saúde para o reposicionamento da AF nos serviços de saúde, destacando a integração com os serviços e população e corresponsabilização com o processo de cuidado ofertado nos diversos níveis de atenção. Neste sentido, o presente trabalho ilustra a reformulação da Política de Custeio da AF (PCAF) em um estado da federação. Objetivos: Descrever a estratégia de reformulação da PCAF enfatizando a promoção do acesso de qualidade a medicamentos, atuando de forma integrada às redes, com foco na implementação e qualificação dos serviços farmacêuticos. Material e Método: A PCAF foi elaborada adotando unidade de análise bottom-up, inserida em racional de coalizões de defesa (4). Iniciou-se coleta e diagnostico online de dados e anseios de profissionais executores da AF nos municípios. Após tabulação foi elaborada revisão bibliográfica e proposta inicial. Posteriormente, oficinas ascendentes e transversais no âmbito dos setores correlacionados à PCAF na estrutura da secretaria de saúde, baseadas em brainstorming e painéis de especialistas apreciaram a proposta inicial, até validação da alta gestão. Finalmente ocorreu pactuação na Comissão Intergestores Bipartite (CIB), com publicação de instrumento normativo (5). Resultados: A PCAF configura a primeira política de abrangência estadual no Brasil que remunera serviços clínico-farmacêuticos com base em produção e tabulação de indicadores em sistemas oficiais. Serviços de assistência farmacêutica (SAF) são indicados pelos municípios, com tetos de acordo com porte populacional e correção remuneratória com base no fator de alocação (6). Conceituou-se SAF como um ou mais equipamentos de saúde, que atuem em rede visando a execução de ações técnico-gerenciais, técnico-pedagógicas e/ou clínico-assistenciais, sob supervisão de profissional farmacêutico devidamente habilitado nos termos legais. Mesmo com recente publicação, no comparativo entre julho de 2022 e março de 2023, há incremento de 230,77% na quantidade de municípios com produção autorizada no sistema ambulatorial (SIA-SUS) e 156,52% de aumento no volume de procedimentos faturados e autorizados no sistema da atenção básica (SISAB). Discussão e Conclusões: A PCAF sustenta-se na premissa de que a AF deve ser entendida para além do seu aparato logístico na provisão de medicamentos, inserindo-se na rede como ponto de convergência do cuidado. Ao se condicionar o repasse financeiro à execução de serviços, espera-se maior integração entre os farmacêuticos e outras equipes, criando ambiente que propicia melhor cuidado em saúde para os usuários

Biochemical studies with DNA polymerase beta and DNA polymerase beta-PAK of Trypanosoma cruzi suggest the involvement of these proteins in mitochondrial DNA maintenance

Mammalian DNA polymerase p is a nuclear enzyme involved in the base excision and single-stranded DNA break repair pathways. in trypanosomatids, this protein does not have a defined cellular localization, and its function is poorly understood. We characterized two Trypanosoma cruzi proteins homologous to mammalian DNA polymerase beta, TcPol beta and TcPol beta PAK, and showed that both enzymes localize to the parasite kinetoplast. in vitro assays with purified proteins showed that they have DNA polymerization and deoxyribose phosphate lyase activities. Optimal conditions for polymerization were different for each protein with respect to dNTP concentration and temperature, and TcPol beta PAK, in comparison to TcPol beta, conducted DNA synthesis over a much broader pH range. TcPol beta was unable to carry out mismatch extension or DNA synthesis across 8-oxodG lesions, and was able to discriminate between dNTP and ddNTP. These specific abilities of TcPol beta were not observed for TcPol beta PAK or other X family members, and are not due to a phenylalanine residue at position 395 in the C-terminal region of TcPol beta, as assessed by a site-directed mutagenesis experiment reversing this residue to a well conserved tyrosine. Our data suggest that both polymerases from T. cruzi could cooperate to maintain mitochondrial DNA integrity through their multiple roles in base excision repair, gap filling and translesion synthesis. (C) 2008 Elsevier B.V. All rights reserved.PRONEXFundação de Amparo à Pesquisa do Estado de Minas Gerais (FAPEMIG)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Univ Fed Minas Gerais, ICB, Dept Bioquim & Imunol, Belo Horizonte, MG, BrazilUniversidade Federal de São Paulo, Dept Microbiol Imunol & Parasitol, São Paulo, BrazilCNRS, Inst Pharmacol & Biol Struct, Grp Genet Instabil & Canc, Toulouse, FranceUniversidade Federal de São Paulo, Dept Microbiol Imunol & Parasitol, São Paulo, BrazilWeb of Scienc

Amino acid substitutions in the DNA-binding domain of the human androgen receptor are a frequei.t cause of receptor binding positive androgen resistance

Snake venom metalloproteinases (SVMPs) belonging to P-I class are able to hydrolyze extracellular matrix proteins and coagulation factors triggering local and systemic reactions by multiple molecular mechanisms that are not fully understood. BmooMP alpha-I, a P-I class SMVP from Bothrops moojeni venom, was active upon neuro- and vaso-active peptides including angiotensin I, bradykinin, neurotensin, oxytocin and substance P. Interestingly, BmooMPa-I showed a strong bias towards hydrolysis after proline residues, which is unusual for most of characterized peptidases. Moreover, the enzyme showed kininogenase activity similar to that observed in plasma and cells by kallikrein. FRET peptide assays indicated a relative promiscuity at its S-2-S '(2) subsites, with proline determining the scissile bond. This unusual post-proline cleaving activity was confirmed by the efficient hydrolysis of the synthetic combinatorial library MCA-GXXPXXQ-EDDnp, described as resistant for canonical peptidases, only after Pro residues. Structural analysis of the tripeptide LPL complexed with BmooMP alpha-I, generated by molecular dynamics simulations, assisted in defining the subsites and provided the structural basis for subsite preferences such as the restriction of basic residues at the S-2 subsite due to repulsive electrostatic effects and the steric impediment for large aliphatic or aromatic side chains at the Si subsite. These new functional and structural findings provided a further understanding of the molecular mechanisms governing the physiological effects of this important class of enzymes in envenomation process. (c) 2014 Elsevier B.V. All rights reserved