Differential diagnosis of hereditary and acquired thrombocytopenia by the immature platelet fraction (IPF) and thrombopoietin plasmatic levels (TPO)

Orientador: Erich Vinícius de PaulaDissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Ciências MédicasResumo: Introdução: Nos últimos anos, novas gerações de analisadores de hematologia expandiram o arsenal de parâmetros hematológicos para a avaliação de pacientes com trombocitopenia. Entre estes, a determinação da fração de contagem de plaquetas imaturas (IPF) figura entre os marcadores mais promissores por relacionar-se com a atividade trombopoiética. De uma forma análoga à contagem de reticulócitos para série vermelha, a IPF aumenta em condições em que há maior atividade trombopoiética, constituindo-se em um potencial auxiliar no diagnóstico diferencial nas diferentes causas de trombocitopenia. Entre estas, o diagnóstico diferencial entre a plaquetopenia imune (PTI) e as trombocitopenias hereditárias (TH) pode ser desafiador, particularmente no grupo de pacientes em que a apresentação clínica não aponte claramente para um destes dois grupos. Além disso, embora o comportamento da IPF tenha sido demonstrando em diferentes etiologias de trombocitopenia, a avaliação da reprodutibilidade deste parâmetro, aspecto essencial para seu uso clínico, é ainda limitada. Metodologia: Neste estudo, avaliamos o IPF, o VPM (volume médio de plaquetas) e a TPO em um coorte de pacientes com trombocitopenia pós-quimioterapia (n= 56), insuficiências medulares (mielodisplasia e anemia aplástica; n = 22), PTI (n = 105) e TH (n = 27). A determinação da IPF foi realizada no analisador hematológico Sysmex XE5000 e, em um subgrupo de pacientes com TH, testes de reprodutibilidade intra e inter-ensaios foram realizados. Para refinamento da análise da atividade trombopoiética de pacientes com PTI e TH, níveis plasmáticos de trombopoietina (TPO) foram dosados por ELISA. Resultados e Conclusões: A contagem da IPF mostrou o comportamento esperado em pacientes com trombocitopenia pós-quimioterapia e insuficiência medular, com valores significativamente menores do que na PTI. Ao contrário do esperado, valores significativamente maiores de IPF foram observados em pacientes com TH, conferindo à IPF uma acurácia diagnóstica significativa para o diagnóstico diferencial entre PTI e TH. Considerando um valor de corte de 17,45%, a IPF foi capaz de diferenciar estas duas condições com uma sensibilidade de 77,78% e uma especificidade de 71,43%. Testes de reprodutibilidade confirmaram que a contagem de IPF é um parâmetro robusto, mesmo no subgrupo com contagens elevadas e diagnóstico de TH. Durante a realização do estudo, resultados de um outro grupo japonês também observaram valores elevados para IPF em uma coorte inferior de pacientes com TH. Em conclusão, a IPF é um parâmetro atrativo para auxiliar no diagnóstico diferencial entre TH e PTIAbstract: Introduction: In recent years, new generations of hematology analyzers expanded the arsenal of hematological parameters for the evaluation of patients with thrombocytopenia. Among these, the determination of the immature platelet fraction (IPF) count figure among the most promising markers for relate with the thrombopoietic activity. In an analogous manner to the reticulocyte count for red series, the IPF increases in conditions where there is greater thrombopoietic activity, thus becoming a potential aid in the differential diagnosis in different causes of thrombocytopenia. Among these, the differential diagnosis of immune thrombocytopenia (ITP) and hereditary thrombocytopenia (HT) can be challenging, particularly in patients where the clinical presentation does not point clearly to one of this two groups. Furthermore, although the IPF behavior has been demonstrated in different etiologies of thrombocytopenia, the evaluation of the reproducibility of this parameter, essential aspect for it clinical use, is still limited. Methodology: In this study, we evaluated the IPF, the MPV (mean platelet volume) and TPO in a cohort of patients with post-chemotherapy thrombocytopenia (n = 56), medullary insufficiencies (myelodysplasia and aplastic anemia; n = 22), ITP (n = 105) and HT (n = 27). The IPF determination was realized in Sysmex XE5000 hematology analyzer and a subgroup of patients with HT, intra- and inter-testing assays were performed. For refinement analysis of thrombopoietin activity in patients with ITP and HT, plasma levels of thrombopoietin (TPO) were measured by ELISA. Results and Conclusions: The IPF count showed a expected behavior in patients with post-chemotherapy thrombocytopenia and bone marrow failure, with significantly lower values than ITP. Contrary to expectations, significantly higher IPF values were observed in patients with HT, giving to IPF a significant diagnostic accuracy for differential diagnosis between PTI and TH. Considering a cutoff value of 17.45%, the IPF was able to differentiate these two conditions with a sensitivity of 77.78% and a specificity of 71.43%. Reproducibility tests confirmed that the IPF count is a robust parameter even in the subgroup with elevated scores and HT diagnostic. During this study, results of another Japanese group also noted high IPF values in a smaller cohort of patients with HT. In conclusion, the IPF is an attractive parameter to aid in the differential diagnosis between TH and PTIMestradoClinica MedicaMestre em Ciências2013/09319-0CAPE

Evaluation and selection of candidates for liver transplantation : an economic perspective

Background – Over the next 20 years, the number of patients on the waiting list for liver transplantation (LTx) is expected to increase by 23%, while pre-LTx costs should raise by 83%. Objective – To evaluate direct medical costs of the pre-LTx period from the perspective of a tertiary care center. Methods – The study included 104 adult patients wait-listed for deceased donor LTx between October 2012 and May 2016 whose treatment was fully provided at the study transplant center. Clinical and economic data were obtained from electronic medical records and from a hospital management software. Outcomes of interest and costs of patients on the waiting list were compared through the Kruskal-Wallis test. A generalized linear model with logit link function was used for multivariate analysis. P-values <0.05 were considered statistically significant. Results – The costs of patients who underwent LTx ($8,879.83; 95$6,464.73; 95% CI 3,845.75–10,867.28; P=0.04) were higher than those of patients who were excluded from the list for any reason except death ($4,647.78; 95% CI 2,469.35–8,748.04; P=0.254) or those who remained on the waiting list at the end of follow-up. Conclusion – Although protocols of inclusion on the waiting list vary among transplant centers, similar approaches exist and common problems should be addressed. The results of this study may help centers with similar socioeconomic realities adjust their transplant policies

AGRESSIVIDADE INFANTIL NO CONTEXTO ESCOLAR: AS POSSIBILIDADES DE AUXÍLIO PROPORCIONADAS PELA LUDICIDADE

O presente artigo trata de um projeto de extensão e pesquisa, realizado em uma escola de Ensino Fundamental da rede pública do município de Presidente Prudente-SP. O referido projeto compreende que conflitos internos e dificuldades emocionais na infância podem ser responsáveis por dificuldades de relacionamento em sala de aula. Nessa perspectiva, se propõe a investigar a possibilidade de se contribuir para a resolução desses conflitos, obtendo‐se uma consequente melhoria no comportamento dos alunos, por meio de atividades lúdicas. Ao participar do referido projeto, deparou‐se com o caso de um garoto considerado agressivo por sua professora e buscou‐se investigar se o brincar e o suporte emocional oferecido ao aluno teria potencial para minimizar sua conduta agressiva. A pesquisa foi realizada com base nos pressupostos da pesquisa qualitativa, tendo como referencial a investigação das manifestações da criança, pensadas a partir das contribuições da teoria psicanalítica winnicottiana

A AGRESSIVIDADE COMO ELEMENTO INTRÍSECO DO DESENVOLVIMENTO EMOCIONAL NA TEORIA WINNICOTTIANA

Este artigo versa sobre a concepção de Winnicott sobre a agressividade infantil, abordando suas raízes e apontando o caráter das manifestações agressivas à medida em que o indivíduo se desenvolve, bem como o modo como o fator destrutivo relaciona-se com o processo de amadurecimento. Para atingir os objetivos propostos, foram realizados, nesta pesquisa de natureza bibliográfica, levantamento e análise de obras de Winnicott. O estudo mostrou que a agressividade, conforme os pressupostos da teoria winnicottiana, não se restringe a uma reação à frustração, sendo entendida como elemento necessário para o transcorrer do processo evolutivo que, quando ocorre de modo saudável, culmina no desenvolvimento de características e capacidades indispensáveis para o bem-estar psíquico e para os relacionamentos interpessoais

Head‐to‐Head comparison of consensus‐recommended platelet function tests to assess P2Y12 Inhibition : insights for multi‐center trials

The vasodilator‐associated stimulated phosphoprotein (VASP) phosphorylation level is a highly specific method to assess P2Y12 receptor inhibition. Traditionally, VASP phosphorylation is analyzed by flow cytometry, which is laborious and restricted to specialized laboratories. Recently, a simple ELISA kit has been commercialized. The primary objective of this study was to compare the performance of VASP assessment by ELISA and flow cytometry in relation to functional platelet aggregation testing by Multiplate® whole‐blood aggregometry. Blood from 24 healthy volunteers was incubated with increasing concentration of a P2Y12 receptor inhibitor (AR‐C 66096). Platelet function testing was carried out simultaneously by Multiplate® aggregometry and by VASP assessment through ELISA and flow cytometry. As expected, increasing concentrations of the P2Y12 receptor inhibitor induced a proportional inhibition of platelet aggregation and P2Y12 receptor activation across the modalities. Platelet reactivity index values of both ELISA‐ and flow cytometry‐ based VASP assessment methods correlated strongly (r = 0.87, p < 0.0001) and showed minimal bias (1.05%). Correlation with Multiplate® was slightly higher for the flow cytometry‐based VASP assay (r = 0.79, p < 0.0001) than for the ELISA‐based assay (r = 0.69, p < 0.0001). Intraclass correlation (ICC) was moderate for all the assays tested (ICC between 0.62 and 0.84). However, categorization into low, optimal, or high platelet reactivity based on these assays was strongly concordant (κ between 0.86 and 0.92). In conclusion, the consensus‐recommended assays with their standardized cut‐offs should not be used interchangeably in multi‐center clinical studies but, rather, they should be standardized throughout sites

Mérito Sob Custódia: Os Limites da Menção aos Fatos da Prisão Durante as Audiências de Custódia

A Resolução do CNJ nº 213/ 2015 não permite que se entre no mérito dos fatos que geraram a prisão em flagrante nas audiências de custódia. Contudo, pesquisas de campo têm revelado que juízes/as compreendem de forma heterogênea a entrada do mérito em algumas situações. Para compreender como os atores jurídicos mobilizam o mérito nessas audiências, propomos realizar uma reflexão sobre essa temática, considerando, sobretudo, ser um tema ainda pouco aprofundado no campo do debate acadêmico. Para isso, realizamos uma revisão da doutrina que aborda a temática do mérito no processo penal, bem como a revisão de pesquisas sobre audiência de custódia que descrevem a menção ao mérito em seus estudos empíricos. Analisamos também entrevistas realizadas com juízes/as que mencionaram seus critérios de entrada ou não no mérito dos casos avaliados em audiência de custódia. Acreditamos que este artigo poderá contribuir com um debate ainda pouco aprofundado sobre a temática do mérito nas audiências de custódia. Apesar de aparecer na Resolução CNJ nº 213/2015 e nas pesquisas sobre audiência de custódia, ainda não há uma discussão mais aprofundada desse tema

Biochemical Characterization of an In-House Coccidioides Antigen: Perspectives for the Immunodiagnosis of Coccidioidomycosis

The objective of this study was to evaluate the reactivity of an in-house antigen, extracted from a strain of C. posadasii isolated in northeastern Brazil, by radial immunodiffusion and Western blotting, as well as to establish its biochemical characterization. the protein antigen was initially extracted with the use of solid ammonium sulfate and characterized by 1-D electrophoresis. Subsequently, it was tested by means of double radial immunodiffusion and Western blotting. A positive reaction was observed against the antigen by both immunodiagnostic techniques tested on sera from patients suffering from coccidioidomycosis. Besides this, two immunoreactive protein bands were observed and were revealed to be a beta-glucosidase and a glutamine synthetase after sequencing of the respective N-terminal regions. Our in-house Coccidioides antigen can be promising as a quick and low-cost diagnostic tool without the risk of direct manipulation of the microorganism.Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Coordination for the Improvement of Higher Education PersonnelUniv Fed Ceara, Specialized Med Mycol Ctr, Sch Med, BR-60430270 Fortaleza, Ceara, BrazilUniv Estadual Ceara, Postgrad Program Vet Sci, Sch Vet Med, BR-60740000 Fortaleza, Ceara, BrazilUniv Fed Ceara, Dept Biochem & Mol Biol, BR-60455760 Fortaleza, Ceara, BrazilUniversidade Federal de São Paulo, Dept Microbiol Immunol & Parasitol, BR-04021001 São Paulo, BrazilUniversidade Federal de São Paulo, Dept Microbiol Immunol & Parasitol, BR-04021001 São Paulo, BrazilCNPq: 302574/2009-3CNPq: 306637/2010-3Coordination for the Improvement of Higher Education Personnel: CAPES/PNPD2103/2009Web of Scienc

Schistosomal Lipids Activate Human Eosinophils via Toll-Like Receptor 2 and PGD2 Receptors: 15-LO Role in Cytokine Secretion

Parasite-derived lipids may play important roles in host-pathogen interactions and immune evasion mechanisms. Remarkable accumulation of eosinophils is a characteristic feature of inflammation associated with parasitic disease, especially caused by helminthes. Infiltrating eosinophils are implicated in the pathogenesis of helminth infection by virtue of their capacity to release an array of tissue-damaging and immunoregulatory mediators. However, the mechanisms involved in the activation of human eosinophils by parasite-derived molecules are not clear. Here we investigated the effects and mechanisms of schistosomal lipids-induced activation of human eosinophils. Our results showed that stimulation of human eosinophils in vitro with total lipid extracts from adult worms of S. mansoni induced direct activation of human eosinophils, eliciting lipid droplet biogenesis, synthesis of leukotriene (LT) C4 and eoxin (EX) C4 (14,15 LTC4) and secretion of eosinophil pre-formed TGFβ. We demonstrated that main eosinophil activating components within S. mansoni lipid extract are schistosomal-derived lysophosphatidylcholine (LPC) and prostaglandin (PG)D2. Moreover, TLR2 is up-regulated in human eosinophils upon stimulation with schistosomal lipids and pre-treatment with anti-TLR2 inhibited both schistosomal lipids- and LPC-, but not PGD2-, induced lipid droplet biogenesis and EXC4 synthesis within eosinophils, indicating that TLR2 mediates LPC-driven human eosinophil activation. By employing PGD2 receptor antagonists, we demonstrated that DP1 receptors are also involved in various parameters of human eosinophil activation induced by schistosomal lipids, but not by schistosomal LPC. In addition, schistosomal lipids and their active components PGD2 and LPC, triggered 15-LO dependent production of EXC4 and secretion of TGFβ. Taken together, our results showed that schistosomal lipids contain at least two components—LPC and PGD2—that are capable of direct activation of human eosinophils acting on distinct eosinophil-expressed receptors, noticeably TLR2 as well as DP1, trigger human eosinophil activation characterized by production/secretion of pro-inflammatory and immunoregulatory mediators