Browsing by Author "Lima, Margarida"
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- Doenças linfoproliferativas crónicas T e NKPublication . Lima, MargaridaAs doenças linfoproliferativas crónicas (DLPC) de linfócitos T e de células NK são doenças raras com características clínico-biológicas ainda pouco definidas. Para além da sua raridade, os maiores obstáculos à sua identificação e caracterização têm sido a diversidade e complexidade fenotípica dos linfócitos T e das células NK, associados à ausência de critérios fenotípicos que permitam estabelecer o diagnóstico de suspeita de (mono) clonalidade e ao desconhecimento sobre a existência e incidência de fenótipos verdadeiramente aberrantes associados a estas DLPC. Este trabalho teve como objectivo aprofundar o conhecimento das DLPC de linfócitos T e de células NK maturas, utilizando uma abordagem baseada na avaliação da utilidade dos estudos imunofenotípicos para confirmação de suspeita de (mono) clonalidade, para a identificação e caracterização de células T e NK neoplásicas e para a compreensão das manifestações clínicobiológicas e da etiopatogenia destas doenças. Com o intuito de estabelecer as bases para definir os critérios fenotípicos de (mono) clonalidade e identificação de linfócitos T e de células NK neoplásicas, efectuamos o estudo imunofenotípico detalhado dos linfócitos T e das células NK do sangue periférico de indivíduos adultos saudáveis e de indivíduos adultos com diferentes condições patológicas associadas à estimulação aguda ou crónica destas células e comparamos o perfil imunofenotípico e o repertório de famílias de regiões variáveis da cadeia b do receptor da célula T de expansões monoclonais de linfócitos T e de células NK com os de expansões policlonais e oligoclonais das mesmas células. Para ultrapassar os obstáculos inerentes à raridade destas DLP, estabelecemos um protocolo multicêntrico centralizado no Hospital Geral de Santo António (Porto) e no Hospital Universitario de Salamanca (Salamanca) e orientamos a nossa atenção para a identificação e caracterização de fenótipos aberrantes associados às neoplasias de células T e NK. Das DLPC estudadas, escolhemos como modelos de estudo três grupos: as linfocitoses crónicas de linfócitos grandes granulares T CD4+ e as linfocitoses crónicas de linfócitos grandes granulares NK, cujas características clínico-biológicas eram, até hoje, praticamente desconhecidas e o Síndrome de Sezary, uma forma particular de linfoma T cutâneo que, embora já bem definido do ponto de vista clínico e histopatológico, estava ainda mal caracterizado do ponto de vista fenotípico. Nestes três tipos de DLPC, fizemos um estudo detalhado do perfil fenotípico e funcional da célula linfóide neoplásica,caracterizamos e interpretamos as manifestações clínicas da doença e avançamos com algumas hipóteses relativamente à sua etiopatogenia.
- Estudo da ativação dos basófilos em doentes com mastocitosePublication . Spínola, Ana; Guimarães, Joana; Santos, Marlene; Lima, Margarida
- Human Peripheral Blood Gamma Delta T Cells: Report on a Series of Healthy Caucasian Portuguese Adults and Comprehensive Review of the LiteraturePublication . Fonseca, Sónia; Pereira, Vanessa; Lau, Catarina; Teixeira, Maria dos Anjos; Bini-Antunes, Marika; Lima, MargaridaGamma delta T cells (Tc) are divided according to the type of Vδ and Vγ chains they express, with two major γδ Tc subsets being recognized in humans: Vδ2Vγ9 and Vδ1. Despite many studies in pathological conditions, only a few have quantified the γδ Tc subsets in healthy adults, and a comprehensive review of the factors influencing its representation in the blood is missing. Here we quantified the total γδ Tc and the Vδ2/Vγ9 and Vδ1 Tc subsets in the blood from 30 healthy, Caucasian, Portuguese adults, we characterized their immunophenotype by 8-color flow cytometry, focusing in a few relevant Tc markers (CD3/TCR-γδ, CD5, CD8), and costimulatory (CD28), cytotoxic (CD16) and adhesion (CD56) molecules, and we examined the impacts of age and gender. Additionally, we reviewed the literature on the influences of race/ethnicity, age, gender, special periods of life, past infections, diet, medications and concomitant diseases on γδ Tc and their subsets. Given the multitude of factors influencing the γδ Tc repertoire and immunophenotype and the high variation observed, caution should be taken in interpreting "abnormal" γδ Tc values and repertoire deviations, and the clinical significance of small populations of "phenotypically abnormal" γδ Tc in the blood.
- IL-31 and IL-8 in Cutaneous T-Cell Lymphoma: Looking for Their Role in ItchPublication . Abreu, Maria; Miranda, Marta; Castro, M; Fernandes, Iolanda; Cabral, Renata; Santos, Ana Helena; Fonseca, Sonia; Rodrigues, João; Leander, Magdalena; Lau, Catarina; Freitas, Inês; Coimbra, Susana; Santos-Silva, Alice; Lima, MargaridaThe itch associated with cutaneous T-cell lymphoma (CTCL), including Mycosis Fungoides (MF) and Sézary syndrome (SS), is often severe and poorly responsive to treatment with antihistamines. Recent studies have highlighted the possible role of interleukins in nonhistaminergic itch. We investigated the role of IL-31 and IL-8 in CTCL, concerning disease severity and associated itch. Serum samples of 27 patients with CTCL (17 MF and 10 SS) and 29 controls (blood donors) were analyzed for interleukin- (IL-) 31 and IL-8; correlations with disease and itch severity were evaluated. IL-31 serum levels were higher in CTCL patients than in controls and higher in SS than in MF. Also, serum IL-31 levels were higher in patients with advanced disease compared to those with early disease, and they correlated positively with lactate dehydrogenase and beta 2-microglobulin levels, as well as with the Sézary cell count. Itch affected 67% of CTCL patients (MF: 47%; SS: 100%). Serum IL-31 levels were higher in itching patients than in controls and in patients without itching. There was no association between serum IL-8 and disease severity, nor with itching. Serum IL-8 levels correlated positively with peripheral blood leukocyte and neutrophil counts in CTCL patients. Our study suggests a role for IL-31 in CTCL-associated itch, especially in advanced disease and SS, offering a rational target for new therapeutic approaches. Increased serum IL-8 observed in some patients may be related to concomitant infections, and its role in exacerbating itch by recruiting neutrophils and promoting the release of neutrophil proteases deserves further investigation
- Isolamento mecânico das células endoteliais e sua caraterização por citometria de fluxoPublication . Torres, Cláudia; Machado, Rui; Lima, Margarida
- Laboratory studies for paroxysmal nocturnal hemoglobinuria, with emphasis on flow cytometryPublication . Lima, MargaridaParoxysmal nocturnal hemoglobinuria (PNH) is a rare acquired clonal hematopoietic stem cell disorder caused by somatic mutations in the PIG-A gene, leading to the production of blood cells with absent or decreased expression of glycosylphosphatidylinositol-anchored proteins, including CD55 and CD59. Clinically, PNH is classified into three variants: classic (hemolytic), in the setting of another specified bone marrow disorder (such as aplastic anemia or myelodysplastic syndrome) and subclinical (asymptomatic). PNH testing is recommended for patients with intravascular hemolysis, acquired bone marrow failure syndromes and thrombosis with unusual features. Despite the availability of consensus guidelines for PNH diagnosis and monitoring, there are still discrepancies on how PNH tests are carried out, and these technical variations may lead to an incorrect diagnosis. Herein, we provide a brief historical overview of PNH, focusing on the laboratory tests available and on the current recommendations for PNH diagnosis and monitoring based in flow cytometry.
- Manifestações Cutâneas nas Mastocitoses: AtualizaçãoPublication . Ferreira, Sandra; Fernandes, Iolanda; Cabral, Renata; Machado, Susana; Lima, Margarida; Selores, ManuelaIntrodução: As mastocitoses caraterizam-se pela expansão clonal de mastócitos, com acumulação de mastócitos morfológica e imunofenotipicamente anormais em diferentes órgãos. A pele é o órgão mais frequentemente envolvido. Virtualmente, todas as crianças e mais de 80% dos adultos com mastocitose apresentam lesões cutâneas. Material e Métodos: O presente artigo descreve os sinais e sintomas associados à mastocitose na pele, tendo por base a revisão das normas de orientação de consenso internacionais, recentemente publicadas. Discussão: De acordo com a classificação proposta pela Organização Mundial de Saúde em 2016, a mastocitose divide-se em mastocitose cutânea, mastocitose sistémica e sarcoma de mastócitos. A mastocitose cutânea pode subdividir-se em três subtipos: a mastocitose cutânea maculopapular (também denominada urticária pigmentosa), mastocitose cutânea difusa e mastocitoma cutâneo. A telangiectasia macular eruptiva perstans já não é considerada uma entidade independente. Conclusão: As manifestações cutâneas da mastocitose são variáveis, dependendo da idade de início da doença. Recentemente a classificação da mastocitose cutânea foi atualizada. Nas crianças, a mastocitose ocorre como mastocitose cutânea que tende à regressão espontânea durante a adolescência. Quando tem início na idade adulta, a mastocitose é geralmente sistémica, sendo a forma mais frequente a mastocitose sistémica indolente, que normalmente também cursa com manifestações cutâneas e tem um curso crónico.
- Metabolic control of T cell immune response through glycans in inflammatory bowel diseasePublication . Dias, A.; Correia, A.; Pereira, M.; Almeida, C.; Alves, I.; Pinto, V.; Catarino, T.; Mendes, N.; Leander, M.; Oliva-Teles, M.; Maia, L.; Delerue-Matos, C.; Taniguchi, N.i; Lima, Margarida; Pedroto, I.; Marcos-Pinto, Ricardo; Lago, P.; Reis, C.; Vilanova, M.; Pinho, S.Mucosal T lymphocytes from patients with ulcerative colitis (UC) were previously shown to display a deficiency in branched N-glycosylation associated with disease severity. However, whether this glycosylation pathway shapes the course of the T cell response constituting a targeted-specific mechanism in UC remains largely unknown. In this study, we demonstrated that metabolic supplementation of ex vivo mucosal T cells from patients with active UC with N-acetylglucosamine (GlcNAc) resulted in enhancement of branched N-glycosylation in the T cell receptor (TCR), leading to suppression of T cell growth, inhibition of the T helper 1 (Th1)/Th17 immune response, and controlled T cell activity. We further demonstrated that mouse models displaying a deficiency in the branched N-glycosylation pathway (MGAT5-/-, MGAT5+/-) exhibited increased susceptibility to severe forms of colitis and early-onset disease. Importantly, the treatment of these mice with GlcNAc reduced disease severity and suppressed disease progression due to a controlled T cell-mediated immune response at the intestinal mucosa. In conclusion, our human ex vivo and preclinical results demonstrate the targeted-specific immunomodulatory properties of this simple glycan, proposing a therapeutic approach for patients with UC.
- N-glicosilação do recetor das células t: um novo mecanismo molecular na patogénese da doença inflamatória intestinalPublication . Dias, Ana; Catarino, Telmo; Lago, Paula; Marcos-Pinto, Ricardo; Salgueiro, Paulo; Almeida, Catarina; Fonseca, Sónia; Lima, Margarida; Vilanova, Manuel; Dinis-Ribeiro, Mário; Reis, Celso; Pinho, Salomé
- αIIbβ3 variants in ten families with autosomal dominant macrothrombocytopenia: Expanding the mutational and clinical spectrumPublication . Morais, Sara; Oliveira, Jorge; Lau, Catarina; Pereira, Mónica; Gonçalves, Marta; Monteiro, Catarina; Gonçalves, Ana; Matos, Rui; Sampaio, Marco; Cruz, Eugénia; Freitas, Inês; Santos, Rosário; Lima, MargaridaBackground: Rare pathogenic variants in either the ITGA2B or ITGB3 genes have been linked to autosomal dominant macrothrombocytopenia associated with abnormal platelet production and function, deserving the designation of Glanzmann Thrombasthenia-Like Syndrome (GTLS) or ITGA2B/ITGB3-related thrombocytopenia. Objectives: To describe a series of patients with familial macrothrombocytopenia and decreased expression of αIIbβ3 integrin due to defects in the ITGA2B or ITGB3 genes. Methods: We reviewed the clinical and laboratory records of 10 Portuguese families with GTLS (33 patients and 11 unaffected relatives), including the functional and genetic defects. Results: Patients had absent to moderate bleeding, macrothrombocytopenia, low αIIbβ3 expression, impaired platelet aggregation/ATP release to physiological agonists and low expression of activation-induced binding sites on αIIbβ3 (PAC-1) and receptor-induced binding sites on its ligand (bound fibrinogen), upon stimulation with TRAP-6 and ADP. Evidence for constitutive αIIbβ3 activation, occurred in 2 out of 9 patients from 8 families studied, but also in 2 out of 12 healthy controls. We identified 7 missense variants: 3 in ITGA2B (5 families), and 4 in ITGB3 (5 families). Three variants (αIIb: p.Arg1026Trp and p.Arg1026Gln and β3: p.Asp749His) were previously reported. The remaining (αIIb: p.Gly1007Val and β3: p.Thr746Pro, p.His748Pro and p.Arg760Cys) are new, expanding the αIIbβ3 defects associated with GTLS. The integration of the clinical and laboratory data allowed the identification of two GTLS subgroups, with distinct disease severity. Conclusions: Previously reported ITGA2B and ITGB3 variants related to thrombocytopenia were clustered in a confined region of the membrane-proximal cytoplasmic domains, the inner membrane clasp. For the first time, variants are reported at the outer membrane clasp, at the transmembrane domain of αIIb, and at the membrane distal cytoplasmic domains of β3. This is the largest single-center series of inherited macrothrombocytopenia associated with αIIbβ3 variants published to date.