RESUMO
BACKGROUND: Animal models have shown that CD47-deficient mice develop severe autoimmune hemolytic anemia (AIHA) because the binding of red blood cell (RBC) CD47 to signal-regulatory protein (SIRP-alpha) on macrophages contributes to the inhibition of phagocytosis. In contrast, complement-inhibitory proteins such as CD35, CD55, and CD59 may protect RBCs against the lysis by complement. STUDY DESIGN AND METHODS: With the use of flow cytometric analyses, the expression of CD47, CD35, CD55, and CD59 on RBCs and of SIRP-alpha,beta on peripheral monocytes of 36 patients with warm AIHA (wAIHA; 23 with active wAIHA, 13 with wAIHA in remission) and 20 healthy subjects was evaluated. RESULTS: The mean fluorescence intensities (MFIs) of the expression of CD47, CD35, CD55, and SIRP-alpha,beta of active wAIHA patients, wAIHA in remission, and healthy subjects were not statistically different. Patients with active wAIHA showed significantly lower CD59 expression on RBCs than healthy individuals (MFI, 512.5 +/- 59.6 vs. 553.7 +/- 36.6; p = 0.009), while the CD59 expression in patients with wAIHA in remission was not significantly different from that of healthy controls (MFI, 538.4 +/- 48.3 vs. 553.7 +/- 36.6; p > 0.05). The expression of CD59 on RBCs of 3 patients who died from the wAIHA was lower than that seen on RBCs of healthy controls (MFI, 433.6 +/- 69.6 vs. 553.74 +/- 36.6; p = 0.0001). CONCLUSIONS: Our data show that the expression of CD47 on RBCs and SIRP-alpha,beta on monocytes of patients with wAIHA is not different from that seen in healthy individuals. In addition, we detected that patients with active wAIHA present low expression of CD59 and normal expression of CD35 and CD55 on their RBCs. Complement-regulatory proteins may play an important role in protecting RBC destruction through the activation of complement.
Assuntos
Anemia Hemolítica Autoimune/metabolismo , Antígenos de Diferenciação/biossíntese , Antígeno CD47/biossíntese , Antígenos CD55/biossíntese , Antígenos CD59/biossíntese , Eritrócitos/metabolismo , Monócitos/metabolismo , Receptores de Complemento 3b/biossíntese , Receptores Imunológicos/biossíntese , Adulto , Idoso , Anemia Hemolítica Autoimune/genética , Animais , Antígenos de Diferenciação/genética , Antígeno CD47/genética , Antígenos CD55/genética , Antígenos CD59/genética , Modelos Animais de Doenças , Feminino , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Macrófagos/metabolismo , Masculino , Camundongos , Camundongos Knockout , Pessoa de Meia-Idade , Fagocitose/genética , Receptores de Complemento 3b/genética , Receptores Imunológicos/genéticaRESUMO
BACKGROUND: The clonal deletion seen in recipients of allogeneic blood transfusion (ABT) refers to the removal of lymphocytes that promote the clearance of transfused alloantigens. Interactions between Fas (CD95) and FasL (CD95L) are involved in the clonal deletion of T cells and in the down regulation of the cytotoxic T-cell activity. STUDY DESIGN AND METHODS: The expression of CD95/95 L on spleen T cells of C57Bl/6 mice infused with unmodified ABT, prestorage leukoreduced ABT (LR-ABT), or saline was investigated by flow cytometry. The numbers of apoptotic spleen cells were evaluated after transfusion using the acridine orange and ethidium bromide uptake technique. RESULTS: Compared with untransfused animals, mice transfused with ABT showed higher expression of CD95 (MFI = 94.4 +/- 8.6 vs. 73.1 +/- 7.9, p = 0.02) and CD95L (23.5 +/- 6.9 vs. 8.1 +/- 2.0, p = 0.008) on CD4+ spleen cells. Expression of CD95 (92.2 +/- 7.5 vs. 64.9 +/- 7.5, p = 0.007) and CD95L (17.7 +/- 3.6 vs. 8.2 +/- 2.2, p = 0.02) was also increased on CD8+ cells of these animals. CD8+ spleen cells from mice transfused with ABT showed higher expression of CD95 (92.2 +/- 7.5 vs. 76.9 +/- 4.0, p = 0.03) and CD95L (17.7 +/- 3.6 vs. 8.3 +/- 1.5, p = 0.03) than cells from mice transfused with LR-ABT. The number of apoptotic spleen cells from mice transfused with ABT was greater than that from mice infused with LR-ABT (10.9 +/- 1.3 vs. 6.6 +/- 1.8, p = 0.01) or saline (10.9 +/- 1.3 vs. 6.5 +/- 0.7, p = 0.001). CONCLUSIONS: The data suggest that ABT up-regulates the expression of Fas/FasL on spleen T cells of mice and may promote their apoptosis. These ABT-associated immunologic alterations can be partially prevented by the leukoreduction of the transfused blood.
Assuntos
Transfusão de Sangue , Glicoproteínas de Membrana/metabolismo , Linfócitos T/citologia , Linfócitos T/metabolismo , Receptor fas/metabolismo , Animais , Apoptose/imunologia , Proteína Ligante Fas , Citometria de Fluxo , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Baço/citologia , Baço/imunologia , Linfócitos T/imunologiaRESUMO
INTRODUÇÃO: O diagnóstico das doenças linfoproliferativas (DLP) tradicionalmente baseia-se no estudo histológico dos linfonodos (LN) acrescido de imuno-histoquímica. A imunofenotipagem (IFT) pela citometria de fluxo (CF) é uma ferramenta sensível e rápida, que pode ser aplicada nas DLP, em material obtido por punção aspirativa por agulha fina (PAAF) de LN. O Bcl-2 é um proto-oncogene que se expressa em várias DLP, porém em níveis especialmente elevados no linfoma folicular (LF). OBJETIVOS: Diagnosticar DLP, através de morfologia e imunofenotipagem por CF, em amostras obtidas por PAAF de LN. MATERIAL e MÉTODO: Amostras de 25 pacientes com adenopatias e de duas tonsilas reacionais foram analisadas pela morfologia e IFT, utilizando um painel inicial de AcMo (CD3, CD4, CD8, CD19, anti-kappa; e anti-lambda;), ampliado conforme a necessidade (CD5, CD10, CD11c, CD23, CD79b, sIgM, FMC-7 e Bcl-2). Os resultados foram comparados com a histologia. RESULTADOS:Dos 25 casos, quatro foram classificados como reacionais e 21 como DLP-B, havendo concordância com resultados histológicos em todos os casos. A intensidade média de fluorescência (IMF) da Bcl-2 no LF (19,92) foi maior que em outras DLP-B (11,93) e que nos controles (3,49) (p = 0,032). CONCLUSÃO:A PAAF de LN combinada com a citomorfologia e a IFT por CF permite uma rápida diferenciação entre os processos reacionais e linfoproliferativos B. A elevada expressão da Bcl-2 nos LFs pela citometria mostra sua utilidade no diagnóstico do tipo mais freqüente das DLP-B. A obtenção de células por PAAF requer treinamento e recomendamos mais de uma punção.
BACKGROUND: The diagnosis of lymphoproliferative disorders (LPD) is routinely made through histological and immunohistochemical analysis of lymph nodes. Immunophenotyping by flow cytometry (FC) is a sensitive and fast tool, which may be applied in samples obtained through fine needle aspiration for the diagnosis of LPD. Bcl-2 is a proto-oncogene that appears in several LPD and it has a significantly high expression in follicular lymphomas. OBJECTIVES: to diagnose LPD in FNA samples through morphology and flow cytometry immunophenotyping. MATERIAL AND METHODS: Samples from 25 patients with lymphadenopathies and 2 reactive tonsils were studied through morphology and immunophenotyping. The antigens expressions were evaluated by using a screening panel of monoclonal antibodies (CD3, CD4, CD8, CD19, light chains kappa; and lambda), followed by CD5, CD10, CD11c, CD23, CD79b, sIgM, FMC-7 and Bcl-2 when required. The results were compared with histology. RESULTS:Four out of 25 samples were reactive processes and 21were B-LPD. In all cases there was consistency with histological results. The mean fluorescence intensity of Bcl-2 in Follicular Lymphoma (19.92) was higher compared with other lymphoproliferative diseases (11.93) and controls (3.49) (p = 0.032). CONCLUSION: Fine needle aspiration of lymph nodes associated with cytomorphology and flow cytometry immunophenotyping allows a fast differentiation between reactive processes and B lymphoproliferative cases. The high expression of Bcl-2 by cytometry shows its usefulness in the diagnosis of the most frequent type of B-LPD. Fine needle aspiration sampling requires training and more than one aspiration is recommended.
Assuntos
Humanos , Masculino , Feminino , Pessoa de Meia-Idade , Biópsia por Agulha Fina/métodos , Citometria de Fluxo/métodos , Linfoma de Células B/diagnóstico , Diagnóstico Diferencial , Imuno-Histoquímica , Imunofenotipagem/métodos , Linfoma de Células B/patologia , Linfonodos/citologia , Linfonodos/patologia , Sensibilidade e Especificidade , Tecido Linfoide/patologiaRESUMO
Chronic lymphocytic leukemia is the most prevalent type of leukemia in the West. It is characterized by an extremely variable clinical course. The aim of the study was to detect the most frequent chromosomal abnormalities in patients with CLL using FISH, and assess them regarding age, gender, clinical stage and CD38 and ZAP-70 expressions. We found 51.7 percent of the patients with chromosome abnormalities. The most frequent one was del 13q14 in 34.5 percent of cases. It was associated to other alterations in 17.2 percent. 17p13 deletions were found in 17.2 percent and trisomy 12 in 13.8 percent (in isolation in 6.9 percent and associated to del 13q14, in 6.9 percent of the cases). An 11q22 deletion was found in one case associated to a 13q14 deletion. To better evaluate the relationship between chromosome aberrations and other prognostic factors in CLL, two cytogenetics groups were considered: favorable (13q deletion in isolation and no alteration) and unfavorable outcomes (trisomy 12, 17p13 deletion, 11q22 deletion and two simultaneous alterations).The unfavorable alterations were more frequently seen among young individuals (<60y). There were more females (70 percent) than males in this group (p=0.04). In relation to the Binet's staging system, patients with unfavorable cytogenetic alterations, tended to be B and C stages, while in the favorable group prevailed patients in stage A. Additionally, patients with poor prognostic cytogenetics tended to express CD38 and ZAP-70 proteins.
A leucemia linfocítica crônica (LLC) é o tipo de leucemia mais prevalente no Ocidente e é caracterizada por curso clínico extremamente variável. O objetivo deste estudo foi detectar as anomalias cromossômicas mais freqüentes em pacientes com LLC, empregando a técnica FISH, e correlacioná-las com idade, sexo, estádio clínico, expressão de CD 38 e ZAP-70. Foram encontradas alterações cromossômicas em 51,7 por cento dos pacientes. A mais freqüente foi a del 13q14, observada em 34,5 por cento dos casos e que esteve associada a outras anomalias em 17,2 por cento. Deleção 17p13 foi encontrada em 17,2 por cento e trissomia 12 em 13,8 por cento (isolada em 6,9 por cento e associada à del 13q14 em 6,9 por cento). Deleção 11q22 foi observada em um caso em concomitância à del 13q14. Para melhor avaliar a relação entre alteração cromossômica e outros fatores prognósticos em LLC, dois grupos citogenéticos foram considerados: favorável (deleção 13q isolada e ausência de alterações) e desfavorável (trissomia 12, deleção 17p13, deleção 11q22 e duas anomalias simultâneas). As alterações desfavoráveis foram mais freqüentemente observadas em indivíduos jovens (<60 anos) e em mulheres (70 por cento)(p=0,04). Em relação ao sistema de estadiamento de Binet, houve tendência dos pacientes com alterações cromossômicas desfavoráveis apresenteram-se nos estágios B e C enquanto no grupo favorável prevaleceram aqueles com estágio A. Em adição, pacientes com achados citogenéticos de prognóstico desfavorável tiveram tendência a expressar proteínas CD 38 e ZAP-70.