RESUMO
BACKGROUND: Monoclonal T-cell receptor (TCR) rearrangement is detected in 57-75% of early-stage mycosis fungoides (MF) at diagnosis. A retrospective study showed molecular residual disease (MRD) in 31% of patients in complete clinical remission (CR) after 1 year of treatment. OBJECTIVES: To confirm the frequency of MRD at 1 year and to determine its prognostic value for further relapse. METHODS: Patients with T1-, T2- or T4-stage MF were prospectively included in this multicentre study. At diagnosis, clinical lesions and healthy skin were biopsied. After 1 year of topical treatment, previously involved skin of patients in CR was biopsied for histology and analysis of TCR-γ gene rearrangement. The results were compared with the clinical status each year for 4 years. RESULTS: We included 214 patients, 133 at T1, 78 at T2 and three at T4 stage. At diagnosis, 126 of 204 cases (61·8%) showed TCR clonality in lesional skin. After 1 year, 83 of 178 patients (46·6%) still being followed up were in CR and 13 of 63 (21%) showed MRD. At 4 years, 55 of 109 patients (50·5%) still being followed up were in CR and 44 of 109 (40·4%) were in T1 stage. MRD did not affect clinical status at 4 years (CR vs. T1/T2, P = 1·0; positive predictive value 36·4%; negative predictive value 67·6%). CONCLUSIONS: T-cell clonality at diagnosis and MRD at 1 year are not prognostic factors of clinical status at 4 years.
Assuntos
Rearranjo Gênico do Linfócito T/genética , Micose Fungoide/tratamento farmacológico , Neoplasia Residual/genética , Neoplasias Cutâneas/tratamento farmacológico , Administração Cutânea , Corticosteroides/administração & dosagem , Adulto , Idoso , Idoso de 80 Anos ou mais , Células Clonais , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Micose Fungoide/genética , Recidiva Local de Neoplasia/genética , Estudos Prospectivos , Neoplasias Cutâneas/genética , Resultado do Tratamento , Adulto JovemRESUMO
The objective of the study was to investigate the association of interleukin (IL)-10 promoter microsatellite polymorphisms, linked with altered IL-10 secretion, with the susceptibility to acute coronary syndrome (ACS) in adult Tunisian patients. We genotyped 291 ACS patients and 291 age-, gender- and ethnically matched control subjects for the microsatellites IL-10R [X78437.2g.5325CA(11_15)] and IL-10G [X78437.2g.8134CA(14_29)] by PCR-based assays. Haplotypes were reconstructed using maximum likelihood method. Regression analysis was used in determining the risk imparted by specific IL-10 genotypes and haplotypes. A significant decrease in IL-10G12 (24 CA repeats) (P<0.001; OR=0.465) and IL-10G15 (27 CA repeats) (P=0.043; OR=0.232), and a significant increase in the low IL-10 producer allele, IL-10R3 (14 CA repeats) (P=0.049; OR=1.461), microsatellites were seen in the ACS group compared with controls. Of the possible 14 haplotypes constructed, there was an enrichment of the R2G9 (13CA vs. 21CA) haplotype in controls [P=0.019; adjusted OR (95% CI)=0.67 (0.48-0.94)] and R2G15 (13CA vs. 27CA) haplotype in cases [P=0.042; adjusted OR (95% CI)=5.29 (1.06-26.30)], thus assigning a protective and susceptible nature to these haplotypes respectively. The differential association of IL-10 microsatellite alleles and haplotypes with ACS suggests that IL-10 contributes to ACS pathogenesis. While the functional attributes of these microsatellite markers remain to be seen, it is likely that they have distinct functional properties (altered IL-10 secretion), which in turn affect the susceptibility to ACS development.
Assuntos
Síndrome Coronariana Aguda/genética , Interleucina-10/genética , Repetições de Microssatélites/genética , Polimorfismo Genético , Adulto , Idoso , Estudos de Casos e Controles , Feminino , Frequência do Gene , Predisposição Genética para Doença , Genótipo , Haplótipos , Humanos , Masculino , Pessoa de Meia-Idade , Fatores de Risco , TunísiaRESUMO
BACKGROUND: The usefulness of T-cell receptor gene rearrangement (TCR-GR) analyses for differentiating cutaneous T-cell lymphoma (CTCL) from benign inflammatory disorders (BID) has been insufficiently studied to date. OBJECTIVES: To evaluate the diagnostic value of TCR-GR analyses, comparing polymerase chain reaction (PCR) with denaturing gradient gel electrophoresis (DGGE) analysis and BIOMED-2 standardized protocol PCR with GeneScan analysis (BIOMED-2-GS). METHODS: Both types of PCR were performed in 157 patients evaluated for initial features suggestive of CTCL between 1996 and 2007. After clinical and histological review, the final diagnosis was CTCL in 77 cases and BID in 80 cases. RESULTS: DGGE and BIOMED-2-GS had a similar diagnostic value for distinguishing CTCL from BID, with a sensitivity of 74% and 77%, respectively, and a specificity of 86%. The observed concordance between both methods was 90% and the kappa coefficient was 0.79. Positivity rates did not depend on the PCR method but varied according to the type of CTCL (73-75% in mycosis fungoides, 90-100% in Sézary syndrome, 40-60% in lymphomatoid papulosis and 100% in other types). The positivity rate in BID was 14% with both methods. The most frequent BID with a monoclonal pattern were drug-induced cutaneous lymphoid hyperplasia, erythrodermic psoriasis and pityriasis lichenoides chronica. CONCLUSIONS: BIOMED-2-GS analysis of the TCRgamma gene is as sensitive and specific as DGGE for CTCL diagnosis. In addition, BIOMED-2-GS is less time-consuming and gives more information concerning the size and nature of TCR-GR.
Assuntos
Eletroforese em Gel de Poliacrilamida/métodos , Rearranjo Gênico do Linfócito T/genética , Linfoma Cutâneo de Células T/genética , Reação em Cadeia da Polimerase/métodos , Neoplasias Cutâneas/genética , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Criança , Feminino , Géis , Humanos , Masculino , Pessoa de Meia-Idade , Reprodutibilidade dos Testes , Sensibilidade e Especificidade , Adulto JovemRESUMO
Recently, we and others described a new chromosomal rearrangement, that is, inv(7)(p15q34) and t(7;7)(p15;q34) involving the T-cell receptor beta (TCRbeta) (7q34) and the HOXA gene locus (7p15) in 5% of T-cell acute lymphoblastic leukemia (T-ALL) patients leading to transcriptional activation of especially HOXA10. To further address the clinical, immunophenotypical and molecular genetic findings of this chromosomal aberration, we studied 330 additional T-ALLs. This revealed TCRbeta-HOXA rearrangements in five additional patients, which brings the total to 14 cases in 424 patients (3.3%). Real-time quantitative PCR analysis for HOXA10 gene expression was performed in 170 T-ALL patients and detected HOXA10 overexpression in 25.2% of cases including all the cases with a TCRbeta-HOXA rearrangement (8.2%). In contrast, expression of the short HOXA10 transcript, HOXA10b, was almost exclusively found in the TCRbeta-HOXA rearranged cases, suggesting a specific role for the HOXA10b short transcript in TCRbeta-HOXA-mediated oncogenesis. Other molecular and/or cytogenetic aberrations frequently found in subtypes of T-ALL (SIL-TAL1, CALM-AF10, HOX11, HOX11L2) were not detected in the TCRbeta-HOXA rearranged cases except for deletion 9p21 and NOTCH1 activating mutations, which were present in 64 and 67%, respectively. In conclusion, this study defines TCRbeta-HOXA rearranged T-ALLs as a distinct cytogenetic subgroup by clinical, immunophenotypical and molecular genetic characteristics.
Assuntos
Proteínas de Homeodomínio/genética , Leucemia-Linfoma de Células T do Adulto/genética , Receptores de Antígenos de Linfócitos T alfa-beta/genética , Adolescente , Adulto , Criança , Deleção Cromossômica , Inversão Cromossômica , Feminino , Rearranjo Gênico do Linfócito T , Proteínas Homeobox A10 , Humanos , Imunofenotipagem , Leucemia-Linfoma de Células T do Adulto/patologia , Leucemia-Linfoma de Células T do Adulto/fisiopatologia , Masculino , Pessoa de Meia-Idade , Receptor Notch1/genética , Ativação Transcricional , Translocação GenéticaRESUMO
A retrospective cytogenetic study of acute myeloid leukemias (AML) and myelodysplastic syndromes (MDS) was conducted by the Groupe Francophone de Cytogénétique Hématologique (GFCH) to evaluate the structural abnormalities of chromosome 5 associated with other chromosomal abnormalities, in particular of chromosome 7, in these pathologies. In all, 110 cases of AML/MDS were recruited based on the presence of chromosome 5 abnormalities under conventional cytogenetics and supplemented by a systematic fluorescence in situ hybridization study of chromosomes 5 and 7. The abnormalities of the long arm of chromosome 5 (5q) were deletions of various sizes and sometimes cryptic. The 5q abnormalities were associated with translocations in 54% of cases and were simple deletions in 46%. In 68% of cases, 5q deletions were associated with chromosome 7 abnormalities, and 90% of these presented a complex karyotype. Of the 110 patients, 28 had a hematopoietic disorder secondary to chemotherapy, radiotherapy, or both. Among 82 patients with de novo AML/MDS, 63 were older than 60 years. Chromosomal abnormalities often associated hypodiploidy and chromosome 5 and 7 abnormalities in complex karyotypes, features resembling those of secondary hemopathies. Systematic investigation of the exposure to mutagens and oncogenes is thus essential to specify the factors potentially involved in MDS/AML with 5q abnormalities.
Assuntos
Aberrações Cromossômicas , Cromossomos Humanos Par 5 , Cromossomos Humanos Par 7 , Leucemia Mieloide/genética , Síndromes Mielodisplásicas/genética , Doença Aguda , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Antineoplásicos/efeitos adversos , Deleção Cromossômica , Feminino , Humanos , Hibridização in Situ Fluorescente , Masculino , Pessoa de Meia-Idade , Neoplasias Induzidas por Radiação , Translocação GenéticaRESUMO
The NUP98 gene is fused with 19 different partner genes in various human hematopoietic malignancies. In order to gain additional clinico-hematological data and to identify new partners of NUP98, the Groupe Francophone de Cytogénétique Hématologique (GFCH) collected cases of hematological malignancies where a 11p15 rearrangement was detected. Fluorescence in situ hybridization (FISH) analysis showed that 35% of these patients (23/66) carried a rearrangement of the NUP98 locus. Genes of the HOXA cluster and the nuclear-receptor set domain (NSD) genes were frequently fused to NUP98, mainly in de novo myeloid malignancies whereas the DDX10 and TOP1 genes were equally rearranged in de novo and in therapy-related myeloid proliferations. Involvement of ADD3 and C6ORF80 genes were detected, respectively, in myeloid disorders and in T-cell acute lymphoblastic leukemia (T-ALL), whereas the RAP1GDS1 gene was fused to NUP98 in T-ALL. Three new chromosomal breakpoints: 3q22.1, 7p15 (in a localization distinct from the HOXA locus) and Xq28 were detected in rearrangements with the NUP98 gene locus. The present study as well as a review of the 73 cases previously reported in the literature allowed us to delineate some chromosomal, clinical and molecular features of patients carrying a NUP98 gene rearrangements.
Assuntos
Neoplasias Hematológicas/genética , Complexo de Proteínas Formadoras de Poros Nucleares/genética , Translocação Genética/genética , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Criança , Pré-Escolar , Análise Citogenética , Feminino , França , Proteínas de Homeodomínio/genética , Humanos , Hibridização in Situ Fluorescente , Lactente , Masculino , Pessoa de Meia-Idade , Receptores Citoplasmáticos e Nucleares/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Sensibilidade e Especificidade , Sociedades MédicasRESUMO
Defects in the fusion of facial buds can result from an anomaly in tissue development or apoptosis, or both. Our working hypothesis was that anomalies in the development of tissues could be caused by a genetic angiogenic defect. Our main objective was to design a reproducible experimental model to study the expression of angiogenic genes in the borders of cleft lips with or without cleft palate. We therefore prospectively studied seven non-syndromic patients, three with a cleft lip (2 right, 1 left), and four with a cleft lip and palate (1 bilateral, 2 right, 1 left), with no CGH (comparative genomic hybridisation) array, who had primary operations to repair their clefts. We also used four controls (cultured fibroblasts from healthy skin samples). The mean (range) age at operation was 44 (13-77) days. We studied the lateral and medial borders histologically and did qPCR (quantitative real-time polymerase chain reaction) analysis for gene expression with 22 genes of interest (and two housekeeping genes) involved in cleft lip and angiogenesis. The qPCR analysis found significant (p<0.05) overexpression of eight genes in the medial border and seven in the lateral border, and underexpression of nine genes in the medial, and ten in the lateral border. The difference in expression between the two borders was not significant. This preliminary study has enabled us to develop a new method to analyse the expression of angiogenic genes in the borders of cleft lips.
Assuntos
Fenda Labial/genética , Fissura Palatina/genética , Expressão Gênica , Neovascularização Patológica/genética , Fenda Labial/cirurgia , Fissura Palatina/cirurgia , Feminino , Humanos , Lactente , Recém-Nascido , Masculino , Estudos Prospectivos , Reação em Cadeia da Polimerase em Tempo RealRESUMO
Chromosomal abnormalities of erythroleukemia (EL) are often described as complex and unspecific. A retrospective study of 75 EL defined following the WHO classification was performed by the Groupe Francophone de Cytogénétique Hématologique (GFCH) in order to reexamine the cytogenetics of this infrequent leukemia subtype. Clonal chromosomal abnormalities were found in 57 patients (76%), distributed in 4 subgroups according to their ploidy status: pseudodiploid (16%), hypodiploid (47%), hyperdiploid (19%), and 18% mixed cases associating 2 different clones (hypodiploid+hyperdiploid) or (pseudodiploid+hyperdiploid). Complex rearrangements and hypodiploid chromosome number were widely dominant (50%). Partial or entire monosomies represented 56% of abnormalities. Chromosomes 5 and 7 were the most frequently involved (41 and 33 times, respectively), followed by chromosomes 8, 16, and 21 (19 times each). Unbalanced abnormalities were more frequent than balanced. All these kinds of abnormalities were observed in de novo as well as in secondary EL. Four out of 7 cases of "pure erythroid" leukemia were associated with a BCR-ABL fusion. Lastly, no chromosome abnormality specific to EL could be established. However, the large overlap of chromosomal abnormality patterns of EL (pure erythroid form excepted) and refractory anemia with excess of blasts in transformation (RAEB-t) favors the hypothesis of similarities between these 2 hematologic disorders.
Assuntos
Aberrações Cromossômicas , Leucemia Eritroblástica Aguda/genética , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Criança , Pré-Escolar , Cromossomos Humanos , Humanos , Pessoa de Meia-Idade , Ploidias , Estudos Retrospectivos , Análise de SobrevidaRESUMO
To accurately estimate the incidence of HOX11L2 expression, and determine the associated cytogenetic features, in T-cell acute lymphoblastic leukemia (T-ALL), the Groupe Français de Cytogénétique Hématologique (GFCH) carried out a retrospective study of both childhood and adult patients. In total, 364 patients were included (211 children =15 years and 153 adults), and 67 (18.5%) [47 children (22.4%) and 20 adults (13.1%)] were shown to either harbor the t(5;14)q35;q32) translocation or express the HOX11L2 gene or both. Most of the common hematological parameters did not show significant differences within positive and negative populations, whereas the incidence of CD1a+/CD10+ and cytoplasmic CD3+ patients was significantly higher in positive than in negative children. Out of the 63 positive patients investigated by conventional cytogenetics, 32 exhibited normal karyotype, whereas the others 31 showed clonal chromosome abnormalities, which did not include classical T-ALL specific translocations. Involvement of the RANBP17/HOX11L2 locus was ascertained by fluorescence in situ hybridization in six variant or alternative (three-way translocation or cytogenetic partner other than 14q32) translocations out of the 223 patients. Our results also show that HOX11L2 expression essentially occurs as a result of a 5q35 rearrangement, but is not associated with another identified T-ALL specific recurrent genetic abnormality, such as SIL-TAL fusion or HOX11 expression.
Assuntos
Cromossomos Humanos Par 14/genética , Cromossomos Humanos Par 5/genética , Proteínas de Homeodomínio/genética , Leucemia-Linfoma de Células T do Adulto/genética , Proteínas Oncogênicas/genética , Translocação Genética , Adolescente , Adulto , Criança , Pré-Escolar , Aberrações Cromossômicas , Células Clonais , Feminino , Humanos , Imunofenotipagem , Hibridização in Situ Fluorescente , Cariotipagem , Masculino , Ploidias , Proteínas Proto-Oncogênicas , Estudos Retrospectivos , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Taxa de SobrevidaRESUMO
Adenosine (Ado) is a potent anti-inflammatory agent acting on a variety of cell functions. However, its effects on human monocytes have been less well characterized. We investigated the effect of Ado and its receptor-specific analogs on NADPH oxidase activity with the use of luminol-enhanced chemiluminescence (CL). Adenosine inhibited fMLP-triggered NADPH oxidase activity with a maximal inhibition of 55+/-5%. IB-MECA, a selective A3 Ado receptor agonist reduced fMLP triggered NADPH oxidase activity more potently than the A2 receptor agonist CGS 2180 HCl (CGS) and the A1 Ado receptor agonist N-2-phenylethyl-adenosine (R-PIA). The inhibitory effect of Ado was reversed by neither the A1 Ado receptor antagonist 1,3-dipropyl-8(2-amino-4chlorophenyl)-xanthine (PACPX) nor the A2 Ado receptor antagonist 3,7-dimethyl-1-(2-propynyl)xanthine (DMPX). It was significantly reversed by the A1/A3 Ado receptor antagonist xanthine amine congener (XAC). Pretreatment of monocytes by cytochalasin B reversed the effect of Ado but not of dibutyryl cAMP (dBcAMP) on fMLP-CL response. KT 5720, a specific cAMP-dependent protein kinase inhibitor completely counteracted the inhibition of NADPH oxidase activity by dBcAMP but not by Ado. Using flow cytometry, we observed that Ado did not inhibit intracellular oxidative metabolism, whereas dBcAMP did. Furthermore, the inhibition of NADPH oxidase activity by Ado was not mediated by changes in cytosolic calcium. These results demonstrated that Ado inhibited NADPH oxidase activity via A3 Ado receptor independently of cAMP elevation or changes in calcium mobilization.
Assuntos
Adenosina/farmacologia , Carbazóis , Monócitos/efeitos dos fármacos , N-Formilmetionina Leucil-Fenilalanina/antagonistas & inibidores , Receptores Purinérgicos P1/efeitos dos fármacos , Explosão Respiratória/efeitos dos fármacos , Bucladesina/farmacologia , Cálcio/fisiologia , AMP Cíclico/fisiologia , Proteínas Quinases Dependentes de AMP Cíclico/antagonistas & inibidores , Proteínas Quinases Dependentes de AMP Cíclico/fisiologia , Citocalasina B/farmacologia , Ativação Enzimática/efeitos dos fármacos , Inibidores Enzimáticos/farmacologia , Citometria de Fluxo , Humanos , Indóis/farmacologia , Medições Luminescentes , Monócitos/enzimologia , N-Formilmetionina Leucil-Fenilalanina/farmacologia , NADPH Oxidases/metabolismo , Consumo de Oxigênio/efeitos dos fármacos , Pirróis/farmacologia , RNA Mensageiro/análise , Receptor A3 de Adenosina , Receptores Purinérgicos P1/genética , Receptores Purinérgicos P1/fisiologia , Sistemas do Segundo Mensageiro/efeitos dos fármacos , Teobromina/análogos & derivados , Teobromina/farmacologia , Xantinas/farmacologiaRESUMO
A simplified prognostic score is presented based on the multivariate analysis of 138 refractory/relapsed acute myeloid leukaemia (AML) patients (median age 55 years, range: 19-70) receiving a combination of intensive chemotherapy+Gemtuzumab as salvage regimen. Overall, 2-year event-free survival (EFS) and overall survival (OS) were 29±4% and 36±4%, respectively. Disease status (relapse <12 months, including refractory patients), FLT3-ITD-positive status and high-risk cytogenetics were the three strongest independent adverse prognostic factors for OS and EFS in this series. We then defined three subgroups with striking different outcomes at 2 years: no adverse factor (favourable, N=36): OS 58%, EFS 45%; one adverse factor (intermediate, N=54): OS 37%, EFS 31%; two or three adverse factors (poor, N=43): OS 12%, EFS 12% (P<10(-4), P=0.001). This new simplified Leukemia Prognostic Scoring System was then validated on an independent cohort of 111 refractory/relapsed AML patients. This new simplified prognostic score, using three clinical and biological parameters routinely applied, allow to discriminate around two third of the patients who should benefit from a salvage intensive regimen in the setting of refractory/relapsed AML patients. The other one third of the patients should receive investigational therapy.
Assuntos
Leucemia Mieloide Aguda/patologia , Prognóstico , Índice de Gravidade de Doença , Adulto , Idoso , Aminoglicosídeos/uso terapêutico , Anticorpos Monoclonais/uso terapêutico , Anticorpos Monoclonais Humanizados , Intervalo Livre de Doença , Gemtuzumab , Humanos , Leucemia Mieloide Aguda/diagnóstico , Pessoa de Meia-Idade , Recidiva , Terapia de Salvação/métodos , Resultado do TratamentoRESUMO
BACKGROUND: Endothelial progenitor cells (EPC) are good candidates for cell-based therapy in cardiovascular diseases. However, concerns have been raised about the potential risks of EPC-based cell therapy, in terms of thrombogenicity particularly in inflammatory conditions, currently observed in such patients. Tissue factor (TF) can trigger coagulation and may support thrombogenicity. TF is also a key receptor in angiogenesis. OBJECTIVE: The present study was designed to (i) evaluate the capacity of resting and tumour necrosis factor-alpha (TNF)-α-stimulated late-outgrowth endothelial colony-forming cells (ECFCs) to express TF and (ii) investigate the effect of TF/FVII(a) interaction on procoagulant and non-procoagulant activities of ECFCs in vitro. METHODS: ECFCs from cord blood (cb) and adult peripheral blood (ab) were analyzed for TF expression and activity using reverse transcription-polymerase chain reaction (RT-PCR), flow cytometry, Western blot and a thrombin generation assay. Non-procoagulant properties of TF-expressing ECFCs were investigated in vitro using wound-healing, cell proliferation, tube formation and spheroid-based assays. RESULTS: ECFCs expressed TF in response to TNF-α. The up-regulation of TF conferred to ECFCs a FVII(a)-dependent thrombin generation activity. Compared with cb-ECFC, ab-ECFCs can display a higher level of constitutive TF expression and activity, with a notable heterogeneity among donors. TF/FVIIa interaction did not modify non-procoagulant properties of TNF-α stimulated cb-ECFCs in vitro. CONCLUSIONS: Proinflammatory conditions up-regulate TF expression in ECFCs. This expression confers to ECFCs a strong thrombin generation capacity without influencing their non-coagulant properties. Our results suggest that EPC-based cell therapy may be associated with prothrombotic risk which could be limited by inhibiting TF without affecting the proangiogenic capacity of the cells.
Assuntos
Endotélio Vascular/citologia , Trombina/metabolismo , Tromboplastina/metabolismo , Regulação para Cima , Western Blotting , Proliferação de Células , Coagulantes/química , Fator VII/metabolismo , Sangue Fetal/citologia , Citometria de Fluxo/métodos , Humanos , Inflamação , Risco , Trombose , Fator de Necrose Tumoral alfa/metabolismo , CicatrizaçãoRESUMO
The t(8;16)(p11;p13) is a rare translocation involved in de novo and therapy-related myelomonocytic and monocytic acute leukemia. It fuses two genes encoding histone acetyltransferases (HATs), MYST3 located at 8p11 to CREBBP located at 16p13. Variant translocations involve other HAT-encoding genes such as EP300, MYST4, NCOA2 or NCOA3. MYST3-linked acute myeloid leukemias (AMLs) share specific clinical and biological features and a poor prognosis. Because of its rarity, the molecular biology of MYST3-linked AMLs remains poorly understood. We have established the genome and gene expression profiles of a multicentric series of 61 M4/M5 AMLs including 18 MYST3-linked AMLs by using array comparative genome hybridization (aCGH) (n=52) and DNA microarrays (n=44), respectively. We show that M4/5 AMLs have a variety of rare genomic alterations. One alteration, a gain of the MYB locus, was found recurrently and only in the MYST3-linked AMLs (7/18 vs 0/34). MYST3-AMLs have also a specific a gene expression profile, which includes overexpression of MYB, CD4 and HOXA genes. These features, reminiscent of T-cell acute lymphoid leukemia (ALL), suggest the targeting of a common T-myeloid progenitor.
Assuntos
Perfilação da Expressão Gênica/métodos , Genes myb/genética , Histona Acetiltransferases/genética , Leucemia Mielomonocítica Aguda/genética , Antígenos CD4/genética , Hibridização Genômica Comparativa , Regulação Neoplásica da Expressão Gênica , Genoma Humano , Proteínas de Homeodomínio/genética , Humanos , Análise de Sequência com Séries de Oligonucleotídeos , Proteínas Proto-Oncogênicas c-myb/genéticaRESUMO
A series of 38 patients with acute myeloblastic leukemia (AML) with 49 or more chromosomes and without structural abnormalities was selected within the Groupe Francophone de Cytogénétique Hématologique (GFCH) to better define their characteristics. The median age of the patients was 65 years, and all FAB subtypes were represented. Although all chromosomes were gained, some seems to prevail: chromosome 8 (68%), 21 (47%), 19 (37%), and 13 and 14 (34% each). Since MLL rearrangement leads patients in a group with an unfavorable prognosis, search for cryptic rearrangements of MLL was performed in 34 patients and showed abnormalities in 5 (15%). When we applied the most frequent definition of complex karyotypes (three or more abnormalities), all patients with high hyperdiploid AML fall in the unfavorable category. Among the 18 patients without MLL rearrangement receiving an induction therapy, 16 (89%) reached CR and 6 (33%) were still alive after a 31-month median follow-up (14-61 months). Although this study was retrospective, these results suggest that high hyperdiploid AML without chromosome rearrangement seems to be a subgroup of uncommon AML (less than 1%), and may be better classified in the intermediate prognostic group.
Assuntos
Aberrações Cromossômicas , Cromossomos Humanos/genética , Leucemia Mieloide/genética , Ploidias , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Feminino , Rearranjo Gênico , Histona-Lisina N-Metiltransferase , Humanos , Hibridização in Situ Fluorescente , Cariotipagem , Leucemia Mieloide/tratamento farmacológico , Leucemia Mieloide/epidemiologia , Masculino , Pessoa de Meia-Idade , Proteína de Leucina Linfoide-Mieloide/genética , Prognóstico , Estudos Prospectivos , Estudos RetrospectivosRESUMO
BACKGROUND: Counterflow centrifugal elutriation is the method of choice for obtaining a large quantity of highly purified monocytes. In spite of the fact that this technique has been used for many years to isolate monocytes for cellular immunotherapy, it is not known whether the process of elutriation can stimulate tissue factor (TF) expression and therefore trigger coagulation in patients receiving these cell preparations. The aim of the present study is thus to identify TF and TF pathway inhibitor (TFPI) in elutriated monocytes and to evaluate their ability to trigger thrombin generation. STUDY DESIGN AND METHODS: Human monocytes are separated by leukapheresis and elutriation in sterile, endotoxin-free conditions. TF and TFPI mRNA is detected by reverse transcription-polymerase chain reaction. TF and TFPI are measured by enzyme-linked immunosorbent assay in cell lysates. TF antigen expression on cell surface is evidenced by direct-flow cytometry. Two functional tests (a chronometric test and an amidolytic assay) assess the capacity of monocytes to trigger thrombin generation. The response to lipopolysaccharide (LPS) is evaluated with each technique. Monocytic cell line THP-1 is used as a positive control. RESULTS: Elutriated monocytes coexpress TF mRNA and TFPI mRNA. The expression of TF mRNA is dramatically increased by LPS activation. This is correlated with a 100-fold increase in the amount of TF antigen in monocyte lysates. Flow immunocytometry confirms the expression of TF antigen on cell membrane in response to LPS stimulation, whereas TFPI mRNA is slightly increased after LPS stimulation. The amount of TFPI antigen in cell lysates is small when compared to that in plasma. Elutriated monocytes have a strong potential to trigger thrombin generation in response to LPS. CONCLUSION: In spite of the coexpression of TF mRNA and TFPI mRNA, elutriated monocytes are capable of supporting prothrombinase activity. This should be taken into account in the evaluation of the safety of adoptive cellular immunotherapy.
Assuntos
Citaferese/métodos , Leucaférese , Lipoproteínas/biossíntese , Monócitos/metabolismo , Tromboplastina/biossíntese , Antígenos/sangue , Adesão Celular/efeitos dos fármacos , Hemostáticos/metabolismo , Humanos , Lipopolissacarídeos/farmacologia , Lipoproteínas/genética , Lipoproteínas/imunologia , Monócitos/citologia , RNA Mensageiro/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Trombina/biossíntese , Tromboplastina/genética , Tromboplastina/imunologia , Tromboplastina/metabolismoRESUMO
Human monocytes may play a central role in haemostasis since they are the only circulating blood cells capable to express tissue factor (TF) and therefore trigger the extrinsic pathway of coagulation. The aim of this review is to illustrate that monocytes also participate in haemostasis independently of TF expression. Indeed, the exposure of anionic phospholipids provides a procoagulant surface. Activation of factor X involves specific membrane receptor EPR-1 (Effector cell Protease Receptor-1). Mac-1 and membrane factor Va also bind factor X/Xa. In addition, monocytic proteases have been reported to modulate the coagulation cascade. Monocyte adhesion and cell-cell interactions represent important mechanisms implicated in TF expression. Furthermore, Tissue factor pathway inhibitor (TFPI), the natural inhibitor of TF activity, is expressed by monocytes. This suggests that monocytes could participate to the local regulation of coagulation initiation. The importance of monocytes in the development of vascular diseases remains to be clarified. In thrombotic disorders, it may be worthwhile not to limit the investigations to the expression of TF.
RESUMO
BACKGROUND: In vitro generation of dendritic cells (DCs) from human monocytes represents a promising tool in immunotherapy. However, it is not known whether the separation of DCs from monocytes induces tissue factor expression and therefore may trigger coagulation in patients receiving these DC preparations. The aim of this study is thus to analyze tissue factor expression on monocyte-derived DCs and to compare their ability to trigger thrombin generation to that of macrophages obtained from the same monocytes. STUDY DESIGN AND METHODS: Human monocytes are separated by leukapheresis and washed by using counterflow centrifugation in sterile, endotoxin-free conditions. Macrophages are grown from human monocytes in the presence of GM-CSF alone and immature DCs are grown in the presence of GM-CSF plus IL-4 for 5 days with fetal calf serum (IDC-FCS). Immature DCs are also grown from human monocytes for 7 days in the presence of GM-CSF plus IL-4 with human group AB serum (IDC-HS). The addition of prostaglandin E(2) and TNFalpha in this culture medium at Day 5 leads to mature DCs (MDC-HS). Tissue factor mRNA expression is studied by RT-PCR analysis. Tissue factor antigen is measured by ELISA in cell lysates and by direct flow cytometry. The procoagulant activity of intact cells is assessed by using an amidolytic assay or a chronometric assay. RESULTS: IDC-FCS express tissue factor mRNA and antigen and trigger thrombin generation. Procoagulant activity of IDC-FCS is dependent on both tissue factor expression and exposure to anionic phospholipid. Monocyte-derived macrophages cultured for 5 days with GM-CSF alone express lower levels of tissue factor mRNA, tissue factor antigen, and procoagulant activity than IDC-FCS. IDC-HS and MDC-HS also express high levels of tissue factor mRNA and antigen and support procoagulant activity. CONCLUSION: Monocyte-derived DCs express a high level of functional tissue factor and support procoagulant activity. This finding should be taken into account in clinical trials.
Assuntos
Separação Celular/métodos , Células Dendríticas/citologia , Monócitos/citologia , Tromboplastina/genética , Animais , Ânions , Antígenos , Bovinos , Células Cultivadas , Centrifugação/métodos , Coagulantes/metabolismo , Sangue Fetal/citologia , Humanos , Fosfolipídeos/biossíntese , RNA Mensageiro/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Tromboplastina/imunologiaRESUMO
Due to the extensive use of unfractionated heparins in France, there is considerable experience with heparin-induced thrombocytopenia (HIT). It is recommended that platelet counts be performed twice a week for three weeks when patients are treated with any form of heparin. A drop in platelet counts can, however, occur not only in HIT patients but also for other unrelated reasons. For diagnosing HIT, all laboratories in France use platelet aggregometry inspite of poor sensitivity. Both false positive and false negative results are obtained. The serotonin release test is not used in France. The ELISA test for HIT does not always correlate with the platelet aggregation test and many patients with a positive ELISA test do not necessarily have other evidence for HIT. This is especially true in patients following cardiopulmonary bypass surgery. None of the available laboratory tests reliably identify patients with HIT. Patients with HIT should not be managed with low-molecular-weight heparins, but danaparoid, argatroban and ancrod are viable options. Also, recombinant hirudin has been employed. All have advantages and disadvantages. At present, the diagnosis and management of patients with HIT remains difficult and properly designed clinical studies are needed to obtain answers to several open questions.