RESUMEN
BACKGROUND: To evaluate the efficacy and toxicity of CHG regimen (low-dose cytarabine, homoharringtonine with G-CSF priming) as an induction chemotherapy for elderly patients with high-risk MDS or acute myeloid leukemia transformed from MDS (MDS-AML). METHODS: Thirty-three untreated patients (21 high-risk MDS and 12 MDS-AML) were enrolled in this study. Each patient was administered with the CHG regimen comprised of low-dose cytarabine (25 mg/day, days 1-14) and homoharringtonine (1 mg/day, days 1-14) by intravenous continuous infusion in combination with G-CSF (300 µg/day) by subcutaneous injection from day 0 until neutrophil count recovery to 2.0 × 10(9)/L. RESULTS: The overall response rate (OR) was 66.7% after one course of the CHG regimen with 19 patients reaching CR (57.6%) and 3 patients reaching partial remission (PR) (9.1%). The median overall survival (OS) was 15.0 months. Patients with normal serum lactate dehydrogenase (LDH) appeared longer median OS when compared to patients with high LDH level (18 months vs. 5 months, P = 0.011). Grade 3/4 thrombocytopenia occurred in 28% of patients, neutropenia in 34%. No treatment-related deaths occurred during the induction therapy. CONCLUSIONS: These data suggest that the CHG priming regimen is effective and safe as a novel induction therapy for elderly patients with high-risk MDS and MDS-AML. The results need to be conformed in further study involving a larger cohort of patients.
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Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Leucemia Mieloide Aguda/tratamiento farmacológico , Síndromes Mielodisplásicos/tratamiento farmacológico , Anciano , Anciano de 80 o más Años , Protocolos de Quimioterapia Combinada Antineoplásica/efectos adversos , Citarabina/administración & dosificación , Femenino , Factor Estimulante de Colonias de Granulocitos/administración & dosificación , Harringtoninas/administración & dosificación , Homoharringtonina , Humanos , Leucemia Mieloide Aguda/mortalidad , Masculino , Persona de Mediana Edad , Síndromes Mielodisplásicos/complicaciones , Síndromes Mielodisplásicos/mortalidad , RiesgoRESUMEN
The aim of study was to explore the influence of the number of megakaryocytes in bone marrow smear and trephine biopsy on clinical outcome of idiopathic thrombocytopenic purpura (ITP). The clinical outcome of 115 patients with ITP was compared by different clinical subtype, number of blood platelet, number of megakaryocyte in bone marrow smear and trephine biopsy. The results showed that: (1) the clinical outcome of acute ITP was better than that of chronic ITP, the short clinical outcome of acute ITP and chronic ITP were 86.6% vs 60.4% respectively (p < 0.01), the long clinical outcome of them were 82.5% vs 68.9% respectively (p < 0.05); (2) different number of blood platelet at occurrence of diseases had no obviously influence on clinical outcome of patients with ITP; (3) all cases were subgrouped according to number of megakaryocyte in bone marrow smear, the number of megakaryocyte in bone marrow smear less than 7/4.5 cm(2) was defined as group I, the number of megakaryocyte between 7/4.5 cm(2) to 35/4.5 cm(2) was defined as group II, and the number of megakaryocyte in bone marrow smear greater than 35/4.5 cm(2) was defined as group III. The effective rates of 3 groups in short term treatment were 53.3%, 73.8% and 86.2% respectively, and there were statistical difference between these 3 groups (p < 0.01), the effective rates of these 3 groups in long term treatment were 42.8%, 84.6% and 85.5% respectively, and there was no statistical different between group II and group III (p > 0.05), but both had statistical difference, as compared with group I (p < 0.01). (4) all cases were subgrouped by the number of megakaryocyte in trephine biopsy on time of disease occurrence, the number of megakaryocyte in bone marrow smear less than 8/mm(2) was defined as group I, the number of megakaryocyte between 8/mm(2) to 15/mm(2) was defined as group II, the number of megakaryocyte greater than 15/mm(2) was defined as group III. The effective rate of these 3 groups in short term treatment were 53.8%, 85.0% and 90.3% respectively, group II and III had no statistical difference each other (p > 0.05), but both groups had statistical difference as compared with group I (p < 0.01). Effective rate of these 3 groups in long term treatment were 33.3%, 63.1% and 87.9% respectively, and there were statistical difference between them (p < 0.01). (5) the number of blood platelet at time of disease occurrence was not related to number of megakaryocyte in bone marrow smear and in trephine biopsy section(r = 0.31, p > 0.05; r = 0.41, p > 0.05). The number of megakaryocyte in bone marrow smear had positive correlation to that in trephine biopsy slides (r = 0.52, p < 0.01). In case to use single factor Logistic regression, the results showed that number of megakaryocyte in bone marrow smear and trephine biopsy had obvious influence on long term treatment of ITP. It is concluded that the number of megakaryocyte in trephine biopsy can be used as a available supplement method for bone smear, and can forecast the therapeutic effect of patients with ITP.
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Células de la Médula Ósea/patología , Megacariocitos/patología , Púrpura Trombocitopénica Idiopática/diagnóstico , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Células de la Médula Ósea/citología , Recuento de Células , Niño , Femenino , Humanos , Masculino , Megacariocitos/citología , Persona de Mediana Edad , Adulto JovenRESUMEN
OBJECTIVE: To determine the accuracy of nonattenuation corrected (NAC) F-fluorodeoxyglucose positron emission tomography (F-FDG PET) images in the evaluation of solitary pulmonary lesion as compared with more established methods. METHODS: Fifty-six patients received F-FDG PET/CT for diagnosing solitary pulmonary nodules or mass lesions based on histopathology (n=39) and clinical follow-up (n=17). Visual pulmonary lesion FDG uptake was graded by consensus of two nuclear medicine physicians on both attenuation corrected (AC) [absent, less than mediastinal blood pool (MBP), equal to MBP, greater than MBP] and NAC (absent, less than skin, equal to skin, greater than skin) images. Standardized uptake values (SUV) were also measured from AC images. SUV, visual AC, and visual NAC methods' diagnostic performances were compared, distinguishing benign from malignant pulmonary nodules. RESULTS: There were 34 malignant and 22 benign lesions. Lesion diameter varied from 5 to 100 mm (mean ± SD, 24.0 ± 17.9 mm). The NAC, AC, and SUV method sensitivities and specificities were 100/64%, 91/59%, and 79/77%, respectively. For lesions less than 3 cm, NAC, AC, and SUV methods yielded accuracies of 85%, 78%, and 73%, respectively. The NAC method was the most sensitive and accurate especially for small nodules. CONCLUSION: Visual assessment of NAC F-FDG PET images alone may provide a more accurate characterization of solitary pulmonary lesions.
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Fluorodesoxiglucosa F18 , Procesamiento de Imagen Asistido por Computador , Neoplasias Pulmonares/diagnóstico por imagen , Tomografía de Emisión de Positrones/métodos , Nódulo Pulmonar Solitario/diagnóstico por imagen , Adulto , Anciano , Anciano de 80 o más Años , Diagnóstico Diferencial , Reacciones Falso Negativas , Femenino , Fluorodesoxiglucosa F18/metabolismo , Humanos , Neoplasias Pulmonares/metabolismo , Masculino , Persona de Mediana Edad , Estudios Retrospectivos , Nódulo Pulmonar Solitario/metabolismoAsunto(s)
Examen de la Médula Ósea/métodos , Neoplasias de la Médula Ósea/secundario , Médula Ósea/patología , Neoplasias Pulmonares/patología , Neoplasias Gástricas/patología , Biopsia , Biopsia con Aguja , Neoplasias de la Médula Ósea/patología , Neoplasias de la Mama/patología , Técnicas Citológicas , Femenino , Humanos , Masculino , Neoplasias de la Próstata/patología , Estudios RetrospectivosRESUMEN
A total of 32 patients (25 with advanced MDS and 7 with t-AML) were enrolled in this study to evaluate the efficacy and toxicity of the low-dose cytarabine and homoharringtonine in combination with granulocyte colony-stimulating factor (G-CSF) (CHG protocol) in patients with advanced myelodysplastic syndromes (MDS) or MDS-transformed acute myeloid leukemia (t-AML). All the patients were administered the CHG regimen comprising low-dose cytarabine (25 mg/day, intravenous continuous infusion, days 1-14), homoharringtonine (1 mg/day, intravenous continuous infusion, days 1-14), and G-CSF (300 microg/day, subcutaneous injection, days 0-14, interrupted when the peripheral white blood cell count reached >20 x 10(9)/L). The overall response rate was 71.9% after the administration of one course of the CHG regimen. Of the 32 patients, 15 (46.9%) achieved complete remission (CR) and 8 (25%) achieved partial remission (PR). This regimen was followed by a post-remission therapy that included conventional chemotherapy, when CR was achieved. Of the patients with CR who just received post-remission regimens as homoharringtonine and cytarabine (HA) and daunorubicin and cytarabine (DA) 6 relapsed rapidly and just had a mean 6.1 months of CR. Otherwise, the other 8 out of 14 patients with CR alternatively received subsequent chemotherapy, which combined mitoxantrone, idarubicin, pirarubicin, or aclarubicin with cytarabine. The mean CR duration of the 8 patients had reached 10.6 months, and 5 of the 8 still kept a continuous CR. The median overall survival (OS) was 18.2 months. There were no statistically significant differences for CR, PR, and OS when the patients were grouped by age, blasts in bone marrow, and karyotypes, respectively. No treatment-related deaths were observed. Myelosuppression was mild to moderate, and no severe non-hematological toxicity was observed. Thus, a CHG priming regimen as an induction therapy was well tolerated and effective in patients with advanced MDS or t-AML. Stronger and alternative subsequent chemotherapy is necessary for patients with CR to maintain longer CR and better OS.
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Protocolos de Quimioterapia Combinada Antineoplásica/administración & dosificación , Citarabina/administración & dosificación , Factor Estimulante de Colonias de Granulocitos/administración & dosificación , Harringtoninas/administración & dosificación , Leucemia Mieloide Aguda/tratamiento farmacológico , Síndromes Mielodisplásicos/tratamiento farmacológico , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Protocolos de Quimioterapia Combinada Antineoplásica/efectos adversos , Transformación Celular Neoplásica/efectos de los fármacos , Transformación Celular Neoplásica/patología , Citarabina/efectos adversos , Progresión de la Enfermedad , Relación Dosis-Respuesta a Droga , Femenino , Factor Estimulante de Colonias de Granulocitos/efectos adversos , Harringtoninas/efectos adversos , Homoharringtonina , Humanos , Leucemia Mieloide Aguda/patología , Masculino , Persona de Mediana Edad , Síndromes Mielodisplásicos/patología , Terapia Neoadyuvante , Resultado del Tratamiento , Adulto JovenRESUMEN
The aim of this paper was to investigate whether the morphological dysplastic cells in myelodysplastic syndromes (MDS) marrows derived from abnormal chromosomal clone. Fluorescence in situ hybridization was used in combination with immunochemistry/immuno-magnetic bead sorting to investigate the penetration of chromosomal abnormalities into dysplastic hematopoietic cells in MDS patients with documented chromosome abnormalities in the bone marrow at diagnosis. Typical granulocyte dysplasia was defined as granulocytes with bilobed pseudo Pelger-Huet anomaly, erythrocyte dysplasia as tri-nucleus or nuclear budding erythrocytes (21 patients were investigated, respectively) and megakaryocytes dysplasia as lymphocytic appearance megakaryocytes (14 patients were investigated). When data were analyzed as a whole, the mean percentage of clonal cells with typical dysplasis was always lower than clonal cell percentage in all nucleated cells no matter whether in granulocytes, erythrocytes or megakaryocytes. For each individual case, the situation was the same with just few exceptions. The chromosome ploidy of micro-megakaryocytes was obviously reduced when compared to reported normal megakaryocytes. Our research suggested that tri-lineage morphological dyshematopoiesis is secondum alteration not a specific feature (imagery) of the abnormal chromosomal clone in MDS.
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Aberraciones Cromosómicas , Eritroblastos/patología , Megacariocitos/patología , Síndromes Mielodisplásicos/genética , Síndromes Mielodisplásicos/patología , Anomalía de Pelger-Huët/patología , Adulto , Anciano , Anciano de 80 o más Años , Estudios de Cohortes , Femenino , Humanos , Hibridación Fluorescente in Situ , Masculino , Persona de Mediana Edad , Anomalía de Pelger-Huët/genéticaRESUMEN
OBJECTIVE: To explore polarization of T lymphocyte and its relationship with apoptosis of marrow cells in patients with myelodysplastic syndromes (MDS). METHODS: Measurements of Th1, Th2, Tc1, Tc2 subsets in bone marrows from 34 patients with MDS and 13 normal controls were performed by flow cytometry. INF-gamma and TNF-alpha in marrow serum were determined by ELISA (Enzyme-linked immunosorbent assay). Apoptosis index of marrow cells was detected by TUNEL (TdT-mediated dUTP nick end labeling). Correlations between Th1, Th2, Tc1, Tc2 subsets and INF-gamma, TNF-alpha levels as well as apoptosis index were analyzed, and relationship between TNF-alpha, INF-gamma levels and apoptosis index was also investigated. RESULTS: (1) The percentage of Th1 cells [(10.1 +/- 1.6)%], Tc1 cells [(24.0 +/- 3.6)%] and Tc1/Tc2 ratio (50.0 +/- 11.1) was significantly increased in patients with MDS than in normal controls [(4.0 +/- 0.5)%, (5.8 +/- 0.6)% and 13.4 +/- 2.7, respectively]. Levels of INF-gamma [(58.6 +/- 21.7) microg/L] and TNF-alpha [(15.7 +/- 3.8) microg/L] in marrow serum of MDS patients was markedly elevated compared to normal controls [0 and (0.3 +/- 0.2) microg/L, respectively]. An increased apoptosis index of nucleated cells was observed in MDS patients [(7.8 +/- 1.5)%] as compared to controls [(2.1 +/- 0.3)%, P < 0.05]. The Th1 cell percentage showed a positive correlation with the levels of INF-gamma and TNF-alpha (r = 0.38, P < 0.05 and r = 0.39, P < 0.05, respectively), and with apoptotic index of nucleated marrow cells in MDS patients (r = 0.33, P < 0.05). Furthermore, a positive correlation was observed between INF-gamma, TNF-alpha levels and apoptotic index of marrow cells (r = 0.74, P < 0.01 and r = 0.73, P < 0.01, respectively). (2) Th1, Tc1 cells and Tc1/Tc2 ratio in MDS-RCMD patients was markedly elevated (P < 0.01) but did not in RCMD-RS, RAEB-1 and RAEB-2 patients as compared to normal controls. (3) An elevation in the percentages of Th1, Tcl and Tc1/ Tc2 ratio was detected in patients with IPSS lower-risk but did not in higher-risk group as compared to controls. (4) Increased Th1 and Tc1 percentages and Th1/Th2 and Tc1/Tc2 ratios were observed in RCMD patients with normal karyotype, but did not in those with abnormal karyotype. Conclusions Th1/Th2 and Tc1/Tc2 in bone marrow of MDS patients were unbalanced, polarizing to type I reaction especially in patients with RCMD subtype, IPSS lower-risk and normal karyotype. The increased Th1 cells in bone marrow may account for the increased apoptosis of nucleated marrow cells in MDS, through proapoptotic cytokines such as INF-gamma and TNF-alpha.
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Síndromes Mielodisplásicos/inmunología , Linfocitos T/inmunología , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Apoptosis/inmunología , Médula Ósea/inmunología , Médula Ósea/metabolismo , Médula Ósea/patología , Femenino , Sistema Hematopoyético/inmunología , Humanos , Interferón gamma/metabolismo , Masculino , Persona de Mediana Edad , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
OBJECTIVE: To investigate the biological difference of clonal cells between myelodysplastic syndromes (MDS) and acute myeloid leukemia (AML). METHOD: Bone marrow (BM) clonal cells (which had cytogenetic markers detected by FISH assay) and blasts were quantitatively analysed in 51 MDS and 11 AML patients. The clonal cell percentage in orthochromatic normoblasts, granulocytes and megakaryocytes were assayed. The biological functions for phagocytosis and oxidation of MDS peripheral blood (PB) neutrophils were compared with that of normal controls. RESULTS: Almost all MDS patients BM had a higher clonal cell percentage (mean 48.2%) than blasts percentage (mean 6.7%) (P < 0.01), but with the subtype of MDS advancing this percentage gap was closing up, and in 11 AML patients no such gap was observed. This gap in MDS patients with + 8 abnormality was smaller than in those with 5q -. In MDS BM, clonal cells were detected in segmented granulocytes (mean 45.9%), orthochromatic normoblasts (mean 46.0%) and mature megakaryocytes (mean 38.0%). In Addition, an approximate amount of clonal cells with the same karyotype abnormality in BM were detected in MDS PB (mean 37.3% in blood vs 48.6% in marrow). Functional analysis showed that the neutrophils in MDS PB could exert nearly normal physiological functions (P > 0.05), but those from AML could not as compared to healthy donors (P < 0.01). CONCLUSION: There is a significant difference in the biological features between MDS and AML clonal cells.
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Leucemia Mieloide Aguda/patología , Síndromes Mielodisplásicos/patología , Adulto , Anciano , Anciano de 80 o más Años , Células de la Médula Ósea/patología , Diferenciación Celular , Células Clonales , Femenino , Humanos , Cariotipificación , Masculino , Persona de Mediana EdadRESUMEN
Studies were performed to obtain evidence of the differentiating and hematopoietic potential of marrow clone-original cells in patients with myelodysplastic syndromes (MDS). First, results of correlation analysis between bone marrow clonal cells and blast percentages for a total of 60 MDS showed that almost all cases had higher clonal (mean 50.1%) than blast proportion (mean 7.0%) (p < 0.001). In contrast, in 16 AML patients, mean clone/blasts disparity was nearly zero. Secondly, the amount of clone-original individual cells were defined in mature hematopoietic cells, mean 47.9% in segmented granulocytes, 47.1% in orthochromatic normoblasts and 37.8% in mature megakaryocytes. In addition, FISH examination showed approximately similar proportions of clonal cells in peripheral blood and marrow for all 22 tested cases (mean 39.1% in blood vs 49.8% in marrow). Moreover, the neutrophils in MDS peripheral blood oxidized dihydrorhodamine123 (DHR) nearly the same as neutrophils in a normal donor's circulation, while obviously poor function was observed in neutrophils from AML blood. Of note, research on clonality had an unexpected outcome as most of the typical morphological dysplastic cells possessed normal karyotypes. These findings lead to the proposal that the biological features of MDS clones are distinctive from those of AML clones.
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Diferenciación Celular , Síndromes Mielodisplásicos/patología , Enfermedad Aguda , Adulto , Anciano , Anciano de 80 o más Años , Crisis Blástica/patología , Células Sanguíneas , Células de la Médula Ósea , Células Clonales/patología , Femenino , Hematopoyesis , Humanos , Cariotipificación , Leucemia Mieloide/genética , Leucemia Mieloide/patología , Masculino , Persona de Mediana Edad , Síndromes Mielodisplásicos/genética , Neutrófilos/metabolismoRESUMEN
To determine the expression of Wilms' tumor gene (WT1) in clonal hematopoietic cells in patients with myelodysplastic syndromes (MDS), immunochemistry labelling (alkaline phosphatase anti-alkaline phosphatase) and fluorescence in situ hybridization (FISH) were coperformed on bone marrow cytospins from 18 patients whose abnormal karyotypes had been determined by G-binding analysis. Compared to 12 healthy donors, WT1 positive nucleated cells in MDS marrows were significantly higher (t = 2.30; P = 0.032). Moreover, WT1 was expressed predominantly in MDS clonal cells (with abnormal FISH signals) rather than in non-clonal cells (residual normal cells) (t = 2.19; P = 0.043).
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Síndromes Mielodisplásicos/genética , Proteínas WT1/genética , Adulto , Anciano , Anciano de 80 o más Años , Anemia Refractaria/genética , Anemia Refractaria/metabolismo , Anemia Refractaria con Exceso de Blastos , Estudios de Casos y Controles , Aberraciones Cromosómicas , Células Clonales , Femenino , Expresión Génica , Sistema Hematopoyético/metabolismo , Humanos , Hibridación Fluorescente in Situ , Cariotipificación , Leucemia Mielógena Crónica BCR-ABL Positiva/genética , Leucemia Mielógena Crónica BCR-ABL Positiva/metabolismo , Masculino , Persona de Mediana Edad , Síndromes Mielodisplásicos/metabolismo , Proteínas WT1/metabolismoRESUMEN
Some of the most prominent "neutral losses" in peptide ion fragmentation are the loss of ammonia and water from N-terminal glutamine. These processes are studied by electrospray ionization mass spectrometry in singly- and doubly-protonated peptide ions undergoing collision-induced dissociation in a triple quadrupole and in an ion trap instrument. For this study, four sets of peptides were synthesized: (1) QLLLPLLLK and similar peptides with K replaced by R, H, or L, and Q replaced by a number of amino acids, (2) QLnK (n = 0, 1, 3, 5, 7, 9, 11), (3) QLnR (n = 0, 1, 3, 5, 7, 9), and (4) QLn (n = 1, 2, 3, 4, 8). The results for QLLLPLLLK and QLLLPLLLR show that the singly protonated ions undergo loss of ammonia and to a smaller extent loss of water, whereas the doubly protonated ions undergo predominant loss of water. The fast fragmentation next to P (forming the y5 ion) occurs to a larger extent than the neutral losses from the singly protonated ions but much less than the water loss from the doubly protonated ions. The results from these and other peptides show that, in general, when N-terminal glutamine peptides have no "mobile protons", that is, the number of charges on the peptide is no greater than the number of basic amino acids (K, R, H), deamination is the predominant neutral loss fragmentation, but when mobile protons are present the predominant process is the loss of water. Both of these processes are faster than backbone fragmentation at the proline. These results are rationalized on the basis of resonance stabilization of the two types of five-membered ring products that would be formed in the neutral loss processes; the singly protonated ion yields the more stable neutral pyrrolidinone ring whereas the doubly protonated ion yields the protonated aminopyrroline ring (see Schemes). The generality of these trends is confirmed by analyzing an MS/MS spectra library of peptides derived from tryptic digests of yeast. In the absence of mobile protons, glutamine deamination is the most rapid neutral loss process. For peptides with mobile protons, dehydration from glutamine is far more rapid than from any other amino acid. Most strikingly, end terminal glutamine is by far the most labile source of neutral loss in excess-proton peptides, but not highly exceptional when mobile protons are not available. In addition, rates of deamination are faster in lysine versus arginine C-terminus peptides and 20 times faster in positively charged than negatively charged peptides, demonstrating that these formal neutral loss reactions are not "neutral reactions" but depend on charge state and stability.
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Desecación , Glutamina/química , Péptidos/química , Protones , Espectrometría de Masa por Ionización de Electrospray/métodos , Secuencia de Aminoácidos , Desaminación , Datos de Secuencia Molecular , Péptidos/síntesis química , Agua/químicaAsunto(s)
Antineoplásicos/uso terapéutico , Leucemia Mielógena Crónica BCR-ABL Positiva/tratamiento farmacológico , Trióxido de Arsénico , Arsenicales/uso terapéutico , Azacitidina/uso terapéutico , Benzamidas , Vacunas contra el Cáncer/uso terapéutico , Humanos , Mesilato de Imatinib , Óxidos/uso terapéutico , Piperazinas/uso terapéutico , Pirimidinas/uso terapéutico , Factor de Necrosis Tumoral alfa/uso terapéuticoRESUMEN
OBJECTIVE: To investigate the clonal origin of the apoptotic hematopoietic cells in myelodysplastic syndromes (MDS). METHODS: Nineteen patients with MDS whose karyotypes had been analysed with G- or R-binding were studied. ISEL (DNA in situ end labelling) and FISH (fluorescence in situ hybridization) (simplified as ISEL/FISH) were performed simultaneously in a single nucleated cells on cytospin slides in 10/19 of the patients. The ratio of FISH positive cells to all nucleated cells and to apoptotic cells were counted. Eight slides from 4 normal donors induced apoptosis by Fas monoclonal antibody (McAb) were used as methodology control. For the rest 9 patients, different fractions of cells (PI-, Annexin V+PI-, Annexin V-PI-) were sorted by flow cytometry and then cytospined for FISH analysis (simplified as FCM/FISH). RESULTS: The mean percentage of abnormal clonal cells in nuclear cells in the 10 cases detected by ISEL/FISH was 37.1%, whereas it was 24.0% in apoptotic cells. Induction by Fas McAb didn't lead to change of karyotype. Moreover, 8/9 patients analysed by FCM/FISH showed higher cell proportion with normal karyotypes in apoptosis-undergoing cells than that in non-apoptotic cells. CONCLUSION: Over apoptotic hematopoietic cells in MDS are mainly residual normal hematopoietic cells.
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Apoptosis , Células de la Médula Ósea/patología , Síndromes Mielodisplásicos/patología , Adulto , Anciano , Anciano de 80 o más Años , Células de la Médula Ósea/metabolismo , Células Cultivadas , Células Clonales/metabolismo , Células Clonales/patología , Femenino , Citometría de Flujo , Humanos , Hibridación Fluorescente in Situ , Etiquetado Corte-Fin in Situ , Cariotipificación , Masculino , Persona de Mediana Edad , Síndromes Mielodisplásicos/genética , Síndromes Mielodisplásicos/metabolismo , Receptor fas/metabolismoRESUMEN
Apart from their role as antigen presenting cells, human peripheral blood monocyte and CD34+ cell-derived dendritic cells (DC), have been demonstrated to exert cytotoxicity against some tumor cells, and their tumoricidal activity can be enhanced by some stimili. However, there have been no reports concerning the tumoricidal activity of human cord blood dendritic cells (CBDC). In this article, we report that human cord blood monocyte-derived DC acquire the ability to kill hematological tumor cells, after activation with lipopolysaccharide (LPS) or gamma-interferon (IFN-gamma), associated with the enhanced TNF-alpha-related apoptosis-inducing ligand (TRAIL) expression in CBDC cytoplasm. The CD14-positive cells collected from cord blood were induced to CBDC in vitro. After activation with IFN-gamma for 12 h, CBDC exhibited cytotoxicity against HL60 and Jurkat cells, while activation with LPS induced cytotoxicity against Daudi and Jurkat cells. However, both LPS- and IFN-gamma-stimulated CBDC showed no cytotoxic activity against normal CD14-negative cord blood mononuclear cells. The formation of umbilical cord hematopoietic progenitor colonies, identified as burst-forming unit-erythroid and colony-forming unit granulocyte-macrophage, was not inhibited by stimulated or unstimulated CBDC. IFN-gamma or LPS stimulation enhanced intracellular but not cellular surface TRAIL, and neither intracellular nor cellular surface tumor necrosing factor-alpha and Fas Ligand as analyzed by flow cytometry. Our results show that activated CBDC can serve as cytotoxic cells against hematological tumor cells without damaging the normal hematopoietic progenitor cells.
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Citotoxicidad Inmunológica , Células Dendríticas/inmunología , Neoplasias/inmunología , Antígenos CD34/metabolismo , Apoptosis , Proteínas Reguladoras de la Apoptosis , Cápsulas Bacterianas , Ensayo de Unidades Formadoras de Colonias , Sangre Fetal , Células HL-60 , Humanos , Interferón gamma/farmacología , Células Jurkat , Receptores de Lipopolisacáridos/metabolismo , Glicoproteínas de Membrana/metabolismo , Polisacáridos Bacterianos/farmacología , Ligando Inductor de Apoptosis Relacionado con TNF , Células Tumorales Cultivadas , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
To investigate the tumoricidal activity of dendritic cell (DC) stimulated by interferon-gamma (IFN-gamma) against freshly isolated myeloid leukemia cells and its mechanism, the peripheral blood monocytes collected from healthy donors were cocultured with interleukin-4 and granulocyte-macrophage colony-stimulating factor in medium to induce DC for 7 days. After 12 hour culture in the absence or presence of IFN-gamma, the changes of costimulatory molecules were analyzed with flow cytometry. To assay the cytotoxicity of DC against freshly isolated acute myeloid cells, they were cocultured at various effector-to-target ratio for 18 hours, then the percentage of tumoricidal activity was measured with (51)Cr release assay. To explore the mechanism of DC-mediated cytotoxicity, the change of DC surface or intracellular protein expression of Fas ligand (Fas L), TNF-alpha and TNF related apoptosis-inducing ligand (TRAIL) were analyzed with flow cytometry. The results showed that IFN-gamma enhanced cytotoxicity of DC against AML cells was (33.8 +/- 1.6)% at E:T as 20:1, compared with unstimulated DC (P < 0.05); IFN-gamma up-regulated expression of costimulatory molecules of DC surface such as CD86 and CD83; after stimulation with IFN-gamma, expression of intracellular TRAIL of DC was significantly enhanced, but expression of TRAIL on cell surface of DC was low; while the significant changes of Fas L and TNF-alpha expression neither on cell surface or in cells were not observed before or after stimulation with IFN-gamma. It is concluded that DC stimulated by IFN-gamma exhibit tumoricidal activity against AML cells. The cytotoxicity is partially related to maturation of DC and TRAIL inducing apoptosis, but not associated with death domain-independent mechanism of Fas L and TNF-alpha.
Asunto(s)
Citotoxicidad Inmunológica/efectos de los fármacos , Células Dendríticas/efectos de los fármacos , Interferón gamma/farmacología , Enfermedad Aguda , Antígenos CD/análisis , Antígeno B7-2/análisis , Técnicas de Cocultivo , Citotoxicidad Inmunológica/inmunología , Células Dendríticas/inmunología , Células Dendríticas/metabolismo , Proteína Ligando Fas/análisis , Citometría de Flujo , Humanos , Inmunoglobulinas/análisis , Leucemia Mieloide/inmunología , Leucemia Mieloide/patología , Glicoproteínas de Membrana/análisis , Ligando Inductor de Apoptosis Relacionado con TNF/análisis , Células Tumorales Cultivadas , Factor de Necrosis Tumoral alfa/análisis , Antígeno CD83RESUMEN
In order to investigate simultaneously the megakaryocytopoiesis and apoptotic characteristics in bone marrow in patients with myelodysplastic syndromes (MDS), we used CD41 immunoenzyme (alkaline phosphatase anti-alkaline phosphatase) and DNA in situ end-labeling techniques on plastic embedded bone marrow biopsy sections of 29 MDS patients. Fourteen patients with iron deficiency anemia served as controls. The results showed that CD41-positive cells in MDS marrow numbered 26.2 +/- 18.2/mm2 (mean +/- standard deviation) compared with 15.6 +/- 7.1/mm2 in controls (P < 0.05). Numbers of cells with the morphology of micro-megakaryocytes in MDS marrow were significantly higher than in controls (P < 0.01). Furthermore, megakaryocytes in MDS marrow were frequently distributed along trabeculae (in 27 cases) and formed clusters (in 25 cases). Apoptotic megakaryocytes in MDS marrow accounted for just 4.4 and 9.3% of all CD41-positive cells and all apoptotic cells, respectively (P > 0.05 compared with controls), but apoptosis occurred only in micro-megakaryocytes. Based on these observations, we conclude that megakaryocytosis and dysmegakaryocytosis are the features of dyshematopoiesis in MDS marrow. Decreased thrombocyte production and thrombocyte release coming from increased dys(micro)megakaryocytes and abnormally located megakaryocytes perhaps play a more important role in peripheral thrombocytopenia than megakaryocytic apoptosis itself. Apoptosis of micro-megakaryocytes may be a protective biological mechanism to remove useless megakaryocytes.
Asunto(s)
Apoptosis , Megacariocitos/patología , Síndromes Mielodisplásicos/patología , Trombopoyesis , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Médula Ósea/patología , Células de la Médula Ósea/patología , Niño , ADN/genética , Femenino , Humanos , Técnicas para Inmunoenzimas , Etiquetado Corte-Fin in Situ , Masculino , Megacariocitos/metabolismo , Persona de Mediana Edad , Glicoproteína IIb de Membrana Plaquetaria/biosíntesisRESUMEN
OBJECTIVE: To investigate the biological changes in myelodysplastic syndromes (MDS) myeloid blast cell line MDS-L after different duration and concentration of As2O3/TRAIL (TNF related apoptosis inducing ligand) treatment. METHODS: MDS-L cells were treated with As2O3 and TRAIL at 9 different concentrations and the treated cells were detected at 24 h, 48 h and 72 h for biologic indexes. The same detections were conducted in untreated MDS-L cells and normal and MDS marrow cells as controls. Apoptosis was assayed by flow cytometry after Annexin V-FITC labelling. Differentiation-induction effect of these drugs on the cells were detected by morphologic examination and CD34(+) proportion analysis after 24 hours treatment and further agar culture for 18 days; P15(ink4b) mRNA expression were detected by RT-PCR and its protein expression by DAB immunocytochemistry, P15(ink4b) DNA methylation by methylation specific PCR (Msp). RESULTS: As2O3/TRAIL treatment promoted MDS-L cells to undergo apoptosis and As2O3 plus TRAIL showed obvious differentiation-induction effect on MDS-L. P15(ink4b) mRNA expression was upregulated in MDS-L cell line after different drug treatment but almost no protein expression increased. Increased P15 expression seemed to be related with DNA demethylation effect of these drugs. CONCLUSIONS: As2O3 or/and TRAIL treatment could promote apoptosis of the clonal cells and induce incomplete cell differentiation. The drugs treatment could also increase P15(ink4b) expression in MDS-L cell line through their demethylation effects.
Asunto(s)
Apoptosis/efectos de los fármacos , Arsenicales/farmacología , Diferenciación Celular/efectos de los fármacos , Óxidos/farmacología , Ligando Inductor de Apoptosis Relacionado con TNF/farmacología , Antígenos CD34/análisis , Trióxido de Arsénico , Línea Celular , Inhibidor p15 de las Quinasas Dependientes de la Ciclina/genética , Inhibidor p15 de las Quinasas Dependientes de la Ciclina/metabolismo , Relación Dosis-Respuesta a Droga , Citometría de Flujo , Humanos , Inmunohistoquímica , Síndromes Mielodisplásicos/genética , Síndromes Mielodisplásicos/metabolismo , Síndromes Mielodisplásicos/patología , ARN Mensajero/genética , ARN Mensajero/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Factores de TiempoRESUMEN
OBJECTIVE: To investigate the apoptotic situation of CD(34) positive cells in myelodysplastic syndromes (MDS). METHOD: In 36 MDS patients, immunocytochemical technique was used for the detection of the expression of CD(34) antigen and DNA in situ end labelling (ISEL) (fluorescein) for the apoptotic signals. Fourteen cases of iron deficiency anemias (IDA) were used as controls. RESULTS: (1) CD(34) expression in MDS group was much higher than that in controls (49.2 +/- 38.5 vs 10.2 +/- 9.7, P < 0.01), and MDS cases had an obviously higher apoptotic rate than control did (69.1 +/- 28.2 vs 17.8 +/- 11.2, P < 0.01). (2) Expression of CD(34) was higher in transforming group (P < 0.05) than in non-transforming and post-transforming groups. Apoptotic rates in both non-transforming/transforming group were higher than in post-transforming group (P < 0.02 and < 0.05 respectively). (3) No apoptosis was found in CD(34) positive cells in MDS; (4) Both CD(34) positive cells and apoptotic cells formed into small or large clusters but did not co-distributed in a given area. CONCLUSION: There is overexpression of CD(34) antigen on hematopoietic cells in MDS. High CD(34) expression accompanied high apoptosis coexisted in the process of transformation from MDS to AML. Apoptosis-resistance of these CD(34) positive cells suggested that they came from malignant hematopoietic cell clones.