RESUMO
Post-graft hematopoiesis is characterized by long-term quantitative deficiency in marrow progenitor cells in both autologous and allogenic settings. In order to evaluate the function of post-graft progenitor cells, the proliferative capacity of marrow CD34(+) cells was evaluated in 10 patients 6 months after autologous bone marrow transplantation (ABMT) for non-Hodgkin's lymphoma and compared to that of 10 patients before ABMT and 10 normal controls. Immuno-selected CD34(+) cells were cultured for 7 days in liquid serum-free medium with a combination of early-acting GF consisting of stem cell factor, IL-3 and IL-1beta. Clonogenic efficiency of unselected cells for CFU-GM and BFU-E was decreased in post-graft patients compared to pre-graft and control patients. However, clonogenic efficiency of selected CD34(+) cells for CFU-GM was not different in post-graft, pre-graft and control patients but BFU-E values of post-graft patients remained lower than those of control patients. Decreased percentages of CD34(+) CD38(-) cells were observed in both post-graft and pre-graft patients while those of CD34(+) c-kit(+) cells were similar in all three patient groups. After 7-day liquid culture, expansion yields of total progenitor cells were significantly lower in post-graft patients (147 +/- 28%) than in pre-graft (255 +/- 27%) and control patients (246 +/- 23%). Post-graft deficiency in progenitor cell expansion was particularly marked for BFU-E (61 +/- 24%) compared to pre-graft patients (220 +/- 82%) and to controls (349 +/- 82%). These results indicate impaired proliferative potential of marrow CD34(+) cells several months after ABMT involving erythroid progenitor cells and/or commitment towards erythroid lineage from a more immature stage (pre-CFU).
Assuntos
Transplante de Medula Óssea , Hematopoese/efeitos dos fármacos , Células-Tronco Hematopoéticas/efeitos dos fármacos , Interleucina-1/farmacologia , Interleucina-3/farmacologia , Fator de Células-Tronco/farmacologia , Adulto , Antígenos CD34/análise , Divisão Celular/efeitos dos fármacos , Células Cultivadas/efeitos dos fármacos , Ensaio de Unidades Formadoras de Colônias , Meios de Cultura Livres de Soro , Células Precursoras Eritroides/efeitos dos fármacos , Células Precursoras Eritroides/patologia , Feminino , Células-Tronco Hematopoéticas/patologia , Humanos , Imunofenotipagem , Linfoma não Hodgkin/terapia , Masculino , Pessoa de Meia-Idade , Proteínas Proto-Oncogênicas c-kit/análise , Transplante AutólogoRESUMO
Marrow stromal cells were evaluated several months after autologous BMT for their capacity to support both normal hemopoiesis and secrete the main growth factors involved in its control, G-CSF, GM-CSF, IL-3 and SCF. Stromal layers (SL) were obtained by long-term marrow cultures (LTMC) established from 15 patients (9 with hematologic malignancies and 6 with solid tumors) 3 months after autologous BMT and were compared to pre-graft patients. After irradiation, both post-graft and pre-graft SL were recharged with the same inoculum of normal marrow cells. As compared to pre-graft values, CFU-GM production on post-graft SL was significantly increased during the first 2 weeks of culture whereas it was decreased from week 3 to week 8. These findings were only observed in patients with hematologic malignancies and not in patients with solid tumors. Growth factor secretion was evaluated by ELISA in the supernatants of unstimulated and IL-1-stimulated SL from 10 post-graft patients, 13 pre-graft patients and 5 normal controls. In any group of patients, IL-3 was undetectable either spontaneously or after IL-1-stimulation. As compared to controls, secretion by IL-1-stimulated SL was not different for GM-CSF in pre- and post-graft patients but tended to be decreased for G-CSF in post-graft patients. SCF secretion, which was not induced by IL-1, appeared dramatically decreased in both pre- and post-graft patients. The capacity of post-graft SL to support CFU-GM growth in LTMC was correlated at week 1 with G-CSF secretion and from week 3 to week 8 with SCF secretion. These results suggest that microenvironment remains qualitatively damaged several months after BMT involving a decreased capacity both to support early hemopoiesis and to secrete SCF, particularly in patients grafted for hemopoietic malignancies.
Assuntos
Células da Medula Óssea/patologia , Transplante de Medula Óssea/patologia , Fatores de Crescimento de Células Hematopoéticas/metabolismo , Células Estromais/patologia , Adulto , Células da Medula Óssea/metabolismo , Técnicas de Cultura de Células/métodos , Células Cultivadas , Criança , Ensaio de Unidades Formadoras de Colônias , Seguimentos , Fator Estimulador de Colônias de Granulócitos/metabolismo , Fator Estimulador de Colônias de Granulócitos e Macrófagos/metabolismo , Neoplasias Hematológicas/patologia , Neoplasias Hematológicas/terapia , Fatores de Crescimento de Células Hematopoéticas/deficiência , Humanos , Interleucina-1/farmacologia , Interleucina-3/metabolismo , Pessoa de Meia-Idade , Neoplasias/patologia , Neoplasias/terapia , Fator de Células-Tronco/metabolismo , Células Estromais/efeitos dos fármacos , Células Estromais/metabolismo , Fatores de Tempo , Condicionamento Pré-TransplanteRESUMO
Marrow stromal cells of patients treated by autologous bone marrow transplantation (ABMT) for malignancies have been assessed for their ability to secrete granulocyte colony-stimulating factor (G-CSF), granulocyte-macrophage colony-stimulating factor (GM-CSF), stem cell factor (SCF), leukemia inhibitory factor (LIF), interleukin-6 (IL-6), transforming growth factor beta1 (TGFbeta1) and macrophage inflammatory protein-1alpha. (MIP-1alpha). Long-term marrow cultures were established from 10 patients prior to and 3 months after ABMT, from 7 patients 1 yr after ABMT and from 11 controls. Cytokines in culture supernatants of stromal layers (SL) were evaluated by enzyme-linked immunosorbent assay (ELISA). Significant differences between patient groups and controls were apparent in baseline production of GM-CSF, SCF, MIP-1alpha and TGFbeta1. After IL-1beta addition in cultures, G-CSF production was reduced in pretransplant and post-transplant patients compared to controls. The production of TGFbeta1, LIF, IL-6 and more particularly SCF were reduced in post-transplant patients, while elevated levels of GM-CSF and MIP-1alpha were observed in these patients only when the values were corrected for the number of cells growing in the SL. These results indicate a prolonged stromal defect in growth factor production following ABMT for the early-stage acting cytokines IL-6, LIF and SCF as well as for G-CSF, but not for GM-CSF, while the production of the 2 inhibitors shows different pathways.
Assuntos
Transplante de Medula Óssea/imunologia , Citocinas/metabolismo , Leucemia Linfocítica Crônica de Células B/imunologia , Linfoma não Hodgkin/imunologia , Adulto , Células da Medula Óssea/metabolismo , Células da Medula Óssea/patologia , Transplante de Medula Óssea/patologia , Contagem de Células , Quimiocina CCL3 , Quimiocina CCL4 , Feminino , Fator Estimulador de Colônias de Granulócitos/metabolismo , Fator Estimulador de Colônias de Granulócitos e Macrófagos/metabolismo , Inibidores do Crescimento/metabolismo , Humanos , Interleucina-6/metabolismo , Fator Inibidor de Leucemia , Leucemia Linfocítica Crônica de Células B/metabolismo , Leucemia Linfocítica Crônica de Células B/patologia , Linfocinas/metabolismo , Linfoma não Hodgkin/metabolismo , Linfoma não Hodgkin/patologia , Proteínas Inflamatórias de Macrófagos/metabolismo , Masculino , Pessoa de Meia-Idade , Fator de Células-Tronco/metabolismo , Células-Tronco/patologia , Células Estromais/imunologia , Células Estromais/metabolismo , Células Estromais/patologia , Fator de Crescimento Transformador beta/metabolismo , Células Tumorais CultivadasRESUMO
We assessed the effect of antitumoural therapy at intensive doses on the haemopoietic system using long-term marrow cultures (LTMC) established from 33 patients (25 with haematological diseases and eight with solid tumours) after autologous bone marrow transplantation (ABMT). When compared to 42 pre-graft patients, a decreased CFU-GM production and a defect in stromal layer (SL) confluence were found after ABMT on day 90 but also on day 365. However, these abnormalities were observed only in patients with haematological diseases and no differences between pre-graft and post-graft results were found in patients with solid tumours. Among the patients with haematological diseases, on day 90 those with acute lymphoid leukaemias showed lower CFU-GM production whereas patients with non-Hodgkin's lymphomas developed more frequently subconfluent or confluent SL. Other factors studied such as sex, patient age, disease status, marrow purging and post-graft administration of growth factors did not appear to influence post-graft LTMC results. Multivariate analysis including all the patients has shown (a) that solid tumours were associated with higher CFU-GM production, and (b) that conditioning regimens with total body irradiation (TBI) or busulfan led more frequently to non-confluent SL. In conclusion, high-dose therapy followed by ABMT can induce a persistent impairment of the stem cell and stromal cell compartments, particularly in patients with haematological diseases conditioned with TBI, despite the absence of any alloimmune reaction and post-graft immunosuppressive therapy.
Assuntos
Antineoplásicos/efeitos adversos , Medula Óssea/efeitos dos fármacos , Hematopoese/efeitos dos fármacos , Leucemia/terapia , Linfoma/terapia , Neoplasias/terapia , Adolescente , Adulto , Medula Óssea/patologia , Transplante de Medula Óssea , Criança , Pré-Escolar , Ensaio de Unidades Formadoras de Colônias , Técnicas de Cultura , Feminino , Humanos , Leucemia/fisiopatologia , Linfoma/fisiopatologia , Masculino , Pessoa de Meia-Idade , Neoplasias/fisiopatologia , Fatores de Tempo , Transplante Autólogo , Irradiação Corporal Total/efeitos adversosRESUMO
Hematopoietic reconstitution has been studied in 180 patients after autologous bone marrow transplantation based on peripheral blood cell (PBC) recovery time and marrow progenitor counts sequentially tested for up to 4 years. Several factors that could influence hematopoietic reconstitution have been analyzed including sex, age, diagnosis, disease status, conditioning regimen, graft progenitor content, graft in vitro purging, and postgrafting administration of growth factors. Before transplantation, marrow progenitor values were normal only for colony-forming unit granulocyte macrophage (CFU-GM) in contrast to colony-forming unit-erythroid (CFU-E), burst-forming unit-erythroid (BFU-E), and colony-forming unit-megakaryocyte (CFU-Meg). After transplantation, as described with allogenic grafts, these values remained low for several years, although PBC counts were nearly normalized within a few weeks. Pregraft values were reached after 2 years for CFU-GM and BFU-E, and after 4 years for CFU-E, while CFU-Meg failed to reach pregraft values after this time. Normal levels were reached after 4 years only by CFU-GM. On univariate and multivariate analysis, the following factors appeared to delay both PBC and marrow progenitor reconstitution: underlying disease (particularly acute myeloid leukemias), graft characteristics such as low stem cell content and in vitro purging, conditioning regimens with total body irradiation or busulfan, and lack of postgraft administration of growth factors. In conclusion, high-dose therapy followed by bone marrow transplantation induces a deep and prolonged impairment of hematopoiesis irrespective of any alloimmune reaction or postgraft immunosuppressive therapy.(ABSTRACT TRUNCATED AT 250 WORDS)