RESUMO
We report on characteristics of the first human cell line, PC-MDS, derived from a bone marrow of a patient with therapy-related myelodysplastic syndrome (t-MDS) who had no overt post-MDS leukemia. Classic cytology analyses, immunophenotyping, cytogenetic and molecular genetic procedures were used for characterization of the cell line. PC-MDS cells are positive for the expression of CD13, CD15, CD30, CD33, and CD45 antigen. Positive cytochemical staining and immunophenotype analyses indicated that PC-MDS cells have some characteristics of the early myeloid precursor cell. The karyotype analysis of PC-MDS cell line revealed various numerical and structural changes including those typically associated with t-MDS: del(5)(q13)[7], der(5)t(5;11)(p11;q11)[13], -7[6], del(7)(q31)[2], +20[3], -20[4]. Evaluation of methylation status in a promoter region of p15, p16 and MGMT genes showed biallelic hypermethylation pattern of 5' promoter region only in MGMT gene. PC-MDS is the first t-MDS derived cell line, and based on its immunological, cytogenetic and molecular characterization could be a new tool in evaluation of complex biology of MDS and a model for methylation studies.
Assuntos
Medula Óssea/patologia , Síndromes Mielodisplásicas/patologia , Células Progenitoras Mieloides/patologia , Adulto , Antineoplásicos/uso terapêutico , Inibidor de Quinase Dependente de Ciclina p15/genética , Inibidor de Quinase Dependente de Ciclina p15/metabolismo , Metilação de DNA , Metilases de Modificação do DNA/genética , Enzimas Reparadoras do DNA/genética , Genes Codificadores dos Receptores de Linfócitos T/genética , Genes p16 , Humanos , Região Variável de Imunoglobulina/genética , Imunofenotipagem , Hibridização in Situ Fluorescente , Cariotipagem , Masculino , Reação em Cadeia da Polimerase , Células Tumorais Cultivadas , Proteínas Supressoras de Tumor/genéticaRESUMO
TNF-alpha is a pleiotropic cytokine produced by activated T-cytotoxic lymphocytes and NK cells that is involved in signal transduction after interacting with the appropriate cell surface receptors. The modulation of signals by TNF-alpha receptor super-family is involved in the regulation of cell activation, proliferation, differentiation and control of the cell survival including cell death by apoptosis and necrosis. We have monitored the kinetics of apoptosis/necrosis on PC cells, after TNF-alpha exposure of pre-treated cells to anti-CD95 and anti-CD45 monoclonal antibodies. The results showed that in comparison with untreated cells, TNF-alpha, after 6-24 h of incubation significantly increased apoptosis and necrosis in PC cells. These effects were significantly different in comparison to both untreated cells and cells pre-treated with anti-CD45 monoclonal antibodies. However, TNF-alpha on PC cells pre-treated with anti-CD95 monoclonal antibody significantly decreased apoptotic and necrotic form of cell death. We concluded that anti-CD45 and CD95 monoclonal antibodies modulates the effect of TNF-alpha on this cell line in vitro, and that these molecules participate in TNF-alpha cytotoxic response.
Assuntos
Antineoplásicos/imunologia , Antineoplásicos/farmacologia , Células da Medula Óssea/patologia , Antígenos Comuns de Leucócito/imunologia , Síndromes Mielodisplásicas/patologia , Fator de Necrose Tumoral alfa/imunologia , Fator de Necrose Tumoral alfa/farmacologia , Receptor fas/imunologia , Anticorpos Monoclonais/imunologia , Apoptose , Humanos , NecroseRESUMO
The platinum (II)complexes, cis-[PtCl(2)(CH(3)SCH(2)CH(2)SCH(3))] (Pt1), cis-[PtCl(2)(dmso)(2)] (dmso is dimethylsulfoxide; Pt2) and cis-[PtCl(2)(NH(3))(2)] (cisplatin), and taxol (T) have been tested at different equimolar concentrations. Cells were exposed to complexes for 2 h and left to recover in fresh medium for 24, 48 or 72 h. Growth inhibition was measured by tetrazolium WST1 assay Analyses of the cell cycle, and apoptosis were performed by flow cytometry, at the same exposure times. The IC50 value of each platinum(II) complex as well as combination index (CI; platinum(II) complex + taxol) for various cytotoxicity levels were determined by median effects analysis.MCF7 cells were found to be sensitive to both Pt1 and Pt2 complexe These cisplatin analogues influenced the cell growth more effectively as compared to cisplatin. Cytotoxic effect was concentration and time-dependent. Profound growth inhibitory effect was observed for Pt1 complex, across all its concentrations at all recovery periods. A plateau effect was achieved three days after treatment at Pt1 concentrations = 1 muM. Pt2, however, decreased MCF7 cells survival only for the first 24 h ranging between 50-55%. Pt2 cytotoxicity sharply decreased thereafter, approaching 2 h - treatment cytotoxicity level. The median IC50 values for Pt1 and Pt2 were similar (0.337 and 0.3051 muM, respectively) but only for the first 24 h. The IC50 values for Pt1 strongly depend on the recovery period. On simultaneos exposure of cells to taxol and platinum(II) complexes no consistent effect was found. The Cls for combinations of taxol with Pt1 or Pt2 revealed cytotoxic effects that were in most Cases synergistic (Pt1) or less than addtiive (Pt2). Flow cytometry analysis has shown that each platinum(II) complex induced apoptosis in MCF7 cells. The level of apoptosis correlated with cytotoxicity level for the range concentrations. Both cisplatin analogues, at IC50 concentrations, increased the number of MCF7 cells in G0G1 phase of cell cycle. Pt2-treated cells remained arrested in G0G1 phase up to 72 h after treatment. Combination of Pt2 and taxol caused further arrest of cells in G0G1 phase (24 h) in parallel with strong decrement of G2M phase cells.