Transfection of the c-myc oncogene into normal Epstein-Barr virus-harboring B cells results in new phenotypic and functional features resembling those of Burkitt lymphoma cells and normal centroblasts.
Activated c-myc gene was introduced into the cells of three normal Epstein-Barr virus (EBV)-positive lymphoblastoid B cell lines (LCL). The cells were monitored for the appearance of new phenotypic and functional features compared with the control LCL cells transfected with plasmid that did not contain the c-myc gene. The LCL-expressing c-myc constitutively did not arrest growth in low serum concentration. However, the cell number in the cultures failed to increase because of substantial cell death. Death was due to apoptosis as demonstrated by flow cytometric analysis of propidium iodide-stained cells, by typical DNA laddering in gel electrophoresis, and by the inspection of Giemsa-stained cell smears. Apoptosis was also induced by exposing the transfected cells to antibodies directed to the immunoglobulin mu chain (a-mu-ab) irrespective of the serum concentration in the culture. Exposure of the cells to CD40 ligand (CD40L) or CD40 monoclonal antibody prevented cell apoptosis. Upon transfection with c-myc, the LCL cells acquired a vacuolated morphology that was never observed in control cells. Moreover, the expression of CD10 and CD38 was upregulated, while that of CD39 and especially CD23 was downregulated. Unlike that observed in certain Burkitt lymphoma (BL) cell lines that share the same surface phenotype (CD10+CD38+CD23-CD39-), the c-myc-transfected cells expressed lymphocyte function-associated (LFA) 1, LFA-3, and intercellular adhesion molecule 1 and grew in large clumps rather than single-cell layers. Expression of CD10 and CD38 was particularly evident on the cells undergoing apoptosis, thus suggesting a correlation between the presence of these markers and the apoptotic process. Cells placed in conditions favoring in vitro apoptosis displayed downregulation of Bcl-2 protein. Bcl-2 expression was, however, upregulated when the cells were exposed to CD40L. These data indicate that the B cells expressing c-myc constitutively acquire some of the features of normal centroblasts and of BL cells, including the expression of CD10 and CD38, and the propensity to undergo apoptosis, which can be prevented by exposure to CD40L. Therefore, these cells can serve as a model system to study both BL lymphomagenesis as well as the process of B cell selection occurring in the germinal centers.
Istituto Nazionale per la Ricerca sul Cancro, IST, Genoa, Italy.
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Cutrona
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Ulivi
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Fais
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Roncella
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Ferrarini
D036541:ADP-ribosyl Cyclase; D051997:ADP-ribosyl Cyclase 1; D015703:Antigens, CD; D000943:Antigens, Differentiation / Q000235:genetics; D017209:Apoptosis / Q000235:genetics; D001402:B-Lymphocytes / Q000473:pathology / Q000821:virology*; D001483:Base Sequence; D002051:Burkitt Lymphoma / Q000235:genetics / Q000276:immunology / Q000473:pathology*; D002454:Cell Differentiation; D017931:DNA Primers; D015536:Down-Regulation; D016259:Genes, myc; D004854:Herpesvirus 4, Human / Q000302:isolation & purification; D006801:Humans; D008562:Membrane Glycoproteins; D008969:Molecular Sequence Data; D015260:Neprilysin / Q000235:genetics; D010641:Phenotype; D011518:Proto-Oncogene Proteins / Q000235:genetics; D019253:Proto-Oncogene Proteins c-bcl-2; D014162:Transfection; D014407:Tumor Cells, Cultured
D015703:Antigens, CD; D000943:Antigens, Differentiation; D017931:DNA Primers; D008562:Membrane Glycoproteins; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D036541:ADP-ribosyl Cyclase; C498154:CD38 protein, human; D051997:ADP-ribosyl Cyclase 1; D015260:Neprilysin