Expression of human fos proto-oncogene (c-fos) was analyzed in primary cells from 50 untreated acute lymphocytic (ALL) and myeloblastic (AML) leukemias. c-fos RNA, analyzed by blot hybridization, was detected virtually only in myelomonocytic (M4) and monocytic (M5) AML. Both M4 and M5 samples show a strong positive correlation between the amount of c-fos transcripts and the percentage of leukemic cells expressing surface antigens specific for mature monocytes and macrophages. Normal mature monocytes exhibit a detectable level of c-fos RNA, which is virtually unaltered on activation to macrophage differentiation, but is always below that observed in M4 through M5 monocyticlike cells. These data provide evidence that c-fos expression is linked to terminal monocyte and macrophage differentiation in normal and leukemic hemopoiesis.

Antigens, Neoplasm↗

PubMed1987

Expression of blood group A antigen during erythroid differentiation in A1 and A2 subjects.

The expression of blood group A antigen on marrow and blood cells from A1 and A2 subjects was investigated by the binding of Helix pomatia and Dolichos biflorus lectins using immunofluorescence. These two lectins stained BFU-E-derived colonies from A subjects in the early days of culture before the expression of glycophorin. The erythroid origin of these cells was ascertained by the coexpression of two other very early erythroid markers. In bone marrow, the ultrastructural immunogold method revealed that the entire erythroid lineage including proerythroblasts was labeled by HPA, whereas no staining was observed on granulomonocytic cells including myeloblasts. Platelets from A subjects were HPA-labeled and so were platelets from an O subject preincubated in A plasma. Megakaryocytes obtained in CFU-MK-derived colonies were weakly and heterogeneously labeled by the HPA lectin. Cultures from A1 and A2 subjects were the reflection of the genetic differences only when investigations were performed on mature erythroblasts. In contrast, the great majority of immature erythroblasts both from A2 and A1 subjects were equally labeled by both lectins; during further erythroid maturation, binding of both lectins markedly diminished only on A2 erythroblasts. When marrow erythroblasts were investigated at electron microscopic level, heterogeneity of labeling among all stages of maturation was clearly observed in A2 subjects, with staining stronger on immature than on mature erythroblasts. Therefore, the genetic differences between A1 and A2 subjects are revealed during terminal erythroid differentiation.

ABO Blood-Group System↗

PubMed1986

Expression of transferrin receptors in phytohemagglutinin-stimulated human T-lymphocytes. Evidence for a three-step model.

Resting human T-lymphocytes show an elevated intracellular concentration of ferritin, whereas transferrin receptors are not detectable. Stimulation by phytohemagglutinin markedly lowers their ferritin content, while inducing the synthesis of transferrin receptors. Addition of iron salts (ferric ammonium citrate) in activated T-lymphocyte cultures causes a marked enhancement of both [3H]uridine and [3H]thymidine incorporation. Nevertheless, it also induces a concentration-dependent decrease in transferrin receptor synthesis, associated with a marked rise of ferritin production. Hemin treatment exerts the same effects. Addition of picolinic acid in phytohemagglutinin-stimulated cultures causes a decrease of [3H]thymidine incorporation, whereas transferrin expression is markedly enhanced. The action of iron salts and chelators is specific for transferrin receptors, since the expression of other membrane markers of activated human T-lymphocytes (interleukin-2 receptor, insulin receptor, and HLA-DR antigen) is not modified by treatment with iron or picolinic acid. These observations suggest that expression of transferrin receptors in activated T-lymphocytes is specifically modulated by their intracellular iron level, rather than their proliferative rate. Addition of picolinic acid to resting T-lymphocytes in the absence of mitogen induces a marked decrease of their ferritin content, but not the appearance of transferrin receptors. On the basis of these results, we suggest a three-step model: (a) in resting T-lymphocytes, the gene for transferrin receptor is apparently "closed," in that it is not expressed under both normal conditions and following iron deprivation. (b) After mitogen stimulus, T-lymphocytes are reprogrammed into cell cycle progression, which necessarily entails synthesis of transferrin receptors (c) Expression of these receptors is modulated by the intracellular iron level, rather than the rate of proliferation per se.

Ferric Compounds↗

PubMed1986

HLA antigens on human hemopoietic progenitors during embryonic-fetal life: expression and in vitro modulation.

We have analyzed the time course of the expression of HLA antigens on hemopoietic progenitors (BFU-E, CFU-E and CFU-GM) from human embryonic-fetal livers (FL). HLA ABC and Ia-like antigens are never detected on progenitor cells at 5-week post-conception. Their expression initiates at 6-week and progressively increases thereafter, up to near-adult levels at 9-week. The time course of HLA antigen expression on erythroid precursors, derived from FL at corresponding gestational ages, parallels that observed on hemopoietic progenitors. Incubation of embryonic progenitors or precursors with medium conditioned by PHA-stimulated adult peripheral blood mononucleated cells induces a marked increase of the expression of both HLA-ABC and Ia-like antigens. Conversely, incubation with recombinant gamma-interferon causes a marked increase of HLA-ABC, but not Ia-like antigen expression, thus in contrast with results observed on adult monocytes.

Antibodies, Monoclonal↗

PubMed1986

Proliferative response and oncogene expression induced by epidermal growth factor in EL2 rat fibroblasts.

Extensive evidence supports a two-step model for the control of fibroblast growth, which includes first the action of a competence factor (e.g., platelet-derived growth factor) followed by the stimulus of a progression factor (e.g., epidermal growth factor [EGF]). We investigated whether this model may be applied to the euploid EL2 fibroblast line recently isolated from rat embryos (E. Liboi, M. Caruso, and C. Basilico, Mol. Cell. Biol. 4:2925-2928, 1984). Our results clearly show that EGF alone leads EL2 cells to proliferate in serum-free conditions at a rate corresponding to 50 to 60% of that observed in the presence of 10% calf serum. It is of interest that, when resting EL2 cells were exposed to EGF, transcription of both c-myc and c-fos was markedly induced. Altogether, these observations suggest that, in contrast with the model of fibroblast growth mentioned above, EL2 cells require the presence of a single growth factor (EGF) for induction of DNA synthesis, and the expression of myc and fos proto-oncogenes may represent an obligatory step in the pathway of commitment of EL2 cells to proliferation. In addition, we showed that EGF may induce EL2 cells to acquire some properties of transformed cells, such as growth in agar and loss of contact inhibition. This suggests that the particular response to EGF of the EL2 line may be strictly connected with the expression of a transformed phenotype.

Animals↗

PubMed1986

Human embryonic hemopoiesis. Kinetics of progenitors and precursors underlying the yolk sac----liver transition.

Human embryonic development involves transition from yolk sac (YS) to liver (L) hemopoiesis. We report the identification of pluripotent, erythroid, and granulo-macrophage progenitors in YS, L, and blood from human embryos. Furthermore, comprehensive studies are presented on the number of hemopoietic progenitors and precursors, as well as of other cell types, in YS, L, and blood at precisely sequential stages in embryos and early fetuses (i.e., at 4.5-8 wk and 9-10 wk postconception, respectively). Our results provide circumstantial support to a monoclonal hypothesis for human embryonic hemopoiesis, based on migration of stem and early progenitor cells from a generation site (YS) to a colonization site (L) via circulating blood. The YS----L transition is associated with development of the differentiation program in proliferating stem cells: their erythroid progeny shows, therefore, parallel switches of multiple parameters, e.g., morphology (megaloblasts----macrocytes) and globin expression (zeta----alpha, epsilon----gamma).

Colony-Forming Units Assay↗

PubMed1986

[Cystosarcoma phyllodes of the breast].

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Adult↗

PubMed1986

Expression of transferrin receptors in human erythroleukemic lines: regulation in the plateau and exponential phase of growth.

The present study was undertaken in an attempt to elucidate the mechanism(s) underlying transferrin (TRF) receptor expression in human erythroleukemic (K562 and HEL) lines during the exponential and the plateau phase of growth. TRF receptor synthesis is enhanced when stationary cells are subcultured at low density in fresh medium. This rise occurs in either the presence or the absence of serum, which is associated with cell proliferation or quiescence, respectively. In the presence of serum, it is not inhibited by the addition of hydroxyurea (i.e., an agent blocking DNA synthesis). Thus, the receptor synthesis is enhanced not only in subcultures of actively proliferating cells (in the presence of serum), but also in subcultures of quiescent elements (in the absence of serum or upon the addition of serum plus hydroxyurea). Conversely, the ferritin content is markedly decreased when stationary cells are subcultured at low density, in either the presence or the absence of serum. These results suggest that stationary cells subcultured in fresh medium undergo a depletion of their intracellular iron pool, which in turn may represent the stimulus triggering TRF receptor synthesis. This hypothesis is supported by two observations: both the depletion of this pool and the rise in TRF receptor synthesis are more marked in the absence than in the presence of serum; addition of excess exogenous iron fully inhibits the rise of TRF receptor synthesis in cells subcultured with fresh medium and serum.

Blood Physiological Phenomena↗

PubMed1986

[2 cases of Mondor's syndrome].

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Adult↗

PubMed1985

Inhibition of transferrin receptor expression by interferon-alpha in human lymphoblastoid cells and mitogen-induced lymphocytes.

125I-Transferrin binding to lymphoblastoid K562 and Daudi cells markedly increased after exposure of the cells to culture conditions that stimulated proliferation. Treatment of these cells with interferon-alpha (IFN-alpha) resulted in concurrent inhibition of cell growth and of the rise in transferrin binding. Scatchard analyses revealed that IFN reduced the number of transferrin receptors without altering the binding constant. When 125I-transferrin binding was measured using permeabilized cells, the IFN-induced reduction of binding was comparable to that observed with intact cells, indicating that IFN diminished the total number of cellular transferrin receptors. We also found that addition of IFN-alpha to phytohemagglutinin-stimulated human lymphocytes inhibited the mitogen-induced enhancement of [3H]thymidine incorporation as well as surface binding of 125I-transferrin. Our findings suggest that the decrease in transferrin receptor expression on IFN-alpha-treated cells may be one of the mechanisms responsible for the antiproliferative action of IFN.

Cell Division↗

PubMed1985

Inhibition of insulin receptor binding by A23187: synergy with phorbol esters.

The ionophore A23187 inhibits the binding of insulin on U-937 monocyte-like cells which had been induced to differentiate by 1 alpha, 25-(OH)2 cholecalciferol, while it remains inactive on the undifferentiated cells. A 50% reduction of the specific binding of 125I-insulin is observed within 20 to 30 min at 37 degrees C. The effect is obtained at 10(-7)M to 10(-6)M A23187. It is reversible in 60 min at 37 degrees C. A suboptimal concentration of the ionophore potentiates the inhibitory action of phorbol esters on insulin binding.

Calcimycin↗

PubMed1985

Molecular mechanisms regulating the synthesis of transferrin receptors and ferritin in human erythroleukemic cell lines.

The effects of either iron salts or iron chelators on the biosynthesis of transferrin receptors in human erythroleukemic lines was investigated. Addition of these compounds induced a rapid and marked decrease (iron salts) or increase (iron chelators) in the level of transferrin receptors synthesis. Both phenomena were inhibited by actinomycin D. In contrast, the increase in the synthesis of ferritin induced in these cells by addition of iron salts was not inhibited by actinomycin D. These results suggest that iron salts modulate the synthesis of transferrin receptors and ferritin via different molecular mechanisms, of transcriptional and translation type, respectively.

Cell Line↗

PubMed1985

Induction of (2'-5') oligoadenylate synthetase activity during granulocyte and monocyte differentiation.

Variations in the (2'-5') oligoadenylate synthetase (2-5 A synthetase) level were examined prior to and during the differentiation in culture of the human monocyte cell line U937 and the promyelocytic cell line HL60 in an attempt to reveal whether the enzyme is actively involved in hematopoietic cell maturation. The basal level of this enzyme was much higher in U937 than in HL60 cells. The activity of 2-5 A synthetase was enhanced in both cell lines in response to alpha, beta interferons. During cell differentiation, ten markers were measured. The level of the enzyme rose during the process of cellular maturation in both cell lines. The 2-5 A synthetase activity observed in differentiated HL60 and U937 cells was comparable to that observed in mature normal granulocytes and monocytes respectively. Induction of U937 differentiation by chemicals was associated with detectable production of IFN. The increase in enzyme activity observed was mostly dependent on endogenous production of interferon, since it was inhibited by interferon antibodies. Kinetic studies showed that in U937 cells 2-5 A synthetase was expressed prior to several of the differentiation markers. The rise in the enzyme's level observed during the differentiation of HL60 cells was independent of endogenous production of interferon, since it was not inhibited by the addition of anti-interferon antibodies. These results suggest that different biochemical and molecular mechanisms are responsible for the induction of 2-5 A synthetase observed during the differentiation of hematopoietic cells. In any case, 2-5 A synthetase can be considered as a biochemical marker of cell status and differentiation in hematopoietic cells.

2',5'-Oligoadenylate Synthetase↗

PubMed1985

Differentiation of U-937 human monocyte-like cell line by 1 alpha,25-dihydroxyvitamin D3 or by retinoic acid. Opposite effects on insulin receptors.

The monocyte-like human cell line U-937 has been differentiated in vitro by incubation with either 1 alpha,25-dihydroxyvitamin D3 or retinoic acid (RA) plus dibutyryl cyclic AMP (db-cAMP). Both methods were effective in inducing the appearance of maturation markers. Their actions on insulin receptors were the opposite, however; 1 alpha,25-dihydroxyvitamin D3 increased the binding of the hormone, while RA plus db-cAMP decreased the binding. These effects were specific for insulin, since the transferrin receptors were reduced by both methods of differentiation. Thus, the changes in insulin receptors during maturation in vitro depend on the inducing agent and are not causally related to the differentiation process.

Bucladesine↗