scholarly journals The growth of Rauscher erythroleukemia cells is mediated by autocrine production of a factor with biological activity similar to interleukin- 3

Blood ◽  
1989 ◽  
Vol 73 (7) ◽  
pp. 1770-1777
Author(s):  
G Migliaccio ◽  
AR Migliaccio ◽  
S Ruscetti ◽  
JW Adamson
Keyword(s):  
Biological Activity ◽  
Growth Factor ◽  
Cell Lines ◽  
Leukemic Cells ◽  
Northern Analysis ◽  
Interleukin 3 ◽  
Gm Csf ◽  
Erythroleukemia Cells ◽  
Defined Culture

Under serum-deprived and chemically defined culture conditions, the growth of Rauscher erythroleukemia cells is mediated by an autocrine mechanism. The growth-promoting activity is produced by fresh or irradiated cells and resembles the activity of interleukin-3 (IL-3) in its ability to sustain colony formation from three of four IL-3- dependent cell lines and to induce formation of granulocyte/macrophage (GM) colonies and, in the presence of erythropoietin (Ep), of erythroid bursts and mixed erythroid colonies. IL-3, IL-1, IL-4, IL-6, GM colony- stimulating factor (GM-CSF), G-CSF, M-CSF, Ep, and media conditioned by concanavalin A-stimulated mouse spleen cells or phytohemagglutinin- stimulated LBRM 33 cells were unable to induce proliferation of the Rauscher erythroleukemia cells. Northern analysis of total and polyA- selected RNA extracted from untreated Rauscher cells or from cells 24 hours after irradiation showed the presence of message for M-CSF but not for IL-3, IL-1, GM-CSF, or G-CSF. The production of IL-6 was excluded by a sensitive bioassay. These results indicate that the autocrine growth of the Rauscher cell line is mediated by a growth factor different from IL-3, but with similar biological activity. Activation of the expression of such a growth factor during viral infection may contribute to the generation of leukemic cells that have the property to grow in vitro and generate Rauscher erythroleukemia cell lines.

scholarly journals The growth of Rauscher erythroleukemia cells is mediated by autocrine production of a factor with biological activity similar to interleukin- 3

Blood ◽  
1989 ◽  
Vol 73 (7) ◽  
pp. 1770-1777 ◽  
Author(s):  
G Migliaccio ◽  
AR Migliaccio ◽  
S Ruscetti ◽  
JW Adamson
Keyword(s):  
Biological Activity ◽  
Growth Factor ◽  
Cell Lines ◽  
Leukemic Cells ◽  
Northern Analysis ◽  
Interleukin 3 ◽  
Gm Csf ◽  
Erythroleukemia Cells ◽  
Defined Culture

Abstract Under serum-deprived and chemically defined culture conditions, the growth of Rauscher erythroleukemia cells is mediated by an autocrine mechanism. The growth-promoting activity is produced by fresh or irradiated cells and resembles the activity of interleukin-3 (IL-3) in its ability to sustain colony formation from three of four IL-3- dependent cell lines and to induce formation of granulocyte/macrophage (GM) colonies and, in the presence of erythropoietin (Ep), of erythroid bursts and mixed erythroid colonies. IL-3, IL-1, IL-4, IL-6, GM colony- stimulating factor (GM-CSF), G-CSF, M-CSF, Ep, and media conditioned by concanavalin A-stimulated mouse spleen cells or phytohemagglutinin- stimulated LBRM 33 cells were unable to induce proliferation of the Rauscher erythroleukemia cells. Northern analysis of total and polyA- selected RNA extracted from untreated Rauscher cells or from cells 24 hours after irradiation showed the presence of message for M-CSF but not for IL-3, IL-1, GM-CSF, or G-CSF. The production of IL-6 was excluded by a sensitive bioassay. These results indicate that the autocrine growth of the Rauscher cell line is mediated by a growth factor different from IL-3, but with similar biological activity. Activation of the expression of such a growth factor during viral infection may contribute to the generation of leukemic cells that have the property to grow in vitro and generate Rauscher erythroleukemia cell lines.

scholarly journals Interleukin-3, GM-CSF, and G-CSF receptor expression on cell lines and primary leukemia cells: receptor heterogeneity and relationship to growth factor responsiveness

Blood ◽  
1989 ◽  
Vol 74 (1) ◽  
pp. 56-65 ◽  
Author(s):  
LS Park ◽  
PE Waldron ◽  
D Friend ◽  
HM Sassenfeld ◽  
V Price ◽  
...  
Keyword(s):  
Growth Factor ◽  
Cell Lines ◽  
Specific Binding ◽  
Absolute Number ◽  
Lymphocytic Leukemia ◽  
Receptor Expression ◽  
Interleukin 3 ◽  
Gm Csf ◽  
In Vitro Cell Lines

Abstract Recombinant human granulocyte-macrophage (GM) colony-stimulating factor (GM-CSF), G-CSF, and interleukin-3 (IL-3) labeled with 125I were used to study the characteristics and distribution of receptors for these factors on in vitro cell lines and on cells from patients with acute nonlymphocytic leukemia (ANL) and acute lymphocytic leukemia (ALL). Receptors for GM-CSF and G-CSF were restricted to a subset of myelomonocytic cell lines whereas IL-3 receptors were also found on pre- B- or early B-cell lines. Receptors for all three CSFs were broadly distributed on ANL cells, with considerable variability in levels of expression. Measurement of the colony-forming ability of ANL cells in response to the CSFs showed that there was no direct correlation between the ability of the cells to respond to a growth factor and the absolute number of receptors expressed for that growth factor. Binding of radiolabeled IL-3 and GM-CSF to ANL cells produced complex biphasic curves. Further analysis showed that both IL-3 and GM-CSF were able to partially compete for specific binding of the heterologous radiolabeled ligand to cells from several ANL patients, suggesting that heterogeneity may exist in human CSF receptors. These results provide new insights into the complex role that CSFs may play in ANL.

scholarly journals Interleukin-3, GM-CSF, and G-CSF receptor expression on cell lines and primary leukemia cells: receptor heterogeneity and relationship to growth factor responsiveness

Blood ◽  
1989 ◽  
Vol 74 (1) ◽  
pp. 56-65 ◽  
Author(s):  
LS Park ◽  
PE Waldron ◽  
D Friend ◽  
HM Sassenfeld ◽  
V Price ◽  
...  
Keyword(s):  
Growth Factor ◽  
Cell Lines ◽  
Specific Binding ◽  
Absolute Number ◽  
Lymphocytic Leukemia ◽  
Receptor Expression ◽  
Interleukin 3 ◽  
Gm Csf ◽  
In Vitro Cell Lines

Recombinant human granulocyte-macrophage (GM) colony-stimulating factor (GM-CSF), G-CSF, and interleukin-3 (IL-3) labeled with 125I were used to study the characteristics and distribution of receptors for these factors on in vitro cell lines and on cells from patients with acute nonlymphocytic leukemia (ANL) and acute lymphocytic leukemia (ALL). Receptors for GM-CSF and G-CSF were restricted to a subset of myelomonocytic cell lines whereas IL-3 receptors were also found on pre- B- or early B-cell lines. Receptors for all three CSFs were broadly distributed on ANL cells, with considerable variability in levels of expression. Measurement of the colony-forming ability of ANL cells in response to the CSFs showed that there was no direct correlation between the ability of the cells to respond to a growth factor and the absolute number of receptors expressed for that growth factor. Binding of radiolabeled IL-3 and GM-CSF to ANL cells produced complex biphasic curves. Further analysis showed that both IL-3 and GM-CSF were able to partially compete for specific binding of the heterologous radiolabeled ligand to cells from several ANL patients, suggesting that heterogeneity may exist in human CSF receptors. These results provide new insights into the complex role that CSFs may play in ANL.

scholarly journals In vivo control of differentiation of myeloid leukemic cells by recombinant granulocyte-macrophage colony-stimulating factor and interleukin 3

Blood ◽  
1988 ◽  
Vol 71 (2) ◽  
pp. 375-382 ◽  
Author(s):  
J Lotem ◽  
L Sachs
Keyword(s):  
Therapeutic Potential ◽  
Regulatory Proteins ◽  
Leukemic Cells ◽  
Cell Multiplication ◽  
Interleukin 3 ◽  
Gm Csf ◽  
Blast Cells ◽  
Macrophage Colony

The normal myeloid hematopoietic regulatory proteins include one class of proteins that induces viability and multiplication of normal myeloid precursor cells to form colonies (colony-stimulating factors [CSF] and interleukin 3 [IL-3], macrophage and granulocyte inducing proteins, type 7 [MGI-1]) and another class (called MGI-2) that induces differentiation of normal myeloid precursors without inducing cell multiplication. Different clones of myeloid leukemic cells can differ in their response to these regulatory proteins. One type of leukemic clone can be differentiated in vitro to mature cells by incubating with the growth-inducing proteins granulocyte-macrophage (GM) CSF or IL-3, and another type of clone can be differentiated in vitro to mature cells by the differentiation-inducing protein MGI-2. We have now studied the ability of different myeloid regulatory proteins to induce the in vivo differentiation of these different types of mouse myeloid leukemic clones in normal and cyclophosphamide-treated mice. The results show that in both types of mice (a) the in vitro GM-CSF- and IL- 3-sensitive leukemic cells were induced to differentiate to mature cells in vivo in mice injected with pure recombinant GM-CSF and IL-3 but not with G-CSF, M-CSF, or MGI-2; (b) the in vitro MGI-2-sensitive leukemic cells differentiated in vivo by injection of MGI-2 and also, presumably indirectly, by GM-CSF and IL-3 but not by M-CSF or G-CSF; (c) in vivo induced differentiation of the leukemic cells was associated with a 20- to 60-fold decrease in the number of blast cells; and (d) all the injected myeloid regulatory proteins stimulated the normal myelopoietic system. Different normal myeloid regulatory proteins can thus induce in vivo terminal differentiation of leukemic cells, and it is suggested that these proteins can have a therapeutic potential for myeloid leukemia in addition to their therapeutic potential in stimulating normal hematopoiesis.

Enantiomer-Specific Transformation by 2HG Is Linked to Opposing Effects on α-Ketoglutarate-Dependent Dioxygenases

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. LBA-4-LBA-4
Author(s):  
Julie-Aurore Losman ◽  
Sungwoo Lee ◽  
Peppi Koivunen ◽  
Ryan E. Looper ◽  
William G. Kaelin
Keyword(s):  
Growth Factor ◽  
Cancer Cell ◽  
Leukemic Cells ◽  
Dependent Manner ◽  
Hydroxylase Activity ◽  
Induced Differentiation ◽  
Gm Csf ◽  
Mutant Idh1 ◽  
Opposing Effects

Abstract LBA-4 Somatic mutations in IDH1 and IDH2 occur frequently in clonal myeloid disorders and result in the neomorphic ability of IDH to convert α-ketoglutarate (2-OG) to the R-enantiomer of 2-hydroxyglutarate (R-2HG) (Dang, et al Nature 462: 739, 2009). 2OG is an essential cofactor for many metabolic enzymes, including the TET family of 5-methylcytosine hydroxylases and the EglN family of prolyl-4-hydroxylases, and 2HG has been shown to inhibit several 2OG-dependent dioxygenases in vitro, including TET2 (Xu, et al Cancer Cell 19: 17, 2011; Figueroa, et al Cancer Cell 18: 1, 2010). We recently showed that the (S) enantiomer of 2HG (S-2HG), but not the (R) enantiomer of 2HG (R-2HG), inhibits the EglN prolyl-4-hydroxylases (Koivunen, et al. Submitted for publication). Moreover, we found that R-2HG can act as a cofactor to promote the hydroxylase activity of EglN1, EglN2 and EglN3. We hypothesized that the qualitatively different effects of R- and S-2HG on the EglN prolyl-4-hydroxylases might influence their transforming activities. In order to elucidate the role of mutant IDH, and R- and S-2HG, in myeloid leukemia, we developed a myeloid transformation assay using TF-1 cells. TF-1 is a human erythroleukemia cell line that requires GM-CSF for growth and undergoes erythrocytic differentiation when stimulated with erythropoietin (EPO). We expressed wild-type IDH1 (WTIDH1), a tumor-derived mutant IDH1 (IDH1R132H), or a catalytically inactive IDH1R132H variant (IDH1R132H/3DN) in TF-1 cells. As expected, cells expressing IDH1R132H, but not cells expressing WTIDH1 or IDH1R132H/3DN, had dramatically elevated levels of 2HG. Furthermore, we found that expression of IDH1R132H, but not WTIDH1 or IDH1R132H/3DN, conferred growth factor-independence to TF-1 cells (Figure 1a), and blocked their EPO-induced differentiation (Figure 1b). In order to determine whether transformation of TF-1 cells by IDH1R132H is mediated by 2HG, we treated TF-1 cells with cell-permeable esterified R-2HG or S-2HG. R-2HG recapitulated the growth and differentiation phenotypes of IDH1R132H expression in a dose-dependent manner. In contrast, S-2HG did not induce these phenotypes at any concentration tested. Next, we examined the effect of loss of TET2 on TF-1 cells. We infected TF-1 cells with shRNAs targeting TET1 or TET2 and found that knockdown of TET2, but not TET1, induced growth factor-independence and blocked EPO-induced differentiation similarly to expression of IDH1R132H or treatment with R-2HG. Interestingly, we found that transformation by IDH1R132H and TET2 knockdown were reversed by inhibition of EglN1 (Figure 2), suggesting that R-2HG, but not S-2HG, transforms leukemic cells by inhibiting targets such as TET2 while preserving, and possibly enhancing, EglN activity. These findings further suggest that therapeutic targeting of EglN prolyl-4-hydroxylase activity might be effective in the treatment of IDH1-mutant and TET2-mutant myeloid leukemias. Disclosures: Kaelin: Fibrogen: Consultancy, Equity Ownership.

scholarly journals Growth factor requirements of childhood acute T-lymphoblastic leukemia: correlation between presence of chromosomal abnormalities and ability to grow permanently in vitro

Blood ◽  
1991 ◽  
Vol 77 (7) ◽  
pp. 1534-1545 ◽  
Author(s):  
R O'Connor ◽  
A Cesano ◽  
B Lange ◽  
J Finan ◽  
PC Nowell ◽  
...  
Keyword(s):  
Growth Factors ◽  
T Cell ◽  
Growth Factor ◽  
Cell Lines ◽  
Chromosomal Abnormalities ◽  
Lymphoblastic Leukemia ◽  
Cell Receptor ◽  
Hematopoietic Growth Factors ◽  
Gm Csf

Cells from 10 cases of childhood acute T-lymphoblastic leukemia (T-ALL) were cultured in the presence of recombinant human interleukins (rhIL) or colony-stimulating factors (CSF) to analyze their growth factor requirements and differentiative potential. Although cells from most leukemic samples displayed a short-term proliferative response to several hematopoietic growth factors, only the ones featuring chromosomal translocations could be established as permanent cell lines. Two cell lines could be initiated only in the presence of IL-3 (TALL-103 and TALL-106), one in granulocyte-macrophage CSF (GM-CSF) (TALL-101), and one in IL-2 (TALL-104); only one cell line (TALL-105) was originated in the absence of growth factors. The TALL-101 and TALL- 103 cell lines, derived from very immature T-ALL cases, underwent growth factor-dependent phenotypic conversion (lymphoid to myeloid). However, the T-cell receptor rearrangement and karyotype of the original leukemic clones were retained. In contrast, the TALL-104, - 105, and -106 cell lines which originated from more mature T-ALL cases, maintained a T-lymphoblastic phenotype regardless of the growth factors in which they were expanded. These data demonstrate in vitro the aggressive nature of T-ALL cases bearing chromosomal abnormalities, and indicate that the lineage commitment of the malignant clone depends on its stage of maturation in T-cell ontogeny.

scholarly journals Growth factor requirements of childhood acute T-lymphoblastic leukemia: correlation between presence of chromosomal abnormalities and ability to grow permanently in vitro

Blood ◽  
1991 ◽  
Vol 77 (7) ◽  
pp. 1534-1545 ◽  
Author(s):  
R O'Connor ◽  
A Cesano ◽  
B Lange ◽  
J Finan ◽  
PC Nowell ◽  
...  
Keyword(s):  
Growth Factors ◽  
T Cell ◽  
Growth Factor ◽  
Cell Lines ◽  
Chromosomal Abnormalities ◽  
Lymphoblastic Leukemia ◽  
Cell Receptor ◽  
Hematopoietic Growth Factors ◽  
Gm Csf

Abstract Cells from 10 cases of childhood acute T-lymphoblastic leukemia (T-ALL) were cultured in the presence of recombinant human interleukins (rhIL) or colony-stimulating factors (CSF) to analyze their growth factor requirements and differentiative potential. Although cells from most leukemic samples displayed a short-term proliferative response to several hematopoietic growth factors, only the ones featuring chromosomal translocations could be established as permanent cell lines. Two cell lines could be initiated only in the presence of IL-3 (TALL-103 and TALL-106), one in granulocyte-macrophage CSF (GM-CSF) (TALL-101), and one in IL-2 (TALL-104); only one cell line (TALL-105) was originated in the absence of growth factors. The TALL-101 and TALL- 103 cell lines, derived from very immature T-ALL cases, underwent growth factor-dependent phenotypic conversion (lymphoid to myeloid). However, the T-cell receptor rearrangement and karyotype of the original leukemic clones were retained. In contrast, the TALL-104, - 105, and -106 cell lines which originated from more mature T-ALL cases, maintained a T-lymphoblastic phenotype regardless of the growth factors in which they were expanded. These data demonstrate in vitro the aggressive nature of T-ALL cases bearing chromosomal abnormalities, and indicate that the lineage commitment of the malignant clone depends on its stage of maturation in T-cell ontogeny.

Perturbed Endocytosis as a Mechanism of CALM-Dependent Leukemogenesis: Identification of Specific CALM Domains Required for Internalization and Demonstration of Prolonged JAK2 Signaling in Response to GM-CSF.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 1817-1817
Author(s):  
Natasha L. Brooks ◽  
David A. Erichsen ◽  
Daniel S. Wechsler
Keyword(s):  
Flow Cytometry ◽  
Growth Factor ◽  
Cell Lines ◽  
Fusion Proteins ◽  
Geometric Mean ◽  
Leukemia Cell ◽  
Growth Factor Signaling ◽  
Gm Csf ◽  
Leukemia Cell Lines

Abstract Background: Clathrin Assembly Lymphoid Myeloid leukemia (CALM) gene rearrangements, in which CALM is fused to MLL or AF10 genes, are found in aggressive leukemias and lymphomas. Expression of MLL-CALM or CALM-AF10 fusion proteins immortalizes murine hematopoietic cells in vitro, correlating with leukemogenesis in vivo. While disrupted MLL or AF10 activity contributes to transformation, perturbation of normal CALM function may also play a role. The native CALM protein is primarily cytoplasmic and functions in Clathrin-Dependent Endocytosis (CDE). We have previously shown that overexpression of CALM-containing fusion proteins in COS7 cells impairs CDE of both Transferrin (TF) and Epidermal Growth Factor (EGF). These initial observations were made using a qualitative (but time-consuming) visual assay of Texas Red (TR)-labeled TF or EGF. We confirmed these results using a semi-quantitative radioactive assay (with 131I-TF or EGF) that, while sensitive, lacked specificity for transfected cells. Here we describe the development of a flow cytometry-based assay of CDE that permits specific quantitation in cells transfected with CALM-containing proteins. We also measure downstream effects of perturbed CDE on growth factor signaling. Objectives: To validate a novel flow cytometry-based CDE assay; To identify CALM domains that play critical roles in CDE; To analyze kinetics of phosphorylation of JAK2, a downstream target of growth factor signaling, during perturbed CDE. Methods: COS7 cells transfected with GFP-tagged CALM-containing constructs were incubated at 4°C for 1h with AlexaFluor (AF) 633-TF or AF647-EGF to allow binding, followed by incubation at 25°C (5, 10, or 15 min) to permit internalization. Following acid buffer wash and fixing (1% formalin/PBS), internalized fluorescence was measured by flow cytometry. Geometric mean fluorescence intensities of GFP+/AF+ populations were normalized to respective 4°C controls. JAK2 phosphorylation was assessed by western blotting at various time intervals following GM-CSF stimulation of six different leukemia cell lines. Results: Overexpression of CALM-containing constructs in COS7 cells resulted in a reduced rate of TF or EGF internalization. Native CALM (N-CALM1–660) overexpression reduced the rate of TF and EGF internalization by 53%. The portion of CALM found in MLL-CALM fusions, CALM256–660, reduced TF and EGF internalization by 66% compared with controls. CALM436–583 overexpression reduced the rate of TF and EGF internalization by 43%. A series of CALM deletion constructs (CALM256–492, CALM256–502, CALM337–531, CALM593–660, and CALM601–660) did not perturb internalization of TF or EGF. Thus, the presence of a 52 amino acid (aa) region from CALM aa 531–583 was required to perturb internalization. Phosphorylation of JAK2 was prolonged in the setting of CALM-AF10 expression (U937 and P31/Fuji cells) compared with cell lines that do not express CALM-AF10 (HL60, K562, MonoMac6, THP1), likely as a consequence of perturbed CDE. Conclusions: We demonstrate that CALM-dependent perturbation of CDE in COS7 cells can be measured by a novel flow cytometry-based approach. Use of this rapid, quantitative assay narrows the specific region of CALM critical to CDE perturbation to 52 aa. We also show that signaling downstream of growth factors is enhanced during perturbed CDE in leukemia cell lines. These observations support an underappreciated role for CDE dysregulation in leukemogenic transformation.

Heparin-Binding Epidermal Growth Factor–Like Growth Factor/Diphtheria Toxin Receptor Expression by Acute Myeloid Leukemia Cells

Blood ◽  
1999 ◽  
Vol 93 (5) ◽  
pp. 1715-1723 ◽  
Author(s):  
Fabrizio Vinante ◽  
Antonella Rigo ◽  
Emanuele Papini ◽  
Marco A. Cassatella ◽  
Giovanni Pizzolo
Keyword(s):  
Growth Factor ◽  
Cell Line ◽  
Cell Lines ◽  
Diphtheria Toxin ◽  
Myeloid Leukemia ◽  
Leukemic Cells ◽  
Heparin Binding ◽  
Gm Csf ◽  
Epidermal Growth ◽  
Acute Myeloid

Abstract Heparin-binding epidermal growth factor–like growth factor (HB-EGF) is an EGF family member expressed by numerous cell types that binds to EGF receptor 1 (HER-1) or 4 (HER-4) inducing mitogenic and/or chemotactic activities. Membrane-bound HB-EGF retains growth activity and adhesion capabilities and the unique property of being the receptor for diphtheria toxin (DT). The interest in studying HB-EGF in acute leukemia stems from these mitogenic, chemotactic, and receptor functions. We analyzed the expression of HB-EGF in L428, Raji, Jurkat, Karpas 299, L540, 2C8, HL-60, U937, THP-1, ML-3, and K562 cell lines and in primary blasts from 12 acute myeloid leukemia (AML) cases, by reverse-transcriptase polymerase chain reaction (RT-PCR) and Northern blot and by the evaluation of sensitivity to DT. The release of functional HB-EGF was assessed by evaluation of its proliferative effects on the HB-EGF–sensitive Balb/c 3T3 cell line. HB-EGF was expressed by all myeloid and T, but not B (L428, Raji), lymphoid cell lines tested, as well as by the majority (8 of 12) of ex vivo AML blasts. Cell lines (except for the K562 cell line) and AML blasts expressing HB-EGF mRNA underwent apoptotic death following exposure to DT, thus demonstrating the presence of the HB-EGF molecule on their membrane. Leukemic cells also released a fully functional HB-EGF molecule that was mitogenic for the Balb/c 3T3 cell line. Factors relevant to the biology of leukemic growth, such as tumor necrosis factor- (TNF-), 1,25-(OH)2D3, and especially all-trans retinoic acid (ATRA), upregulated HB-EGF mRNA in HL-60 or ML-3 cells. Granulocyte-macrophage colony-stimulating factor (GM-CSF) induced HB-EGF mRNA and acquisition of sensitivity to DT in one previously HB-EGF–negative leukemia case. Moreover, the U937 and Karpas 299 cell lines expressed HER-4 mRNA. This work shows that HB-EGF is a growth factor produced by primary leukemic cells and regulated by ATRA, 1,25-(OH)2D3, and GM-CSF.

Heparin-Binding Epidermal Growth Factor–Like Growth Factor/Diphtheria Toxin Receptor Expression by Acute Myeloid Leukemia Cells

Blood ◽  
1999 ◽  
Vol 93 (5) ◽  
pp. 1715-1723 ◽  
Author(s):  
Fabrizio Vinante ◽  
Antonella Rigo ◽  
Emanuele Papini ◽  
Marco A. Cassatella ◽  
Giovanni Pizzolo
Keyword(s):  
Growth Factor ◽  
Cell Line ◽  
Cell Lines ◽  
Diphtheria Toxin ◽  
Myeloid Leukemia ◽  
Leukemic Cells ◽  
Heparin Binding ◽  
Gm Csf ◽  
Epidermal Growth ◽  
Acute Myeloid

Heparin-binding epidermal growth factor–like growth factor (HB-EGF) is an EGF family member expressed by numerous cell types that binds to EGF receptor 1 (HER-1) or 4 (HER-4) inducing mitogenic and/or chemotactic activities. Membrane-bound HB-EGF retains growth activity and adhesion capabilities and the unique property of being the receptor for diphtheria toxin (DT). The interest in studying HB-EGF in acute leukemia stems from these mitogenic, chemotactic, and receptor functions. We analyzed the expression of HB-EGF in L428, Raji, Jurkat, Karpas 299, L540, 2C8, HL-60, U937, THP-1, ML-3, and K562 cell lines and in primary blasts from 12 acute myeloid leukemia (AML) cases, by reverse-transcriptase polymerase chain reaction (RT-PCR) and Northern blot and by the evaluation of sensitivity to DT. The release of functional HB-EGF was assessed by evaluation of its proliferative effects on the HB-EGF–sensitive Balb/c 3T3 cell line. HB-EGF was expressed by all myeloid and T, but not B (L428, Raji), lymphoid cell lines tested, as well as by the majority (8 of 12) of ex vivo AML blasts. Cell lines (except for the K562 cell line) and AML blasts expressing HB-EGF mRNA underwent apoptotic death following exposure to DT, thus demonstrating the presence of the HB-EGF molecule on their membrane. Leukemic cells also released a fully functional HB-EGF molecule that was mitogenic for the Balb/c 3T3 cell line. Factors relevant to the biology of leukemic growth, such as tumor necrosis factor- (TNF-), 1,25-(OH)2D3, and especially all-trans retinoic acid (ATRA), upregulated HB-EGF mRNA in HL-60 or ML-3 cells. Granulocyte-macrophage colony-stimulating factor (GM-CSF) induced HB-EGF mRNA and acquisition of sensitivity to DT in one previously HB-EGF–negative leukemia case. Moreover, the U937 and Karpas 299 cell lines expressed HER-4 mRNA. This work shows that HB-EGF is a growth factor produced by primary leukemic cells and regulated by ATRA, 1,25-(OH)2D3, and GM-CSF.

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