transforming-growth-factor-beta and Leukemia--Monocytic--Acute

The intestine-specific caudal-related homeobox gene-2 (CDX2) homeobox gene, while being a tumor suppressor in the gut, is ectopically expressed in a large proportion of acute leukemia and is associated with poor prognosis. Here, we report that turning on human CDX2 expression in the hematopoietic lineage of mice induces acute monoblastic leukemia, characterized by the decrease in erythroid and lymphoid cells at the benefit of immature monocytic and granulocytic cells. One of the highly stimulated genes in leukemic bone marrow cells was BMP and activin membrane-bound inhibitor (Bambi), an inhibitor of transforming growth factor-β (TGF-β) signaling. The CDX2 protein was shown to bind to and activate the transcription of the human BAMBI promoter. Moreover, in a leukemic cell line established from CDX2-expressing mice, reducing the levels of CDX2 or Bambi stimulated the TGF-β-dependent expression of Cd11b, a marker of monocyte maturation. Taken together, this work demonstrates the strong oncogenic potential of the homeobox gene CDX2 in the hematopoietic lineage, in contrast with its physiological tumor suppressor activity exerted in the gut. It also reveals, through BAMBI and TGF-β signaling, the involvement of CDX2 in the perturbation of the interactions between leukemia cells and their microenvironment.

Topics: Animals; CD11b Antigen; CDX2 Transcription Factor; Cell Lineage; Humans; Leukemia, Monocytic, Acute; Membrane Proteins; Mice; Signal Transduction; Transforming Growth Factor beta; Tumor Microenvironment

TGF-beta (transforming growth factor-beta) plays a critical role in modulating the inflammatory response and other biological processes through its regulation of the production of MMPs (matrix metalloproteinases). In both Mono-Mac-6 and RAW264.7 monocyte/macrophage cells, TGF-beta abrogated lipopolysaccharide-induced increases in the enzymic activity and mRNA level of MMP-9. A fragment of the human MMP-9 promoter was used to characterize its regulation by TGF-beta signalling. In RAW264.7 cells, TGF-beta or its downstream signalling protein, Smad3 (Sma- and Mad-related protein 3), inhibited lipopolysaccharide-stimulated promoter activity. The suppressive activity of TGF-beta on the MMP-9 promoter was abrogated by an inhibitory Smad, Smad7. The MMP-9 promoter contains a putative TIE (TGF-beta inhibitory element). However, neither mutation nor deletion of the TIE had any effect on the inhibitory activity of TGF-beta on MMP-9 transcription, indicating that the consensus TIE is not required for this effect of TGF-beta. Analysis using a series of deletion mutants of the MMP-9 promoter revealed that a region containing a consensus NF-kappaB (nuclear factor-kappaB) site is required for the basal activity and TGF-beta-mediated suppression of the promoter. Mutation of the putative NF-kappaB site not only markedly reduced the basal transcriptional activity of the promoter, but also abrogated the responsiveness of the promoter to TGF-beta. In addition, a minimal promoter containing one copy of the NF-kappaB sequence was responsive to TGF-beta treatment. Furthermore, an electrophoretic mobility shift assay was performed with the nuclear extracts from RAW264.7 cells, and it was found that TGF-beta treatment did not disrupt the binding of NF-kappaB p50 and p65 proteins to the NF-kappaB sequence. Taken together, these studies indicate that the NF-kappaB site is indispensable for the suppressive activity of TGF-beta in the regulation of MMP-9 transcription.

Topics: Binding Sites; Cell Line; Cell Line, Tumor; Gene Expression Regulation; Humans; Leukemia, Monocytic, Acute; Macrophages; Matrix Metalloproteinase 9; Monocytes; NF-kappa B; NF-kappa B p50 Subunit; Promoter Regions, Genetic; Protein Binding; Signal Transduction; Transcription Factor RelA; Transcription, Genetic; Transforming Growth Factor beta

The goal of this study was to determine whether cytokines modulate leukotriene C4 (LTC4) synthase expression in mononuclear phagocytes. A panel of cytokines was surveyed for changes in LTC4 synthase mRNA in THP-1 cells. TGF-beta1, -2, and -3 had significant stimulatory effects. The addition of TGF-beta resulted in a time-dependent increase in LTC4 synthase mRNA at 6 h, which persisted through 48 h. Furthermore, this conditioning resulted in an increase in immunoreactive protein for LTC4 synthase through 7 days. TGF-beta conditioning of cells resulted in a time- and dose-dependent increase in stimulated LTC4 synthase activity. Following transient transfection of THP-1 cells with a promoter-reporter construct containing 1.2 kb of the LTC4 synthase promoter, TGF-beta treatment resulted in a 2-fold increase in reporter activity. Conditioning with TGF-beta did not prolong the half-life of LTC4 synthase mRNA, as assessed by RNase protection assays in actinomycin D-treated cells. Cycloheximide exposure experiments revealed that new protein synthesis was not required for the observed stimulatory effect of TGF-beta on LTC4 synthase mRNA. We conclude that LTC4 synthase expression is increased at a transcriptional level by TGF-beta in mononuclear phagocytes.

Topics: Antigen-Antibody Reactions; Culture Media, Conditioned; Cycloheximide; Cytokines; Enzyme Activation; Glutathione Transferase; Half-Life; Humans; Immunoblotting; Leukemia, Monocytic, Acute; Monocytes; Promoter Regions, Genetic; Protein Synthesis Inhibitors; RNA, Messenger; Time Factors; Transfection; Transforming Growth Factor beta; Tumor Cells, Cultured

IL-12 is a heterodimeric cytokine produced by phagocytic and other cells with important physiologic and pathologic properties. Regulated IL-12 production is crucial for the generation of protective Th1 responses to infectious agents. In contrast, IL-12 excess contributes to the pathogenesis of a variety of autoimmune and inflammatory disorders. To further understand the processes regulating IL-12 production, we determined whether IL-11 regulated monocyte/macrophage production of this cytokine moiety. IL-11 did not alter the IL-12 (p70) production of unstimulated THP-1 monocytic cells or human blood monocytes. It did, however, inhibit, in a dose-dependent fashion, the IL-12 production of IFN-gamma plus Staphylococcus aureus Cowan strain 1-stimulated THP-1 cells and stimulated blood monocytes. This inhibition of IL-12 protein production was associated with a proportionate decrease in IL-12 p35 and p40 mRNA accumulation. Nuclear run-on assays revealed comparable decreases in IL-12 p35 and p40 gene transcription. IL-11 did not similarly regulate monocyte/macrophage production of IL-8 or macrophage inflammatory protein-1alpha (MIP-1alpha) and IL-6 did not similarly inhibit IL-12 elaboration. These studies demonstrate that IL-11 is a potent inhibitor of monocyte/macrophage IL-12 production and that this inhibitory effect is, at least in part, transcriptionally mediated. They also demonstrate that this inhibition is not the result of a generalized suppression of macrophage effector function and that the ability to inhibit monocyte/macrophage IL-12 production is not a generalized property of all IL-6-type cytokines.

Topics: Depression, Chemical; Dose-Response Relationship, Immunologic; Gene Expression Regulation; Humans; Interferon-gamma; Interleukin-11; Interleukin-12; Interleukins; Leukemia, Monocytic, Acute; Lipopolysaccharides; Monocytes; Recombinant Proteins; RNA, Messenger; Transcription, Genetic; Transforming Growth Factor beta; Tumor Cells, Cultured

The leukocyte integrin Mac-1 (CD11b/CD18) and the urokinase receptor (uPAR, CD87) mediate complementary functions in myelomonocytic cells. Both receptors promote degradation of fibrin(ogen) and also confer adhesive properties on cells because Mac-1 and uPAR bind fibrin and vitronectin, respectively. Staining of lung biopsy specimens from patients with acute lung injury indicated that fibrin and vitronectin colocalize at exudative sites in which macrophages bearing these receptors accumulate. Because of the parallel roles and physical proximity of Mac-1 and uPAR, the capacity of these receptors to functionally interact was explored. Induction of Mac-1 and uPAR expression on monocytic cell lines by transforming growth factor- beta 1 and 1.25-(OH)2 vitamin D3 conferred urokinase and uPAR-dependent adhesion to vitronectin, which was further promoted by engagement of Mac-1. Vitronectin attachment promoted subsequent Mac-1-mediated fibrinogen degradation threefold to fourfold. In contrast, enhancement of uPAR occupancy by exogenous urokinase or receptor binding fragments thereof inhibited Mac-1 function. Addition of urokinase progressively inhibited Mac-1-mediated fibrinogen binding and degradation (maximal inhibition, 91% +/- 14% and 72% +/- 15%, respectively). Saturation of uPAR with urokinase also inhibited binding of the procoagulant Mac-1 ligand, Factor X. These inhibitory effects of uPAR were reproduced in fresh monocytes, cultured monocytic cells, and in Chinese hamster ovary (CHO) cells transfected with both human Mac-1 and human uPAR. These data show that the procoagulant and fibrinolytic potential of monocytic cells is co-ordinately regulated by ligand binding to both Mac-1 and uPAR and identify uPAR as a regulator of integrin function. Vitronectin-enhanced fibrin(ogen) turnover by Mac-1 may operate as a salvage pathway in the setting of urokinase and plasmin inhibitors to promote clearance of the provisional matrix and subsequent healing.

Topics: Animals; Blood Coagulation; Calcitriol; CD18 Antigens; Cell Adhesion; CHO Cells; Cricetinae; Factor X; Fibrin; Fibrinogen; Fibrinolysis; Humans; Leukemia, Monocytic, Acute; Lung; Lung Injury; Lymphoma, Large B-Cell, Diffuse; Macromolecular Substances; Macrophage-1 Antigen; Macrophages; Monocytes; Neoplasm Proteins; Receptors, Cell Surface; Receptors, Urokinase Plasminogen Activator; Recombinant Proteins; Transfection; Transforming Growth Factor beta; Tumor Cells, Cultured; Urokinase-Type Plasminogen Activator; Vitronectin

The protease gamma-glutamyl transpeptidase (gamma-GT) activity was detected at the surface of human blood granulocytes and monocytes and myeloblastic HL-60 and monoblastic U937 leukemia cell lines using an enzymatic assay (cleavage of gamma-glu-p-nitroanilide and inhibition by the specific irreversible inhibitor of gamma-GT, i.e., acivicin). Flow cytometric analysis of gamma-GT expression and detection of a 2.4-kb gamma-GT mRNA species by Northern blot analysis confirmed the presence of gamma-GT in cells of the monocytic-granulocytic lineage. Differentiation of HL-60, U937 cells, and blood monocytes along the macrophage pathway or granulocytic maturation of HL-60 cells was accompanied by an increase in gamma-GT mRNA levels without modulation of cell surface gamma-GT activity and protein. When added to leukemic cell cultures, acivicin produced a dose- and time-dependent inhibitory growth effect associated with the induction of morphological features characteristic of macrophage maturation and enhanced surface expression of phenotypic markers CD11b and CD71 characteristic of monocyte development. When cultured in the presence of acivicin, freshly isolated monocytes also underwent characteristic changes in morphology and antigenic phenotype (increase in CD71 and HLA-DR class II) consistent with their differentiation into macrophages. In parallel, a marked production of latent transforming growth factor (TGF)-beta was observed in supernatants of cells cultured with acivicin, although TGF-beta 1 mRNA species were expressed in these cells at a level almost similar to that in unstimulated cell cultures. Moreover, acivicin-treated cells still differentiated into macrophages in the presence of a neutralizing antibody to TGF-beta 1/beta 2.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Cell Differentiation; Cell Membrane; Cells, Cultured; Cellular Senescence; gamma-Glutamyltransferase; Humans; Leukemia, Monocytic, Acute; Leukemia, Myeloid; Macrophages; Transforming Growth Factor beta; Tumor Cells, Cultured

By using fresh leukemia cells from 5 cases of acute monocytic leukemia M5 as in vitro model, we investigated the effects of recombinant human transforming growth factor beta 1 (rhTGF-beta 1) on differentiation induction of fresh leukemia cells. The results indicated that after 6 days of induction with TGF-beta 1 in a concentration of 10 ng/ml, leukemia cells in 5 AML-M5 patients differentiated obviously to maturation. The proportion of monoblasts and premonocytes was reduced, while that of mature mononuclear cells elevated. Following administration of TGF-beta 1, alpha-nonspecific esterase (alpha-NSE), whose expression could be inhibited by sodium fluoride, remained positive and peroxidase (POX) was shown to be weakly positive. These results demonstrated that TGF-beta 1 may induce in vitro differentiation of fresh leukaemia cells, but the reactions to TGF-beta 1 may vary in different cases.

Topics: Adult; Aged; Cell Transformation, Neoplastic; Female; Humans; Leukemia, Monocytic, Acute; Male; Middle Aged; Recombinant Proteins; Transforming Growth Factor beta

We have studied the expression and regulation of the interleukin-6 receptor (gp80) and its signal transducer gp130 in primary human blood monocytes. Here, we show that freshly isolated human monocytes express mRNAs for gp80 and gp130. In contrast to a previous report [(1989) FEBS Lett. 249, 27-30] we find that neither lipopolysaccharide nor interleukin-6 (IL-6) lead to a down-regulation of IL-6 receptor mRNA in monocytes. Also in the human monocytic cell line Mono Mac 6 no effect of IL-6 on receptor mRNA levels was observed. For signal transducer gp130 mRNA in monocytes a small and transient up-regulation by IL-6 was found.

Topics: Blotting, Northern; Cell Line; Cells, Cultured; Down-Regulation; Gene Expression Regulation; Humans; Interleukin-6; Leukemia, Monocytic, Acute; Lipopolysaccharides; Monocytes; Receptors, Interleukin; Receptors, Interleukin-6; Recombinant Proteins; RNA, Messenger; Signal Transduction; Transcription, Genetic; Transforming Growth Factor beta; Tumor Cells, Cultured

Previously we reported that transforming growth factor-beta 1 (TGF-beta 1) remarkably enhanced the differentiation of human leukemic cell lines, HL-60 and THP-1, in the presence of 1 alpha,25-dihydroxyvitamin D3 (VD3) and also that it induced Fc receptor for immunoglobulin G (Fc gamma R), type IIIB, in the presence of retinoic acid (RA). The present study revealed that TGF-beta 1 enhanced the Fc gamma RI- and Fc gamma RII-mediated antibody-dependent cellular cytotoxicity (ADCC) of the cells differentiated in the presence of VD3 and RA. However, production of active oxygen molecules was suppressed by TGF-beta 1. On the other hand, IL-6 stimulated production of active oxygen molecules and ADCC of the cells treated with VD3 and tumor necrosis factor-alpha (TNF-alpha). Furthermore, the levels of cell surface Fc gamma RI and Fc gamma RII were not clearly correlated with the ADCC. The TGF-beta 1/VD3-treated HL-60 cells were able to synthesize mRNAs for TGF-beta 1 and TNF-alpha, although TNF-alpha protein was not detectable. These results suggest that TGF-beta 1 has a bifunctional role, either stimulatory or inhibitory, in the modulation of macrophage activities through Fc gamma Rs and that IL-6 stimulates certain macrophage activities in mature cells.

Topics: Antibody-Dependent Cell Cytotoxicity; Blotting, Northern; Calcitriol; Cell Differentiation; Cell Line; Cell Membrane; Humans; Interleukin-6; Leukemia, Monocytic, Acute; Leukemia, Promyelocytic, Acute; Luminescent Measurements; Receptors, IgG; Recombinant Proteins; RNA, Messenger; Superoxides; Transforming Growth Factor beta; Tretinoin; Tumor Cells, Cultured; Tumor Necrosis Factor-alpha

Transforming growth factor-beta 1 (TGF-beta 1) has been implicated in a variety of responses associated with wound healing and inflammation. Thus, TGF-beta 1 enhances production of several extracellular matrix proteins both in vitro and in vivo, is chemotactic for monocytes, and alters the functioning of lymphocytes. We have examined the ability of TGF-beta 1 to affect the behavior of human THP-1 promonocytic leukemia cells, a cell line with the capacity to differentiate into macrophage-like cells. TGF-beta 1 reduces the growth rate of these cells, induces morphologic changes, and promotes adherence to culture surfaces. In addition, the adherent cell population expresses high levels of esterase activity, acquires the ability to ingest latex beads, and releases elevated levels of interleukin 1. TGF-beta 1-treated cells also express elevated levels of the beta 2 family of integrins. Taken together, these results suggest that TGF-beta 1 is capable of promoting the maturation of promonocytic cells into macrophages. This outcome has implications at wound sites where TGF-beta 1 and a myriad of other factors interact with many cell types to facilitate healing.

Topics: Cell Adhesion Molecules; Cell Differentiation; Cell Division; Fluorescent Antibody Technique; Gene Expression Regulation, Neoplastic; Humans; Interleukin-1; Leukemia, Monocytic, Acute; Macrophages; Transforming Growth Factor beta; Tumor Cells, Cultured

We have previously demonstrated (Guan X.-P., Hromchak, R. A., and Bloch, A. (1989) Cancer Commun. 1, 111-115) that ML-1 human myeloblastic leukemia cells differentiate to monocyte/macrophage-like cells by the sequential action of competence and progression factors. Tumor necrosis factor-alpha, transforming growth factor-beta, and the phorbol ester tetradecanoylphorbol acetate were found to induce competence, whereas a 77-kDa glycoprotein (DF77) isolated from mitogen-stimulated human leukocyte-conditioned medium initiated progression. In this communication we show DF77 to be an isoform of human transferrin. Hemin or soluble iron complexes did not induce differentiation progression, suggesting that the participation of transferrin in ML-1 cell differentiation may not be related to its iron-carrying capacity.

Topics: Amino Acid Sequence; Cell Differentiation; Humans; Leukemia, Monocytic, Acute; Molecular Sequence Data; Peptide Mapping; Stereoisomerism; Tetradecanoylphorbol Acetate; Transferrin; Transforming Growth Factor beta; Tumor Cells, Cultured; Tumor Necrosis Factor-alpha