Human interleukin-4 inhibits proliferation of megakaryocyte progenitor cells in culture

Abstract We studied the effects of recombinant human interleukin-4 (rhIL-4) on megakaryocyte colony formation from enriched hematopoietic progenitors. IL-4 strongly inhibited pure and mixed megakaryocyte colony formation in a dose-dependent manner. Formation of erythroid bursts, eosinophil colonies, and erythrocyte-containing mixed colonies was not affected by the addition of IL-4 as reported previously (Sonoda Y, et al; Blood 75:1615, 1990). Delayed addition experiments suggested that IL-4 acts on an early stage of proliferation of megakaryocyte progenitors. Neutralizing antibodies (antisera) prepared against transforming growth factor beta, tumor necrosis factor alpha, interferon alpha (IFN alpha), and IFN gamma did not affect the inhibitory effects of IL-4 on pure and mixed megakaryocyte colony formation. In addition, the inhibitory effects of IL-4 was also seen in serum-free cultures and in cultures containing highly enriched CD34+, HLA-DR+ cells as a target population. These results indicate that IL-4 may function as one of the negative regulators in human megakaryocytopoiesis in vitro.

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Human interleukin-4 inhibits proliferation of megakaryocyte progenitor cells in culture

Blood ◽

10.1182/blood.v81.3.624.bloodjournal813624 ◽

1993 ◽

Vol 81 (3) ◽

pp. 624-630

Author(s):

Y Sonoda ◽

Y Kuzuyama ◽

S Tanaka ◽

S Yokota ◽

T Maekawa ◽

...

Keyword(s):

Transforming Growth Factor Beta ◽

Colony Formation ◽

Neutralizing Antibodies ◽

Transforming Growth Factor ◽

Early Stage ◽

Interleukin 4 ◽

Dependent Manner ◽

Inhibitory Effects ◽

Necrosis Factor Alpha

We studied the effects of recombinant human interleukin-4 (rhIL-4) on megakaryocyte colony formation from enriched hematopoietic progenitors. IL-4 strongly inhibited pure and mixed megakaryocyte colony formation in a dose-dependent manner. Formation of erythroid bursts, eosinophil colonies, and erythrocyte-containing mixed colonies was not affected by the addition of IL-4 as reported previously (Sonoda Y, et al; Blood 75:1615, 1990). Delayed addition experiments suggested that IL-4 acts on an early stage of proliferation of megakaryocyte progenitors. Neutralizing antibodies (antisera) prepared against transforming growth factor beta, tumor necrosis factor alpha, interferon alpha (IFN alpha), and IFN gamma did not affect the inhibitory effects of IL-4 on pure and mixed megakaryocyte colony formation. In addition, the inhibitory effects of IL-4 was also seen in serum-free cultures and in cultures containing highly enriched CD34+, HLA-DR+ cells as a target population. These results indicate that IL-4 may function as one of the negative regulators in human megakaryocytopoiesis in vitro.

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Interacting cytokines regulate in vitro human megakaryocytopoiesis

Blood ◽

10.1182/blood.v73.3.671.bloodjournal733671 ◽

1989 ◽

Vol 73 (3) ◽

pp. 671-677 ◽

Cited By ~ 1

Author(s):

E Bruno ◽

ME Miller ◽

R Hoffman

Keyword(s):

Transforming Growth Factor Beta ◽

Colony Formation ◽

Transforming Growth Factor ◽

Interleukin 1 ◽

Interleukin 4 ◽

Recombinant Erythropoietin ◽

Hematopoietic Growth Factors ◽

Macrophage Colony ◽

Stimulating Factor

The effects of hematopoietic growth factors on in vitro human megakaryocytopoiesis were studied using a serum-depleted culture system. Both recombinant interleukin-3 (r-IL-3) and recombinant granulocyte-macrophage colony-stimulating factor (rGM-CSF) increased megakaryocyte (MK) colony formation (P less than .01) above that observed in baseline cultures. Recombinant interleukin-4 (rIL-4) and interleukin 1 alpha (rIL-1 alpha) failed either to promote MK colony formation alone or to increase rIL-3 or rGM-CSF promoted colony formation. Recombinant erythropoietin (rEpo) and purified thrombocytopoiesis-stimulating factor (TSF) did not increase (P greater than .05) MK colony formation when added alone but synergized with rIL- 1 alpha, leading to a twofold increase in MK colony formation. Such a synergistic relationship was not observed between rIL-4 and rEpo. In addition, TSF enhanced the ability of rIL-3 but not rGM-CSF to promote MK colony formation. Addition of rEpo to optimal or suboptimal concentrations of rGM-CSF or suboptimal concentrations of rIL-3 resulted in a significant increase (P less than .05) in the total number of MK-containing colonies, due to the appearance of multilineage colonies containing MKs. The addition of rEpo to optimal concentrations of rIL-3 resulted in increased numbers of multilineage colonies containing MKs; however, the number of total MK-containing colonies was not significantly increased when compared to assays containing rIL-3 alone. By contrast, transforming growth factor-beta (TGF-beta) inhibited both rIL-3, and rGM-CSF promoted MK colony formation, with optimal inhibition resulting in a 35%-45% reduction of MK colony formation.

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Interacting cytokines regulate in vitro human megakaryocytopoiesis

Blood ◽

10.1182/blood.v73.3.671.671 ◽

1989 ◽

Vol 73 (3) ◽

pp. 671-677 ◽

Cited By ~ 79

Author(s):

E Bruno ◽

ME Miller ◽

R Hoffman

Keyword(s):

Transforming Growth Factor Beta ◽

Colony Formation ◽

Transforming Growth Factor ◽

Interleukin 1 ◽

Interleukin 4 ◽

Recombinant Erythropoietin ◽

Hematopoietic Growth Factors ◽

Macrophage Colony ◽

Stimulating Factor

Abstract The effects of hematopoietic growth factors on in vitro human megakaryocytopoiesis were studied using a serum-depleted culture system. Both recombinant interleukin-3 (r-IL-3) and recombinant granulocyte-macrophage colony-stimulating factor (rGM-CSF) increased megakaryocyte (MK) colony formation (P less than .01) above that observed in baseline cultures. Recombinant interleukin-4 (rIL-4) and interleukin 1 alpha (rIL-1 alpha) failed either to promote MK colony formation alone or to increase rIL-3 or rGM-CSF promoted colony formation. Recombinant erythropoietin (rEpo) and purified thrombocytopoiesis-stimulating factor (TSF) did not increase (P greater than .05) MK colony formation when added alone but synergized with rIL- 1 alpha, leading to a twofold increase in MK colony formation. Such a synergistic relationship was not observed between rIL-4 and rEpo. In addition, TSF enhanced the ability of rIL-3 but not rGM-CSF to promote MK colony formation. Addition of rEpo to optimal or suboptimal concentrations of rGM-CSF or suboptimal concentrations of rIL-3 resulted in a significant increase (P less than .05) in the total number of MK-containing colonies, due to the appearance of multilineage colonies containing MKs. The addition of rEpo to optimal concentrations of rIL-3 resulted in increased numbers of multilineage colonies containing MKs; however, the number of total MK-containing colonies was not significantly increased when compared to assays containing rIL-3 alone. By contrast, transforming growth factor-beta (TGF-beta) inhibited both rIL-3, and rGM-CSF promoted MK colony formation, with optimal inhibition resulting in a 35%-45% reduction of MK colony formation.

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Interleukin 4 protects chronic lymphocytic leukemic B cells from death by apoptosis and upregulates Bcl-2 expression.

Journal of Experimental Medicine ◽

10.1084/jem.176.5.1319 ◽

1992 ◽

Vol 176 (5) ◽

pp. 1319-1326 ◽

Cited By ~ 235

Author(s):

M Dancescu ◽

M Rubio-Trujillo ◽

G Biron ◽

D Bron ◽

G Delespesse ◽

...

Keyword(s):

B Cells ◽

Transforming Growth Factor Beta ◽

Transforming Growth Factor ◽

Protein A ◽

Intermediate Stage ◽

Lymphocytic Leukemia ◽

Interleukin 4 ◽

Necrosis Factor Alpha

B chronic lymphocytic leukemia (B-CLL) is characterized by the accumulation of slow-dividing and long-lived monoclonal B cells arrested at the intermediate stage of their differentiation. We previously showed that interleukin 4 (IL-4) not only inhibits but also prevents the proliferation of B-CLL cells. We report here that IL-4 protects the B-CLL cells from death by apoptosis (programmed cell death [PCD]). IL-4 inhibits spontaneous and hydrocortisone (HC)-induced PCD of highly purified B cells from 12 unselected CLL patients, as shown by sustained cell viability and lack of DNA fragmentation. IL-1, -2, -3, -5, -6, -7, tumor necrosis factor alpha, and transforming growth factor beta have no protective effect. The in vitro rescue from apoptosis by IL-4 is reflected by an increased expression of Bcl-2 protein, a proto-oncogene directly involved in the prolongation of cell survival in vivo and in vitro. Hence, IL-4-treated B-CLL cells express significantly more Bcl-2 than unstimulated, HC-treated, or fresh B-CLL cells. Furthermore, subcutaneous injection of IL-4 into one CLL patient enhances Bcl-2 protein expression in the leukemic B cells. These data may suggest that IL-4 prevents apoptosis of B-CLL cells using a Bcl-2-dependent pathway. Given our recent observations that fresh T cells from B-CLL patients express IL-4 mRNA, we propose that IL-4 has an essential role in the pathogenesis of CLL disease, by preventing both the death and the proliferation of the malignant B cells.

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Foxp3+ T Regulatory Cells as a Potential Target for Immunotherapy against Primary Infection with Echinococcus multilocularis Eggs

Infection and Immunity ◽

10.1128/iai.00542-18 ◽

2018 ◽

Vol 86 (10) ◽

Cited By ~ 9

Author(s):

Junhua Wang ◽

Rita Cardoso ◽

Nelson Marreros ◽

Norbert Müller ◽

Britta Lundström-Stadelmann ◽

...

Keyword(s):

Transforming Growth Factor Beta ◽

Echinococcus Multilocularis ◽

Transforming Growth Factor ◽

Early Stage ◽

Expression Profiles ◽

Growth Potential ◽

Interleukin 4 ◽

Content Type ◽

Hepatic Expression ◽

Th2 Immune Response

ABSTRACT Alveolar echinococcosis (AE) is a lethal disease caused by infection with the metacestode stage of the helminth Echinococcus multilocularis, which develops into a tumorlike mass in susceptible intermediate hosts. The growth potential of this parasite stage is directly linked to the nature of the surrounding periparasitic immune-mediated processes. In a first step (experiment 1), mice were orally infected with E. multilocularis eggs, to be used for assessing the hepatic expression profiles of 15 selected cytokine and chemokine genes related to acquired immunity from 21 to 120 days postinfection. The early stage of infection in immunocompetent animals was marked by a mixed Th1/Th2 immune response, as characterized by the concomitant presence of gamma interferon (IFN-γ) and interleukin-4 (IL-4) and their related chemokines. At the late stage of AE, the profile extended to a combined tolerogenic mode including Foxp3, IL-10, and transforming growth factor beta (TGF-β) as key components. In a second step (experiment 2), the effect of T regulatory cell (Treg) deficiency on metacestode growth was assessed in E. multilocularis-infected DEREG (depletion of regulatory T cells) mice upon induction of Treg deficiency with diphtheria toxin (DT). The parasite lesions were significantly smaller in the livers of treated mice than in corresponding control groups. Foxp3+ Tregs appear to be one of the key players in immune-regulatory processes favoring metacestode survival by affecting antigen presentation and suppressing Th1-type immune responses. For these reasons, we suggest that affecting Foxp3+ Tregs could offer an attractive target in the development of an immunotherapy against AE.

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Expression and secretion of transforming growth factor-beta by bleomycin-stimulated rat alveolar macrophages

AJP Lung Cellular and Molecular Physiology ◽

10.1152/ajplung.1993.264.1.l36 ◽

1993 ◽

Vol 264 (1) ◽

pp. L36-L42 ◽

Cited By ~ 2

Author(s):

E. M. Denholm ◽

S. M. Rollins

Keyword(s):

Growth Factor ◽

Alveolar Macrophages ◽

Transforming Growth Factor Beta ◽

Transforming Growth Factor ◽

Chemotactic Activity ◽

Beta Activity ◽

Dependent Manner ◽

Tgf Beta ◽

Growth Factor Beta

Bleomycin-induced fibrosis in rodents has been used extensively as a model of human pulmonary fibrosis. The influx of monocytes observed during the early stages of fibrosis is at least partially regulated by the elaboration of chemotactic factors in the lung. Exposure of alveolar macrophages (AM phi) to bleomycin either in vivo or in vitro stimulated secretion of monocyte chemotactic activity (MCA). This MCA has been previously characterized as being primarily due to fibronectin fragments. The present experiments revealed that bleomycin also induced AM phi to secrete a second chemotactic factor, transforming growth factor-beta (TGF-beta). However, the TGF-beta secreted by macrophages was in latent form, since no TGF-beta activity was detected unless AM phi conditioned medium (CM) was acid-activated. After acidification, chemotactic activity in CM from AM phi stimulated with bleomycin in vitro was increased by 3.6, whereas activity in AM phi CM from fibrotic rats increased by 2 and that of a bleomycin-stimulated AM phi cell line increased by 1.6. This acid-activatable chemotactic activity was inhibited by antibody to TGF-beta. Bleomycin-stimulated AM phi s secreted significantly more TGF-beta than did unstimulated controls. Further, in vitro exposure of AM phi to bleomycin induced TGF-beta mRNA expression in a time- and concentration-dependent manner, with maximal mRNA being detected following a 16-h incubation with 1 microgram/ml bleomycin.

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TGF-β Promotes the Proliferation of Microglia In Vitro

Brain Sciences ◽

10.3390/brainsci10010020 ◽

2019 ◽

Vol 10 (1) ◽

pp. 20 ◽

Cited By ~ 1

Author(s):

Costansia Bureta ◽

Takao Setoguchi ◽

Yoshinobu Saitoh ◽

Hiroyuki Tominaga ◽

Shingo Maeda ◽

...

Keyword(s):

Cell Death ◽

Cell Lines ◽

Transforming Growth Factor Beta ◽

Microglial Cell ◽

Transforming Growth Factor ◽

Dependent Manner ◽

In Vitro Proliferation ◽

Inhibition Of Proliferation ◽

Significant Difference

The activation and proliferation of microglia is characteristic of the early stages of brain pathologies. In this study, we aimed to identify a factor that promotes microglial activation and proliferation and examined the in vitro effects on these processes. We cultured microglial cell lines, EOC 2 and SIM-A9, with various growth factors and evaluated cell proliferation, death, and viability. The results showed that only transforming growth factor beta (TGF-β) caused an increase in the in vitro proliferation of both microglial cell lines. It has been reported that colony-stimulating factor 1 promotes the proliferation of microglia, while TGF-β promotes both proliferation and inhibition of cell death of microglia. However, upon comparing the most effective doses of both (assessed from the proliferation assay), we identified no statistically significant difference between the two factors in terms of cell death; thus, both have a proliferative effect on microglial cells. In addition, a TGF-β receptor 1 inhibitor, galunisertib, caused marked inhibition of proliferation in a dose-dependent manner, indicating that inhibition of TGF-β signalling reduces the proliferation of microglia. Therefore, galunisertib may represent a promising therapeutic agent for the treatment of neurodegenerative diseases via inhibition of nerve injury-induced microglial proliferation, which may result in reduced inflammatory and neuropathic and cancer pain.

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Ability of flt3 ligand to stimulate the in vitro growth of primitive murine hematopoietic progenitors is potently and directly inhibited by transforming growth factor-beta and tumor necrosis factor-alpha

Blood ◽

10.1182/blood.v87.12.5016.bloodjournal87125016 ◽

1996 ◽

Vol 87 (12) ◽

pp. 5016-5026 ◽

Cited By ~ 29

Author(s):

SE Jacobsen ◽

OP Veiby ◽

J Myklebust ◽

C Okkenhaug ◽

SD Lyman

Keyword(s):

Cell Growth ◽

Transforming Growth Factor Beta ◽

Transforming Growth Factor ◽

Tnf Alpha ◽

Tgf Beta ◽

Necrosis Factor Alpha ◽

Factor Alpha ◽

Growth Factor Beta ◽

Necrosis Factor

The recently cloned flt3 ligand (FL) stimulates the growth of primitive hematopoietic progenitor cells through synergistic interactions with multiple other cytokines. The present study is the first demonstrating cytokines capable of inhibiting FL-stimulated hematopoietic cell growth. Tumor necrosis factor-alpha (TNF-alpha) and transforming growth factor-beta 1 (TGF-beta l) potently inhibited the clonal growth of murine Lin-Sca-l+ bone marrow progenitors stimulated by FL alone or in combination with granulocyte colony-stimulating factor (G-CSF), stem cell factor (SCF), interleukin (IL)-3, IL-6, IL-11, or IL-12. TGF-beta 1 inhibited more than 96% of the myeloid colony formation in response to these cytokine combinations, whereas TNF-alpha reduced the number of colonies by 58% to 96% depending on the cytokine by which FL was combined. In addition, both TNF-alpha and TGF-beta 1 inhibited more than 90% of B220+ cell production from B220- bone marrow cells stimulated by FL + IL-7. The effects of TNF-alpha and TGF-beta 1 appeared to be due to a direct effect and on the early progenitors because the inhibition was observed at the single cell level, and because delayed addition of the two inhibitors for only 48 hours dramatically reduced their inhibitory effects. A neutralizing anti-TGF- beta antibody showed the presence of endogenous TGF-beta in the cultures and potently enhanced the ability of FL to stimulate progenitor cell growth in the absence of other cytokines. Agonistic antibodies specifically activating the p75 TNF receptors were more efficient than wild type murine TNF-alpha in signaling growth inhibition of Lin-Sca-l+ progenitor cells, whereas the p55 agonist had less effect than murine TNF-alpha. Finally, TGF-beta increased the number of FL + IL-11-stimulated Lin-Sca-1+ cells in the G1 phase of the cell cycle with 76%, whereas TNF-alpha only had a marginal effect on cell cycle distribution. Thus, TGF-beta, TNF-alpha, and p75 TNF receptor agonists are potent direct inhibitors of FL-stimulated progenitor cell growth in vitro.

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Cytokine production by primary bone marrow megakaryocytes

Blood ◽

10.1182/blood.v84.12.4151.bloodjournal84124151 ◽

1994 ◽

Vol 84 (12) ◽

pp. 4151-4156 ◽

Cited By ~ 56

Author(s):

S Jiang ◽

JD Levine ◽

Y Fu ◽

B Deng ◽

R London ◽

...

Keyword(s):

Bone Marrow ◽

Transforming Growth Factor Beta ◽

Transforming Growth Factor ◽

Interleukin 1 ◽

Human Bone ◽

Human Bone Marrow ◽

Tnf Alpha ◽

Necrosis Factor Alpha ◽

Gm Csf

Primary human bone marrow megakaryocytes were studied for their ability to express and release cytokines potentially relevant to their proliferation and/or differentiation. The purity of the bone marrow megakaryocytes was assessed by morphologic and immunocytochemical criteria. Unstimulated marrow megakaryocytes constitutively expressed genes for interleukin-1 beta (IL-1 beta), IL-6, granulocyte-macrophage colony-stimulating factor (GM-CSF), and tumor necrosis factor-alpha (TNF-alpha), by the polymerase chain reaction (PCR) and Northern blot analysis. At the protein level, megakaryocytes secreted significant amounts of IL-1 beta (53.6 +/- 3.6 pg/mL), IL-6 (57.6 +/- 15.6 pg/mL), and GM-CSF (24 +/- 4 pg/mL) but not TNF-alpha. Exposure of human marrow megakaryocytes to IL-1 beta increased the levels of IL-6 (87.3 +/- 2.3 pg/mL) detected in the culture supernatants. Transforming growth factor- beta was also able to stimulate IL-6, IL-1 beta, and GM-CSF secretion, but was less potent than stimulation with phorbol-12-myristate-13- acetate (PMA). The secreted cytokines acted additively to maintain and increase the number of colony-forming unit-megakaryocytes colonies (approximately 35%). These studies demonstrate the production of multiple cytokines by isolated human bone marrow megakaryocytes constitutively or stimulated in vitro. The capacity of human megakaryocytes to synthesize several cytokines known to modulate hematopoietic cells supports the concept that there may be an autocrine mechanism operative in the regulation of megakaryocytopoiesis.

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Transforming growth factor beta directly regulates primitive murine hematopoietic cell proliferation

Blood ◽

10.1182/blood.v75.3.596.596 ◽

1990 ◽

Vol 75 (3) ◽

pp. 596-602 ◽

Cited By ~ 112

Author(s):

JR Keller ◽

IK Mcniece ◽

KT Sill ◽

LR Ellingsworth ◽

PJ Quesenberry ◽

...

Keyword(s):

Bone Marrow ◽

Transforming Growth Factor Beta ◽

Transforming Growth Factor ◽

Bone Marrow Cells ◽

Dependent Manner ◽

Tgf Beta ◽

Hematopoietic Progenitor ◽

Growth Factor Beta ◽

Dose Dependent

Abstract We previously reported that transforming growth factor beta (TGF-beta) selectively inhibits colony-stimulating factor-driven hematopoietic progenitor cell growth. We report here that TGF-beta 1 can act directly on hematopoietic progenitors to inhibit the growth of the most primitive progenitors measurable in vitro. Highly enriched populations of hematopoietic progenitor cells were obtained by isolating lineage negative (Lin-), Thy-1-positive (Thy-1+) fresh bone marrow cells, or by isolating cells from interleukin-3 (IL-3) supplemented bone marrow cultures expressing Thy-1 antigen with the fluorescent activated cell sorter. TGF-beta 1 inhibited IL-3-induced Thy-1 expression on Thy-1- negative (Thy-1-) bone marrow cells in a dose-dependent manner with an ED50 of 5 to 10 pmol/L. In addition, TGF-beta 1 inhibited the formation of multipotent and mixed colonies by isolated Thy-1+ cells, while single lineage granulocyte and macrophage colonies were not affected. The growth of Thy-1+ Lin- cells incubated as single cells in Terasaki plates in medium supplemented with IL-3 were inhibited by TGF-beta, demonstrating a direct inhibitory effect. Hematopoietic stem cells, which have a high proliferative potential (HPP) when responding to combinations of growth factors in vitro, have been detected in the bone marrow of normal mice and mice surviving a single injection of 5- fluorouracil. TGF-beta 1 inhibited the growth of all subpopulations of HPP colony forming cells (CFC) in a dose-dependent manner with an ED50 of 5 to 10 pmol/L. Thus, TGF-beta directly inhibits the growth of the most immature hematopoietic cells measurable in vitro.

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