scholarly journals Limiting-dilution analysis of the effects of colony-stimulating factors, phytohemagglutinin, and hydrocortisone on hematopoietic progenitor cell growth

Blood ◽  
1987 ◽  
Vol 70 (5) ◽  
pp. 1611-1618 ◽  
Author(s):  
Y Takaue ◽  
CL Reading ◽  
AJ Roome ◽  
KA Dicke ◽  
S Tindle ◽  
...  
Keyword(s):  
Bone Marrow ◽  
T Cell ◽  
Cell Growth ◽  
Cell Line ◽  
Conditioned Medium ◽  
Hematopoietic Progenitor ◽  
Colony Stimulating Factors ◽  
Gm Csf ◽  
Percoll Gradients ◽  
Limiting Dilution

Abstract The effects of colony-stimulating factors (CSFs), phytohemagglutinin (PHA), and hydrocortisone on the growth of human bone marrow hematopoietic progenitor cells (granulocyte-macrophage; GM) were analyzed in a limiting-dilution assay (LDA). Both low-density bone marrow cells separated by discontinuous Percoll gradients and a T cell- depleted and progenitor-enriched cell fraction obtained by the combination of counterflow elutriation centrifugation and Percoll gradients were examined in LDA. GCT (monocytoid cell line-conditioned medium containing GM-CSF), human placenta-conditioned medium, bladder carcinoma cell line 5637-conditioned medium (containing GM- and G-CSF), and recombinant CSF (G-CSF) directly induced proliferation of progenitors with single-hit kinetics. In some instances, however, PHA- stimulated lymphocyte-conditioned medium (containing G- and GM-CSF) showed deviation from single-hit kinetics, which demonstrated the presence of factor(s) suppressive to progenitor growth. In a T cell- depleted, progenitor-enriched fraction, PHA alone was found to suppress progenitor growth at a level as low as 100 ng/mL. The addition of hydrocortisone (10(-6) mol/L) increased the progenitor frequency but suppressed progenitor growth at 10(-4) mol/L. LDA appears to be a valuable method for exploring mechanisms of factors regulating hematopoietic cell growth.

scholarly journals Limiting-dilution analysis of the effects of colony-stimulating factors, phytohemagglutinin, and hydrocortisone on hematopoietic progenitor cell growth

Blood ◽  
1987 ◽  
Vol 70 (5) ◽  
pp. 1611-1618
Author(s):  
Y Takaue ◽  
CL Reading ◽  
AJ Roome ◽  
KA Dicke ◽  
S Tindle ◽  
...  
Keyword(s):  
Bone Marrow ◽  
T Cell ◽  
Cell Growth ◽  
Cell Line ◽  
Conditioned Medium ◽  
Hematopoietic Progenitor ◽  
Colony Stimulating Factors ◽  
Gm Csf ◽  
Percoll Gradients ◽  
Limiting Dilution

The effects of colony-stimulating factors (CSFs), phytohemagglutinin (PHA), and hydrocortisone on the growth of human bone marrow hematopoietic progenitor cells (granulocyte-macrophage; GM) were analyzed in a limiting-dilution assay (LDA). Both low-density bone marrow cells separated by discontinuous Percoll gradients and a T cell- depleted and progenitor-enriched cell fraction obtained by the combination of counterflow elutriation centrifugation and Percoll gradients were examined in LDA. GCT (monocytoid cell line-conditioned medium containing GM-CSF), human placenta-conditioned medium, bladder carcinoma cell line 5637-conditioned medium (containing GM- and G-CSF), and recombinant CSF (G-CSF) directly induced proliferation of progenitors with single-hit kinetics. In some instances, however, PHA- stimulated lymphocyte-conditioned medium (containing G- and GM-CSF) showed deviation from single-hit kinetics, which demonstrated the presence of factor(s) suppressive to progenitor growth. In a T cell- depleted, progenitor-enriched fraction, PHA alone was found to suppress progenitor growth at a level as low as 100 ng/mL. The addition of hydrocortisone (10(-6) mol/L) increased the progenitor frequency but suppressed progenitor growth at 10(-4) mol/L. LDA appears to be a valuable method for exploring mechanisms of factors regulating hematopoietic cell growth.

scholarly journals Interleukin 6 is a permissive factor for monocytic colony formation by human hematopoietic progenitor cells.

1992 ◽  
Vol 175 (4) ◽  
pp. 1151-1154 ◽  
Author(s):  
J H Jansen ◽  
J C Kluin-Nelemans ◽  
J Van Damme ◽  
G J Wientjens ◽  
R Willemze ◽  
...  
Keyword(s):  
Bone Marrow ◽  
T Cell ◽  
Progenitor Cells ◽  
Interleukin 6 ◽  
Colony Formation ◽  
Bone Marrow Cells ◽  
Hematopoietic Progenitor Cells ◽  
Hematopoietic Progenitor ◽  
Gm Csf ◽  
Marrow Cells

Since monocytes and macrophages that arise during the culture of bone marrow progenitor cells are potential sources of interleukin 6 (IL-6), we investigated whether auto- or paracrine production of this factor is involved in colony formation by normal hematopoietic progenitor cells. We added a polyclonal anti-IL-6 antiserum and a monoclonal anti-IL-6 antibody to cultures of monocyte- and T cell-depleted bone marrow cells. Colony formation was stimulated with granulocyte/monocyte-colony-stimulating factor (GM-CSF), monocyte-CSF, or IL-3. Addition of anti-IL-6 antibody resulted in decreased numbers of monocytic colonies to 40-50% of control values, whereas the numbers of granulocytic colonies were not altered. The inhibitory effect was preserved in cultures of CD34(+)-enriched bone marrow cells. As a second approach, we added a monoclonal antibody directed against the IL-6 receptor to cultures of monocyte- and T cell-depleted bone marrow cells. This antibody almost completely inhibited the growth of monocytic colonies, again without decreasing the number of granulocytic colonies. Finally, the importance of IL-6 in monocytopoiesis was demonstrated in serum-deprived bone marrow cultures: addition of exogenous IL-6 to cultures stimulated with GM-CSF resulted in increased numbers of monocytic colonies. Our results indicate that the permissive presence of IL-6 is required for optimal monocytic colony formation by bone marrow progenitor cells.

scholarly journals Human P40 T-cell growth factor (interleukin-9) supports erythroid colony formation

Blood ◽  
1990 ◽  
Vol 75 (12) ◽  
pp. 2271-2275 ◽  
Author(s):  
RE Donahue ◽  
YC Yang ◽  
SC Clark
Keyword(s):  
Bone Marrow ◽  
T Cell ◽  
Growth Factor ◽  
Cell Growth ◽  
Cell Line ◽  
Peripheral Blood ◽  
Colony Formation ◽  
T Cell Growth ◽  
Interleukin 9 ◽  
T Cell Growth Factor

Abstract Because human P40 T-cell growth factor, tentatively designated interleukin-9 (IL-9), was isolated through its ability to stimulate a human IL-3-dependent leukemic cell line (M-O7E), we tested the ability of IL-9 to support the growth and differentiation of normal hematopoietic progenitor cells from peripheral blood and bone marrow. Although the M-O7E cell line was derived from a patient with megakaryoblastic leukemia, IL-9 has not proved to be a growth or maturation factor for megakaryocytes, but instead has proved to be effective in supporting the development of erythroid bursts (BFU-E) in cultures supplemented with erythropoietin. Using highly purified progenitors from peripheral blood, IL-3 showed a BFU-E plating efficiency of 46% compared with 20% for IL-9. Because of the purity of these cell preparations and the low cell density in culture, IL-9 is likely to interact directly with erythroid progenitors. Analysis of mixing experiments and of the morphology of the BFU-E in culture indicated that IL-9 interacts preferentially with a relatively early population of IL-3-responsive BFU-E. In cultures of human bone marrow or cord blood, IL-9 selectively supported erythroid colony formation, while IL-3 and granulocyte/macrophage colony-stimulating factor additionally yielded granulocyte/macrophage colonies. Therefore, IL-9 represents a new T cell-derived cytokine with the potential for selectively stimulating erythroid development in the hematopoietic system.

scholarly journals A recombinant murine granulocyte/macrophage (GM) colony-stimulating factor derived from an inducer T cell line (IH5.5). Functional restriction to GM progenitor cells.

1986 ◽  
Vol 164 (4) ◽  
pp. 1102-1113 ◽  
Author(s):  
S Kajigaya ◽  
T Suda ◽  
J Suda ◽  
M Saito ◽  
Y Miura ◽  
...  
Keyword(s):  
Stem Cells ◽  
T Cell ◽  
Cell Line ◽  
Conditioned Medium ◽  
Fetal Liver ◽  
Plasmid Vector ◽  
Colony Stimulating Factor ◽  
Gm Csf ◽  
Blast Cells ◽  
Stimulating Factor

The cDNA for the murine granulocyte/macrophage colony-stimulating factor (GM-CSF) was cloned from a cDNA library obtained from a murine T cell line, IH5.5, by using two synthetic probes that encoded two parts of the GM-CSF from murine lung. The cDNA inserted into the plasmid vector pcDV1 was transfected into monkey COS-1 cells and the conditioned medium was used to investigate the hemopoietic activities of the resultant product, recombinant GM-CSF (rGM-CSF), by means of various colony assays. rGM-CSF stimulated only neutrophil/macrophage colonies in the cultures of murine normal bone marrow and fetal liver cells. No other colony stimulating activities (CSA) were seen in the preparation including burst-promoting activity, eosinophil-CSA, megakaryocyte-CSA and mast cell-CSA. rGM-CSF could not support colony formation of 5-fluorouracil-treated mouse spleen cells, in which only the primitive population of stem cells survived. However, after culture of these cells with PWM-spleen cell-conditioned medium (PWM-SCM), the colonies consisting of blast cells were formed. These blast cells could now be induced to form neutrophil/macrophage colonies in the presence of rGM-CSF. Pure neutrophil colonies, pure macrophage colonies, as well as mixed neutrophil/macrophage colonies, were formed from these single blast cells in the presence of rGM-CSF by micromanipulation. rGM-CSF did not act on pluripotent hemopoietic stem cells, but did act directly and selectively on neutrophil/macrophage progenitors. Moreover, striking heterogeneities were noted in the size of the colonies and the proportion of components. GM-CSF is, therefore, considered to play a noninstructive role in the differentiation of the GM pathway.

scholarly journals Human P40 T-cell growth factor (interleukin-9) supports erythroid colony formation

Blood ◽  
1990 ◽  
Vol 75 (12) ◽  
pp. 2271-2275 ◽  
Author(s):  
RE Donahue ◽  
YC Yang ◽  
SC Clark
Keyword(s):  
Bone Marrow ◽  
T Cell ◽  
Growth Factor ◽  
Cell Growth ◽  
Cell Line ◽  
Peripheral Blood ◽  
Colony Formation ◽  
T Cell Growth ◽  
Interleukin 9 ◽  
T Cell Growth Factor

Because human P40 T-cell growth factor, tentatively designated interleukin-9 (IL-9), was isolated through its ability to stimulate a human IL-3-dependent leukemic cell line (M-O7E), we tested the ability of IL-9 to support the growth and differentiation of normal hematopoietic progenitor cells from peripheral blood and bone marrow. Although the M-O7E cell line was derived from a patient with megakaryoblastic leukemia, IL-9 has not proved to be a growth or maturation factor for megakaryocytes, but instead has proved to be effective in supporting the development of erythroid bursts (BFU-E) in cultures supplemented with erythropoietin. Using highly purified progenitors from peripheral blood, IL-3 showed a BFU-E plating efficiency of 46% compared with 20% for IL-9. Because of the purity of these cell preparations and the low cell density in culture, IL-9 is likely to interact directly with erythroid progenitors. Analysis of mixing experiments and of the morphology of the BFU-E in culture indicated that IL-9 interacts preferentially with a relatively early population of IL-3-responsive BFU-E. In cultures of human bone marrow or cord blood, IL-9 selectively supported erythroid colony formation, while IL-3 and granulocyte/macrophage colony-stimulating factor additionally yielded granulocyte/macrophage colonies. Therefore, IL-9 represents a new T cell-derived cytokine with the potential for selectively stimulating erythroid development in the hematopoietic system.

IL2/IL-4, OX40L and FDC-like cell line support the in vitro tumor cell growth of adult T-cell leukemia/lymphoma

2014 ◽  
Vol 38 (5) ◽  
pp. 608-612 ◽  
Author(s):  
Dai Chihara ◽  
Yoshitoyo Kagami ◽  
Harumi Kato ◽  
Noriaki Yoshida ◽  
Tohru Kiyono ◽  
...  
Keyword(s):  
T Cell ◽  
Cell Growth ◽  
Tumor Cell ◽  
Cell Line ◽  
Tumor Cell Growth ◽  
T Cell Leukemia ◽  
Cell Leukemia ◽  
Adult T Cell Leukemia

Overexpression of hTLX3 in Association with Mutant IL7Rα Is Sufficient to Generate T-ALL In Vivo and to Transform Thymocytes in Vitro

Blood ◽  
2020 ◽  
Vol 136 (Supplement 1) ◽  
pp. 21-21
Author(s):  
Gisele Olinto Libanio Rodrigues ◽  
Julie Hixon ◽  
Hila Winer ◽  
Erica Matich ◽  
Caroline Andrews ◽  
...  
Keyword(s):  
Bone Marrow ◽  
T Cells ◽  
T Cell ◽  
Cell Line ◽  
Lymphoblastic Leukemia ◽  
T Cell Leukemia ◽  
Cell Leukemia ◽  
Cell Counts

Mutations of the IL-7Rα chain occur in approximately 10% of pediatric T-cell acute lymphoblastic leukemia cases. While we have shown that mutant IL7Ra is sufficient to transform an immortalized thymocyte cell line, mutation of IL7Ra alone was insufficient to cause transformation of primary T cells, suggesting that additional genetic lesions may be present contributing to initiate leukemia. Studies addressing the combinations of mutant IL7Ra plus TLX3 overexpression indicates in vitro growth advantage, suggesting this gene as potential collaborative candidate. Furthermore, patients with mutated IL7R were more likely to have TLX3 or HOXA subgroup leukemia. We sought to determine whether combination of mutant hIL7Ra plus TLX3 overexpression is sufficient to generate T-cell leukemia in vivo. Double negative thymocytes were isolated from C57BL/6J mice and transduced with retroviral vectors containing mutant hIL7R plus hTLX3, or the genes alone. The combination mutant hIL7R wild type and hTLX3 was also tested. Transduced thymocytes were cultured on the OP9-DL4 bone marrow stromal cell line for 5-13 days and accessed for expression of transduced constructs and then injected into sublethally irradiated Rag-/- mice. Mice were euthanized at onset of clinical signs, and cells were immunophenotyped by flow cytometry. Thymocytes transduced with muthIL-7R-hTLX3 transformed to cytokine-independent growth and expanded over 30 days in the absence of all cytokines. Mice injected with muthIL7R-hTLX3 cells, but not the controls (wthIL7R-hTLX3or mutIL7R alone) developed leukemia approximately 3 weeks post injection, characterized by GFP expressing T-cells in blood, spleen, liver, lymph nodes and bone marrow. Furthermore, leukemic mice had increased white blood cell counts and presented with splenomegaly. Phenotypic analysis revealed a higher CD4-CD8- T cell population in the blood, bone marrow, liver and spleen compared in the mutant hIL7R + hTLX3 mice compared with mice injected with mutant IL7R alone indicating that the resulting leukemia from the combination mutant hIL7R plus hTLX3 shows early arrest in T-cell development. Taken together, these data show that oncogenic IL7R activation is sufficient for cooperation with hTLX3 in ex vivo thymocyte cell transformation, and that cells expressing the combination muthIL7R-hTLX3 is sufficient to trigger T-cell leukemia in vivo. Figure Disclosures No relevant conflicts of interest to declare.

scholarly journals CAL-1 as Cellular Model System to Study CCR7-Guided Human Dendritic Cell Migration

2021 ◽  
Vol 12 ◽  
Author(s):  
Edith Uetz-von Allmen ◽  
Guerric P. B. Samson ◽  
Vladimir Purvanov ◽  
Takahiro Maeda ◽  
Daniel F. Legler
Keyword(s):  
T Cell ◽  
Cell Line ◽  
Neoplastic Cell ◽  
Model System ◽  
Model Systems ◽  
Cellular Model ◽  
Gm Csf ◽  
Endogenous Expression ◽  
Dendritic Cell Migration ◽  
Spatio Temporal

Dendritic cells (DCs) are potent and versatile professional antigen-presenting cells and central for the induction of adaptive immunity. The ability to migrate and transport peripherally acquired antigens to draining lymph nodes for subsequent cognate T cell priming is a key feature of DCs. Consequently, DC-based immunotherapies are used to elicit tumor-antigen specific T cell responses in cancer patients. Understanding chemokine-guided DC migration is critical to explore DCs as cellular vaccines for immunotherapeutic approaches. Currently, research is hampered by the lack of appropriate human cellular model systems to effectively study spatio-temporal signaling and CCR7-driven migration of human DCs. Here, we report that the previously established human neoplastic cell line CAL-1 expresses the human DC surface antigens CD11c and HLA-DR together with co-stimulatory molecules. Importantly, if exposed for three days to GM-CSF, CAL-1 cells induce the endogenous expression of the chemokine receptor CCR7 upon encountering the clinically approved TLR7/8 agonist Resiquimod R848 and readily migrate along chemokine gradients. Further, we demonstrate that CAL-1 cells can be genetically modified to express fluorescent (GFP)-tagged reporter proteins to study and visualize signaling or can be gene-edited using CRISPR/Cas9. Hence, we herein present the human CAL-1 cell line as versatile and valuable cellular model system to effectively study human DC migration and signaling.

scholarly journals The immunosuppressant rapamycin blocks in vitro responses to hematopoietic cytokines and inhibits recovering but not steady-state hematopoiesis in vivo

Blood ◽  
1994 ◽  
Vol 84 (5) ◽  
pp. 1543-1552 ◽  
Author(s):  
VF Quesniaux ◽  
S Wehrli ◽  
C Steiner ◽  
J Joergensen ◽  
HJ Schuurman ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Steady State ◽  
T Cell ◽  
T Cell Response ◽  
Colony Stimulating Factor ◽  
Gm Csf ◽  
Hematopoietic Recovery ◽  
Stimulating Factor

Abstract The immunosuppressive drug rapamycin suppresses T-cell activation by impairing the T-cell response to lymphokines such as interleukin-2 (IL- 2) and interleukin-4 (IL-4). In addition, rapamycin blocks the proliferative response of cell lines to a variety of hematopoietic growth factors, including interleukin-3 (IL-3), interleukin-6 (IL-6), granulocyte-colony stimulating factor (G-CSF), granulocyte macrophage- colony stimulating factor (GM-CSF), and kit ligand (KL), suggesting that it should be a strong inhibitor of hematopoiesis. In this report, we studied the effects of rapamycin on different hematopoietic cell populations in vitro and in vivo. In vitro, rapamycin inhibited the proliferation of primary bone marrow cells induced by IL-3, GM-CSF, KL, or a complex mixture of factors present in cell-conditioned media. Rapamycin also inhibited the multiplication of colony-forming cells in suspension cultures containing IL-3 plus interleukin-1 (IL-1) or interleukin-11 (IL-11) plus KL. In vivo, treatment for 10 to 28 days with high doses of rapamycin (50 mg/kg/d, orally) had no effect on myelopoiesis in normal mice, as measured by bone marrow cellularity, proliferative capacity, and number of colony-forming progenitors. In contrast, the same treatment strongly suppressed the hematopoietic recovery normally seen 10 days after an injection of 5-fluorouracil (5- FU; 150 mg/kg, intravenously [i.v.]). Thus, rapamycin may be detrimental in myelocompromised individuals. In addition, the results suggest that the rapamycin-sensitive cytokine-driven pathways are essential for hematopoietic recovery after myelodepression, but not for steady-state hematopoiesis.

Sign in / Sign up

Export Citation Format

Share Document