Effect of Thrombospondin-1 on Apoptosis of Human Megakaryocytic Leukemia Cells

Abstract Background: Thrombospondin 1 (TSP-1) is an extracellular matrix protein that interacts with a wide array of ligands including cell receptors, growth factors, cytokines, and proteases to regulate various physiological and pathological processes. TSP-1 induces apoptosis of endothelial and cancer cells via its receptor CD36. This study was to investigate the effect of TSP-1 on apoptosis of human megakaryocytic leukemia cell line Meg-01 and its possible mechanism. Methods: The expression of CD36 antigen in Meg-01 cells was detected by flow cytometry and immunocytochemistry. Meg-01 cells were cultured for 48 hours with TSP-1 and CD36 antibody FA6-152 at different concentrations, to investigate the effect of TSP-1 on the growth of megakaryocytes. The early apoptosis and the activity of the apoptotic protease caspase-3 were detected by flow cytometry. Bone marrow cells of mice were cultured for CFU-MK and stained with acetylcholine to detect the differentiation of megakaryocytes. Results: CD36 antigen was detected on the surface of Meg-01 cells by flow cytometry and immunocytochemistry. TSP-1 (5 μg/mL) inhibited the proliferation of Meg-01 cells, but not M-07e cells (CD36 -). After the addition of CD36 antibody FA6-152 (5, 10 and 25 μg/mL), the inhibitory effect of TSP-1 was significantly reduced. TSP-1 (2.5, 5 and 7.5 μg/mL) exerted a pro-apoptotic effect by increasing the expression of Annexin V (P<0.01) and caspase-3 activation (P<0.01). Addition of FA6-152 (25 μg/mL) can significantly reduce the apoptosis induced by TSP-1 in Meg-01 cells. In addition, TSP-1 (5, 10 and 25 μg/mL) repress the formation of CFU-MK in mouse bone marrow cells, while β-TG did not. This repression was relieved efficiently by FA6-152 (25 μg/mL). Conclusion: TSP-1 could inhibit the proliferation of Megakaryocyte cell line Meg-01 cells, and induce it`s apoptosis. TSP-1 may induce apoptosis of Meg-01 cells via CD36 or caspase-3, which provides a potential new drug choice for clinical treatment of megakaryocytic leukemia. Disclosures No relevant conflicts of interest to declare.

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The use of cationized ferritin to measure cell surface charge of mouse bone marrow cells by flow cytometry

Histochemistry ◽

10.1007/bf00482978 ◽

1986 ◽

Vol 84 (4-6) ◽

pp. 454-461 ◽

Cited By ~ 3

Author(s):

J. G. J. Bauman ◽

E. Bouwman

Keyword(s):

Bone Marrow ◽

Flow Cytometry ◽

Cell Surface ◽

Surface Charge ◽

Bone Marrow Cells ◽

Mouse Bone Marrow ◽

Cationized Ferritin ◽

Cell Surface Charge ◽

Mouse Bone Marrow Cells ◽

Marrow Cells

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The Growth Inhibition and Apoptosis Induction of Acute Myeloid Leukemia Blast Population by the Blocking IGF-IR Pathway.

Blood ◽

10.1182/blood.v108.11.4515.4515 ◽

2006 ◽

Vol 108 (11) ◽

pp. 4515-4515

Author(s):

Si Sun ◽

Yanli He ◽

Xingbing Wang ◽

Wei Liu ◽

Jun Liu ◽

...

Keyword(s):

Bone Marrow ◽

Flow Cytometry ◽

Acute Myeloid Leukemia ◽

Myeloid Leukemia ◽

Bone Marrow Cells ◽

Pi3k Pathway ◽

Leukemia Cell Line ◽

Free System ◽

Acute Myeloid ◽

Marrow Cells

Abstract The insulin-like growth factor-1receptor (IGF-1R) is overexpressed in a variety of tumors and has been associated with cancer development. Here, we analysis the IGF-IR expression on the bone marrow cells from 45 newly diagnosed patients with acute myeloid leukemia (AML) by flow cytometry. IGF-1R universally expressed on AML blasts and the leukemia cell line HL-60, did not show significant correlation with FAB subtypes. However, the bone marrow cells from AML patients with high myeloblast counts (>80%) generally showed brighter IGF-IR expressions, which indicated the IGF-IR pathway might play an important role for AML blast proliferation and survival. Indeed, blocking the IGF-1R pathway by neutralizing monoclonal antibodies could reduce the proliferation of HL-60 cells by 38.28% at 48 hr. This inhibitory effect on blast growth was observed in 4 of 5 AML samples. In the same IGF-1R blocking treatment, the apoptosis of HL-60 cells was significantly induced, resulting in apoptosis of 57% of the cell population with the measurement of Annexin V vs PI staining by flow cytometry. The control contained only 20% apoptotic cells. We also demonstrated that the blockade of the IGF-1R pathway inhibited the phophorylation of the PI3K pathway component Akt in HL-60 cells when cultured in a serum free system with a supplement of 50ng/ml exogenous IGF. Since PI3K pathway activation greatly contributes to the proliferation, survival and drug resistance of AML, it is of interest to study whether blockading IGF-IR could also inhibit the PI3K pathway in primary AML blasts and synergize other anti-leukemia agents to improve the therapeutic effectiveness. Conclusions: IGF-IR may play an important role in the proliferation and survival of the AML blast population; Blocking the IGF-IR pathway could significantly inhibit the growth of AML blasts and considerably induce the apoptosis of AML blasts; IGF-IR could become a critical molecular target in anti-leukemia drug discovery.

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Lipid accumulation and production of colony-stimulating activity by the 266AD cell line derived from mouse bone marrow

Blood ◽

10.1182/blood.v61.2.397.397 ◽

1983 ◽

Vol 61 (2) ◽

pp. 397-402 ◽

Cited By ~ 15

Author(s):

DL Hines

Keyword(s):

Bone Marrow ◽

Cell Line ◽

Stromal Cells ◽

Bone Marrow Cells ◽

Collagen Gel ◽

Mouse Bone Marrow ◽

Cell Sheets ◽

Colony Stimulating Activity ◽

Confluent Cell ◽

Marrow Cells

Abstract The availability of cloned lines of bone marrow stromal cells could facilitate the analysis of their role in hemopoietic cell development. The 266AD cell line was isolated from a colony of lipid-accumulating bone marrow cells growing in a collagen gel. 266AD cells have subsequently been maintained by passage in tissue culture plastic flasks about every 10 days for greater than 10 mo. Subconfluent cultures of cells are fibroblast-appearing, but in confluent cell sheets, prominent foci of lipid-containing cells develop in both uncloned and four separate cloned cell lines. Supernatants from confluent cultures containing lipid-laden cells contain granulocyte- macrophage colony-stimulating activity (GM-CSA) for normal bone marrow cells and can induce differentiation of Abelson virus transformed murine promonocytic leukemia cells. 266AD cells were originally isolated in the presence of hydrocortisone, but hydrocortisone is not necessary for lipogenesis to occur. Growth of bone marrow cells in a collagen gel matrix provided a way to isolate stromal cells, and the 266AD cell line provides a means to examine the relationships between stromal cell lipogenesis and regulation of granulopoiesis.

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A novel growth-factor–dependent myeloid cell line derived from mouse bone marrow cells contains progenitors endowed with high proliferative potential

Experimental Hematology ◽

10.1016/s0301-472x(98)00013-7 ◽

1999 ◽

Vol 27 (1) ◽

pp. 88-98 ◽

Cited By ~ 2

Author(s):

V Revol

Keyword(s):

Bone Marrow ◽

Growth Factor ◽

Cell Line ◽

Bone Marrow Cells ◽

Myeloid Cell ◽

Proliferative Potential ◽

Mouse Bone Marrow ◽

Myeloid Cell Line ◽

Mouse Bone Marrow Cells ◽

Marrow Cells

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Lipid accumulation and production of colony-stimulating activity by the 266AD cell line derived from mouse bone marrow

Blood ◽

10.1182/blood.v61.2.397.bloodjournal612397 ◽

1983 ◽

Vol 61 (2) ◽

pp. 397-402

Author(s):

DL Hines

Keyword(s):

Bone Marrow ◽

Cell Line ◽

Stromal Cells ◽

Bone Marrow Cells ◽

Collagen Gel ◽

Mouse Bone Marrow ◽

Cell Sheets ◽

Colony Stimulating Activity ◽

Confluent Cell ◽

Marrow Cells

The availability of cloned lines of bone marrow stromal cells could facilitate the analysis of their role in hemopoietic cell development. The 266AD cell line was isolated from a colony of lipid-accumulating bone marrow cells growing in a collagen gel. 266AD cells have subsequently been maintained by passage in tissue culture plastic flasks about every 10 days for greater than 10 mo. Subconfluent cultures of cells are fibroblast-appearing, but in confluent cell sheets, prominent foci of lipid-containing cells develop in both uncloned and four separate cloned cell lines. Supernatants from confluent cultures containing lipid-laden cells contain granulocyte- macrophage colony-stimulating activity (GM-CSA) for normal bone marrow cells and can induce differentiation of Abelson virus transformed murine promonocytic leukemia cells. 266AD cells were originally isolated in the presence of hydrocortisone, but hydrocortisone is not necessary for lipogenesis to occur. Growth of bone marrow cells in a collagen gel matrix provided a way to isolate stromal cells, and the 266AD cell line provides a means to examine the relationships between stromal cell lipogenesis and regulation of granulopoiesis.

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