Immune Netw.  2009 Feb;9(1):27-33. 10.4110/in.2009.9.1.27.

Production of TGF-beta1 as a Mechanism for Defective Antigen-presenting Cell Function of Macrophages Generated in vitro with M-CSF

• Lee JK
• Lee YR
• Lee YH
• Kim K
• Lee CK

• KMID: 2167994
• DOI: http://doi.org/10.4110/in.2009.9.1.27

Abstract

BACKGROUND: Macrophages generated in vitro using macrophage-colony stimulating factor (M-CSF) and interleukin (IL)-6 from bone marrow cells (BM-Mp) are defective in antigen presenting cell (APC) function as shown by their ability to induce the proliferation of anti-CD3 mAb-primed syngeneic T cells. However, they do express major histocompatibility (MHC) class I and II molecules, accessory molecules and intracellular adhesion molecules. Here we demonstrate that the defective APC function of macrophages is mainly due to production of TGF-beta1 by BM-Mp.
METHODS
Microarray analysis showed that TGF-beta1 was highly expressed in BM-Mp, compared to a macrophage cell line, B6D, which exerted efficient APC function. Production of TGF-beta1 by BM-Mp was confirmed by neutralization experiments of TGF-beta1 as well as by real time-polymerase chain reaction (PCR).
RESULTS
Addition of anti-TGF-beta1 monoclonal antibody to cultures of BM-Mp and anti-CD3 mAb-primed syngeneic T cells efficiently induced the proliferation of syngeneic T cells. Conversely, the APC function of B6D cells was almost completely suppressed by addition of TGF-beta1. Quantitative real time-PCR analysis also confirmed the enhanced expression of TGF-beta1 in BM-Mp.
CONCLUSION
The defective APC function of macrophages generated in vitro with M-CSF and IL-6 was mainly due to the production of TGF-beta1 by macrophages.

Keyword

macrophage; M-CSF; APC function; TGF-beta1

MeSH Terms

Antigen-Presenting Cells
Bone Marrow Cells
Cell Line
Histocompatibility
Interleukin-6
Interleukins
Macrophage Colony-Stimulating Factor
Macrophages
Microarray Analysis
T-Lymphocytes
Transforming Growth Factor beta1
Interleukin-6
Interleukins
Macrophage Colony-Stimulating Factor
Transforming Growth Factor beta1

Figure

• 

  Figure 1 Effects of various cytokines on the growth of B6D cells. B6D cells were cultured in the presence of the indicated amounts of the culture supernatant of TFGD cells (A) or with the indicated cytokines (B) for 3 days. The proliferation of B6D cells was determined by an XTT assay. The results show the mean±S.D. of three independent experiments.

• 

  Figure 2 Phenotypic differences between BM-Mp and B6D cells. The cells were stained with the indicated mAbs, washed and analyzed by flow cytometry. Levels of expression (thin line) were illustrated in comparison to isotype control (dotted line).

• 

  Figure 3 Comparison of the APC functions of BM-Mp and B6D cells. Syngeneic anti-CD3 mAb-primed T cells (1×105 cells/well) were cultured with the indicated number of BM-Mp or B6D cells. Proliferation of T cells was measured by [3H]-thymidine incorporation for the final 8 h of the culture period of 3 days. The results show the mean±S.D. of three independent experiments.

• 

  Figure 4 Recovery of APC function of BM-Mp by blocking with anti-TGF-β1 mAb. Syngeneic anti-CD3 mAb-primed T cells (1×105 cells/well) were cultured with BM-Mp (1×104 cells/well) in the presence of the indicated amounts of anti-TGF-β1 mAb. IgG is an isotype control for anti-TGF-β1 mAb. The results show the mean±S.D. of three independent experiments.

• 

  Figure 5 Inhibition of the APC function of B6D cells by addition of TGF-β1. Syngeneic anti-CD3 mAb-primed T cells (1×105 cells/well) were culture with B6D cells (1×104 cells/well) in the presence of the indicated amounts of rhTGF-β1. The results show the mean±S.D. of three independent experiments.

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