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In Vitro Differentiation of Mouse Granulocyte-macrophage-colony-stimulating Factor GM-CSF-producing T Helper THGM Cells
Published on: September 10, 2018
Extended Culture of Bone Marrow with Granulocyte Macrophage-Colony Stimulating Factor Generates Immunosuppressive
Hye Young Na1, Moah Sohn1,2, Seul Hye Ryu1,2
1Laboratory of Immunology, Severance Biomedical Science Institute, Yonsei University College of Medicine, Seoul 03722, Korea.
Abstract:
Bone marrow-derived dendritic cells (BM-DCs) are generated from bone marrow (BM) cells cultured with granulocyte macrophage-colony stimulating factor (GM-CSF) for a week. In this study we investigated the effect of duration on the BM culture with GM-CSF. Within several months, the cells in the BM culture gradually expressed homogeneous levels of CD11c and major histocompatibility complex II on surface, and they became unable to stimulate allogeneic naïve T cells in mixed lymphocyte reaction (MLR). In addition, when the BM culture were sustained for 32 wk or longer, the BM cells acquired ability to suppress the proliferation of allogeneic T cells in MLR as well as the response of ovalbumin-specific OT-I transgenic T cells in antigen-dependent manner. We found that, except for programmed death-ligand 1, most cell surface molecules were expressed lower in the BM cells cultured with GM-CSF for the extended duration. These results indicate that BM cells in the extended culture with GM-CSF undergo 2 distinct steps of functional change; first, they lose the immunostimulatory capacity; and next, they gain the immunosuppressive ability.
Insights
Extended culture of bone marrow cells with GM-CSF initially impairs their ability to stimulate T cells. Prolonged culture then induces immunosuppressive properties in these cells.
Area of Science:
- Immunology
- Cell Biology
Background:
- Bone marrow-derived dendritic cells (BM-DCs) are crucial for immune responses.
- Standard protocols involve culturing bone marrow (BM) cells with granulocyte macrophage-colony stimulating factor (GM-CSF).
Purpose of the Study:
- To investigate the impact of extended culture duration on BM cells stimulated with GM-CSF.
- To characterize functional changes in BM cells with prolonged GM-CSF exposure.
Main Methods:
- Culturing BM cells with GM-CSF for extended periods (several months, including 32 weeks).
- Assessing cell surface molecule expression (e.g., CD11c, MHC II, PD-L1).
- Evaluating T cell stimulatory and suppressive capacities using mixed lymphocyte reactions (MLR) and antigen-specific T cell assays.
Main Results:
- Extended GM-CSF culture led to homogeneous expression of CD11c and MHC II.
- Cells lost the ability to stimulate allogeneic naive T cells in MLR.
- BM cells cultured for ≥32 weeks gained the capacity to suppress allogeneic T cell proliferation in an antigen-dependent manner.
- Most cell surface molecules, except PD-L1, were downregulated with extended culture.
Conclusions:
- BM cells cultured with GM-CSF undergo distinct functional transitions over time.
- Initial prolonged culture leads to loss of immunostimulatory function.
- Further extended culture confers immunosuppressive properties on BM cells.
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