Granulocyte-macrophage colony-stimulating factor (GM-CSF) promotes dendritic cell differentiation from precursors, and consequently, enhances the antigen presentation process and adaptive immune responses. With such functions, GM-CSF has been used as immunotherapy in combination with radiotherapy for cancer treatment to augment the survival and activity of immune cells. However, an immune-suppressive tumor microenvironment may cause anergy of T cells. It has also been reported that GM-CSF contributes to the development of myeloid-derived suppressor cells from the precursors. In this study, to analyze the combined effect of GM-CSF and released factors from cancer cells after gamma-ray irradiation on bone marrow cell differentiation and dynamics, we established an in vitro culture system using mouse bone marrow cells, GM-CSF, and conditioned medium from gamma ray irradiated mouse melanoma B16 cells at 24 Gy. We analyzed the gene expression changes of the bone marrow-derived cells on day 6. The results showed that GM-CSF dose-dependently enhanced the differentiation of macrophages from bone marrow cells, their antigen-presenting function and polarization to type I. The results implied the induced macrophages from the bone marrow may potentially contribute to tumor immune responses in a systemic manner when GM-CSF is boosted during photon-beam radiation therapy.