To develop a culture system for bone marrow (BM) cell expansion, we examined the effect of growth factors (GFs) on the proliferation and differentiation of BM cells cultured in three-dimensional (3D) scaffolds of porous polyvinyl formal (PVF) resin. Murine BM cells were cultured for 2 weeks in the PVF resin or in culture dishes as a control, in the presence or absence of 4 GFs (erythropoietin, stem cell factor, interleukin [IL]-3, and IL-6). These GFs remarkably stimulated cell proliferation both in PVF and dish cultures. In addition, the PVF cultures showed enhanced cell proliferation in comparison with the corresponding dish cultures. Moreover, PVF cultures with GFs revealed the highest number of colony-forming units and the highest percentage of hematopoietic progenitor cells (HPCs) among all the cultures examined. Therefore, this 3D PVF culture system with GFs is considered as a potential alternative method for the ex vivo expansion of HPCs.