OPN gene expression in PBMC, primary stroma, and stromal cell lines. (A) Total RNA was immediately isolated from peripheral blood mononuclear cells (PBMCs) and the buffy coat layer of normal bone marrow cells (BM). Remaining cells from the buffy coat were cultured for 2 to 3 weeks to establish primary LTCs, which contain both stroma and macrophages, at which time total RNA from the LTCs was isolated. OPN gene expression was determined by a Syber Green assay. (B) CD14+ cells (0.2 × 106 cells) were cultured alone or cocultured with 0.4 × 106 cells of HS-27a, HS-5, or human foreskin fibroblasts (HFFs) for 3 days. Total RNA of nonadherent CD14+ cells was harvested, and OPN gene expression was measured. The data represent the average ± SD for at least 2 samples.
