Tetracyclines, including tetracycline (TC), minocycline (MINO), and doxycycline (DOXY), were widely used as topical therapy in treating periodontitis, which is an infectious disease of the gingival crevice caused by periodontopathic bacteria. In addition, TC is used during bone grafting procedures because of its anticollagenase activity, antibacterial effect, and fibroblast-stimulatory properties.In this study, the effects of TC on osteoprecursor cells were evaluated. The cytotoxic effect was determined by testing cell viability. Differentiation and mineralization were evaluated using an alkaline phosphatase activity test and alizarin red S staining. In addition, protein expressions related to bone formation, such as bone morphogenetic protein 2 (BMP-2), estrogen receptor α (ER-α), and estrogen receptor β (ER-β), were evaluated by using Western blot analysis.The morphology of the cells seemed normal, and their viability was not significantly affected when they were treated with 10 μM MINO and 10 μM DOXY. Cells treated with 10 μM DOXY showed alkaline phosphatase activity that was comparable to the control. Results from the Western blot analysis showed that TC, MINO, and DOXY reduced the expression of BMP-2 and ER-β. Normalization of the protein expressions showed that 10 μM DOXY retained 87% in BMP-2 and 85% in ER-β.Higher levels of these agents led to a dose-dependent decrease of cellular differentiation and protein expression. There are several commercially available agents for TC, which has to be considered when applying the TC in local delivery applications.