Tissue engineering of bone may supersede the need in the future of autograft procedures to treat bone defects resulting from trauma or developmental diseases. A Rotational Oxygen-Permeable Bioreactor System (ROBS) has recently been developed in our laboratory to reproduce dynamic and gas-permeable culture conditions that would supply optimal oxygen and continuous loading to cell/polymer constructs in culture. The cell culture media in ROBS were examined at 1, 24 and 48 h to evaluate the kinetics of pO₂, pCO₂ and pH without culturing cells. The results were compared to the kinetics in 100 mm diameter cell culture dishes (Control I: static, gas permeable) and 50 ml centrifuge tubes (Control II: dynamic, non-gas permeable). The results showed the same kinetics in ROBS and Control I, whereas Control II failed to maintain the gas conditions of the media. Next, osteoblasts derived from mesenchymal stromal cells (MSCs) of neonatal rats were cultured in three-dimensional poly(dl-lactide-co-glycolide) (PLGA) foams using ROBS to study the effectiveness of this bioreactor system to support cell growth and differentiation. Mineralization was observed within 2 weeks of culture and was shown throughout the polymer at 7 weeks with embedded osteocytic cells. This study demonstrates the usefulness of ROBS for in vitro bone tissue engineering.