Mesenchymal stem cells (MSCs) has been used as one of the new cell sources in osteochondral tissue engineering. It has been well known that control of their differentiation into chondrocytes plays a key role in developing engineered cartilages. Therefore, this study aims to develop a fundamental protocol to control the differentiation and proliferation of MSCs to construct an engineered cartilage. We compared the effects of three different culture conditions on cell proliferation, extracellular matrix formation and the mechanical response of engineered cartilage constructed using a collagen-based hybrid scaffold and human MSCs. The experimental results clearly showed that the combined culture condition of the chondrogenic differentiation culture and the chondrocyte growth culture exhibited statistically significant cell proliferation, ECM formation and stiffness responses as compared to the other two combinations. It is thus concluded that the combination of the differentiation culture with the subsequent growth culture is recommended as the culture condition for chondrogenic tissue engineering using hMSCs.