The study was to produce a novel hybrid poly-(lactic-co-glycolic acid) (PLGA)-gelatin/chondroitin/hyaluronate (PLGA-GCH) scaffold and evaluate its potentials in cartilage repair. The porous PLGA-GCH scaffold was developed to mimic the natural extra cellular matrix of cartilage. The differentiated mesenchymal stem cells (MSCs) seeded on PLGA-GCH or PLGA scaffold were incubated in vitro and showed that, compared to PLGA scaffold, the PLGA-GCH scaffold significantly augmented the proliferation of MSCs and GAG synthesis. Then autologous differentiated MSCs/PLGA-GCH was implanted to repair full-thickness cartilage defect in rabbit, while MSCs/PLGA for the contra lateral cartilage defect (n=30). Fifteen additional rabbits without treatment for defects were used as control. Histology observation showed the MSCs/PLGA-GCH repair group had better chondrocyte morphology, integration, continuous subchondral bone, and much thicker newly formed cartilage compared with MSCs/PLGA repair group 12 and 24 weeks postoperatively. There was a significant difference in histological grading score between these two groups, which both showed much better repair than control. The present study implied that the hybrid PLGA-GCH scaffold might serve as a new way to keep the differentiation of MSCs for enhancing cartilage repair.