Non-porous poly(lactic-co-glycolic acid) (PLGA) microspheres have been previously proposed as an injectable scaffold for in vivo cartilage tissue engineering. In this study, we tested whether using open macroporous PLGA microspheres as an injectable scaffold for in vivo cartilage tissue engineering provides a larger surface area for cell adhesion and a larger void space for cartilage tissue regeneration and, thus, regeneration of higher quality cartilage than non-porous PLGA microspheres. Rabbit chondrocytes were mixed with non-porous or macroporous PLGA microspheres and injected immediately through 18-gauge needles into subcutaneous sites in athymic mice. Six weeks after implantation, chondrocytes implanted using both types of PLGA microspheres formed solid, white cartilaginous tissues. Histological analysis of the implants with hematoxylin and eosin, safranin O and Masson's trichrome staining confirmed cartilaginous tissue formation. The portion of cartilage tissue area in the implant cross-sectional area was significantly higher in the macroporous PLGA microsphere group than in the non-porous PLGA microsphere group (88.9% versus 34.6%, P < 0.001). Importantly, the collagen (P < 0.01) and glycosaminoglycan (P < 0.01) contents of the implants were significantly higher in the macroporous PLGA microsphere group than in the non-porous PLGA microsphere group. We conclude that an open macroporous PLGA microsphere scaffold may be useful in cartilage regeneration through minimally-invasive surgical procedures in orthopedic applications.