The proteolytic functions of the matrix-degrading enzymes, such as ADAMTS (A disintegrin and metalloproteinases) and MMP (Matrix metalloproteinase), play a crucial role in development. This study investigated the proteolytic activity of different types of bone cells, focusing on the expression of MMP-9, throughout the development of the femur in rabbit embryos. The femur formed the physeal growth cartilage, which facilitated the process of endochondral bone development. Osteoblasts are situated on the outer layer of bone spicules and within the periosteum. Osteoclasts are found within the lacunae of the bone matrix, and they have also been observed in the ossification center. Osteoclasts, osteocytes, and osteoblasts all exhibit MMP-9 expression with different intensity of staining with the perichondral stem cells showing the greatest value. MMP-9 is essential in bone formation. Utilizing MMP-9 to modify the extracellular matrix could potentially serve as a favorable approach for identifying treatment targets in bone diseases.