The authors investigated the process of endochondral ossification quantitatively and objectively in respect to proliferation and apoptosis.

Fractures were made on the left tibiae of 72 male Sprague-Dawley rats. The fracture callus was harvested at the 5th, 7th, 9th, 11th, 14th, and the 21st day after fracture. Cellular DNA content was analyzed with image cytometry, and proliferative index was determined from the data. The Ki-67 antigen expression was semiquantitatively measured by the immunohisto-chemical method. TUNEL was used for in situ localization of apoptotic cells. The expression of cell cycle inhibitors, P21 and P27, was investigated with Northern blotting.

The proliferation index was highest on the 5th day, then gradually decreased until the 11th day. The expression of Ki-67 antigen gradually decreased with time. Apoptotic cells increased in accordance with enhanced bone formation within chondroid callus. The expression of p21 and p27 was highest on the 11th and the 14th day.

These findings show that proliferative activity decreased with the reduction of mesenchymal tissue and the appearance of mature chondroid tissue. The apoptosis of hypertrophic chondrocytes occurred in accordance with enhanced bone formation. P21 and P27 had a certain role in the differentiation of chondrocytes.