Production of collagenase inhibitor by the growth cartilage of embryonic chick bone: isolation and partial characterization.

Production of collagenase and collagenase inhibitors by the explants of epiphyseal, metaphyseal and diaphyseal regions of embryonic chick limbs during development has been investigated. Collagenase-inhibitory activity was first detected in the culture medium of the diaphyseal region of limbs at stage 36 where cartilage matrix erosion began to occur. However, neither active nor latent collagenase was detected in the media of any regions examined. At stage 38 and later, the maximum production of the inhibitory activity was observed in the explants of metaphyseal region (growth cartilage), while collagenase production was only in the diaphyseal region. Two collagenase inhibitors were isolated and purified approximately 150 fold from the culture medium of stage 38 and 43 metaphyseal regions of limbs by anion- and cation-exchange chromatography followed by gel filtration. The inhibitors are cationic proteins with the same molecular weight of approximately 25,000, but slight difference in molecular charge. They are heat-stable and inhibit collagenases from tadpole skin, chick bone and skin and human granulocytes and gelatinases from human granulocytes and chick skin as well as trypsin, but not Clostridium histolyticum collagenase. A possible function of the inhibitors in multistep regulations of the collagen-degrading enzyme system at the region of osteo-chondral transition is discussed.