PRODUCTION OF BOTH 92-KDA AND 72-KDA GELATINASES BY BONE-CELLS

We investigated the ability of murine bone organ cultures and osteoblast-like bone cells to produce 72- and 92-kDa gelatinase. 4-6 day newborn mouse calvaria cultures were found to release gelatinase activity into their conditioned medium (CM). This activity was increased by four stimulators of resorption, tumor necrosis factor alpha (TNF), interleukin-1 alpha (IL-1), parathyroid hormone (PTH) and the active phorbol ester, 12-O-tetradecanoylphorbol-13-acetate (TPA). Both the 72- and 92-kDa forms of gelatinase were produced by murine bone cultures. In unstimulated bones 72-kDa gelatinase activity was approximately equal to that of the 92-kDa enzyme. IL-1, TNF, PTH and TPA all increased 92-kDa gelatinase activity in the CM of the bone cultures by about 2- to 2.5-fold. In addition TPA and IL-1 also increased 72-kDa gelatinase activity. In unstimulated osteoblast-like MC3T3-E1 cell cultures 72-kDa gelatinase enzyme activity was much greater than 92-kDa activity and was not substantially regulated (<40% change) by IL-1, TNF or PTH. In contrast, these agents stimulated 92-kDa gelatinase activity by 2- to 5-fold. As with the MC3T3-E1 cells, primary cells constitutively produced both 72-kDa and 92-kDa gelatinase. This was true for cells with both the most differentiated osteoblast-like phenotype (populations 3 and 4) and the least osteoblast-like phenotype (populations 1 and 2). In unstimulated cultures of all 4-primary populations, 92-kDa gelatinase production was less than 72-kDa and IL-1, TNF and PTH had only small effects on 72-kDa production in any of the populations (<60% change). However, IL-1, and TNF-stimulated 92-kDa gelatinase production in all populations by 2.7 to 5.7 fold; while PTH completely inhibited 92-kDa gelatinase activity in population 1 and had no effect on populations 2-4. These results demonstrate that gelatinase activity is released by bone organ cultures and osteoblast-like cells in a regulated manner. This finding suggests that these enzymes are involved in the mechanisms regulating bone turnover.