We have compared the effects of a general
matrix metalloproteinase (
MMP) inhibitor (
CT435) with those of a concentration-dependent specific
gelatinase inhibitor (
CT543; Ki < 20 nM) on
bone resorption in vitro. The test systems consisted of measuring: (i) the release of 45Ca2+ from prelabelled mouse calvarial explants; (ii) the release of 45Ca2+ from prelabelled osteoid-free calvarial explants co-cultured with purified chicken osteoclasts; and (iii) lacunar resorption by isolated rat osteoclasts cultured on ivory slices. Both
CT435 and
CT543 dose-dependently inhibited the release of 45Ca2+ from neonatal calvarial bones stimulated by either
parathyroid hormone or
1,25-dihydroxyvitamin D3. Moreover,
CT543 produced a 40% inhibition at a concentration (10(-8) M) selective for the inhibition of human
gelatinases A and B.
CT435 (10(-5) M) and
CT543 (10(-5) M) partially inhibited the release of 45Ca2+ from osteoid-free calvarial explants by chicken osteoclasts with a maximum of approximately 25% for unstimulated cultures, and approximately 36% for cultures stimulated by
interleukin-1 alpha (IL-1 alpha; 10(-10) M). Neither inhibitor prevented lacunar resorption on ivory by unstimulated rat osteoclasts, but the compounds produced a partial reduction in both the number and total surface area of lacunae in
IL-1 alpha-stimulated cultures, with maximal action
at 10(-5) M. Neither of the inhibitors affected
protein or
DNA synthesis, nor the
IL-1 alpha-stimulated secretion of the lysosomal
enzyme beta-glucuronidase. Immunocytochemistry demonstrated that isolated rabbit osteoclasts constitutively expressed
gelatinase A and synthesized
gelatinase B,
collagenase and
stromelysin, as well as the tissue inhibitor of matrix metalloproteinases-1 (TIMP-1) following
IL-1 alpha stimulation. These experiments have shown that in addition to
collagenase,
gelatinases A and B are likely to play a significant role in
bone resorption. They further suggest that
MMPs produced by osteoclasts are released into the sub-osteoclastic resorption zone where they participate in bone
collagen degradation.