INHIBITION OF BONE-RESORPTION IN-VITRO BY SELECTIVE INHIBITORS OF GELATINASE AND COLLAGENASE

Two low-molecular-mass inhibitors of matrix metalloproteinases (MMPs), CT1166, a concentration-dependent selective inhibitor of gelatinases A and B, and Ro 31-7467, a concentration-dependent selective inhibitor of collagenase, were examined for their effects on bone resorption an d type-I collagenolysis. The test systems consisted of measuring (1) t he release of [H-3]proline from prelabelled mouse calvarial explants; (2) the release of C-14 from prelabelled type-I collagen films by mous e calvarial osteoblasts; and (3) lacunar resorption by isolated rat os teoclasts cultured on ivory slices, In 24 h cultures, CT1166 and Ro 31 -7467 inhibited both interleukin-1 alpha-(IL-1 alpha; 10(-10) M) and 1 ,25-dihydroxyvitamin D-3 (10(-8) M)-stimulated bone resorption in cult ured neonatal mouse calvariae at concentrations selective for the inhi bition of gelatinase (10(-9) M for CT1166) and collagenase (10(-8) M f or Ro 31-7467) respectively. For each compound the inhibition was dose -dependent, reversible, and complete at a 10(-7) M concentration, Howe ver, CT1166 (10(-9) M) and Ro 31-7467 (10(-8) M) in combination were r equired to completely abolish a 96 h culture period. Neither of the in hibitors affected protein synthesis, DNA synthesis nor the IL-1 alpha- stimulated secretion of the lysosomal enzyme, beta-glucuronidase, Both CT1166 and Ro 31-7467 partially inhibited IL-1 alpha-stimulated lacun ar resorption by isolated osteoclasts, but were without effect on unst imulated lacunar resorption, Rodent osteoclasts produced collagenase a nd gelatinases-A and -B activity. In contrast the substrate used to as sess osteoclast lacunar resorption contained no detectable collagenase or gelatinase activity, Both compounds dose-dependently inhibited 1,2 5-dihydroxyvitamin D-3 (10(-8) M)-stimulated degradation of type-I col lagen by mouse calvarial osteoblasts; however, complete inhibition of collagenolysis was only achieved at concentrations at which CT1166 and Ro 31-7467 act as general MMP inhibitors. This study demonstrates tha t collagenase and gelatinases A and/or B participate in bone resorptio n. While these MMPs may be primarily involved in osteoid removal, we c onclude that they may also be released by osteoclasts, where they part icipate in bone collagen degradation within the resorption lacunae.