A Tissue Inhibitor of Metalloproteinases and α-Macroglobulins in the Ovulating Rat Ovary: Possible Regulators of Collagen Matrix Breakdown1

Abstract

Immature female Sprague-Dawley rats were primed with 20 IU eCG at 28 days of age and treated with 10 IU hCG 48 h later. Ovulation followed at 12–14 h. Ovaries were extracted at various times after hCG by use of Triton X-100 and 10 mM CaCl₂ (Triton extract) followed by heating to 60°C for 6 min with 0.1 M CaCl₂ in 50 mM Tris/0.15 M NaCl, pH 7.5 (heat extract). These extracts were tested for their ability to inhibit tissue metalloproteinases by use of the small uterine metalloproteinase (UMP) of the rat. The ovary contains three plasma-derived inhibitors (α₁-macroglobulin [α₁M], α₂-macroglobulin [α₂M], and α₁ inhibitor₃ [α₁I₃]) and one tissue-derived inhibitor of the tissue inhibitor of metalloproteinases (TIMP) family. α₁I₃ was shown to inhibit UMP and rat collagenase. The concentration of the tissue inhibitor rose 5-fold from 0.6 μg UMP blocked per gram wet tissue in ovaries not primed with eCG to 3.2 μg UMP blocked at 8 h after hCG. Over this same time interval, the sum of α₁M + α₂M per gram of ovary rose 7-fold from 3.2 to 22.4 μg UMP inhibited and α₁I₃ rose 2-fold from 4.4 to 10.7 μg UMP inhibited. The increases in the tissue inhibitor are interpreted to be due to increased synthesis by the tissue, whereas the changes in α-macroglobulins are postulated to be due to increased vascularity and increased permeability of the vessels. Since the increase in tissue inhibitor is proportionally about one half of the increase previously reported for collagenase (Curry et al., Biol Reprod 1985; 33:981), it is hypothesized that collagenase and other metalloproteinases may escape the control of TIMP, at least in focal regions, thereby digesting collagen of the follicle wall and permitting escape of the ovum.