MOLECULAR-CLONING OF AN OVINE OVARIAN TISSUE INHIBITOR OF METALLOPROTEINASES - ONTOGENY OF MESSENGER-RIBONUCLEIC-ACID EXPRESSION AND IN-SITU LOCALIZATION WITHIN PREOVULATORY FOLLICLES AND LUTEAL TISSUE

Secretion of a tissue inhibitor of metalloproteinases (TIMP-1) is initiated by ovine preovulatory follicles after the gonadotropin surge. In addition, TIMP-1 is a major secretory product of ovine corpora lutea. We have isolated an approximately full-length cDNA clone for TIMP-1 from an ovine luteal cDNA library. The 887-basepair cDNA obtained was 95%, 86%, and 77% identical to the reported nucleotide sequences of bovine, human, and mouse TIMP-1 cDNAs, respectively. Total cellular RNA was isolated from preovulatory follicles collected before (presurge; n = 5) or 12-14 h after (postsurge; n = 4) a LHRH-induced gonadotropin surge (36 h after prostaglandin F-2 alpha-induced luteolysis); from luteal tissue collected on days 3, 7, 10, 13, and 16 postestrus (n = 5, 5, 4, 5, and 5, respectively); and from purified populations of small (n = 4) and large (n = 3) luteal cells. Concentrations of TIMP-1 mRNA (picograms per pg tissue DNA) were increased in preovulatory follicles after exposure to a gonadotropin surge (P less than or equal to 0.01). TIMP-1 mRNA was localized primarily to the granulosa layer of postsurge follicles by in situ hybridization. Concentrations of TIMP-1 mRNA in luteal tissue did not differ throughout the luteal phase (P = 0.07). However, TIMP-1 mRNA was localized predominantly to specific cells located in the connective tissue surrounding and within day 3 corpora lutes. In situ hybridization of day 10 corpora lutes localized TIMP-1 mRNA predominantly to specific cells that were randomly dispersed throughout the luteal tissue. TIMP-1 mRNA was expressed by purified populations of both small and large luteal cells collected from day 10 corpora lutes. Concentrations of TIMP-1 mRNA (picograms per mu g DNA) were greater in the large luteal cell populations (P less than or equal to 0.0001). We conclude that 1) expression of TIMP-1 mRNA by the granulosa layer of ovine preovulatory follicles increased after the gonadotropin surge, whereas TIMP-1 mRNA concentrations during the luteal phase remained constant; 2) during the luteal phase, TIMP-1 mRNA was localized to specific cells surrounding (day 3) or located within (day 10) the corpus luteum; and 3) expression of TIMP-1 mRNA was greatest in large luteal cells.