Expression of messenger ribonucleic acid encoding tissue inhibitor of metalloproteinases-2 within ovine follicles and corpora lutea.

Abstract

Metalloproteinase inhibitors, such as tissue inhibitors of metalloproteinases 1 and 2 (TIMP-1 and -2), play a key role in the regulation of metalloproteinases that modify extracellular matrix composition. Although expression of TIMP-1 within ovarian tissues has been well characterized, little information is available regarding expression of TIMP-2. The objective of the present studies was to characterize the ontogeny and localization of TIMP-2 messenger RNA (mRNA) within ovine preovulatory follicles and luteal tissue. Total cellular RNA was isolated from preovulatory follicles collected before (presurge; n = 3), or 12-14 h after (post surge; n = 4) an LHRH-induced gonadotropin surge, and from luteal tissue collected on days 3, 7, 10, 13, and 16 post estrus (n = 5, 5, 4, 5, and 5, respectively). TIMP-2 mRNA was expressed by both presurge and postsurge follicles, and expression did not increase after the gonadotropin surge (P = 0.44). In situ hybridization localized TIMP-2 mRNA primarily to the thecal layer of post-surge follicles (n = 3). TIMP-2 mRNA was also localized in a heterogeneous distribution within corpora lutea collected on days 3 and 10 post estrus (n = 3 each). Concentrations of TIMP-2 mRNA (picograms per microgram tissue DNA) were greater in corpora lutea collected during the early luteal phase (days 3 and 7) than the late luteal phase (day 16; P < 0.05). TIMP-2 mRNA was detected in purified populations of both small (n = 4) and large (n = 3) luteal cells, and mRNA concentrations (picograms per microgram DNA) were greater in the large luteal cells (P < or = 0.0002). In addition, immunoreactive TIMP-2 (approximately 21,000 M(r)) was detected by Western blot analysis of ovine luteal cell secreted proteins. We conclude that 1) TIMP-2 mRNA is expressed by the thecal layer of ovine preovulatory follicles and expression is not increased by the preovulatory gonadotropin surge; 2) expression of TIMP-2 mRNA is maximal during the early luteal phase; and 3) expression of TIMP-2 mRNA is greatest in large luteal cells.