Molecular characterization and hormonal regulation of tissue inhibitor of metalloproteinase 1 in goat ovarian granulosa cells.

Tissue inhibitor of metalloproteinase 1 (TIMP1) belongs to a group of endogenous inhibitors that control the activity of matrix metalloproteinases and other metalloproteinases. TIMP1 is ubiquitously expressed and implicated in many physiological and pathologic processes. In this study, the full-length complementary DNA of goat (Capra hircus) Timp1 was cloned from adult goat ovary for the first time to better understand the regulatory role of TIMP1. The putative TIMP1 protein shared a high amino acid sequence identity with other species. Real-time polymerase chain reaction results showed that Timp1 was widely expressed in adult goat tissues, and messenger RNA expression was higher in the ovary than in other tissues; meanwhile, increasing expression of Timp1 was also discovered during the process of follicle growth and corpus luteum. We then investigated Timp1 expression patterns in different types of ovarian follicular cells from goats. In small or large antral follicles, Timp1 expression was higher (P < 0.05) in theca cells than in granulosa cells, cumulus cells, and oocytes. Increasing expression of Timp1 in theca and granulosa cells was observed as the variation of the follicle size. Immunohistochemical analyses further revealed the presence of the TIMP1 proteins in follicles at all antral stages of development. The most intense staining for TIMP1 was observed in the theca cells and granulosa cells of large antral follicles and corpus luteum. Timp1 was highly (P < 0.05) induced in granulosa cells in vitro after treatment with the luteinizing hormone agonist, human chorionic gonadotropin. Treatments with forskolin, phorbol 12-myristate 13-acetate, or phorbol 12-myristate 13-acetate + forskolin could also stimulate Timp1 messenger RNA expression. The effects of human chorionic gonadotropin were reduced (P < 0.05) by the inhibitors of protein kinase A, protein kinase C, MAPK kinase, or p38 kinase, indicating that Timp1 expression could be adjusted by luteinizing hormone-initiated activation of these signaling mediators. Our results suggested that TIMP1 may be involved in regulating ovarian follicle development and ovulation.