The following proposal is based upon the hypothesis that TIMP-1 and TIMP-3 display specific patterns of expression in uterine tissue during the reproductive cycle, that these patterns of expression are modulated by estradiol and/or progesterone, and that the steroid-induced expression of TIMP-3 is mediated at least in part through TIMP-1. To test this hypothesis, two specific aims are proposed. The experiments in Specific Aim 1 are designed to examine the effect of endogenous and exogenous estradiol-17beta and progesterone on the pattern of uterine TIMP- 1 and TIMP-3 expression as well as examine the effect of disruption of the TIMP-1 gene product on uterine TIMP-3 expression under these hormonal milieus. To accomplish this goal, uteri will be obtained from mature, reproductively cycling, and steroid-primed, immature female TIMP-1 deficient and wild- type mice and TIMP-1 and TIMP-3 mRNA localization and expression will be examined. Dual in-situ hybridization will be used to co- localize the expression of estrogen and progesterone receptors with that of TIMP-1 and TIMP-3 within specific uterine cell-types during the estrous cycle (mature mice) or during exposure to exogenous steroids (immature mice). Northern analysis will then be performed using uterine tissues from these same animals to provide quantitative analysis of the aforementioned parameters. As such, the studies outlined in Specific Aim 1 will allow us to qualitatively and quantitatively determine the effect of steroids and TIMP-1 on the spatiotemporal relationship among the steroid receptors and TIMP-1 and TIMP-3 in the uterus.
In Specific Aim 2, the mechanisms for steroidal regulation of uterine TIMP-1 and TIMP-3, as well as the involvement of steroid receptors and TIMP- 1 in this process, will be assessed using a cell culture model. Uterine stromal cells will be obtained from TIMP-1 deficient and wild-type mice, cells will be treated with estradiol-17beta and progesterone, and the spatiotemporal relationship among estrogen and progesterone receptors and TIMP-1 and TIMP-3 will be quantitated by Northern analysis. To further support the role of TIMP-1 as a regulator of TIMP-3 expression, stromal cells obtained from TIMP-1 deficient mice will be treated with exogenous TIMP-1 to restore the normal pattern of TIMP-3 expression detected in the wild-type mice. Taken together, these experiments will provide important information on the mechanisms of TIMP regulation and function in the mouse uterus. These experiments are timely and unique as they incorporate a novel model to examine the functions of TIMP-1, as well as provide information on a newly-described function of TIMP-1 which will have broad impact on the fields of uterine and TIMP biology.
| Nothnick, W B (2001) Reduction in reproductive lifespan of tissue inhibitor of metalloproteinase 1 (TIMP-1)-deficient female mice. Reproduction 122:923-7 |
| Nothnick, W B (2001) Disruption of the tissue inhibitor of metalloproteinase-1 gene in reproductive-age female mice is associated with estrous cycle stage-specific increases in stromelysin messenger RNA expression and activity. Biol Reprod 65:1780-8 |
| Nothnick, W B (2000) Disruption of the tissue inhibitor of metalloproteinase-1 gene results in altered reproductive cyclicity and uterine morphology in reproductive-age female mice. Biol Reprod 63:905-12 |
