In experiment 3, PF (peritoneal fluid) was collected from Endo and Sham rats from experiment 2. Sham rats than in ovaries from Sham or TIMP1 antibody-treated Endo rats. In experiment three, control rats (no surgery) treated with Endo PF experienced fewer follicles and CLs and increased TIMP1 localization in the ovarian theca whereas treatment with Endo PF stripped SYP-5 of TIMP1 or with Sham PF experienced no effect, providing further evidence that endometriotic TIMP1 sequesters in the ovary and inhibits MMPs necessary for ovulation. Collectively, these results showed that excessive TIMP1 was deleterious to ovulation and embryo development. Thus, novel TIMP1-modulating therapies may be developed to alleviate infertility in women with endometriosis. Keywords: embryo,; endometriosis,; infertility,; ovary,; TIMP1 Modulating peritoneal levels of TIMP1 affects ovarian function and embryo development in rats with surgically induced endometriosis. INTRODUCTION Endometriosis is usually a gynecological disease that causes pain and infertility in women of reproductive age. Clear mechanisms causing the endometriosis-associated infertility have not been strongly established. Infertility in women with endometriosis may be associated with delicate, explicit or multifaceted abnormalities [1C8]. Anomalies have been recognized in the ovary such as reduced rates of follicular growth, functional capacity of the preovulatory follicle, and early luteal function [1, 2, 4, 5]; in gametes and embryos, including reduced rates of fertilization and defects in embryo development [1, 6, 8C10]; and in endometrial function [7, 11]. Because of the ethical limitations of performing controlled studies of infertility in women with endometriosis, animal models provide a useful tool to study risk factors, prevalence, and the pathogenesis and pathophysiologies of endometriosis [12, 13]. Rats with surgically induced endometriosis (Endo) display pathophysiologies much like those of primates and humans with the disease, including pain and infertility [12C14]. An association between the presence of ectopic endometriotic implants and reduced fecundity in rats has been explained [13, 15C17]. Expression of matrix metalloproteinases (MMPs) and their inhibitors (tissue inhibitors of metalloproteinases, TIMPs) is usually reportedly involved in the pathogenesis and pathophysiologies of endometriosis occurring in women and in animal models [17C22]. The MMPs are a family of enzymes that degrade the extracellular matrix, including basement membrane components [23]. TIMPs inhibit MMPs to facilitate tightly controlled tissue remodeling and other biological functions. A 1:1 stoichiometric balance of these enzymes and their inhibitors is required for normal follicular development, ovulation, formation and regression of the corpora lutea (CL), embryo development, and embryo implantation [24C32]. The MMPs and TIMPs are synthesized and secreted by both eutopic and ectopic endometrium in both the human and the rat [19C21, 33, 34]. TIMP1 represents at least 10% to 15% of proteins secreted into the peritoneal cavity by both rat implants and human endometriotic lesions [33, 35]. Because products from endometriotic lesions in the peritoneal fluid (PF) bathe the SYP-5 ovary and enter the uterus through the oviducts, endometriotic TIMP1 may influence the entire reproductive system, including ovulation, oocyte quality, embryo development, and early spontaneous pregnancy loss [17]. Our long-term goal is to understand the affects of endometriotic lesion-secreted TIMP1 on reduced fecundity. In these studies, we used a well-established rat SDF-5 model of endometriosis to evaluate the impact of modulation of TIMP1 on ovarian function and preimplantation embryo development. MATERIALS AND METHODS Three experiments were performed (Fig. 1). 1) TIMP1 was modulated in vitro to determine its effect on embryo development. 2) TIMP1 was modulated in vivo in Endo and Sham rats to determine the effect on embryo development and ovulation. 3) Control rats were treated with peritoneal fluid from Endo and Sham rats to determine the specificity of TIMP1 effects on ovulation. Open in a separate windows FIG. 1. In vitro and in vivo effects of modulating TIMP1 on preimplantation embryo development and ovulation. In experiment 1, zygotes were collected from control rats and cultured for 24 h in the presence SYP-5 and absence of TIMP1. In experiment 2, Endo rats were treated with a TIMP1 function-blocking antibody or vehicle, and Sham rats were treated with TIMP1 or vehicle. Zygotes were examined for embryo quality, and ovaries were evaluated for anomalies in ovulation. In experiment 3, PF (peritoneal fluid) was collected from Endo and Sham rats from experiment 2. TIMP1 was immunoprecipitated out of a portion of Endo.