Despite the documented importance of the protein hormone relaxin in reproduction in various mammalian species, the role of relaxin in human reproduction is poorly understood, largely because of the lack of studies in women or in suitable non-human primate models. Here we describe the establishment of a nonhuman primate model of early human pregnancy and its use in defining the actions of relaxin. Results demonstrate that relaxin exerts dramatic uterine effects including pronounced increase in uterine weight and stimulation of endometrial angiogenesis and resident endometrial lymphocyte number. In addition, relaxin decreases endometrial levels of matrix metalloproteinases 1 and 3 and increases levels of their endogenous inhibitor, tissue inhibitor of metalloproteinase 1, resulting in maintenance of endometrial collagen content. Relaxin significantly inhibits endometrial levels of estrogen receptor ␣, but not ␤, and of progesterone receptor isoforms A and B. The findings that relaxin stimulates new blood vessel formation and increases cytokine-containing lymphocyte number while maintaining endometrial connective tissue integrity are consistent with a significant role of relaxin in the establishment and͞or maintenance of early pregnancy.R elaxin is a 6-kDa protein hormone member of the insulinlike growth factor family present in circulation in women during the latter part of the menstrual cycle and throughout pregnancy (1). In many mammalian species, relaxin exerts pronounced effects on the female reproductive tract that are involved in the maintenance of pregnancy and successful parturition (2, 3). Relaxin is important for normal delivery in several mammalian species because of its marked rearrangement of reproductive tract connective tissue (2-4). Despite the documented importance of relaxin in various mammalian species, the role of relaxin in human reproduction is, to date, an important, yet unanswered, question. Elucidation of the role of relaxin in women has been hampered by the inability to perform studies in women and by the lack of studies performed in suitable primate models of human pregnancy. Here we describe the establishment of a non-human primate model of early human pregnancy and its use in defining the actions of relaxin. Results demonstrate that relaxin exerts dramatic uterine effects including pronounced increase in uterine weight, stimulation of endometrial angiogenesis and resident lymphocyte number, maintenance of endometrial connective tissue integrity, and inhibition of endometrial estrogen and progesterone action. These effects are the developmental changes that occur in the human uterus during the late secretory phase of the menstrual cycle and early pregnancy. These findings support the thesis that relaxin acts as an important factor in uterine accommodation to and maintenance of early pregnancy in women.The dramatic species differences in the sources, secretion patterns, and target organs of relaxin have contributed greatly to the lack of understanding of the role of relaxin in human repr...

Extensive evidence demonstrates pronounced effects of relaxin on the differentiation of human endometrial cells in vitro. In vivo data in rhesus monkeys suggest a role for relaxin in the development of endometrial vascular architecture. In women, pregnancy can be established and maintained in the absence of circulating relaxin. Thus, local synthesis by the endometrium is necessary if relaxin plays a physiological role in human endometrial function. Although relaxin protein and the prorelaxin C peptide have been localized to human endometrium, no data for relaxin synthesis have been provided to date. We therefore assessed relaxin mRNA and protein levels in cultured, defined human endometrial cells. Reverse transcriptase-polymerase chain reaction (RT-PCR) techniques were used to demonstrate the presence of relaxin mRNA in human stromal and glandular epithelial cells. Secretion of the protein into the media of cultured cells of both types was also detected. Relaxin stimulated the expression of vascular endothelial growth factor in glandular epithelial and stromal cells that were isolated from tissue that had been taken during the secretory phase of the cycle. Relaxin inhibited the expression of procollagenase from both glandular epithelial cells, with a more marked inhibition demonstrated from cells that were isolated from tissue that had been taken during the secretory phase, and from stromal cells. These data demonstrate that human endometrial cells synthesize relaxin, and they support the concept that relaxin fosters endometrial conditions that are required for implantation in women.

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Narrative AbstractOur studies demonstrate significantly lower expression of relaxin and its receptor in ectopic endometriotic tissues than their expression in eutopic endometrium and in endometrium from normal controls. These data suggest that in normal and eutopic endometrium, relaxin may exert a protective effect against endometriosis.The etiology of endometriosis, defined as the presence of endometrial stromal and glandular tissue growing at an extrauterine site, is poorly understood. The Sampson theory (1) does not sufficiently explain the etiology of this disease, since almost all women of reproductive age exhibit some degree of retrograde menstruation, but only 10-15% develop endometriosis (2). Additional factors must contribute to the development and progression of endometriosis.Relaxin is locally synthesized by human endometrium (3) and has potent effects in the endometrium, including decidualization (4). In human and rhesus monkey endometrium, relaxin is a powerful inhibitor of matrix metalloproteinases (MMPs) (3,5,6), which play an important role in the invasive process of endometriosis (7,8). Relaxin's action in target tissues is mediated by binding to its specific receptor, leucine-rich G protein-coupled receptor 7 (LGR7). Both relaxin and its LGR7 receptor are expressed in human endometrium (3,9,10) and human decidua (11,12).The current studies were performed to determine whether relaxin and/or its LGR7 receptor are expressed in human endometriotic tissues and whether their expression differ from that in endometrium from normal controls. We compared expression of relaxin mRNA and LGR7 mRNA in human endometriotic tissues to their expression in endometrium from normal controls. Publisher's Disclaimer: This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final citable form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. Of the 40 patients, 21 were diagnosed with ovarian endometriosis on the basis of surgical pathology. These patients were classified by laparoscopy as having stage III (n = 18) or IV (n = 3) endometriosis using the revised American Society for Reproductive Medicine classification of endometriosis (15). Patients with laparoscopic evidence and histological confirmation of extra-ovarian endometriosis were not included in the study. Of the 21 endometriosis patients, 7 had only an endometrial biopsy available (eutopic endometrium), 8 had only ectopic endometrial tissue available, and 6 had both ectopic and eutopic endometrial tissue available for this study, providing a total of 13 eutopic and 14 ectopic endometrial samples. NIH Public AccessThe remaining 19 patients were found upon laparoscopy to have non-endometriotic ovarian cysts and thus ser...

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Estrogen regulates LGR7 (RXFP1) mRNA expression in an in vitro model of human term pregnancy cervix that utilizes lower uterine segment fibroblasts. LGR7 mRNA levels were increased by estradiol to mean levels of 152%+/- 5.9% above those in untreated control cells. Therefore, estradiol may amplify relaxin's actions in the cervix.

In women, the corpus luteum is the source of circulating relaxin. No previous studies have addressed whether the corpus luteum is also a relaxin target organ. We determined relaxin receptor LGR7 mRNA expression in human term pregnancy corpora lutea and non-human primate corpora lutea obtained during the menstrual cycle. Real-time RT-PCR demonstrated the expression of LGR 7 mRNA in both human and rhesus monkey corpora lutea. Rhesus monkey corpora lutea were obtained from naturally cycling animals following documented LH surges at early, mid, mid-late and late luteal phases. Luteal expression of LGR7 mRNA did not show temporal variation. Since the primate corpus luteum is LH dependent, we assessed LGR7 mRNA expression in corpora lutea from rhesus monkeys treated with a GnRH antagonist which significantly suppressed pituitary LH levels. GnRH antagonist treatment, which also inhibits both progesterone and relaxin production, resulted in a five fold increase in luteal LGR7 mRNA expression. These data suggest that luteal LGR7 mRNA expression may be regulated by relaxin and/or LH, and that the primate corpus luteum is a target organ for relaxin.

To determine whether the effect of endothelin upon in vitro uterine contractility requires estrogen, immature female Long Evans rats were subcutaneously injected daily for 3 days with either estradiol benzoate or vehicle. The uterine contractile response to endothelin (5 or 100 nM) was measured. A response to endothelin in vehicle-treated animals exposed to physiologic (5.6 mM) or high (70.6 mM) potassium levels was also noted. Vehicle-treated uteri did not respond to endothelin; the contractions in estrogen-treated uteri increased by 288%. With high potassium, and without estrogen, no response to endothelin was seen. These data indicate that the contractile effect of endothelin on the uterus requires estrogen.