The history and future of in vitro fertilization in the United States: the complex interrelationships among basic science, human medicine, and politics.

Although much of the foundational basic scientific and clinical research was conducted in the United States, the first in vitro fertilization (IVF) birth occurred in the United Kingdom. Why? For centuries, all research surrounding the field of "reproduction" has elicited bipolar passionate responses by the American public, and the issue of "test tube babies" has been no different. The history of conception in the United States is defined by complex interrelationships among scientists, clinicians, and politically charged decisions by various branches of the US government. With a focus on research in the United States, this review summarizes the early scientific and clinical advances important to the development of IVF and then addresses the potential future developments in IVF. We also consider what future advances are possible in the United States given the current regulations, laws, and funding.

Application of a Patient Derived Xenograft Model for Predicative Study of Uterine Fibroid Disease.

Human uterine fibroids, benign tumors derived from the smooth muscle layers of the uterus, impose a major health burden to up to 50% of premenopausal women in their daily life. To improve our understanding of this disease, we developed and characterized a patient-derived xenograft model by subcutaneous transplantation of pieces of human uterine fibroid tissue into three different strains of severe combined immunodeficient mice. Engrafted uterine fibroid tissue preserved the classical morphology with interwoven bundles of smooth muscle cells and an abundant deposition of collagenous matrix, similar to uterine fibroids in situ. The grafts expressed both estrogen receptor 1 and progesterone receptor. Additionally, both receptors were up-regulated by estrogen treatment. Growth of the fibroid grafts was dependent on 17ß-estradiol and progesterone supplementation at levels similar to women with the disease and was studied for up to 60 days at maximum. Co-treatment with the antiprogestin mifepristone reduced graft growth (four independent donors, p<0.0001 two-sided t-test), as did treatment with the mTOR inhibitor rapamycin (three independent donors, p<0.0001 two-sided t-test). This in vivo animal model preserves the main histological and functional characteristics of human uterine fibroids, is amenable to intervention by pharmacological treatment, and can thus serve as an adequate model for the development of novel therapies.

Alterations in progesterone receptor membrane component 2 (PGRMC2) in the endometrium of macaques afflicted with advanced endometriosis.

The hormonally driven expression and cell-specific localization patterns of the progesterone receptor membrane components (PGRMC1 and PGRMC2) in the macaque endometrium during the menstrual cycle are unknown. Additionally, the expression and localization patterns of PGRMC1 and PGRMC2 in the secretory eutopic endometrium of primates afflicted with endometriosis are also unknown. Therefore, we used real-time PCR to quantify transcript expression levels of the PGRMCs in well-defined samples of endometrium collected from artificially cycled macaques during the menstrual cycle, and in the secretory phase endometrium of naturally cycling macaques afflicted with endometriosis. In situ hybridization and immunocytochemistry were used to localize PGRMC1 and PGRMC2 mRNA and protein, respectively. We compared the patterns of expression and localization of the PGRMCs with the expression and localization patterns of nuclear progesterone receptor (PGR). PGRMC1 and PGR were elevated during the proliferative phases of the cycle, and then declined to nearly undetectable levels during the late secretory phase of the cycle. Levels of PGRMC2 were lowest during the proliferative phases of the cycle and then increased markedly during the secretory phases. Strong staining for PGRMC2 was localized to the luminal and glandular epithelia during the secretory phases. When compared with artificially cycled disease-free animals, macaques with endometriosis exhibited no changes in the expression or localization patterns for PGR and PGRMC1 but exhibited strikingly reduced levels of PGRMC2 transcript and altered intracellular staining patterns for the PGRMC2 protein. Collectively, these results suggest that membrane-bound PGRMC2 may provide a pathway of action that could potentially mediate the non-genomic effects of progesterone on the glandular epithelia during the secretory phase of the cycle. Further, reduced levels of membrane-bound PGRMC2 may be associated with the progesterone insensitivity often observed in the endometrium of primates afflicted with endometriosis.

Estrogen and progesterone regulate expression of the endothelins in the rhesus macaque endometrium.

BACKGROUND: Endothelins (EDNs) are thought to modulate endometrial blood flow during menses, stromal healing and endometrial growth during the proliferative phase. Our goal was to assess the effects of estrogen and progesterone on the EDN paracrine system in the endometrium of rhesus macaques. METHODS: In this study, archived samples were used. These samples were collected from oophorectomized rhesus macaques that were treated sequentially with estradiol (E(2)) and then E(2) plus progesterone to create artificial menstrual cycles. Endometrium from animals in the menstrual, proliferative and secretory phases of the artificial cycle were analyzed by real-time PCR, in situ hybridization and immunocytochemistry to detect changes in EDN peptides (EDN1, EDN2, EDN3), EDN receptors (EDNRA, EDNRB), EDN-converting enzyme 1 (ECE1) and membrane metalloendopeptidase (MME)-an enzyme that degrades the EDNs. RESULTS: Compared with the late secretory phase, progesterone withdrawal at the end of the artificial menstrual cycle triggered an increase (P< 0.05) in EDN1, EDNRB and ECE1 in the upper functionalis zone during menses of the next cycle. Treatment with E(2) alone in the proliferative phase increased (P< 0.05) EDNRA transcript, which was confined predominantly to the stromal cells. E(2) plus progesterone in the artificial secretory phase suppressed (P< 0.05) the expression of EDN3 in the functionalis zone stroma and epithelia, tended (P= 0.08) to attenuate levels of epithelial EDN2 and markedly up-regulated (P< 0.05) the stromal expression of MME. CONCLUSIONS: Our results indicate that estrogen and progesterone regulate the EDN family during the menstrual cycle. The changes in the EDN paracrine system during the mid-secretory phase may indicate a role for EDN during embryo implantation.

Contrast-enhanced ultrasound reveals real-time spatial changes in vascular perfusion during early implantation in the macaque uterus.

OBJECTIVE: To use contrast-enhanced ultrasound (CEU) to quantify blood flow in the macaque uterus during early pregnancy. DESIGN: Prospective nonhuman primate study. SETTING: Oregon National Primate Research Center. ANIMALS: Naturally cycling female rhesus macaques (Macaca mulatta). INTERVENTION(S): Female macaques were mated on days 11-14 of the cycle. Females were then imaged by CEU and Doppler ultrasound once every 3 days from day 21 through day 39 of the fertile cycle. MAIN OUTCOME MEASURE(S): Visualization and quantification of uterine vascular perfusion. RESULT(S): CEU identified the primary placental disc and underlying vessels approximately 2 days earlier than Doppler ultrasound was able to observe endometrial thickening. CEU revealed spatial differences in vascular perfusion between the endometrium, myometrium, and endometrial-myometrial (junctional) zone. Myometrium displayed the highest rate of blood flow (>10 mL/min/g tissue). There was less blood flow in the endometrium and junctional zone (<3 mL/min/g). A brief fall in progesterone was observed during early implantation, which was correlated with reduced blood flow to all three uterine compartments, but did not reduce flow to the placenta. CONCLUSIONS: CEU provides a sensitive, noninvasive method to assess vascular perfusion of the uterus during embryo implantation in macaques. We propose CEU as a new diagnostic tool to monitor vascular changes associated with early pregnancy in women.

Luteotrophic and luteolytic effects of nitric oxide in sheep are dose-dependent in vivo.

It has been suggested that nitric oxide (NO) acts in either an anti-luteolytic or in a luteolytic manner, but the mechanism for these opposing roles is unclear. We hypothesized that NO may act in a dose-dependent manner to regulate luteal function, whereby low concentrations of NO might stimulate luteal progesterone production (i.e. luteotrophic) and high concentrations of NO might reduce concentrations of plasma progesterone (i.e. luteolytic). To test this hypothesis we infused increasing concentrations of the fast-acting NO donor, dipropylenetriamine NONOate (DPTA), into the arterial supply of sheep with ovarian transplants bearing a corpus luteum (CL). Infusions were performed on sheep with CL 11 days of age (n=9) or over 30 days of age (n=15). We measured changes in the concentration of progesterone in ovarian venous plasma during the 1-h infusion and for 24h after the infusion, and then compared the mean concentration of progesterone between treatment groups for effects by dose and dose by period interactions. Compared with saline-treated controls (n=6), the highest dose of 1000 microg/min DPTA (n=6) reduced (P0.05) in sheep infused with the lowest dose of 1 microg/min DPTA (n=6) compared with controls. We conclude that NO regulates luteal function in a dose-dependent manner in sheep in vivo.

Intrauterine infusion of BQ-610, an endothelin type A receptor antagonist, delays luteolysis in dairy heifers.

Three separate in vivo experiments were conducted to evaluate the putative role of endothelin-1 (ET-1) during luteal regression in heifers. In Experiment 1, a single intraluteal injection of 500 microg BQ-610 [(N,N-hexamethylene) carbamoyl-Leu-D-Trp (CHO)-D-Trp], a highly specific endothelin A (ETA) receptor antagonist, did not diminish the decline in plasma progesterone following a single exogenous injection of 25 mg prostaglandin F2 alpha (PGF2alpha) administered at midcycle of the estrous cycle. In Experiment 2, six intrauterine infusions of 500 microg BQ-610 given every 12 h on days 16-18 delayed spontaneous luteolysis, as evidenced by an extended elevation (P=0.054) of plasma progesterone concentration. In Experiment 3, heifers were administered six intrauterine infusions of BQ-610 or saline on days 16-19, and peripheral blood samples were collected from day 11 to 16 (before infusion), hourly on days 16-19 (during infusion), and on days 20-25 (after infusion). BQ-610 treated heifers had markedly higher (P<0.0001) levels of plasma progesterone compared with saline controls, and this effect was most notable during the infusion period (treatment by period interaction; Por=0.05) between treatments. These results indicate that the in vivo antagonism of the ETA receptor can delay functional luteolysis, and supports the theory that ET-1 regulates luteal function in ruminants.

Role of progesterone in nonhuman primate implantation.

Herein we review the morphological and physiological effects of estradiol and progesterone (P) on the nonhuman primate uterus. Progesterone action acts to prepare the endometrium for embryo implantation, which normally occurs only during a brief period in the mid-luteal phase of the menstrual cycle. During this window of implantation, P stimulates secretory morphological differentiation and suppresses estrogen receptor alpha (ERalpha), in the endometrial functionalis zone. Reduced endometrial ERalpha is a definitive physiological marker for the onset of endometrial receptivity in primates. These actions of P are specific for the functionalis zones, and P does not fully inhibit ERalpha in the glands of basalis zone of nonhuman primates. Paradoxically, during the secretory phase of the cycle, progesterone receptor (PR) is also reduced in the glandular epithelium of the progestin-responsive functionalis zone. Therefore, P action on the epithelium in the functionalis zone may be mediated by paracrine factors arising from the PR-positive cells in the stroma. Genomic analysis of the endometrium of women and nonhuman primates has revealed numerous secretory phase genes that may contribute to differentiation of the endometrium. However, the exact nature and function of these putative factors have been elusive. We propose that nonhuman primates, especially macaques, can provide a valuable animal model for experimentally testing the functional role of P-regulated genes on endometrial receptivity.