Endometrial prostaglandin E2 binding: characterization in rats sensitized for the decidual cell reaction and changes during pseudopregnancy

As an initial step in testing the hypothesis that uterine receptivity for blastocyst implantation and sensitivity for decidualization are controlled in part by the presence of functional receptors for prostaglandin E2 (PGE2) in the endometrium, we have characterized the high-affinity binding of [3H]PGE2 to an endometrial membrane preparation from ovariectomized rats treated with progesterone and estradiol so that their uteri were sensitized for the decidual cell reaction. As determined by Scatchard analysis, a single class of [3H]PGE2 binding sites with an apparent Kd ranging from 2 to 6 nM and a capacity of approximately 100 fmol/mg protein was found. Prostaglandins E1 and E2 competed equally for binding while relative cross-reactivity of other prostanoids and compounds tested was less than 3%. Binding was temperature-dependent and reversible. Under the assay conditions used, no metabolism of [3H]PGE2 was detectable. Pretreatment of the membrane preparation with proteolytic enzymes, or by heating, reduced subsequent specific [3H]PGE2 binding. These data are consistent with the presence of endometrial PGE receptors in the sensitized endometrium. The binding of [3H]PGE2 to endometrial membrane preparations from rats on Days 2 to 7 pseudopregnancy was determined. No specific binding could be detected on Day 2. A low binding capacity was found on Days 3 and 4; this increased markedly on Day 5 and reached a maximum on Day 6. These data indicate that the onset of uterine receptivity/sensitivity is temporally correlated with the appearance of endometrial PGE binding sites.     

Uterine sensitization-associated gene-1: a novel gene induced within the rat endometrium at the time of uterine receptivity/sensitization for the decidual cell reaction

For successful implantation, the embryo must develop to the blastocyst stage and the endometrium must attain a state that is receptive to the implanting blastocyst. In rodents, the timing, duration, and hormonal regulation of this receptive state has been well defined. However, the molecular cascade of events involved in the onset of the receptive phase remains unclear. In the present study, we sought to identify genes involved in the onset of the receptivity using the technique of suppressive subtraction hybridization. Herein we report the isolation, cloning, and characterization of a novel gene, uterine sensitization-associated gene-1 (UASG-1), that is preferentially expressed within the maximally sensitized/receptive rat endometrium. USAG-1 mRNA encodes a putative protein of 206 amino acids that contains a possible N-terminal secretion signal and a C-terminal cystine knotlike motif. Northern blot analysis revealed that induction of USAG-1 mRNA was restricted to the Day 5 pregnant or pseudopregnant uterus. In situ hybridization experiments demonstrated that this induction was restricted to the uterine glandular epithelial cells. Given the remarkably tight restriction of its expression, USAG-1 may be involved in the onset of endometrial receptivity for implantation/sensitization for the decidual cell reaction.     

Characterization of temporal and cell-specific changes in transcripts for prostaglandin E(2) receptors in pseudopregnant rat endometrium

In the rodent uterus, prostaglandin E(2) (PGE(2)) is believed to have a major role in implantation and decidualization. The present study investigated the temporal and hormonal control of mRNA expression for the four E-prostanoid (EP(1-4)) receptors in the rat endometrium. For Northern blot analysis and in situ hybridization, samples were obtained from rats on Days 1-10 of pseudopregnancy or from rats differentially sensitized for the decidual cell reaction with estradiol. No EP(1) mRNA signal was detected. Endometrial EP(2) and EP(3) mRNA levels increased to a maximum on Day 5, and the mRNAs were localized to the luminal epithelium at the antimesometrial pole, and in the endometrial stroma and glandular epithelium, respectively. Endometrial EP(4) mRNA levels were unchanged on Days 1-5, but the mRNA was concentrated in the antimesometrial endometrial stroma on Day 5. Cell-specific expression of EP(2), EP(3), and EP(4) on Day 5 was dependent upon a dose of estradiol given on Day 4 that induced differential uterine sensitization on Day 5. After the application of a deciduogenic stimulus on Day 5, mRNA levels for these receptors decreased significantly, while in nonstimulated horns they remained elevated. Overall, these results support a role for PGE(2) in the onset of receptivity and initiation of decidualization in the rat.     

Decreased ANGPTL4 impairs endometrial angiogenesis during peri‐implantation period in patients with recurrent implantation failure

Insufficient endometrial angiogenesis during peri‐implantation impairs endometrial receptivity (ER), which contributes to recurrent implantation failure (RIF) during in vitro fertilization and embryo transfer (IVF‐ET). Angiopoietin‐like protein 4 (ANGPTL4) acts as a multifunctional secretory protein and is involved in the regulation of lipid metabolism and angiogenesis in various tissues including the endometrium. Herein, we found decreased ANGPTL4 expression in endometrial tissue and serum during peri‐implantation period in 18 RIF‐affected women with elevated uterine arterial impedance (UAI) compared with the pregnancy controls. ANGPTL4 and peroxisome proliferator‐activated receptor gamma (PPARγ) expression were up‐regulated upon decidualization on human endometrial stromal cells (HESCs). Rosiglitazone promoted the expression of ANGPTL4 in HESCs and human umbilical vein endothelial cells (HUVECs) via PPARγ. ANGPTL4 promoted the proliferation, migration and angiogenesis of HUVECs in vitro. Our results suggest that decreased abundance of ANGPTL4 in endometrial tissues impairs the endometrial receptivity via restraining endometrial angiogenesis during decidualization; while rosiglitazone‐induced ANGPTL4 up‐regulation in hESCs and HUVECs through PPARγ. Therefore, ANGPTL4 could be a potential therapeutic approach for some RIF‐affected women with elevated UAI.     

Expression of matrix metalloproteinases 2 and 9 in the mouse uterus during implantation and oil-induced decidualization

During implantation, matrix metalloproteinases are believed to play roles in the tissue remodelling that accompanies decidualization in the endometrium and in embryo invasion. The objective of this study was to characterize further the expression of matrix metalloproteinases 2 and 9 in the mouse uterus during early pregnancy and oil-induced decidualization. mRNA encoding matrix metalloproteinase 2 was detected in pregnant uteri and uteri undergoing oil-induced decidualization by northern blot analyses. The steady-state concentrations of mRNA encoding matrix metalloproteinase 2 did not change significantly in implantation compared with inter-implantation areas on days 5-8 of pregnancy but were significantly lower in stimulated compared with non-stimulated uterine horns during artificially induced decidualization. mRNA encoding matrix metalloproteinase 9 was also detected in uteri undergoing oil-induced decidualization but not in pregnant uteri. Its concentration was significantly greater in uterine horns undergoing oil-induced decidualization compared with control horns. Immunoreactive matrix metalloproteinases 2 and 9 were detected in the uterus during early pregnancy and oil-induced decidualization by immunohistochemistry, localized to the endometrial stroma, but the staining progressively became weaker and was absent in areas that had undergone decidualization. By day 8 of pregnancy and 72 h after the induction of decidualization, matrix metalloproteinase 2 and 9 proteins remained mainly in the region of non-decidualized stromal cells adjacent to the myometrium. In implantation segments, they were also localized to the region of the trophoblast giant cells. The second objective of the present study was to determine whether endometrial stromal cells isolated from uteri sensitized for decidualization express matrix metalloproteinases 2 and 9. Northern blot analyses and gelatin zymography showed that these cultured cells expressed matrix metalloproteinase 2 and 9, and that transforming growth factor beta1 significantly increased matrix metalloproteinase 9 expression. The results of the present study further characterize matrix metalloproteinases 2 and 9 expression in the uterus during implantation and artificially induced decidualization.     

LncRNA882 regulates leukemia inhibitory factor (LIF) by sponging miR-15b in the endometrial epithelium cells of dairy goat

Despite the fact that long noncoding RNAs (lncRNAs) play roles in almost all biological processes, little is known about their biological function in the endometrium during the formation of endometrial receptivity. In this study, a comprehensive analysis of lncRNAs in goat endometrial tissues on Day 5 (prereceptive endometrium, PE) and Day 15 (receptive endometrium, RE) of pregnancy was performed by using RNA-Seq. As a result, 668 differentially expressed lncRNAs (DELs) were found between the PE and RE. Further study showed that lncRNA882, regulated by estrogen (E2) and progestin (P4), could act as competing endogenous RNAs (ceRNAs) for miR-15b, which inhibited the expression of transforming growth factor-b-activated kinase 1 binding protein 3 (TAB3) and then indirectly regulated the level of leukemia inhibitory factor (LIF). This was helpful for the formation of endometrial receptivity in dairy goats. In conclusion, we elucidated the endometrium lncRNA profiles of PE and RE in dairy goats; lncRNA882 acted as a ceRNA for miR-15b and then indirectly regulated the level of LIF in goat endometrial epithelium cells. Thus, this study helped us to better understand the molecular regulation of endometrial receptivity in dairy goats.     

Gastrin-releasing peptide (GRP) in the ovine uterus: regulation by interferon tau and progesterone

Gastrin-releasing peptide (GRP) is abundantly expressed by endometrial glands of the ovine uterus and processed into different bioactive peptides, including GRP1-27, GRP18-27, and a C-terminus, that affect cell proliferation and migration. However, little information is available concerning the hormonal regulation of endometrial GRP and expression of GRP receptors in the ovine endometrium and conceptus. These studies determined the effects of pregnancy, progesterone (P4), interferon tau (IFNT), placental lactogen (CSH1), and growth hormone (GH) on expression of GRP in the endometrium and GRP receptors (GRPR, NMBR, BRS3) in the endometrium, conceptus, and placenta. In pregnant ewes, GRP mRNA and protein were first detected predominantly in endometrial glands after Day 10 and were abundant from Days 18 through 120 of gestation. Treatment with IFNT and progesterone but not CSH1 or GH stimulated GRP expression in the endometrial glands. Western blot analyses identified proGRP in uterine luminal fluid and allantoic fluid from Day 80 unilateral pregnant ewes but not in uterine luminal fluid of either cyclic or early pregnant ewes. GRPR mRNA was very low in the Day 18 conceptus and undetectable in the endometrium and placenta; NMBR and BRS3 mRNAs were undetectable in ovine uteroplacental tissues. Collectively, the present studies validate GRP as a novel IFNT-stimulated gene in the glands of the ovine uterus, revealed that IFNT induction of GRP is dependent on P4, and found that exposure of the ovine uterus to P4 for 20 days induces GRP expression in endometrial glands.     

A morphological study of uterine alterations in mice due to exposure to cadmium

We investigated the morphologic and molecular effects of exposure to cadmium (Cd) for 30 and 60 days on the uteri of mice. We assessed uterine morphometric measurements, eosinophilia, mast cell numbers, endometrial apoptosis, proliferation and estrogen receptor alpha (ERα) immunoreactivity. We examined vaginal smears that reflected the hormonal alterations in the female reproductive tract. Because the female reproductive tract exhibits different morphology at each stage of the estrous cycle, we sacrificed all animals at estrus to make appropriate comparisons. Female BALB/c mice were exposed to 200 ppm Cd in their drinking water for either 30 or 60 days. Cd exposure caused significant decreases in endometrial thickness and number of glands in estrus phase uteri. The endometrial eosinophilia in the groups exposed to Cd also decreased compared to controls. Cd exposure increased the number of mast cells. Luminal and glandular epithelia were examined using the terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay and by immunostaining proliferating cell nuclear antigen (PCNA) and estrogen receptor α (ERα). Compared to controls, the apoptotic index increased with time in both Cd exposed groups, while the proliferation index decreased. ERα immunoreactivity was decreased in both Cd exposed groups compared to controls; the decrease was most apparent in the 30 day Cd group. We found that 60 day Cd exposure increased apoptosis in the endometrium, which may affect the receptivity of the uterus for implantation.     

Estrogen receptor-alpha (ER-alpha) and defects in uterine receptivity in women

Endometriosis is a disorder that affects 5% of the normal population but is present in up to 40% of women with pelvic pain and/or infertility. Recent evidence suggests that the endometrium of women with endometriosis exhibits progesterone insensitivity. One endometrial protein that fluctuates in response to progesterone is the estrogen receptor-alpha (ER alpha), being down-regulated at the time of peak progesterone secretion during the window of implantation. Here we demonstrate that the biomarker of uterine receptivity, beta 3 integrin subunit, is reduced or absent in some women with endometriosis and that such defects are accompanied by inappropriate over-expression of ER alpha during the mid-secretory phase. Using a well-differentiated endometrial cell line we showed that the beta 3 integrin protein is negatively regulated by estrogen and positively regulated by epidermal growth factor (EGF). By competing against estrogen with various selective estrogen receptor modulators (SERMs) and estrogen receptor agonists and antagonists, inhibition of expression of the beta 3 integrin by estrogen can be mitigated. In conclusion, we hypothesize that certain types of uterine receptivity defects may be caused by the loss of appropriate ER alpha down-regulation in the mid-secretory phase, leading to defects in uterine receptivity. Such changes might be effectively treated by timely administration of the appropriate anti-estrogens to artificially block ER alpha and restore normal patterns of gene expression. Such treatments will require further clinical studies.     

Spatial and temporal patterns of deoxyribonucleic acid synthesis and mitosis in the endometrial stroma during decidualization in the pseudopregnant rat

A quantitative study was made of the spatial patterns of stromal cell mitosis and DNA synthesis in the endometrium of the pseudopregnant rat before and during decidualization. A colchicine block was used for mitotic counts, and DNA synthesis was studied by [3H] thymidine autoradiography. Observations were also made on the subsequent fates of [3H] thymidine-labeled stromal cells. Before the onset of decidualization, on Days 3 and 4 (vaginal cornification = Day 0), mitosis was largely confined to the subepithelial stroma along the sides and around the antimesometrial pole of the lumen. [3H] thymidine labeling and stromal mitosis following a decidualizing stimulus at noon on Day 4 of pseudopregnancy were first seen close to the uterine lumen, with subsequent spread to deeper layers of the endometrium. At noon on Day 5, mitotic figures were numerous on all sides of the lumen and at all depths in the endometrium. At later stages, mitosis and the development of polyploidy continued in the decidual tissue, but little DNA synthesis or mitosis occurred in the basal zone of the stroma adjacent to the myometrium. In this zone, many cells in animals given [3H] thymidine 18 to 24 h after induction of decidualization remained heavily labeled throughout the growth and regression of deciduomata. Labeled cells derived from the basal zone and outer edge of the decidual capsule were present in the stroma of the regenerated endometrium following the regression of deciduomata. It was concluded that although cells at all depths in the endometrial stroma undergo DNA synthesis and mitosis in the early stages of response to a decidualizing stimulus, their subsequent behavior and fate depend upon their position in the endometrium.     

DHA induces apoptosis by altering the expression and cellular location of GRP78 in colon cancer cell lines

n-3 polyunsaturated fatty acids exert growth-inhibitory and pro-apoptotic effects in colon cancer cells. We hypothesized that the anti-apoptotic glucose related protein of 78kDa (GRP78), originally described as a component of the unfolded protein response in endoplasmic reticulum (ER), could be a molecular target for docosahexaenoic acid (DHA) in these cells. GRP78 total and surface overexpression was previously associated with a poor prognosis in several cancers, whereas its down-regulation with decreased cancer growth in animal models. DHA treatment induced apoptosis in three colon cancer cell lines (HT-29, HCT116 and SW480), and inhibited their total and surface GRP78 expression. The cell ability to undergo DHA-induced apoptosis was inversely related to their level of GRP78 expression. The transfection of the low GRP78-expressing SW480 cells with GRP78-GFP cDNA significantly induced cell growth and inhibited the DHA-driven apoptosis, thus supporting the essential role of GRP78 in DHA pro-apoptotic effect. We suggest that pERK1/2 could be the first upstream target for DHA, and demonstrate that, downstream of GRP78, DHA may exert its proapoptotic role by augmenting the expression of the ER resident factors ERdj5 and inhibiting the phosphorylation of PKR-like ER kinase (PERK), known to be both physically associated with GRP78, and by activating caspase-4. Overall, the regulation of cellular GRP78 expression and location is suggested as a possible route through which DHA can exert pro-apoptotic and antitumoral effects in colon cancer cells.     

Endometrial contraception: modulation of molecular determinants of uterine receptivity

Modulation of endometrial receptivity is a promising approach for fertility regulation since it allows a contraceptive to act specifically at the endometrium. This was corroborated by our previous observations that treatment with low doses of a pure progesterone antagonist (PA, antiprogestin), onapristone (ZK 98299), in bonnet monkeys inhibited fertility by selectively retarding endometrial development, without affecting the hypophyseal-hypothalamic function. In the present study, further investigations, undertaken to analyze the molecular repertoire of a nonreceptive primate endometrium, determined expression of: steroid hormone receptors, i.e. progesterone receptor (PR) and estrogen receptor (ER); cytokines, i.e. leukemia inhibitory factor (LIF): transforming growth factor beta (TGFbeta) and its receptor (TGFbetaR); and cell adhesion molecules, i.e. integrins (alpha(v)beta(3), alpha(1)beta(1)). These studies were conducted during the different phases of the normal menstrual cycle and following treatment with different doses of onapristone (2.5 mg, 5 mg, or 10 mg every third day for one cycle) in bonnet monkeys. The molecules were analysed collectively to explore the possibility of a correlation between expression of these markers and endometrial receptivity and to investigate whether there exists a regulatory link between expression of these molecules under in vivo conditions. Three types of expression patterns of endometrial factors were observed during the peri-implantation period following onapristone treatment: 1) LIF, alpha(v)beta(3), and alpha(1)beta(1) showed significant (P < 0.02) down regulation in glandular epithelium of endometria in animals treated with all three doses of onapristone as compared to the control group. This was indicative of their critical role in the progesterone-driven cascade leading to implantation. 2) PR, TGFbeta, and TGFbetaR remained unaffected in the endometria from 2.5 mg treated animals and showed down regulation in animals treated with 5 and 10 mg onapristone as compared to the control group, thereby suggesting that the expression of these markers may not truely reflect endometrial receptivity per se. However, their facilitatory role in preparing the endometrium for implantation can not be ruled out since continued perturbation in the expression of these molecules may affect endometrial growth, remodelling, and differentiation, which in turn may render the endometrium nonreceptive; 3) ER remained unaltered in endometria of animals rendered infertile with 2.5, 5, and 10 mg onapristone. This observation indirectly suggests that onapristone-induced endometrial changes are mediated via some specific mechanisms. The present study clearly demonstrates that endometrial non-receptivity induced at low doses of onapristone is associated with changes in the expression pattern of specific molecular markers. However, no direct correlation was observed between in vivo expression of TGFbeta, LIF, and integrins, thereby lending support to the concept that there exists redundancy or multiple pathways which regulate implantation events.     

Immunocytochemical localization and messenger ribonucleic acid levels of a progesterone-dependent endometrial secretory protein (cathepsin L) in the pregnant cat uterus.

The purpose of this study was to localize immunocytochemically a progesterone-dependent protein (PDP) and to determine PDP mRNA levels during the initial stage of the implantation period. Uterine tissue was collected from Day 0-18 postcoital animals. The tissue was processed for immunocytochemical localization of PDP, and the endometrial RNA was isolated and analyzed for PDP gene expression by slot-blot hybridization. PDP was detected immunocytochemically as early as Day 5 postcoitus in the epithelial cells of the deep uterine glands, and the intensity of immunostaining appeared to peak by Day 12 postcoitus. PDP was absent in the endometrium obtained from implantation sites after Day 16 postcoitus, but the synthesis of PDP was maintained in the endometrium obtained from nonimplantation sites. Immunogold electron microscopy demonstrated that PDP was present in electron-dense granules of the glandular epithelial cells. PDP mRNA was detectable in the endometrium at Day 5 postcoitus and peaked around Day 10 postcoitus. PDP mRNA was absent in the endometrium from implantation sites after Day 16 postcoitus, but was maintained in the endometrium from nonimplantation sites. In summary, the results of this study illustrate that PDP is synthesized within the epithelial cells of the deep uterine glands, packaged within membrane-bound secretory granules, and released into the uterine lumen. Also, the process of implantation alters the gene expression in a very localized way since PDP mRNA and PDP-positive granules were absent in the endometrial glands obtained from the implantation site within 1-2 days of the onset of implantation, whereas both PDP mRNA and PDP-positive granules were maintained in the endometrial glands from nonimplantation-site regions.(ABSTRACT TRUNCATED AT 250 WORDS)     

Role of interleukin 1 in the regulation of cyclooxygenase gene expression in rat endometrial stromal cells.

Interleukin 1 alpha (IL-1 alpha) stimulates prostaglandin production and cyclooxygenase activity in endometrial stromal cells isolated from the uteri of ovariectomized rats that have been sensitized for the decidual cell reaction. The aim of the present study was to examine the effect of IL-1 alpha on the amount of cyclooxygenase mRNA and protein in these cells. Treatment with IL-1 alpha (20 ng ml-1) for 24 h significantly increased steady-state concentrations of cyclooxygenase 2 (COX-2) mRNA and protein in the cells, as determined by northern and western blot analyses, respectively. Cyclooxygenase 1 (COX-1) mRNA and protein were not detected. Dexamethasone (5 mumol l-1) prevented the IL-1 alpha-induced increase in COX-2 steady-state mRNA. Immunocytochemical staining of COX-2 in the treated cells indicated that IL-1 alpha increased staining, while dexamethasone inhibited this increase. Furthermore, the changes in staining were generalized and not confined to a small subpopulation of cells. These data demonstrate that IL-1 alpha increases steady-state concentrations of COX-2 mRNA and protein in endometrial stromal cells isolated from the uteri of rats that have been sensitized for decidualization.     

Secretion of plasminogen activator by cultured rat endometrial stromal cells from uteri differentially sensitized for the decidual cell reaction

Endometrial stromal cells from rat uteri differentially sensitized for the decidual cell reaction in vivo and which undergo differing degrees of decidualization in vitro were cultured and plasminogen activator (PA) in the medium determined. The cells were obtained by enzymatic dispersion from the uteri of ovariectomized, steroid-treated rats at the equivalent of day 4, 5, or 6 of pseudopregnancy or on day 5 from rats treated on day 4 with 0, 0.3, or 1.0 μg estradiol (low, intermediate, or high dose of estradiol, respectively) and cultured for 24, 48, or 72 hr. For cells from day 4, 5, and 6 uteri cultured under control conditions, PA activity in the medium was greatest for day 5 cells, which were from uteri maximally sensitized for decidualization both in vivo and in vitro. By contrast, for cells from low-, intermediate-, and high-estradiol uteri, PA activity in the medium was greatest for the high-estradiol cells; these cells do not undergo decidualization in vivo or in vitro to the same extent as intermediate-estradiol cells. Indomethacin, an inhibitor of prostaglandin (PG) synthesis, reduced PGE₂ accumulation to nondetectable amounts and for most cultures decreased PA activity in the medium, suggesting that endogenous PG production regulated in part PA secretion under control conditions. The addition of PGE₂ with indomethacin increased PA activities above those under control conditions, but activities were still lower for day 4 and 6 cells compared with day 5 cells, and for low- and intermediate-estradiol cells compared with high-estradiol cells. This indicates that the differences in PA secretion are not explainable by differences in PGE₂ production. Northern blot analysis of RNA from cells cultured for 72 hr under control conditions did not reveal significant differences in steady-state concentrations of mRNA for urokinase-type PA or plasminogen activator inhibitor 1, but those for tissue-type PA were lower in day 6 cells compared with day 4 and 5 cells. It is concluded that PA activity secreted by the cultured endometrial stromal cells, although controlled in part by the endocrine milieu to which they were exposed prior to culture, does not simulate decidualization in vitro and, therefore, that PA activity is not a marker for decidualization in vitro. Mol. Reprod. Dev. 49:268–276, 1998. © 1998 Wiley-Liss, Inc.     

Changes in the temporal and spatial expression of H beta 58 during formation and maturation of the chorioallantoic placenta in the Rat

Cloning and sequencing of a cDNA amplified by RNA fingerprinting at the implantation site of pregnant rats revealed 80% similarity with H beta 58, previously shown to be essential for formation of the chorioallantoic placenta in the mouse. H beta 58 mRNA was detected in the endometrium of hormonally sensitized rats stimulated to undergo decidualization and in the contralateral uterine horns lacking a decidual stimulus, indicating that uterine expression of H beta 58 mRNA did not require decidualization or the presence of a blastocyst. Immunodetection in the early postimplantation uterus (Days 6-8 of pregnancy) showed H beta 58 localized in the luminal and glandular epithelia and some stromal cells. Decidual cells at Day 6 of pregnancy expressed H beta 58, and by Day 9 of pregnancy, the protein localized throughout the maternal decidua. The temporal and spatial distribution of H beta 58 in the developing chorioallantoic placenta was assessed at Days 10, 12, and 14 of pregnancy. Immunoreactive H beta 58 localized to erythroid cells within the developing fetal vasculature of the chorioallantoic primordia at Day 10 of pregnancy. By Day 12, the fetal vasculature extended into the placental labyrinth, and the erythroid stem cells continued to strongly express H beta 58. At Day 14 of pregnancy, immunoreactivity became evident in the trophoblast giant cells and syncytiotrophoblast of the fetal placenta. As the chorioallantoic placenta matured (Day 18), H beta 58 mRNA was 3.6-fold higher in the labyrinth compared with the junctional region. Stable cell lines (HRP/LRP) isolated from the rat labyrinthine placenta expressed H beta 58 mRNA and protein. The expression pattern of H beta maternal and fetal placental tissues and its early expression in fetal erythroid stem cells during formation and maturation of the chorioallantoic placenta suggest that H beta 58 plays key roles in the regulatory networks that control hematopoietic development and placentation.     

Temporal relationships among uterine pituitary adenylate cyclase-activating polypeptide, decidual prolactin-related protein and progesterone receptor mRNAs expressions during decidualization and gestation in rats

Pituitary adenylate cyclase-activating peptide (PACAP), a novel compound with vasoactive intestinal polypeptide-like activity, was recently shown to be localized in the neuronal endings of the human uterus. The purpose of the present study was to assess the functional presence of PACAP mRNA in the decidual endometrium and its relationship to the expression levels of decidual prolactin-related protein (dPRP) and the progesterone receptor mRNAs during decidualization and pregnancy in Sprague-Dawley rats. PACAP was constitutively and temporally expressed in the decidual endometrium and gravid uterus. The time-dependent correlated expression levels of PACAP, dPRP and the progesterone receptor were induced by the neurogenic reproductive signals, i.e. the vagino-cervical/deciduogenic stimuli of decidualization and by the normal equivalent stimuli of mating/blastocyst implantation of gestation. Correlation among the mRNA expression levels of PACAP, dPRP and the progesterone receptor and the coordinated inhibitory actions of the anti-progesterone (RU-486) suggest that there is also correlated time-dependent steroid regulation of the mRNA levels of PACAP, dPRP and the progesterone receptor in the decidual and pregnant uteri. One possible functional meaning for the time-related localization of endometrial/uterine PACAP could be to facilitate endometrial blood flow and increase the availability of metabolic substrates to the developing deciduoma or embryo. The study demonstrates the potential importance of PACAP expression in the regulation of the maternal feto-placental component and suggests a prominent reproductive role for the neuropeptide in mammalian pregnancy.