Porcine endometrial and conceptus tissue kallikrein 1, 4, 11, and 14 gene expression

Previous studies have suggested that the porcine endometrium may express several tissue kallikreins during the estrous cycle and early pregnancy. The present study investigated porcine endometrial and conceptus tissue kallikrein 1, 4, 11, and 14 mRNA expression during the estrous cycle and early pregnancy. Tissue kallikrein (KLK) gene expression was evaluated using quantitative RT-PCR and in situ hybridization. KLK1 expression was similar across the estrous cycle and early pregnancy, and localized to the endometrial luminal (L) and glandular (G) epithelium. KLK4 endometrial mRNA expression was greatest on days 0, 5, and 10 when compared with days 12, 15, and 17 of the estrous cycle and greater in cyclic compared with pregnant gilts. Expression of KLK4 was more intense in the stroma and uterine epithelium from days 0 to 10 of the estrous cycle. Endometrial KLK11 mRNA was not different between cyclic and pregnant gilts but the expression was greatest on days 10 and 12 compared with all other days evaluated. There was an increased intensity of KLK11 gene expression in the stratum compactum on day 10 of the estrous cycle and early pregnancy. Endometrial KLK14 mRNA expression was not detectable on days 5 and 10 but was expressed on days 0, 12, 15, and 17 of the estrous cycle and pregnancy. KLK14 expression was localized in the uterine L and G epithelium, and stroma throughout the endometrium after day 10. Conceptus KLK1 mRNA did not change from days 10 to 17 of gestation. However, conceptus KLK4, and 14 mRNA expression was greatest on day 10 with expression declining after day 14 of gestation. Expression of the various tissue kallikreins in the endometrium and conceptus during the estrous cycle and early pregnancy in the pig can serve in the activation of growth factors and tissue remodeling during the establishment of pregnancy.     

Complex expression patterns support potential roles for maternally derived activins in the establishment of pregnancy in mouse

Maternal-fetal communications are critical for the establishment of pregnancy. Embryonic growth and differentiation factors produced by the oviduct and uterus play essential roles during the pre- and early post-implantation phases. Although several studies indicate roles for activin in embryonic development, gene-knockout studies have failed to identify a critical role in mammalian embryogenesis. We hypothesized that activin is produced by maternal tissues during the establishment of pregnancy, and thus maternally derived activin could compensate for the absence of embryonic activin in null homozygotes during critical developmental stages. We investigated the expression of inhibin alpha, activin betaA, and betaB subunits in the mouse oviduct and uterus during the estrous cycle and early pregnancy, and in the early conceptus. Inhibin alpha subunit was weakly expressed, while activin betaA and betaB subunits were strongly expressed in oviduct and uterus at estrous, and dramatically upregulated in the uterus on each day of pregnancy between days 3.5 and 8.5 post coitum. Prior to implantation, activin betaA and betaB subunits were immunolocalized to oviductal and uterine epithelial cells; following implantation they were expressed in the stroma, in a wave preceding decidualization. Later in pregnancy, activin betaA and betaB subunits were present in decidua basalis, trophoblast giant cells, and labyrinth zone of the developing placenta. Expression of activin betaA subunit was also detected in blastocysts and early post-implantation embryos. These data are consistent with a role for maternally derived activins in the support of the pre-implantation embryo, and during gastrulation and embryogenesis.     

Temporal and spatial associations of oestrogen receptor alpha and progesterone receptor in the endometrium of cyclic and early pregnant mares

Uterine function is primarily controlled by the combined actions of oestrogen and progesterone working through their cognate nuclear receptors. The mechanism of establishment of pregnancy in the mare is of interest because it involves prolonged pre-attachment and conceptus migration phases, and both invasive and non-invasive placental cell types, and as such has been an important comparative model. This study characterised regulation of oestrogen (ER) and progesterone (PR) receptors in the endometrium of the mare during the oestrous cycle and early pregnancy. Endometrial tissues collected during the oestrous cycle and early pregnancy were analysed for steady-state levels of ER and PR mRNA and protein. Steady-state levels of ER and PR mRNA were highest on days 0, 17 and 20 in cyclic mares and lowest on days 11 and 14. A day-by-status interaction was detected, indicating that day 17 and day 20 pregnant mares exhibited low levels of ER and PR compared with the corresponding days of the oestrous cycle. In situ hybridisation analyses showed receptor mRNA localisation primarily in the luminal epithelium (LE), glandular epithelium (GE) and stroma around oestrus. During dioestrus and early pregnancy, receptors were not detected in the LE, and were lower in the stroma and deeper GE. Changes in hybridisation intensity in these cell types were consistent with changes in mRNA levels detected by slot-blot hybridisation. ER and PR proteins were detected in the nuclei of LE, GE and stromal cells. Consistent with results from in situ hybridisation, levels of ER and PR immunoreactivity were higher around oestrus, declined to low levels during dioestrus and remained low during early pregnancy. Results described here for temporal and spatial changes in steroid receptor gene expression in mares show the greatest similarities with those described for cattle and sheep.     

The IGF system in the neonatal ovine uterus

Postnatal development of the ovine uterus primarily involves uterine gland morphogenesis or adenogenesis. Adenogenesis involves the budding differentiation of the glandular epithelium (GE) from the luminal epithelium (LE) and then GE proliferation and coiling/branching morphogenetic development within the stroma between birth (postnatal day or PND 0) and PND 56. Insulin-like growth factor (IGF)-I and IGF-II mRNAs were previously found to be expressed only in the endometrial stroma, whereas the IGF receptor (IGF-1R) mRNA was most abundant in epithelia and in stroma, suggesting that an intrinsic IGF system regulates postnatal development of the uterus. Given that the biological activities of IGFs are modulated by a family of six IGF binding proteins (IGFBPs) and specific proteases, the objective was to determine the effects of age and estrogen disruption on expression of IGFs, IGFBPs and pregnancy-associated plasma protein A (PAPP-A or IGFBP-4 protease) in the ovine uterus. In Study One, circulating levels of IGF-I and IGF-II in the serum of neonatal ewes did not change between PND 0 and PND 56. Levels of immunoreactive IGF-I, IGF-II and IGF-1R protein were most abundant on the apical surface of the endometrial LE and GE. RT-PCR analyses detected expression of IGFBPs (3, 4, 5 and 6) as well as PAPP-A mRNAs in the uterus, but not IGFBP-1 and IGFBP-2 mRNAs. IGFBP-3 and IGFBP-4 mRNAs were expressed specifically in the endometrial stroma and myometrium and increased after birth. PAPP-A mRNA was expressed specifically in the endometrial stroma and increased after birth. In Study Two, ewes were treated from birth with estradiol-17beta valerate (EV), which reduces uterine growth and inhibits endometrial adenogenesis. On PNDs 14 and 56, IGFBP-3 mRNA was decreased in the uterus of EV-treated ewes, but IGF-1R and IGFBP-4 mRNAs were not affected. PAPP-A mRNA was increased by EV treatment on PND 14, but decreased on PND 56. These results support the hypothesis that an intrinsic IGF system in the uterus regulates epithelial-stromal interactions important for postnatal uterine growth and endometrial gland morphogenesis in the sheep.     

Galectin-15 in ovine uteroplacental tissues

Galectin-15 is the newest member of a secreted beta-galactoside-binding lectin family. The galectin-15 gene is expressed specifically by the endometrial luminal epithelium (LE) and superficial ductal glandular epithelium (sGE) of the ovine uterus. The proposed extracellular role of secreted galec7tin-15 is to regulate implantation and placentation by functioning as a heterophilic cell adhesion molecule between the conceptus trophectoderm and endometrial LE, while that of intracellular galectin-15 is to regulate cell survival, differentiation and function. The present study determined galectin-15 expression in uteroplacental tissues during gestation and in the postpartum uterus. In the uterine lumen, secreted galectin-15 was found as multimers, particularly on days 14 and 16 of pregnancy. In the endometrial epithelium and conceptus trophectoderm, intracellular galectin-15 protein was found associated with crystalline structures. Between days 20 and 120 of pregnancy, galectin-15 mRNA was expressed specifically by the LE and sGE of the intercaruncular endometrium of ewes. Immunoreactive galectin-15 protein was most abundant in the trophectoderm with lower levels in the endometrial LE and sGE. Galectin-15 protein was detected in allantoic fluid, but not in amniotic fluid. After parturition, galectin-15 mRNA declined in the endometrium from postpartum day (PPD) 1 to 28 and exhibited a variegated expression pattern in the LE and sGE. These results indicate that galectin-15 is synthesized and secreted throughout gestation by the endometrial LE/sGE and is absorbed by the placenta and forms crystals within the trophectoderm, whereas the remainder is cleared into the allantois after being transported into the fetal circulation via the placental areolae. Based on the biological properties of other galectin family members, galectin-15 is hypothesized to have biological roles in conceptus-endometrial interactions, uterine immune and inflammatory responses, and placental morphogenesis and function.     

Redistribution of aquaporins 1 and 5 in the rat uterus is dependent on progesterone: a study with light and electron microscopy

During early pregnancy in the rat there is a dramatic reduction in luminal fluid which is associated with uterine receptivity for blastocyst implantation. This study investigates the presence and distributional changes of several members of the aquaporin (AQP) family in the rat uterus in response to hormonal regime. An increase in apical AQP5 protein expression was found in response to progesterone alone or in combination with oestrogen, which is similar to that seen at the time of implantation. AQP1 was found in endothelial cells of the endometrium and in the inner circular layer of smooth muscle, with maximal protein expression seen after three doses of progesterone plus 8 hr of oestrogen treatment. These results, for the first time, show that the up-regulation of AQP5 in the apical plasma membrane of uterine epithelial cells and AQP1 in the inner circular layer of myometrium, is dependent on progesterone. Furthermore, unlike during normal pregnancy, there is no differential gradient of AQP5 expression between mesometrial and antimesometrial poles of the progesterone treated uterus. Hence it is suggested that the differential gradient of AQP5 is dependent on the presence of a blastocyst, in addition to the appropriate hormonal environment.     

Implantation mechanisms: insights from the sheep

Implantation in all mammals involves shedding of the zona pellucida, followed by orientation, apposition, attachment and adhesion of the blastocyst to the endometrium. Endometrial invasion does not occur in domestic ruminants; thus, definitive implantation is achieved by adhesion of the mononuclear trophoblast cells to the endometrial lumenal epithelium (LE) and formation of syncytia by the fusion of trophoblast binucleate cells with the LE. This review highlights new information on mechanisms regulating the implantation cascade in sheep. The embryo enters the uterus on day 4 at the morula stage of development and then develops into a blastocyst by day 6. The blastocyst sheds the zona pellucida (day 8), elongates to a filamentous form (days 11-16), and adheres to the endometrial LE (day 16). Between days 14 and 16, the binucleate cells begin to differentiate in the trophoblast and subsequently migrate and fuse with the endometrial LE to form syncytia. Continuous exposure of the endometrium to progesterone in early pregnancy downregulates the progesterone receptors in the epithelia, a process which is associated with loss of the cell-surface mucin MUC1 and induction of several secreted adhesion proteins. Recurrent early pregnancy loss in the uterine gland knockout ewe model indicates that secretions of the endometrial epithelia have a physiologic role in blastocyst elongation and implantation. A number of endometrial proteins have been identified as potential regulators of blastocyst development and implantation in sheep, including glycosylated cell adhesion molecule 1 (GlyCAM-1), galectin-15, integrins and osteopontin. The epithelial derived secreted adhesion proteins (GlyCAM-1, galectin-15 and osteopontin) are expressed in a dynamic temporal and spatial manner and regulated by progesterone and/or interferon tau, which is the pregnancy recognition signal produced by the trophoblast during blastocyst elongation. The noninvasive and protracted nature of implantation in domestic animals provides valuable opportunities to investigate fundamental processes of implantation that are shared among all mammals. Understanding of the cellular and molecular signals that regulate uterine receptivity and implantation can be used to diagnose and identify causes of recurrent pregnancy loss and to improve pregnancy outcome in domestic animals and humans.     

Up-regulation of expression of interferon-stimulated gene 15 in the bovine corpus luteum during early pregnancy

Interferon-τ (IFNT), the pregnancy recognition signal in ruminant species, is secreted by conceptus trophectoderm cells and induces expression of IFN-stimulated gene 15 (ISG15) in the uterus and corpus luteum (CL) in ewes. Expression of ISG15 in ovine CL is speculated to be through an endocrine pathway, but it is unclear whether expression of ISG15 in bovine CL is via such a pathway. In this study, CL were obtained from cows on d 16, 25, 60, 120, 180, and 270 of pregnancy, and endometrium, mammary gland, ovarian stroma, and CL were also collected from cows on d 18 of pregnancy and on d 15 and 18 of the estrous cycle. All tissue explants from d 15 of the estrous cycle were cultured in the absence or presence of 100 ng/mL of recombinant bovine IFNT for 24 h. The results indicated that ISG15 and conjugated proteins were expressed in CL of both cyclic and pregnant cows regardless of pregnancy status and were upregulated during early pregnancy. The mammary gland from d 18 of pregnancy did not express ISG15, but explants of the mammary gland from d 15 of the estrous cycle did express ISG15 after being treated with IFNT. However, luteal explants from d 15 of the estrous cycle did not express ISG15 after being cultured for 24 h. In conclusion, ISG15 expression is upregulated in the bovine CL during early pregnancy. Interestingly, cultured CL cells do not respond to IFNT, suggesting that the pregnancy-dependent stimulation of ISG15 expression is controlled by something other than IFNT in the bloodstream.     

Differential expression of peroxisome proliferator-activated receptor delta at implantation sites and in decidual cells of rat uterus

The aim of this study was to examine the expression and regulation of peroxisome proliferator-activated receptor delta (PPARdelta) gene in rat uterus during early pregnancy by in situ hybridization and immunohistochemistry. PPARdelta mRNA expression in the luminal epithelium was high on day 1 of pregnancy, gradually declined from day 2 and was undetectable on day 5 of pregnancy. However, expression in the glandular epithelium began to increase from day 2 and was high on day 5 of pregnancy. There was no detectable PPARdelta immunostaining in the luminal and glandular epithelium from day 1 to day 5. On day 6 of pregnancy when embryos implanted, PPARdelta mRNA and immunostaining were intense in the subluminal stroma at implantation sites. On days 7 and 8, there was strong expression of both PPARdelta mRNA and intense immunostaining in the decidualized area near the lumen. There was low expression of PPARdelta in the subluminal stroma and glandular epithelium under delayed implantation. After delayed implantation was terminated by oestrogen treatment and embryo implantation was initiated, both PPARdelta mRNA and immunostaining were strongly induced in the subluminal stroma. Intense PPARdelta immunostaining was observed in the decidua under artificial decidualization, while no detectable immunostaining was seen in the uninjected control horn. Retinoid X receptor (RXRalpha) immunostaining was seen in the subluminal stroma surrounding the implanting blastocyst on day 6 and in the decidual cells on days 7 and 8 of pregnancy. In conclusion, the high PPARdelta expression at implantation sites and in the decidual cells in rat uterus indicates that PPARdelta may play an important role during implantation and decidualization.     

Expression and regulation of stanniocalcin 1 and 2 in rat uterus during embryo implantation and decidualization

Stanniocalcin-1 (STC-1) is a recently discovered polypeptide hormone, while stanniocalcin-2 (STC-2) is a subsequently identified homologue of stanniocalcin-1. Although previous studies have shown that both STC-1 and -2 are involved in various physiological processes, such as ion transport, reproduction and development, their expression in the uterus and roles in implantation and early pregnancy are unclear. Here we have investigated the expression and regulation of both STC-1 and STC-2 in rat uterus during early pregnancy under various physiological conditions. We show that only basal levels of STC-1 and STC-2 mRNA were detected in the uterus from day one (D1) to day five (D5) of pregnancy. STC-2 immunostaining was gradually increased in the glandular epithelium from day two (D2), with a peak occurring on D5. High levels of both STC-1 and STC-2 mRNA were observed in the stoma cells at the implantation site on day six (D6) of pregnancy, whereas their immunostaining signals were also significant in the luminal epithelium. Basal levels of both STC-1 and STC-2 mRNA and STC-1 immunostaining were detected in the uterus with delayed implantation. After the delayed implantation was terminated by estrogen treatment, both STC-1 and STC-2 mRNA signals were significantly induced in the stroma underlying the luminal epithelium at the implantation site, and STC-2 immunostaining was also observed in the luminal epithelium surrounding the implanting blastocyst. Embryo transfer experiments further confirmed that STC-1 and STC-2 expression at the implantation sites was induced by the implanting blastocyst. Both STC-1 mRNA and immunostaining were seen in the decidualized cells from day seven (D7) to day nine (D9) of pregnancy. STC-2 mRNA was also found in the whole decidua from D7 to D9 of pregnancy; STC-2 protein, however, was strictly localized to the primary deciduas on D7 and D8, with a weak expression in the whole deciduas on D9. Consistent with the normal pregnancy process, strong STC-1 and STC-2 mRNA signals were detected in the decidualized cells under artificial decidualization, whereas only basal levels of STC-1 mRNA and immunostaining were observed in the control horn. These data suggest, for the first time, that STC-1 together with STC-2 may play important roles in the processes of implantation and decidualization in the rat.     

Symposium review: Immunological detection of the bovine conceptus during early pregnancy

Infertility and subfertility reduce the economic viability of dairy production. Inflammation reduces conception rates in dairy cattle, but surprisingly little information exists about the populations and the functions of immune cells at the conceptus–maternal interface during the periattachment period in dairy cattle. Early pregnancy is accompanied by immune stimulation at insemination and conceptus secretion of IFN-τ, pregnancy-associated glycoproteins, prostaglandins, and other molecules whose effects on immune function during early pregnancy have not been determined. Our working hypothesis is that pregnancy induces changes in immune cell populations and functions that are biased toward immunological tolerance, tissue remodeling, and angiogenesis. This review summarizes current knowledge, starting with insemination and proceeding through early pregnancy, as this is the period of maximal embryo loss. Results indicated that early pregnancy is accompanied by a marked increase in the proportion of endometrial immune cells expressing markers for natural killer (CD335) cells and cytotoxic T cells (CD8) along with an increase in cells expressing major histocompatibility class II antigens (macrophages and dendritic cells). This is accompanied by increased abundance of mRNA for IL-15, a natural killer growth factor, and IL-10 in the endometrium during early pregnancy. Furthermore, expression of indoleamine 2,3 dioxygenase was 15-fold greater in pregnant compared with cyclic heifers at d 17, but then declined by d 20. This enzyme converts tryptophan to kynurenine, which alters immune function by creating a localized tryptophan deficiency and by activation of the aryl hydrocarbon receptor and induction of downstream tolerogenic mediators. Expression of the aryl hydrocarbon receptor is abundant in the bovine uterus, but its temporal and spatial regulation during early pregnancy have not been characterized. Pregnancy is also associated with increased expression of proteins known to inhibit immune activation, including programed cell death ligand-1 (CD274), lymphocyte activation gene-3 (CD223), and cytotoxic T-lymphocyte associated protein-4 (CD152). These molecules interact with receptors on antigen-presenting cells and induce lymphocyte tolerance. Current results support the hypothesis that early pregnancy signaling in dairy heifers involves changes in the proportions of immune cells in the endometrium as well as induction of molecules known to mediate tolerance. These changes are likely essential for uterine wall remodeling, placentation, and successful pregnancy.     

Possible involvement of IFNT in lymphangiogenesis in the corpus luteum during the maternal recognition period in the cow

The corpus luteum (CL), which secretes large amounts of progesterone and is thus essential for establishing pregnancy, contains various types of immune cells that may play essential roles in CL function by generating immune responses. The lymphatic system is the second circulation system and is necessary for immune function, but the lymphatic system of the bovine CL has not been characterized in detail. We collected bovine CLs on days 12 and 16 of the estrous cycle (C12 and C16) and days 16 and 40 of early pregnancy (P16 and P40). Lymphatic endothelial hyaluronan receptor 1 (LYVE1) protein was detected in the CL by immunohistochemistry and western blotting and increased at P40 compared with C16. The mRNA expression levels of lymphangiogenic factors, such as vascular endothelial growth factor-C (VEGFC), VEGFD, and their common receptor VEGFR3, as well as the lymphatic endothelial cell (LyEC) marker podoplanin, increased in P16 and P40 CLs. Thus, it is suggested that the lymphatic system of the bovine CL reconstitutes during early pregnancy. Interferon tau (IFNT) from the conceptus in the uterus is a candidate for activating luteal lymphangiogenesis during the maternal recognition period (MRP). We found that treatment of LyECs isolated from internal iliac lymphatic vessels with IFNT stimulated LyEC proliferation and significantly increased mRNA expression of VEGFC and IFN-stimulated gene 15. Moreover, both IFNT and VEGFC induced LyECs to form capillary-like tubes in vitro. In conclusion, it is suggested that new lymphangiogenesis in the bovine CL begins during the MRP and that IFNT may mediate this novel phenomenon.     

Glycosylation dependent cell adhesion molecule 1-like protein and L-selectin expression in sheep interplacentomal and placentomal endometrium

Glycosylation dependent cell adhesion molecule 1 (GlyCAM-1), a mucin component of sheep histotroph produced by glandular epithelium (GE) during early pregnancy, is hypothesized to function in implantation. However, GlyCAM-1 is present in uterine tissues subsequent to implantation suggesting additional functions of this l-selectin-binding ligand. This study focused on uterine GlyCAM-1 expression during placentome development in sheep. Western blot analysis of day 50 pregnant sheep identified 45, 40, and 25 kDa bands in interplacentomal endometrium, 40 and 25 kDa bands in placentomes, and 80 and 40 kDa bands in chorioallantois. The GlyCAM-1 proteins in interplacentomal regions were comparable to those detected in day 15-19 pregnant sheep, however, the 80 kDa form was unique to chorioallantois, and the absence of the 45 kDa GlyCAM-1 in placentomes indicated differences between interplacentomal and placentomal endometrium. Immunofluorescence identified GlyCAM-1 in lumenal epithelium (LE), stromal fibroblasts, and vascular smooth muscle cells. To better define its cellular distribution, GlyCAM-1 was co-localized with either epithelium-specific cytokeratin, smooth muscle-specific alpha-smooth muscle actin (alpha SMA), or stromal-specific vimentin. In interplacentomal endometrium, GlyCAM-1 co-localized with cytokeratin in LE but not in GE. GlyCAM-1 did not co-localize with alpha SMA, and was localized in the extracellular matrix of vimentin-positive stroma. In placentomes, GlyCAM-1 did not co-localize with cytokeratin, but did co-localize with alpha SMA and vimentin. Thus, in contrast to interplacentomal regions, GlyCAM-1 in placentomes was predominantly localized in vasculature rather than epithelial cells. Further, leukocytes expressing L-selectin were localized to the endothelial surface of GlyCAM-1-expressing vessels within placentomes. These data suggest that GlyCAM-1 assumes distinct functions in compartment-specific regions of the sheep uterus.     

The expression of transforming growth factor beta in pregnant rat myometrium is hormone and stretch dependent

From a quiescent state in early pregnancy to a highly contractile state in labor, the myometrium displays tremendous growth and remodeling. We hypothesize that the transforming growth factor beta (TGFbeta) system is involved in the differentiation of pregnant myometrium throughout gestation and labor. Furthermore, we propose that during pregnancy the mechanical and hormonal stimuli play a role in regulating myometrial TGFbetas. The expression of TGFbeta1-3 mRNAs and proteins was examined by real-time PCR, Western immunoblot, and localized with immunohistochemistry in the rat uterus throughout pregnancy and labor. Tgfbeta1-3 genes were expressed differentially in pregnant myometrium. Tgfbeta2 gene was not affected by pregnancy, whereas the Tgfbeta1 gene showed a threefold increase during the second half of gestation. In contrast, we observed a dramatic bimodal change in Tgfbeta3 gene expression throughout pregnancy. Tgfbeta3 mRNA levels first transiently increased at mid-gestation (11-fold on day 14) and later at term (45-fold at labor, day 23). Protein expression levels paralleled the changes in mRNA. Treatment of pregnant rats with the progesterone (P4) receptor antagonist RU486 induced premature labor on day 19 and increased Tgfbeta3 mRNA, whereas artificial maintenance of elevated P4 levels at late gestation (days 20-23) caused a significant decrease in the expression of Tgfbeta3 gene. In addition, Tgfbeta3 was up-regulated specifically in the gravid horn of unilaterally pregnant rats subjected to a passive biological stretch imposed by the growing fetuses, but not in the empty horn. Collectively, these data indicate that the TGFbeta family contributes in the regulation of myometrial activation at term integrating mechanical and endocrine signals for successful labor contraction.     

Relationship of the estrous cycle to milk composition.

The relationship of the estrous cycle to milk composition was examined in 15 Holstein cows in first lactation which were approximately 45 days postpartum. Composite milk samples were collected twice daily and analyzed for progesterone, fat, protein, total solids, sodium, potassium, magnesium, calcium, and somatic cells. Milk progesterone was minimal (less than ng/ml) from day -1 to day 2 (day 0 = estrus) of the estrous cycle and then increased at a relatively constant rate through day 15. Milk yield and sodium and magnesium concentrations of the milk varied with days of the estrous cycle. Mean milk yield was highest and sodium and magnesium concentrations were lowest on day 1. However, sodium concentration of the milk was the only component that varied significantly during the 3 days centered on estrus (days -1, 0, and 1). No other milk component changed significantly during the estrous cycle. Although milk yield and composition varied during the estrous cycle, none of these components appeared to be a practical indicator of estrus.     

Prostaglandin E(2) and F(2 alpha) production by equine conceptuses and concentrations in conceptus fluids and uterine flushings recovered from early pregnant and dioestrous mares

A growing equine conceptus must suppress the cyclical release of PGF(2 alpha) from the endometrium to effect maternal recognition of its presence in the uterus. Paradoxically, the conceptus itself secretes PGF(2 alpha), together with other prostaglandins. In this study, the PGF(2 alpha) and PGE(2) content of, and production in vitro by, day 10-32 equine conceptuses were measured and the influence of pregnancy on the concentrations of these prostaglandins in the uterine lumen was examined. In vitro, the release of both prostaglandins per mg conceptus tissue was very high on day 10 after ovulation and lower thereafter. However, while PGF(2 alpha) production decreased further after day 18 of gestation, PGE(2) production remained high until day 32. Prostaglandin concentrations in yolk sac fluid were unaffected by gestational age and PGE(2) concentrations in this compartment were two to five times higher than PGF(2a) concentrations. PGF(2 alpha) concentrations reached high values in uterine flushings recovered from cyclic mares during days 14-16 after ovulation, the expected time of luteolysis, but were negligible in flushings recovered from pregnant mares at this time. Beyond day 18 of gestation, PGF(2 alpha) concentrations in uterine flushings were high and strikingly similar to those recorded during cyclical luteolysis. It is concluded that the equine conceptus effects maternal recognition of pregnancy primarily by inhibiting the ability of the endometrium to release PGF(2 alpha) during days 12-16 after ovulation. However, the conceptus appears to delay, rather than prevent, the development of the uterine PGF(2 alpha) release pathway and an alternative mechanism must prevent luteolysis from being triggered during days 18-32 of gestation.     

Progesterone concentration in the marsupial Sminthopsis macroura: relationship with the conceptus, uterine glandular regeneration and body weight

Close examination of hormonal profiles and uterine morphology in the marsupial reproductive cycle highlights significant differences between pregnant and non-pregnant cycles. In the polyovular dasyurid marsupial Sminthopsis macroura, we identified changes associated with gestation by comparing ovarian and plasma progesterone concentrations, uterine weights, uterine epithelial mitoses, body weights and gestation lengths between pregnant and non-pregnant luteal phases. The plasma progesterone profile of S. macroura was biphasic, peaking during unilaminar blastocyst expansion and on the day of implantation. Periods of rapid embryonic development were associated with increasing plasma progesterone concentrations and animal body weight. For the first time in a polyovular marsupial, we identified 1) a correlation between ovarian progesterone concentration and conceptus number during the luteal phase just prior to implantation (total ovarian progesterone), indicating a conceptus influence on progesterone concentration; 2) a pulse of uterine epithelial mitotic activity at the time of implantation and 3) increased mitotic activity in pregnant animals during unilaminar blastocyst formation compared with non-pregnant animals. Gestation length was reduced by up to 15%, due to the loss of, or reduction in, the four-cell arrest and more rapid definitive blastocyst expansion. This is the first time a conceptus influence on gestation length has been identified in a dasyurid. This study provides further evidence for the modification of the luteal phase by pregnancy in S. macroura.