Select nutrients in the ovine uterine lumen. VII. Effects of arginine, leucine, glutamine, and glucose on trophectoderm cell signaling, proliferation, and migration.

Histotroph is required for survival and development of ovine conceptuses (embryo and extraembryonic membranes). Results from our laboratory indicate that arginine (Arg), leucine (Leu), glutamine (Gln), and glucose increase in the uterine lumen between Days 10 and 15 of pregnancy, coincident with increases in expression of amino acid and glucose transporters by uterine epithelia as well as trophectoderm and yolk sac of conceptuses and elongation of the conceptus trophectoderm. Therefore, we hypothesized that Arg, Leu, Gln, and glucose have differential effects on hypertrophy, hyperplasia, and differentiated functions of trophectoderm cells that are critical to conceptus development.

Select nutrients in the ovine uterine lumen. VIII. Arginine stimulates proliferation of ovine trophectoderm cells through MTOR-RPS6K-RPS6 signaling cascade and synthesis of nitric oxide and polyamines.

During the peri-implantation period in sheep, L-arginine (L-Arg) in the uterine lumen is an essential substrate for the synthesis of nitric oxide (NO), by nitric oxide synthase (NOS), and polyamines, via arginase and ornithine decarboxylase, that are required for survival and development of ovine conceptuses (embryo and its extraembryonic membranes). L-Arginine can stimulate hypertrophy, hyperplasia, and differentiation of the ovine conceptus trophectoderm; however, the responsible signal transduction cascade has not been determined. Therefore, this study examined possible signaling pathways mediated by L-Arg, as well as the effects of two NO donors (S-nitroso-N-acetyl-DL-penicillamine and diethylenetriamine) and putrescine (precursor for spermidine and spermine) on oTr cell proliferation.

Viral particles of endogenous betaretroviruses are released in the sheep uterus and infect the conceptus trophectoderm in a transspecies embryo transfer model.

The sheep genome contains multiple copies of endogenous betaretroviruses highly related to the exogenous and oncogenic jaagsiekte sheep retrovirus (JSRV). The endogenous JSRVs (enJSRVs) are abundantly expressed in the uterine luminal and glandular epithelia as well as in the conceptus trophectoderm and are essential for conceptus elongation and trophectoderm growth and development. Of note, enJSRVs are present in sheep and goats but not cattle.

Endogenous retroviruses in trophoblast differentiation and placental development.

Endogenous retroviruses (ERVs) are present in the genome of all vertebrates and originated from infections of the germline of the host by exogenous retroviruses. ERVs have coevolved with their hosts for millions of years and are recognized to contribute to genome plasticity, protect the host against infection of related pathogenic and exogenous retroviruses, and play a vital role in development of the placenta. Consequently, some ERVs have been positively selected and maintained in the host genome throughout evolution.

Secreted phosphoprotein 1 binds integrins to initiate multiple cell signaling pathways, including FRAP1/mTOR, to support attachment and force-generated migration of trophectoderm cells.

Attachment and migration of trophectoderm (Tr) cells, hallmarks of blastocyst implantation in mammals, are unique uterine events. Secreted phosphoprotein 1 (SPP1) in the uterus binds integrins on conceptus Tr and uterine luminal epithelium (LE), affecting cell-cell and cell-matrix interactions. The signal transduction pathways activated by SPP1 and integrins in conceptuses have not been elucidated.

Cathepsin B, cathepsin L, and cystatin C in the porcine uterus and placenta: potential roles in endometrial/placental remodeling and in fluid-phase transport of proteins secreted by uterine epithelia across placental areolae.

Cathepsins (CTSB and CTSL1) and their inhibitor, cystatin C (CST3), remodel uterine endometrium and placenta for transport of gases, micronutrients, and macromolecules essential for development and growth of the conceptus (embryo/fetus and placental membranes). We examined the temporal/spatial control of expression for CTSB, CTSL1, and CST3 mRNAs in endometria and placentae of pigs using three developmental models: 1) pigs were hysterectomized during the estrous cycle or pregnancy; 2) cyclic pigs were injected with estrogen to induce pseudopregnancy and were hysterectomized; and 3) pigs were ovariectomized, injected with progesterone, and hysterectomized. The abundance of CTSB, CTSL1, and CST3 mRNAs increased in endometrial epithelia during pregnancy and in response to exogenous progesterone but not estrogen.

Endogenous retroviruses of sheep: a model system for understanding physiological adaptation to an evolving ruminant genome.

Endogenous retroviruses (ERVs) are present in the genome of all vertebrates and are remnants of ancient exogenous retroviral infections of the host germline transmitted vertically from generation to generation. Sheep betaretroviruses offer a unique model system to study the complex interaction between retroviruses and their host. The sheep genome contains 27 endogenous betaretroviruses (enJSRVs) related to the exogenous and pathogenic Jaagsiekte sheep retrovirus (JSRV), the causative agent of a transmissible lung cancer in sheep.

Select nutrients and their associated transporters are increased in the ovine uterus following early progesterone administration.

The intrauterine milieu is a complex mixture of substances originating from serum and endometrium that support blastocyst growth and development. The present study identified alterations in glucose and amino acids in response to an early rise in progesterone (P4), which accelerates blastocyst growth and development. Bred ewes received daily injections of either corn oil (CO) vehicle or P4 from 36 h postmating (Day 0) to either Day 9 or Day 12.

Progesterone and interferon tau-regulated genes in the ovine uterine endometrium: identification of periostin as a potential mediator of conceptus elongation.

During early pregnancy in ruminants, progesterone (P(4)) and interferon tau (IFNT) act on the endometrium to regulate genes hypothesized to be important for conceptus development and implantation. The present study was conducted to verify several candidate genes (actin alpha-2, smooth muscle, aorta (ACTA2), collagen, type III, alpha-1 (COL3A1), periostin (POSTN), secreted protein acidic cysteine-rich (SPARC), S100 calcium-binding protein A2 (S100A2), STAT5A and transgelin (TAGLN)) regulated by pregnancy, P(4), and/or IFNT in the endometrium determined using a custom ovine cDNA array. S100A2 mRNA was detected primarily in endometrial epithelia and conceptuses.

Select nutrients in the ovine uterine lumen. VI. Expression of FK506-binding protein 12-rapamycin complex-associated protein 1 (FRAP1) and regulators and effectors of mTORC1 and mTORC2 complexes in ovine uteri and conceptuses.

FRAP1 (FK506-binding protein 12-rapamycin complex-associated protein 1), a component of the nutrient-sensing cell signaling pathway, is critical for cell growth and metabolism. The present study determined expression of FRAP1 and associated members of the mTORC1 and mTORC2 cell signaling pathways in uteri of cyclic and pregnant ewes and conceptuses, as well as effects of pregnancy, progesterone (P4), and interferon tau (IFNT) on their expression. The mRNAs for FRAP1, LST8, MAPKAP1, RAPTOR, RICTOR, TSC1, TSC2, RHEB, and EIF4EBP1 were localized to luminal, superficial glandular, and glandular epithelia and stromal cells of uteri from cyclic and pregnant ewes, as well as trophectoderm and endoderm of conceptuses between Days 13 and 18 of pregnancy.

Select nutrients in the ovine uterine lumen. V. Nitric oxide synthase, GTP cyclohydrolase, and ornithine decarboxylase in ovine uteri and peri-implantation conceptuses.

Nitric oxide (NO) and polyamines are critical for implantation and development of conceptuses (embryo and extraembryonic membranes), but mechanisms regulating their biosynthesis in uteri and conceptuses are largely unknown. This study determined the effects of the estrous cycle, pregnancy, progesterone, and interferon tau (IFNT) on expression of NO synthases (NOS1, NOS2, and NOS3), guanosine triphosphate (GTP) cyclohydrolase (GCH1, the key enzyme in de novo synthesis of tetrahydrobiopterin, a cofactor for NO production), and ornithine decarboxylase (ODC1) in uterine endometria in cyclic ewes (Days 10-16) and pregnant ewes (Days 10-20). The mRNAs and proteins for NOS1 and ODC1 were most abundant in uterine luminal (LE) and superficial glandular (sGE) epithelia, and abundance was affected by day of estrous cycle and early pregnancy.

Interferons and uterine receptivity.

This article focuses on the potential roles of interferons (IFNs) in establishing uterine receptivity to implantation. A common feature of the peri-implantation period of pregnancy in most mammals is production of type I and/or type II IFNs by trophoblasts that induce and/or stimulate expression of an array of IFN-stimulate genes (ISGs). These effects range from pregnancy recognition signaling in ruminants through IFN tau to effects on cellular functions of the uterus and uterine vasculature.

Select nutrients in the ovine uterine lumen. IV. Expression of neutral and acidic amino acid transporters in ovine uteri and peri-implantation conceptuses.

The availability of specific neutral and acidic amino acids in the uterine lumen of ewes increased significantly during the peri-implantation period, but mechanisms for their transport into the uterine lumen and uptake by conceptuses are not established in any species. In this study, effects of pregnancy, progesterone (P4), and interferon tau (IFNT) on expression of neutral and acidic amino acid transporters in uteri of cyclic and pregnant ewes and conceptuses were studied. SLC1A2, SLC1A3, SLC3A1, SLC6A14, SLC6A19, SLC7A6, SLC38A3, and SLC38A6 mRNAs were only weakly expressed in the ovine endometrium.

Progesterone and interferon tau regulate leukemia inhibitory factor receptor and IL6ST in the ovine uterus during early pregnancy.

The actions of leukemia inhibitory factor (LIF) via LIF receptor (LIFR) and its co-receptor, IL6 signal transducer (IL6ST), are implicated in uterine receptivity to conceptus implantation in a number of species including sheep. The present study determined the effects of the estrous cycle, pregnancy, progesterone (P4), and interferon tau (IFNT) on the expression of LIFR and IL6ST in the ovine uterus. LIFR mRNA and protein were localized to the endometrial luminal (LE) and superficial glandular epithelia (sGE), whereas IL6ST mRNA and protein were localized primarily in the middle to deep GE.

Select nutrients in the ovine uterine lumen. III. Cationic amino acid transporters in the ovine uterus and peri-implantation conceptuses.

Arginine is an essential amino acid for conceptus (embryo/fetus and trophoblast/placenta) growth and development; however, the mechanisms for arginine transport into the uterine lumen and uptake by conceptuses are largely unknown. In this study, expression of System y(+) (SLC7A1, SLC7A2, and SLC7A3) cationic amino acid transporters in uteri of cyclic and pregnant ewes and conceptuses was studied, and effects of pregnancy, progesterone (P4), and interferon tau (IFNT) on their expression were investigated. SLC7A1 mRNA was most abundant in endometrial luminal (LE) and superficial glandular (sGE) epithelia on Day 16 of the estrous cycle and on Days 16-20 of pregnancy, whereas SLC7A2 mRNA was most abundant in LE and mid to deep glandular (GE) epithelia on Days 14-20 of gestation.

Interferon regulatory factor 6 (IRF6) is expressed in the ovine uterus and functions as a transcriptional activator.

Interferon tau (IFNT), the maternal recognition of pregnancy signal in sheep and other ruminants, is secreted by the conceptus and regulates the expression of a number of genes in a cell-specific manner within the uterus. The response of different endometrial cell types to IFNT appears to be specified by IFN regulatory factors (IRFs). IRF2, a potent repressor of gene transcription, is expressed only by luminal (LE) and superficial glandular epithelia (sGE), whereas IRF1 and IRF9, activators of gene transcription, are expressed only in GE and stromal cells of the uterus during early pregnancy.

Progesterone regulation of the endometrial WNT system in the ovine uterus.

WNT signalling regulates cell proliferation, differentiation, polarity and organisation. The present study investigated the effects of progesterone (P4) on the endometrial WNT system in relation to blastocyst development and growth in sheep. Ewes received daily intramuscular injections of either corn oil (CO) vehicle or 25 mg P4 from 36 h after mating (Day 0) until hysterectomy on Day 9 or 12.

Stanniocalcin 1 is a luminal epithelial marker for implantation in pigs regulated by progesterone and estradiol.

Stanniocalcin 1 (STC1) is a glycoprotein that decreases calcium and increases phosphate in cells/tissues. This investigation examined endocrine regulation of STC1 in endometria of pigs during the estrous cycle and pregnancy. STC1 mRNA was present exclusively in luminal epithelium (LE) between d 12 and 15 of the estrous cycle, increased between d 12 and d 20, and was not detectable by d 30 of pregnancy.

Select nutrients in the ovine uterine lumen. I. Amino acids, glucose, and ions in uterine lumenal flushings of cyclic and pregnant ewes.

Nutrients in uterine secretions are essential for development and survival of conceptuses (embryo and associated extraembryonic membranes) during pregnancy; however, little is known about changes in the amounts of specific nutrients in the uterine fluids of cyclic and pregnant ruminants. This study determined quantities of glucose, amino acids, glutathione, calcium, sodium, and potassium in uterine lumenal fluid from cyclic (Days 3-16) and pregnant (Days 10-16) ewes. Total recoverable glucose, Arg, Gln, Leu, Asp, Glu, Asn, His, beta-Ala, Tyr, Trp, Met, Val, Phe, Ile, Lys, Cys, Pro, glutathione, calcium, and sodium were greater in the uterine fluid of pregnant compared with cyclic ewes between Days 10 and 16.

Select nutrients in the ovine uterine lumen. ii. glucose transporters in the uterus and peri-implantation conceptuses.

Total glucose in ovine uterine lumenal fluid increases 6-fold between Days 10 and 15 of gestation, but not the estrous cycle; however, mechanisms for glucose transport into the uterine lumen and uptake by conceptuses (embryo/fetus and associated membranes) are not established. This study determined the effects of the estrous cycle, pregnancy, progesterone (P4), and interferon tau (IFNT) on expression of both facilitative (SLC2A1, SLC2A3, and SLC2A4) and sodium-dependent (SLC5A1 and SLC5A11) glucose transporters in ovine uterine endometria from Days 10 to 16 of the estrous cycle and Days 10 to 20 of pregnancy, as well as in conceptuses from Days 10 to 20 of pregnancy. The SLC2A1 and SLC5A1 mRNAs and proteins were most abundant in uterine luminal epithelia and superficial glandular epithelia (LE/sGE), whereas SLC2A4 was present in stromal cells and glandular epithelia (GE).

Progesterone regulates FGF10, MET, IGFBP1, and IGFBP3 in the endometrium of the ovine uterus.

Progesterone (P4) is unequivocally required to maintain a uterine environment conducive to pregnancy. This study investigated the effects of P4 treatment on expression of selected growth factors (fibroblast growth factor 7 [FGF7], FGF10, hepatocyte growth factor [HGF], and insulin-like growth factors [IGF1 and IGF2]), their receptors (MET, FGFR2(IIIB), and IGF1R), and IGF binding proteins (IGFBPs) in the ovine uterus. Ewes received daily injections of corn oil vehicle (CO) or 25 mg of P4 in vehicle from 36 h after mating (Day 0) to hysterectomy on Day 9 or Day 12.

Uterine MHC class I molecules and beta 2-microglobulin are regulated by progesterone and conceptus interferons during pig pregnancy.

MHC class I molecules and beta(2)-microglobulin (beta(2)m) are membrane glycoproteins that present peptide Ags to TCRs, and bind to inhibitory and activating receptors on NK cells and other leukocytes. They are involved in the discrimination of self from non-self. Modification of these molecules in the placenta benefits pregnancy, but little is known about their genes in the uterus.

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.

Insulin-like growth factor II activates phosphatidylinositol 3-kinase-protooncogenic protein kinase 1 and mitogen-activated protein kinase cell Signaling pathways, and stimulates migration of ovine trophectoderm cells.

IGF-II, a potent stimulator of cellular proliferation, differentiation, and development, regulates uterine function and conceptus growth in several species. In situ hybridization analyses found that IGF-II mRNA was most abundant in the caruncular endometrial stroma of both cyclical and pregnant ewes. In the intercaruncular endometrium, IGF-II mRNA transitioned from stroma to luminal epithelium between d 14 and 20 of pregnancy.