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a Departments of Animal Science and Veterinary Anatomy & Public Health,
b Center for Animal Biotechnology and Genomics,
c Institute of Biosciences and Technology, Texas A&M University System Health Science Center, College Station, Texas 77843-2471
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ABSTRACT |
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 TOP ABSTRACT INTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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) acts on the endometrium to increase secretion of several proteins during the pregnancy recognition period in ruminants. One of these is a 70-kDa acidic protein that has not been identified. Our hypothesis was that the 70-kDa acidic protein is osteopontin (OPN). OPN is an acidic glycoprotein that fragments upon freezing and thawing or treatment with proteases including thrombin. OPN contains a Gly-Arg-Gly-Asp-Ser (GRGDS) sequence that binds to cell surface integrins to promote cell-cell attachment and cell spreading. Using antisera to recombinant human OPN, both 70-kDa and 45-kDa proteins were identified in uterine flushings from pregnant ewes by Western blotting. A clone containing the entire ovine OPN cDNA coding sequence was isolated by screening a Day 15 pregnant ovine endometrial cDNA library with a partial ovine OPN cDNA. In pregnant ewes, steady-state levels of OPN endometrial mRNA increased (P < 0.01) after Day 17. In both cyclic and pregnant ewes, in situ hybridization analysis showed that OPN mRNA was localized on unidentified immune cells within the stratum compactum of the endometrium. In pregnant ewes, OPN mRNA was also expressed by the glandular epithelium. Results suggest that progesterone and/or IFN induce expression and secretion of OPN by uterine glands during the periimplantation period and that OPN may induce adhesion between luminal epithelium and trophectoderm to facilitate superficial implantation.
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INTRODUCTION |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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), the signal for maternal recognition of pregnancy. Ovine IFN is secreted by the conceptus trophectoderm between Days 10 and 21 of gestation and blocks transcription of genes for ER and OTR in sheep uterine epithelium [1]. These events attenuate pulsatile release of PGF, prevent regression of the CL, and ensure maintenance of a progestational environment that is required for endometrial support of superficial implantation, placentation, and fetal/placental development.
In addition to signaling pregnancy recognition, IFN increases secretion of several endometrial proteins. These proteins are hypothesized to support conceptus development during the periimplantation period [2]. Specifically, they may act as nutrients, growth factors, immunomodulatory proteins for protection against conceptus allograft rejection, protease inhibitors, enzymes, or molecules that promote adhesion between conceptus trophectoderm and uterine LE. The secretion of at least 11 endometrial proteins is known to be up-regulated by IFN [3], including 1) ß2 microglobulin [4], 2) ubiquitin cross-reactive protein [5], 3) granulocyte chemotactic protein-2 [6], 4) Mx [7], and 5) 2',5'-oligoadenylate synthetase [8]. Another unidentified, secreted 70-kDa protein has a pI of approximately 4 [3].
Osteopontin (OPN), also known as the early T-cell activation-1 (Eta-1) cytokine, is a 70-kDa, acidic (pI 4–5) glycoprotein [9, 10]. Upon freezing and thawing or treatment with proteases, the 70-kDa protein gives rise to 45-kDa and 24-kDa fragments [11]. OPN has been localized to the LE of both mouse and human uterine endometrium [12, 13] and is secreted by a number of tissues [13–15]. OPN/Eta-1 binds sheep red blood cell (SRBC) glycophorin and inhibits the ability of SRBC to interact with mouse T-lymphocytes that express surface receptors specific for SRBC glycophorin [11]. This rosette inhibition activity has been used as an indicator of a viable conceptus [16] and is attributed to early pregnancy factor (EPF) [17, 18].
Our hypothesis was that the 70-kDa IFN-regulated protein was OPN. Therefore, the objectives were to determine 1) if OPN was secreted into the uterine lumen of pregnant ewes, 2) clone a full-length ovine OPN cDNA, and 3) determine temporal and spatial alterations in OPN mRNA expression during the ovine estrous cycle and early pregnancy.
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MATERIALS AND METHODS |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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All experimental and surgical procedures involving animals were approved by the Agricultural Animal Care and Use Committee, Texas A&M University (Animal Use Protocols 7-286 and AG-239AG).
Mature western-range ewes of primarily Rambouillet breeding were observed daily for estrous behavior in the presence of vasectomized rams. After experiencing at least two estrous cycles of normal duration (16–18 days), ewes were assigned randomly on Day 0 (estrus/mating) to cyclic or pregnant status. Ewes assigned to pregnant status were mated to intact rams three times at 12-h intervals beginning at estrus. Fifty-two ewes were ovariohysterectomized (n = 4 ewes/day) on Day 1, 3, 5, 7, 9, 11, 13, or 15 of the estrous cycle or Day 11, 13, 15, 17, or 19 of gestation. At hysterectomy, uteri (excluding those from Day 19) were flushed with 0.9% NaCl, and flushes were frozen at -80°C. Pregnancy was verified by recovery of an apparently normal conceptus in uterine flushes. Several sections (1–1.5 cm) of uterine wall from the middle of each uterine horn were fixed in fresh 4% paraformaldehyde in PBS (pH 7.2). After 24 h, fixed tissues were changed to 70% ethanol for 24 h and then embedded in Paraplast-Plus (Oxford Labware, St. Louis, MO). The remaining endometrium was physically separated by dissection from myometrium, frozen in liquid nitrogen, and stored at -80°C.
Western Blot Analysis
Uterine flushings from cyclic and pregnant ewes were dialyzed against 3 liters of cold dialysis buffer (10 mM Tris, pH 8.2) in Spectrapor 3 dialysis membrane (cutoff, Mr 3500; Spectrum, Houston, TX) for 24 h with stirring at 4°C. Dialysis buffer was changed three times. Samples were then concentrated by snap-freezing in liquid nitrogen followed by lyophilization. Proteins were then resuspended in 10 mM Tris (pH 8.2). Endometrium was thawed and immediately homogenized in extraction buffer (10 mM Tris [pH 7], 1 mM EDTA, 1 mM dithiothreitol [DTT], 100 µg/ml PMSF) at a ratio of 1 g tissue per 5 ml buffer. Homogenates were sonicated for 30 sec with a Mini Ultrasonic Cell Disrupter (Sonics & Materials, Inc., Danbury, CT) and clarified by centrifugation (10 000 x g for 15 min at 4°C). Concentrations of protein in dialyzed uterine flushings and endometrial extracts were determined using a Bradford protein assay (Bio-Rad Laboratories, Hercules, CA) with BSA as the standard. Proteins in uterine flushings (8 µg) and uterine extracts (120 µg) were denatured in Laemmli buffer, separated on 10% (total monomer) 1D or 2D-SDS-PAGE and transferred to nitrocellulose [19]. Blots were blocked overnight in TBST (20 mM Tris [pH 7.5], 137 mM NaCl, 0.05% Tween-20) containing 5% dried milk. Blots were washed three times for 5 min each in TBST and then incubated with a cocktail containing rabbit anti-human OPN IgG (LF-123 and LF-124; 5 µg/ml) [20], or normal rabbit serum (5 µg/ml) in TBST containing 2% dried milk while rocking overnight at 4°C. Blots were then washed three times for 10 min each in TBST with goat anti-rabbit IgG-horseradish peroxidase conjugate (1:15 000 dilution of 1 mg/ml stock; KPL, Bethesda, MD) for 1 h at room temperature while rocking. Blots were washed three times for 10 min each in TBST, and immunoreactive proteins were detected using enhanced chemiluminescence (Amersham Life Sciences, Arlington Heights, Rochester, NY).
Cloning of Partial Ovine OPN cDNA
The bovine OPN sequence [21] was used to derive a forward primer beginning at base 225 (5'-TGATGATAACAGCCAGGACGA-3') and a reverse primer beginning at base 500 (5'-GTGAAGTCCTCCTCTGTGGC-3'). A 291-base pair (bp) OPN cDNA fragment was then amplified by polymerase chain reaction (PCR) from a Day 15 pregnant ovine endometrial cDNA library (Lambda ZAP II library; Stratagene Cloning Systems, La Jolla, CA) as described previously [22, 23]. The 291-bp OPN cDNA was then cloned into the pCR-II vector (InVitrogen, Carlsbad, CA) and sequenced.
cDNA Library Screening
A Day 15 pregnant ovine endometrial Lambda Zap II cDNA library (Stratagene) was screened with the partial ovine OPN cDNA using standard methods [22]. Positive plaques from the initial library screen were subsequently rescreened three times until plaque pure. One clone (10.2.1) containing the entire ovine OPN cDNA coding region was sequenced in both directions using forward and reversed primers as well as internal primers.
Northern Blot Analysis
Total cellular RNA was isolated from cyclic and pregnant endometrial samples using the Trizol reagent (Gibco-BRL, Grand Island, NY). The quantity of RNA was assessed spectrophotometrically, and integrity of RNA was examined by gel electrophoresis in a 1% denaturing agarose gel [22]. Total RNA (20 µg) was loaded onto a 1.2% agarose gel, electrophoresed, and transferred to a 0.2-µm nylon membrane as previously described [23]. The Northern blot was then hybridized with a radiolabeled antisense cRNA probe generated from linearized ovine OPN (10.2.1) plasmid template, washed and visualized by autoradiography for 16 h at -80°C (X-OMAT AR Film; Kodak, Rochester, NY) as previously described [23]. The antisense OPN cRNA probe was made against the OPN 10.2.1 plasmid template in pCR-II, linearized by restriction with NotI, using T7 RNA polymerase, [
-32P]UTP (Amersham), and the Riboprobe Gemini kit (Promega, Madison, WI).
Slot Blot Hybridization Analysis
Steady-state levels of OPN mRNA were measured in cyclic and pregnant endometrial samples using slot blot hybridization analysis. For each ewe, denatured total cellular RNA (20 µg) was hybridized with radiolabeled antisense cRNA probes generated by in vitro transcription with [-32P]UTP (Amersham) as described above. Plasmid templates for ovine OPN (10.2.1) and 18S rRNA (pT718S; Ambion, Austin, TX) were used. The radioactivity in each slot was quantitated using an Instant Imager (Packard Instruments, Meridan, CT) and expressed as total counts.
In Situ Hybridization Analysis
The OPN mRNA was localized in uterine tissue sections by in situ hybridization analysis. Uterine tissue sections were deparaffinized in xylene and then rehydrated to water through a graded series of alcohol. Tissue sections were postfixed in 4% paraformaldehyde in PBS and then digested with Proteinase K (20 µg/ml) in PK digestion buffer (50 mM Tris, 5 mM EDTA, pH 8) for 8 min at 37°C. Sections were then refixed for 5 min in 4% paraformaldehyde, rinsed twice for 5 min each in PBS, dehydrated through a graded series of alcohol, and then dried at room temperature for 30 min. Sections were hybridized with radiolabeled antisense or sense cRNA probes generated from a linearized ovine OPN (10.2.1) plasmid template using in vitro transcription with [-35S]UTP (specific activity: 3000 Ci/mmol; Amersham). Antisense and sense cRNA probes were made from the OPN 10.2.1 plasmid restricted with XhoI and T3 RNA polymerase for sense and restricted with NotI and T7 RNA polymerase for antisense. Radiolabeled cRNA probe (5 x 106 cpm/slide) was denatured in 75 µl hybridization buffer (50% formamide, 0.3 M NaCl, 20 mM Tris-HCL [pH 8], 5 mM EDTA [pH 8], 10 mM sodium phosphate [pH 8], single-strength Denhardt's, 10% dextran sulfate, 0.5 mg/ml yeast RNA, 100 mM DTT) at 70°C for 10 min. Hybridization solution was applied to the middle of each slide and a coverslip placed gently on top. Slides were then incubated in a humidified chamber containing 50% formamide/5-strength SSC and hybridized overnight at 55°C. Coverslips were removed by placing slides in 5-strength SSC/10 mM ßME for 30 min at 55°C. Sections were then washed as follows: 50% formamide/double-strength SSC/50 mM ßME for 20 min at 65°C; single-strength TEN (0.05 M NaCl/10 mM Tris [pH 8]/5 M EDTA) for 10 min at room temperature; and then three times in single-strength TEN for 10 min at 37°C. Sections were then digested with DNase-free RNase (10 µg/ml) in single-strength TEN for 30 min at 37°C to remove nonspecifically bound probe and washed as follows: single-strength TEN for 30 min at 37°C; 50% formamide/double-strength SSC/50 mM ßME for 20 min at 65°C; double-strength SSC for 15 min at room temperature; 0.1-strength SSC for 12 min at room temperature; 70% ethanol containing 0.3 M ammonium acetate for 5 min at room temperature; 95% ethanol containing 0.03 M ammonium acetate for 1 min at room temperature; twice in 100% ethanol; and three times in single-strength TEN for 10 min at 37°C. Liquid film emulsion autoradiography was performed using Kodak NTB-2 liquid photographic emulsion [22]. Slides were stored at 4°C for 5 days, developed in Kodak D-19 developer, counterstained with Harris' modified hematoxylin in acetic acid (Fisher, Fairlawn, NJ), dehydrated through a graded series of alcohol to xylene, coverslipped, and evaluated by both brightfield and darkfield microscopy with a Zeiss Photomicroscope III (Carl Zeiss Inc., Thornwood, NY).
Stainsall Detection of OPN Protein
Endometrium was thawed and homogenized, and the concentration of protein was determined as described for Western blotting. Protein (120 µg) in endometrial extracts was denatured in Laemmli buffer and separated in 10% gels by two-dimensional (2D) SDS-PAGE. After electrophoresis, gels were fixed (10% acetic acid:45% methanol:45% H2O) for 1 h at room temperature, and then soaked in 50% methanol:50% H2O for 4 h with two changes of this solution to remove acetic acid. Phosphoproteins were detected by staining overnight at room temperature (in the dark) with Stainsall [24]: 5 mg Stainsall (1-ethyl-2-[3-(1-ethylnaphtho[1,2-D]thiazolin-2-ylidene)-2-methylpropenyl]naphtho-[1,2-D]thiazolium bromide; Sigma, St. Louis, MO) in 5 ml formamide, 25 ml isopropanol, 0.5 ml 3 M Tris-HCl (pH 8.8), and 69.5 ml H2O. Gels were then soaked in 40% methanol/60% H2O, placed on a gel light-box, and photographed through transmitted white light.
Statistical Analysis
Data were subjected to least-squares analysis of variance (ANOVA) using the general Linear Models (GLM) procedures of the Statistical Analysis System (Users guide 1990, Ver. 6; SAS Institute, Cary, NC). Slot blot hybridization data (total counts) were adjusted for differences in sample loading using the 18S rRNA data as a covariate [25]. All tests of statistical significance were performed using the appropriate error terms according to the expectation of mean squares. Data are presented as least-square means (LSM) with standard errors (SE).
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RESULTS |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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Hybridization and screening of the ovine endometrial cDNA library with the 291-bp ovine OPN cDNA fragment resulted in isolation of 20 positive plaques. Several plaques were purified and cDNA inserts recovered by self excision. One clone, found to be large enough to potentially contain the entire coding sequence, was sequenced in both directions and the nucleotide sequence for ovine OPN cDNA (clone 10.2.1) is shown in Figure 2. The inferred amino acid sequence predicts that ovine OPN is a single chain protein of 278 amino acids with a hydrophobic leader sequence of 16 amino acids characteristic of secreted proteins. Ovine OPN contains 1) a potential calcium phosphate apatite binding region of consecutive Asp residues beginning at amino acid 86, 2) a cell-attachment Gly-Arg-Gly-Asp-Ser (GRGDS) sequence (residues 151–155), 3) a thrombin KS cleavage site (residues 161 and 162), and 4) two glutamines that are recognized substrates for transglutaminase in other species.
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Nucleotide and amino acid similarities of ovine OPN with bovine, rat, mouse, and porcine OPN are summarized in Table 1. Sequences that are highly conserved in all of these species include 1) four of the 9 or 10 residues in the poly-Asp region, 2) the GRGDS, 3) 15 serines that include an SSEEK sequence (residues 26–30), 4) three threonines (residues 131, 140, and 145), 5) an NES sequence (residues 79–81), and 6) glutamines at positions 50 and 52. A feature of the ovine OPN sequence, shared with bovine OPN only, is deletion of 22 amino acids that would otherwise be inserted between residues 196 and 197. Also, ovine and bovine OPN have a KS, rather than RS putative thrombin cleavage site.
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The OPN cDNA detected a ~1-kilobase (kb) mRNA in Northern blot analysis of ovine endometrial total RNA (Fig. 3A). The amount of OPN mRNA increased in pregnant ewes. Steady-state levels (Fig. 3B) did not increase in cyclic ewes but did increase in pregnant ewes after Day 17 (P < 0.05).
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In situ hybridization analysis of cyclic and pregnant uteri revealed two distinct patterns of OPN mRNA expression (Fig. 4). During the estrous cycle and early pregnancy, expression of transcript was observed in a small percentage of cells scattered throughout the endometrium. The cells that expressed OPN mRNA became concentrated in the stratum compactum immediately beneath the LE on Days 1, 5, and 7 of the estrous cycle. This expression was intermittent, and extended along only a small percentage of the total circumference of the uterine wall. In pregnant ewes, OPN mRNA was present in the GE, where expression was first detected in some ewes on Day 13 of pregnancy. Expression of OPN mRNA by GE increased markedly in all ewes between Days 15 and 19 of pregnancy. However, the temporal rise in glandular OPN mRNA varied between ewes. Day 19 conceptuses did not express OPN mRNA (Fig. 4).
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Western blot and Stainsall analysis of endometrial proteins from Day 17 pregnant ewes separated by 2D-PAGE detected a 70-kDa protein (Fig. 5). Western blots were performed using affinity purified rabbit polyclonal antibody raised against recombinant human OPN (rhOPN), whereas the Stainsall reagent stains phosphoproteins and is established as the method of choice for detection of OPN [24, 26, 27]. The stained protein exhibited a range (~5–6) of isoelectric species (Fig. 5).
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DISCUSSION |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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-regulated acidic 70-kDa secretory protein. This hypothesis was reinforced by the presence of increased OPN protein in uterine flushings from pregnant ewes. However, when endometrial protein extracts from Day 17 pregnant ewes were examined by 2D-PAGE, the 70-kDa OPN protein exhibited a number of isoelectric variants ranging in pI from 5–6. Because the IFN-regulated 70-kDa protein has a single pI of 4, identification of this protein as OPN is questionable. The cDNA sequence for ovine OPN shares high homology with those for other mammalian species [28–31] and is most similar to bovine OPN (91.7% amino acid sequence similarity) [21]. Features of OPN are that it 1) is a glycosylated phosphoprotein, 2) contains an RGD peptide integrin binding sequence, 3) is secreted, and 4) is susceptible to cleavage by proteases [9]. The inferred OPN amino acid sequence is consistent with these properties. Ovine OPN has a 16 amino acid hydrophobic signal peptide, GRGDS cell attachment and KS thrombin cleavage sites, 41 serines of which 13 phosphoserines are found in the same location in all species, and three threonine O-glycosylation sites as well as a potential N-glycosylation (NES) site.
Consistent with the presence of OPN protein in uterine flushings, steady-state levels of endometrial OPN mRNA increased during pregnancy. In situ hybridization analysis indicated an increase in OPN mRNA in the GE of some pregnant ewes as early as Day 13, and all ewes by Day 19. In cyclic and pregnant ovine uteri, OPN mRNA was localized to different cell types. In cyclic ewes, OPN mRNA was detected in scattered immune cells that represent a small percentage of total cells in the endometrium. Secretion of OPN by these cells may be responsible for the presence of the 70- and 45-kDa OPN forms in uterine flushings from cyclic ewes. In pregnant ewes, there was significant induction of OPN gene transcript in GE. This increase of GE expression is likely responsible for increased OPN protein in uterine flushings from pregnant ewes.
Pregnancy-specific expression of OPN mRNA in ovine uterine epithelial cells is not surprising. In humans, OPN mRNA has been localized to luminal epithelial cells of the gastrointestinal tract, gall bladder, pancreatic ducts, distal tubules of the kidney, lung, thyroid, GE of the breast, mucinous epithelium of the endocervix, and fallopian tubes [13]. The secretory phase glands of both nonpregnant and pregnant human uteri express OPN [13, 32]. In mice, the uterine epithelium, adjacent to deciduoma elicited in pseudopregnant females, expresses OPN, while the epithelium in the contralateral horn does not [12]. Results of the present study indicate that OPN mRNA is not present in LE or the conceptus but restricted to the endometrial glands of Day 13–19 pregnant ewes.
The role of OPN during the estrous cycle is unknown. The OPN-positive cells in uterine stroma are probably immune cells because 1) OPN/Eta-1 has been cloned and sequenced from activated T helper lymphocytes [33] and may be the most abundant protein secreted by various T lymphocyte populations [11, 34, 35]; 2) OPN/Eta-1 is expressed by monocytes and macrophages after tissue injury [36]; 3) osteoclasts, which deposit OPN/Eta-1 into lacunae of resorbing bone, are specialized tissue macrophages [37]; and 4) OPN/Eta-1 has been detected in activated CD8- CD4- natural killer cells, including granulated metrial gland cells [39]. The immunological role of OPN/Eta-1 may be significant since it binds the CD44 receptor on lymphocytes and monocytes to induce chemotaxis of these cells out of the bloodstream and into sites of inflammation [10]. The OPN/Eta-1 gives rise to 24- and 45-kDa proteins that bind antigen and suppress T-helper cells, respectively [11]. And, the 45-kDa fragment increases immunoglobulin production by B lymphocytes [39].
During early pregnancy, OPN may act as an adhesion molecule. It binds primarily to vß3 integrin heterodimer on tissues via its GRGDS sequence to promote cell-cell attachment, cell spreading, and cell-extracellular matrix communication [40, 41]. In women [42], expression of vß3 is unique to the "implantation window" and increases in response to progesterone in uteri of baboons [43]. Temporal and spatial alterations in expression of extracellular matrix proteins (ECMs) and integrins by the uterus and conceptuses of pigs during the periimplantation period have also been reported [44]. Thus, our working hypothesis is that modulation of expression of OPN by progesterone and/or sheep trophoblast interferons may induce expression and secretion of OPN by uterine epithelium. This OPN then binds vß3 integrin heterodimer expressed by trophectoderm and/or uterus to 1) stimulate changes in morphology of trophectoderm and extra-embryonic endoderm that result in elongation of the conceptus and 2) induce adhesion between luminal epithelium and trophectoderm essential for attachment and superficial implantation. Experiments designed to determine if OPN expression is regulated in vivo by progesterone and/or IFN are in progress.
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ACKNOWLEDGMENTS |
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FOOTNOTES |
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2 Correspondence: Fuller W. Bazer, Department of Animal Science and Center for Animal Biotechnology and Genomics, 442D Kleberg Center, Texas A&M University, College Station, TX 77843-2471. FAX: 409 862 2662; fbazer{at}cvm.tamu.edu
Accepted: May 24, 1999.
Received: February 19, 1999.
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REFERENCES |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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). Endocrine 1997; 6:31–37.[Medline]) alters oestrogen receptor and oxytocin receptor expression in the endometrium of cyclic ewes. J Mol Endocrinol 1995; 15:203–220.vß3 integrin stimulates immediate cell signals in osteoclasts. J Biol Chem 1991; 266:20369–20374.vß3 potentiate bone resorption. J Biol Chem 1993; 263:19433–19436.This article has been cited by other articles:
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  A. Banerjee, R. Rose, G. A. Johnson, R. C. Burghardt, and S. K. Ramaiah The Influence of Estrogen on Hepatobiliary Osteopontin (SPP1) Expression in a Female Rodent Model of Alcoholic Steatohepatitis Toxicol Pathol, June 1, 2009; 37(4): 492 - 501. [Abstract] [Full Text] [PDF]
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 R. C Burghardt, J. R Burghardt, J. D Taylor II, A. T Reeder, B. T Nguen, T. E Spencer, K. J Bayless, and G. A Johnson Enhanced focal adhesion assembly reflects increased mechanosensation and mechanotransduction at maternal-conceptus interface and uterine wall during ovine pregnancy Reproduction, March 1, 2009; 137(3): 567 - 582. [Abstract] [Full Text] [PDF]
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 G. Song, K. A. Dunlap, J. Kim, D. W. Bailey, T. E. Spencer, R. C. Burghardt, G. F. Wagner, G. A. Johnson, and F. W. Bazer Stanniocalcin 1 Is a Luminal Epithelial Marker for Implantation in Pigs Regulated by Progesterone and Estradiol Endocrinology, February 1, 2009; 150(2): 936 - 945. [Abstract] [Full Text] [PDF]
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 K. A. Dunlap, D. W. Erikson, R. C. Burghardt, F. J. White, K. M. Reed, J. L. Farmer, T. E. Spencer, R. R. Magness, F. W. Bazer, K. J. Bayless, et al. Progesterone and Placentation Increase Secreted Phosphoprotein One (SPP1 or Osteopontin) in Uterine Glands and Stroma for Histotrophic and Hematotrophic Support of Ovine Pregnancy Biol Reprod, November 1, 2008; 79(5): 983 - 990. [Abstract] [Full Text] [PDF]
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J. L Herington and B. M Bany The conceptus increases secreted phosphoprotein 1 gene expression in the mouse uterus during the progression of decidualization mainly due to its effects on uterine natural killer cells Reproduction, June 1, 2007; 133(6): 1213 - 1221. [Abstract] [Full Text] [PDF]
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F. J White, R. C Burghardt, J. Hu, M. M Joyce, T. E Spencer, and G. A Johnson Secreted phosphoprotein 1 (osteopontin) is expressed by stromal macrophages in cyclic and pregnant endometrium of mice, but is induced by estrogen in luminal epithelium during conceptus attachment for implantation. Reproduction, December 1, 2006; 132(6): 919 - 929. [Abstract] [Full Text] [PDF]
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A. Banerjee, R. C. Burghardt, G. A. Johnson, F. J. White, and S. K. Ramaiah The Temporal Expression of Osteopontin (SPP-1) in the Rodent Model of Alcoholic Steatohepatitis: A Potential Biomarker Toxicol Pathol, June 1, 2006; 34(4): 373 - 384. [Abstract] [Full Text] [PDF]
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M. D. Ashworth, J. W. Ross, J. Hu, F. J. White, D. R. Stein, U. DeSilva, G. A. Johnson, T. E. Spencer, and R. D. Geisert Expression of Porcine Endometrial Prostaglandin Synthase During the Estrous Cycle and Early Pregnancy, and Following Endocrine Disruption of Pregnancy Biol Reprod, June 1, 2006; 74(6): 1007 - 1015. [Abstract] [Full Text] [PDF]
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G. Song, F. W. Bazer, G. F. Wagner, and T. E. Spencer Stanniocalcin (STC) in the Endometrial Glands of the Ovine Uterus: Regulation by Progesterone and Placental Hormones Biol Reprod, May 1, 2006; 74(5): 913 - 922. [Abstract] [Full Text] [PDF]
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C. A. Gray, C. A. Abbey, P. D. Beremand, Y. Choi, J. L. Farmer, D. L. Adelson, T. L. Thomas, F. W. Bazer, and T. E. Spencer Identification of Endometrial Genes Regulated by Early Pregnancy, Progesterone, and Interferon Tau in the Ovine Uterus Biol Reprod, February 1, 2006; 74(2): 383 - 394. [Abstract] [Full Text] [PDF]
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F. J. White, J. W. Ross, M. M. Joyce, R. D. Geisert, R. C. Burghardt, and G. A. Johnson Steroid Regulation of Cell Specific Secreted Phosphoprotein 1 (Osteopontin) Expression in the Pregnant Porcine Uterus Biol Reprod, December 1, 2005; 73(6): 1294 - 1301. [Abstract] [Full Text] [PDF]
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T. E Spencer, G. A Johnson, F. W Bazer, and R. C Burghardt Implantation mechanisms: insights from the sheep Reproduction, December 1, 2004; 128(6): 657 - 668. [Abstract] [Full Text] [PDF]
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T. E. Spencer, G. A. Johnson, R. C. Burghardt, and F. W. Bazer Progesterone and Placental Hormone Actions on the Uterus: Insights from Domestic Animals Biol Reprod, July 1, 2004; 71(1): 2 - 10. [Abstract] [Full Text] [PDF]
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G. A. Johnson, R. C. Burghardt, F. W. Bazer, and T. E. Spencer Osteopontin: Roles in Implantation and Placentation Biol Reprod, November 1, 2003; 69(5): 1458 - 1471. [Abstract] [Full Text] [PDF]
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G. A. Johnson, R. C. Burghardt, M. M. Joyce, T. E. Spencer, F. W. Bazer, C. A. Gray, and C. Pfarrer Osteopontin Is Synthesized by Uterine Glands and a 45-kDa Cleavage Fragment Is Localized at the Uterine-Placental Interface Throughout Ovine Pregnancy Biol Reprod, July 1, 2003; 69(1): 92 - 98. [Abstract] [Full Text] [PDF]
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G. A. Johnson, R. C. Burghardt, M. M. Joyce, T. E. Spencer, F. W. Bazer, C. Pfarrer, and C. A. Gray Osteopontin Expression in Uterine Stroma Indicates a Decidualization-Like Differentiation During Ovine Pregnancy Biol Reprod, June 1, 2003; 68(6): 1951 - 1958. [Abstract] [Full Text] [PDF]
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K.B.C. Apparao, M. J. Illera, S. A. Beyler, G. E. Olson, K. G. Osteen, M. H. Corjay, K. Boggess, and B. A. Lessey Regulated Expression of Osteopontin in the Peri-Implantation Rabbit Uterus Biol Reprod, May 1, 2003; 68(5): 1484 - 1490. [Abstract] [Full Text] [PDF]
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J. G. Kim, J. H. Song, J. L. Vallet, G. A. Rohrer, G. A. Johnson, M. M. Joyce, and R. K. Christenson Molecular Characterization and Expression of Porcine Bone Morphogenetic Protein Receptor-IB in the Uterus of Cyclic and Pregnant Gilts Biol Reprod, March 1, 2003; 68(3): 735 - 743. [Abstract] [Full Text] [PDF]
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S. Noel, A. Herman, G. A. Johnson, C. A. Gray, M. D. Stewart, F. W. Bazer, A. Gertler, and T. E. Spencer Ovine Placental Lactogen Specifically Binds to Endometrial Glands of the Ovine Uterus Biol Reprod, March 1, 2003; 68(3): 772 - 780. [Abstract] [Full Text] [PDF]
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J. E. Garlow, H. Ka, G. A. Johnson, R. C. Burghardt, L. A. Jaeger, and F. W. Bazer Analysis of Osteopontin at the Maternal-Placental Interface in Pigs Biol Reprod, March 1, 2002; 66(3): 718 - 725. [Abstract] [Full Text] [PDF]
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 K. B. C. Apparao, M. J. Murray, M. A. Fritz, W. R. Meyer, A. F. Chambers, P. R. Truong, and B. A. Lessey Osteopontin and Its Receptor {alpha}v{beta}3 Integrin Are Coexpressed in the Human Endometrium during the Menstrual Cycle But Regulated Differentially J. Clin. Endocrinol. Metab., October 1, 2001; 86(10): 4991 - 5000. [Abstract] [Full Text] [PDF]
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G. A. Johnson, F. W. Bazer, L. A. Jaeger, H. Ka, J. E. Garlow, C. Pfarrer, T. E. Spencer, and R. C. Burghardt Muc-1, Integrin, and Osteopontin Expression During the Implantation Cascade in Sheep Biol Reprod, September 1, 2001; 65(3): 820 - 828. [Abstract] [Full Text] [PDF]
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C. A. Gray, K. M. Taylor, W. S. Ramsey, J. R. Hill, F. W. Bazer, F. F. Bartol, and T. E. Spencer Endometrial Glands Are Required for Preimplantation Conceptus Elongation and Survival Biol Reprod, June 1, 2001; 64(6): 1608 - 1613. [Abstract] [Full Text]
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G. A. Johnson, M. D. Stewart, C. Allison Gray, Y. Choi, R. C. Burghardt, L.-Y. Yu-Lee, F. W. Bazer, and T. E. Spencer Effects of the Estrous Cycle, Pregnancy, and Interferon Tau on 2',5'-Oligoadenylate Synthetase Expression in the Ovine Uterus Biol Reprod, May 1, 2001; 64(5): 1392 - 1399. [Abstract] [Full Text]
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M. D. Stewart, G. A. Johnson, C. A. Gray, R. C. Burghardt, L. A. Schuler, M. M. Joyce, F. W. Bazer, and T. E. Spencer Prolactin Receptor and Uterine Milk Protein Expression in the Ovine Endometrium During the Estrous Cycle and Pregnancy Biol Reprod, June 1, 2000; 62(6): 1779 - 1789. [Abstract] [Full Text]
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G. A. Johnson, T. E. Spencer, R. C. Burghardt, K. M. Taylor, C. A. Gray, and F. W. Bazer Progesterone Modulation of Osteopontin Gene Expression in the Ovine Uterus Biol Reprod, May 1, 2000; 62(5): 1315 - 1321. [Abstract] [Full Text]
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K. M. Taylor, C. A. Gray, M. M. Joyce, M. D. Stewart, F. W. Bazer, and T. E. Spencer Neonatal Ovine Uterine Development Involves Alterations in Expression of Receptors for Estrogen, Progesterone, and Prolactin Biol Reprod, April 1, 2000; 63(4): 1192 - 1204. [Abstract] [Full Text]
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T. E. Spencer, A. Gray, G. A. Johnson, K. M. Taylor, A. Gertler, E. Gootwine, T. L. Ott, and F. W. Bazer Effects of Recombinant Ovine Interferon Tau, Placental Lactogen, and Growth Hormone on the Ovine Uterus Biol Reprod, December 1, 1999; 61(6): 1409 - 1418. [Abstract] [Full Text]
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