Onset of secretion of proteins with antiviral activity by pig conceptuses

In Exp. 1, antiviral activity was detected in Day-15 pregnant uterine flushings (6222 +/- 2167 units/ml) and in conceptus culture medium collected at 0, 1, 2, 4, 8, 16, 24, 32, 40, and 48 h (95, 375, 650, 1216, 1600, 2100, 2017, 2083, 3500 and 5000 units/ml, respectively; R2 = 0.81, P less than 0.01; y = 190.0 + 252.7x - 11.2x2 + 0.2x3. In Exp. 2, antiviral activity of Day-15 conceptus culture medium was reduced 99% after boiling for 20 min (P less than 0.01) and, after 18 h dialysis (6000-8000 Mr cut-off), 100% of the activity was in the retentate. In Exp. 3, antiviral activity was not detected in cultures of conceptuses from Days 10 and 11 and activity was maximal for Day 14 and Day 15 conceptuses (2100 and 2083 units/ml, respectively). Effects of day were best described by a quadratic regression equation (y = 17,652 - 3263x + 150x2; R2 = 0.55, P less than 0.01). In Exp. 4, changes in antiviral activity detected in uterine flushings from pregnant gilts on Days 8, 10, 11, 12, 14 and 15 (1.3, 0, 6.7, 63.3, 580 and 1663 units/ml, respectively) were described by the equation y = -20,743 + 6189x - 606x2 + 20x3 (R2 = 0.85, P less than 0.01). In Exp. 5, low antiviral activities (5-30 units/ml) were detected in all plasma samples collected from the uterine artery and uterine vein of pregnant and cyclic gilts, but values were not significantly influenced by pregnancy status, day or site of collection.(ABSTRACT TRUNCATED AT 250 WORDS)     

Steroid metabolism by endometrial and conceptus tissues during early pregnancy and pseudopregnancy in gilts

Endometrial and conceptus tissues were obtained on Days 10.5, 11, 12, 16 and 25 of pregnancy and Day 25 of pseudopregnancy of gilts and incubated for 6 h in Minimal Essential Medium (5 ml) containing 35 ng [3H]progesterone. Metabolism of [3H]progesterone to oestrone, oestradiol and oestriol was determined by gas and high-pressure liquid chromatography and successive recrystallizations with unlabelled standards. Conceptuses collected between Days 10.5 and 12 were spherical, tubular or filamentous and incubated with 500 mg endometrium and [3H]progesterone. Production of oestrone by spherical conceptuses was not detected, but was 44-47 pg/tubular conceptus and 21 pg/filamentous conceptus. A similar trend was observed for oestradiol. Conceptus tissues from Days 16 and 25 (chorion) were most active in producing oestrone (123 and 520 pg/mg tissue, respectively) and oestradiol (277 and 876 pg/mg tissue, respectively). Endometrial oestrogen production was less than that for conceptus tissue for oestrone and oestradiol on Days 16 and 25 of gestation. Coincubations of endometrium and conceptus tissues had lower oestrogen production than conceptus alone. Endometrium from Day 25 of pseudopregnancy metabolized [3H]progesterone to several non-polar metabolites, but no oestrogens were detected. An unidentified phenolic metabolite of [3H]progesterone was detected in higher quantities than either oestrone or oestradiol; 445 to 461 pg/conceptus at the tubular stage. These results indicate temporal changes in the conversion of [3H]progesterone to oestrogens by conceptus and endometrial tissue from pregnant gilts, but not endometrium from pseudopregnant gilts.     

Pattern of medium proteins radiolabelled after culture of Day 13 to 16 pig conceptus tissue with [3H]leucine and absence of chorionic gonadotrophin-like activity

Day 13-16 pig conceptus tissue was cultured for 24 h in medium containing [3H]leucine. The patterns of radioactively labelled proteins that were released into the medium during culture were analysed by SDS-polyacrylamide gel electrophoresis and fluorography. Day-13 conceptuses released two major radiolabelled proteins of Mr 23 000 and 26 000 and Day 14-16 conceptuses released these as well as proteins of Mr 14 000, 19 000, 44 000, 50 000 and 88 000. Various immunological and biological tests for a human chorionic gonadotrophin-like activity were performed on tissue extracts and on culture medium, but there was no evidence for its presence in the pig conceptus at Day 13-16 of gestation.     

Endometrial surface and secretory alterations associated with embryonic mortality in gilts administered estradiol valerate on days 9 and 10 of gestation

Administration of estrogen to gilts on Days 9 and 10 of pregnancy results in total embryonic loss by Day 18. The present study examined changes in the uterine endometrial surface and secretion during conceptus attachment in control and estrogen-treated (Days 9 and 10) pregnant gilts. Gilts were unilaterally hysterectomized on either Days 12 and 14 or Days 16 and 18 of gestation. Uterine horns were flushed with saline and conceptuses were evaluated. Intact conceptuses were recovered from all control gilts, whereas estrogen-treated gilts contained normal intact conceptuses only on Day 12 of gestation. Antiviral activity, which reflects conceptus viability, was reduced (p less than 0.01) in uterine flushings after Day 14 in estrogen-treated gilts. Culture of endometrial explants with [3H]glucosamine revealed several glycoproteins that are synthesized during the period of conceptus attachment; however, no difference in glycoprotein synthesis between treatment groups was detected by analysis with two-dimensional PAGE and fluorography. Analyses of the uterine epithelium by scanning and transmission electron microscopy demonstrated that estrogen administration caused an alteration in the uterine surface, a thinning of the uterine epithelial glycocalyx, and a reduction of cationic ferritin binding to the microvilli of the uterine epithelium. Results indicate that conceptus mortality after early administration of estrogen is associated with alterations in the uterine endometrial surface during the period of conceptus attachment in the pig.     

Effect of porcine conceptus secretory proteins on in vitro secretion of prostaglandins-F2 alpha and -E2 from luminal and myometrial surfaces of endometrium from cyclic and pseudopregnant gilts

Uterine endometrium collected from pseudopregnant (PP) and cyclic gilts on day (D) 15 after estrus were perifused in vitro with 10 ug/ml of porcine conceptus secretory proteins (pCSP) or serum proteins (SP) in Krebs ringer bicarbonate (KRB) buffer. In Experiment 1, samples were collected from luminal and myometrial surfaces of endometrium and concentrations of prostaglandin F2 alpha (PGF) determined by radioimmunoassay (RIA). Secretion of PGF by endometrium from cyclic gilts was stimulated (P less than .05) by pCSP. In Experiment 2, endometrium from D 14 cyclic and PP gilts was perifused and concentrations of PGF and prostaglandin E2 (PGE) in perfusate were determined by RIA. Across both statuses, luminal surface secretion of PGF was stimulated (P less than .05) by pCSP. Treatment with pCSP decreased secretion of PGE from myometrial surface of endometrium from cyclic gilts and increased (P less than .01) secretion of PGE from the myometrial surface of endometrium from PP gilts. In Experiment 3, pCSP were separated into acidic and basic fractions by anion exchange chromatography and each fraction was perifused separately over the luminal surface of endometrium from cyclic and PP gilts. Perifusion with acidic pCSP suppressed secretion of PGF by endometrium from cyclic or PP gilts; while basic pCSP did not influence secretion of PGF. These results demonstrated that products secreted by Day 15 pig conceptuses stimulate release of PGF and PGE from porcine uterine endometrium.     

Proteins secreted by the sheep conceptus suppress induction of uterine prostaglandin F-2 alpha release by oestradiol and oxytocin

In Exp. I, 0.5 mg oestradiol or vehicle (0.5 ml absolute ethanol + 0.5 ml 0.9% NaCl) was injected i.v. at 08:00 h on Day 14 (onset of oestrus = Day 0). Blood samples were obtained via a jugular catheter at 30 and 1 min before oestradiol and every 30 min for 10 h afterwards. Plasma was obtained and assayed for 15-keto-13,14-dihydro-PGF-2 alpha (PGFM) by radioimmunoassay. Before oestradiol, PGFM basal values were higher (P less than 0.01) in pregnant (N = 10) than nonpregnant (N = 6) ewes (193 +/- 30 vs 67 +/- 8 pg/ml). However, at 4-10 h after oestradiol, pregnant ewes (N = 5) had less variable (P less than 0.01) PGFM values than did nonpregnant ewes (N = 5). In Exp II, conceptus secretory proteins (CSP) were obtained by pooling medium from cultures of Day-16 sheep conceptuses (N = 40). Ewes received 750 micrograms CSP + 750 micrograms plasma protein (N = 6) or 1500 micrograms plasma protein (N = 6) per uterine horn at 08:00 h and 18:00 h on Days 12-14. All ewes received 0.5 mg oestradiol at 08:00 h on Day 14 and blood samples were collected as in Exp. I and assayed for PGFM. On Day 15, 3 ewes in each group received 10 i.u. oxytocin and 3 received saline i.v. at 08:00 h and blood samples were taken continuously from 10 min before to 60 min after treatment. Mean PGFM response to oestradiol was suppressed (P = 0.05) in CSP- vs plasma protein-treated ewes (371 +/- 129 vs 1188 +/- 139 pg/ml).(ABSTRACT TRUNCATED AT 250 WORDS)     

Porcine conceptuses secrete an interferon during the preattachment period of early pregnancy

Maternal recognition of pregnancy in sheep and cattle is thought to be initiated by the conceptus secretory proteins ovine trophoblast protein-1 (oTP-1) and bovine trophoblast protein-1 (bTP-1), respectively. Recently, these proteins have been shown to be members of the interferon-alpha (IFN-alpha) family. In this study, we have examined whether pig conceptuses also produce IFN during early pregnancy. conceptuses were collected at Days 8, 10, 11, 12, 13, 15, and 17 of pregnancy and cultured in serum-free medium for 24 h. Day 11 conceptuses secreted a dominant 22,000-24,000 Mr cluster of acidic proteins (pI 5.2-5.4), that appeared to cross-react on immunoblots with antiserum against human IFN-alpha but not against oTP-1. Antiviral activity characteristic of an IFN was present in conceptus culture medium and uterine flushings from Day 11 through Day 17 of pregnancy, but was absent in flushings prior to Day 11 of pregnancy and in flushings from Day 12 nonpregnant gilts. The antiviral activity coeluted with a 22,000-24,000 Mr protein during partial purification through a gel filtration column. The activity was extremely labile, but could be restored by sequential protein denaturation, reduction, and renaturation. We conclude that production of IFN by early conceptuses is not restricted to ruminant species, and may therefore represent a more general phenomenon.     

Regulation of expression and role of leukemia inhibitory factor and interleukin-6 in the uterus of early pregnant pigs

Cytokines, which are generally involved in the process of inflammation, may also play a critical role in conceptus implantation. We examined: (1) the expression profiles of leukemia inhibitory factor (LIF) and interleukin (IL)-6 mRNA and their protein content in the endometrium of cyclic and pregnant gilts on Days 10 to 18 after estrus; (2) the effect of conceptus-exposed medium on LIF and IL-6 synthesis in the endometrium; (3) the profiles of IL6R and LIFR mRNA expression in pig conceptuses collected on Days 10 to 18 of pregnancy; and (4) the effect of LIF and IL-6 on the attachment and proliferation of porcine trophoblast cells. The expression of LIF mRNA in the endometrium increased between Days 10 and 12 in both cyclic and pregnant gilts, and tended to be higher in Day 12 pregnant animals compared with nonpregnant ones. The LIF protein content in the uterine lumen peaked on Day 12 of pregnancy, and was higher than on Day 12 of the estrous cycle. Endometrial IL-6 mRNA expression was upregulated on Day 12 in pregnant gilts compared with nonpregnant animals. Moreover, a higher content of IL-6 protein was observed in pregnant than in cyclic gilts. The addition of conceptus-exposed medium resulted in up-regulation of LIF and IL6 mRNA expression, and increased IL-6 content in endometrial slices. In conceptuses, increased mRNA expression was detected on Days 10 to 14 for IL6R and on Day 14 for LIFR, when compared with other days studied. LIF and IL-6 stimulated the attachment and proliferation of trophoblast cells in vitro. In summary, LIF and IL-6 are important components of embryo-uterine interactions during early pregnancy in the pig, and may contribute to successful conceptus implantation.     

Insulin-like growth factor-I expression during early conceptus development in the pig

Porcine conceptuses (embryo and associated membranes) in utero undergo developmental morphological transformations coincident with structural and biochemical changes in the uterine endometrium during early gestation. To elucidate a possible role for insulin-like growth factor-I (IGF-I) in these events, porcine endometrial (Days 8, 10, 11, 12, 14, and 30) and conceptus (Days 12, 14, and 16) tissues were characterized for the presence of IGF-I peptide and mRNAs. The corresponding uterine luminal fluids (ULF) at these stages of pregnancy were also analyzed for immunoreactive IGF-I concentration. ULF IGF-I was lowest on Day 8, highest on Day 12, and declined by Day 14. In contrast, endometrial tissue IGF-I content remained constant during this period. Conceptus tissues contained less IGF-I than endometrial tissues; however, conceptus IGF-I values were maximum on Day 12 coincident with peak values for ULF IGF-I. Dot-blot hybridization analyses revealed temporal variation in steady-state levels of IGF-I mRNAs in endometrium. Highest levels of endometrial IGF-I mRNA were detected on Day 12 and were about 4-fold greater than on Day 30 of pregnancy. IGF-I mRNA expression in conceptus tissues on Days 12, 14, and 16 was the same and was significantly less than that in endometrium on Day 12. These results demonstrate the temporal variation of IGF-I mRNA abundance in uterine endometrium and of immunoreactive IGF-I in ULF and in conceptus tissues, with the developmental processes occurring in the conceptuses at early pregnancy.(ABSTRACT TRUNCATED AT 250 WORDS)     

Characterization of the interleukin-1beta system during porcine trophoblastic elongation and early placental attachment

Conceptus-uterine communication is established during trophoblastic elongation when the conceptus synthesizes and releases estrogen, the maternal recognition signal in the pig. Interleukin-1beta (IL-1beta) is a differentially expressed gene during rapid trophoblastic elongation in the pig. The current investigation determined conceptus and endometrial changes in gene expression for IL-1beta, IL-1 receptor antagonist (IL-1Rant), IL-1 receptor type 1 (IL-1RT1), and IL-1 receptor accessory protein (IL-1RAP) in developing peri- and postimplantation conceptuses as well as uterine endometrium collected from cyclic and pregnant gilts. Conceptus IL-1beta gene expression was enhanced during the period of rapid trophoblastic elongation compared with earlier spherical conceptuses, followed by a dramatic decrease in elongated Day 15 conceptuses. IL-1RT1 and IL-1RAP gene expression was greater in Day 12 and 15 filamentous conceptuses compared with earlier morphologies while IL-1Rant gene expression was unchanged by conceptus development. The uterine lumenal content of IL-1beta increased during the process of trophoblastic elongation on Day 12. Uterine IL-1beta content declined on Day 15, reaching a nadir by Day 18 of pregnancy. IL-1beta gene expression in porcine conceptuses was temporally associated with an increase in endometrial IL-1RT1 and IL-1RAP gene expression in pregnant gilts. Endometrial IL-1beta and IL-1Rant gene expression were lowest during Days 10-15 of the estrous cycle and pregnancy. The temporal expression of IL-1beta during conceptus development and the initiation of conceptus-uterine communication suggests conceptus IL-1beta synthesis plays an important role in porcine conceptus elongation and the establishment of pregnancy in the pig.     

Effect of recombinant alpha interferons on fertility and interestrous interval in sheep

In Experiment 1, 12 unmated cyclic ewes received twice-daily intrauterine injections on Days 12 to 14 of one of the following treatments: 1) ovine conceptus secretory proteins (oCSP) containing 25 mug of ovine trophoblast protein-1 (oTP-1) as determined by RIA; 2) 25 or 50 mug recombinant human interferon alpha1 (rhlFN); or 3) 1500 ug of serum proteins (oSP) from a Day-16 pregnant ewe (estrus = Day 0) per uterine horn. Ewes receiving oCSP had longer interestrous intervals (27 +/- 2 days; P<0.05) than ewes receiving oSP (17 +/- 2 days). Ewes receiving either dose of rhlFN had an interestrous interval of 16 +/- 2 days which did not differ (P>0.10) from that of oSP-treated ewes. In Experiment 2, 59 normally cycling ewes, mated on Day 0, received twice-daily intramuscular injections of either 2 mg recombinant bovine interferon alpha1 (rblFN) or placebo on Days 12 to 15 post estrus. On Day 16, pregnancy was confirmed by flushing a morphologically normal conceptus from the uterus. Pregnancy rates for rblFN-treated (80%) and placebo-treated (62%) ewes were not different (P>0.10). Uterine flushings and conceptus-conditioned medium were assayed for oTP-1. Total oTP-1 in conceptus-conditioned culture medium was higher (P<0.02) when conceptuses were from placebo-treated (104 +/- 14 mug/conceptus) than from rblFN-treated (56 +/- 12 mug/conceptus) ewes; while total oTP-1 in uterine flushings was similar (P>0.10) for placebo-treated (132 +/- 15 mug/conceptus) and rblFN-treated (147 +/- 17 mug/conceptus) ewes. The interval from mating to subsequent estrus following conceptus removal was 31 +/- 1 and 28 +/- 1 days for pregnant ewes treated with rblFN and placebo, respectively. Interestrous intervals for nonpregnant ewes were longer (P<0.02) for rblFN-treated (27 +/- 3 days) than for placebo-treated (18 +/- 2 days) ewes.     

Swine leukocyte antigen-DQ expression and its regulation by interferon-gamma at the maternal-fetal interface in pigs

Successful pregnancy requires an appropriate intrauterine immune response to the conceptus, which is a semiallograft within the uterus. We reported that swine leukocyte antigen-DQA (SLA-DQA), a major histocompatibility complex (MHC) class II gene, is expressed in the uterine endometrium at the time of conceptus implantation in pigs. Because MHC molecules play critical roles in the immune system, SLA-DQ was hypothesized to be involved in immune regulation during pregnancy. Therefore, we examined expression of SLA-DQ in uterine endometrial tissues obtained during the estrous cycle and pregnancy. SLA-DQA and SLA-DQB mRNAs were detected as 1.3-kb and 1.2-kb bands, respectively. Real-time RT-PCR analysis indicated that SLA-DQA and SLA-DQB mRNA expression was affected by day and pregnancy status, with the highest expression on Day 15 of pregnancy. SLA-DQ was localized primarily to subepithelial stromal cells and endothelial cells of the uterus. Using endometrial explant cultures from Day 12 of the estrous cycle, we determined that expression of SLA-DQA and SLA-DQB mRNAs increased in response to interferon-gamma (IFNG), which is produced by pig conceptus trophectoderm between Days 14 and 18 of pregnancy. The abundance of SLA-DQ protein was less in endometria from gilts with conceptuses resulting from somatic cell nuclear transfer compared with endometria from gilts with conceptuses resulting from natural mating. These results support our hypothesis that SLA-DQ is expressed in response to IFNG from the conceptus, and likely regulates immune response at the maternal-fetal interface to support the maintenance of pregnancy in pigs.     

Incorporation of [14C] arachidonic acid into ovine conceptus and endometrial lipids.

The purpose of this study was to quantitate conceptus and endometrial incorporation of [14C]arachidonic acid (AA) into individual neutral and polar lipids. Endometrium and conceptuses from pregnant ewes and endometrium from nonbred ewes were collected 14 and 16 d after onset of estrus (d 0). Tissues were incubated for 8 h at 37 degrees C in medium containing 1 microCi of [14C]AA. Thin-layer chromatographic procedures were used to separate 12 lipids. Radioactivity was measured in each lipid, and the amount (ng) of [14C]AA incorporated into each lipid was calculated. Conceptuses and endometrium incorporated more [14C]AA into triacylglycerols than into any other lipid. Day and tissue type affected differentially (i.e., day X tissue interaction) the incorporation of [14C]AA into several lipids; d-14 conceptuses incorporated [14C]AA more actively than did any other day-tissue combination. Results indicate that triacylglycerols may be an important reservoir for conceptus and endometrial AA. The remarkable ability of d-14 conceptuses to incorporate [14C]AA into various lipids may be important for their accelerated elongation and active prostaglandin synthetic system.     

Conceptus development in large white and prolific Chinese Meishan pigs

Large White (LW) and Meishan (MS) gilts were killed on Days 8, 10, 11, 12, 14 and 30 of gestation. Mean diameters (mm) for MS and LW conceptuses, respectively, were: Day 8, 0.45 and 0.69; Day 10, 2.7 and 1.9; Day 11, 5.3 and 2.7, with the differences among days being affected by breed (P less than 0.01). Variation in diameter among conceptuses from LW gilts was greater (P less than 0.01) than that for MS gilts on Days 8-11, respectively: Day 8, 20 and 46%; Day 10, 29 and 38%; and Day 11, 22 and 44%. Conceptuses had elongated in 3 of 5 MS and 1 of 4 LW gilts on Day 11, 6 and 6 MS and 2 of 4 LW gilts on Day 12 and all gilts of both breeds on Day 14. These results indicate that conceptuses of MS gilts develop more rapidly and more uniformly between Days 8 and 14 of gestation. Overall, embryonic survival for Days 8-12 for gilts not having elongated conceptuses was 90.2% for MS and 73.2% for LW gilts (P less than 0.01). On Day 30 of gestation, embryonic survival was also higher (P less than 0.01) for MS (89%) than LW (55%) gilts. However, embryonic weight, crown-rump length, placental length, allantoic fluid volume, amniotic fluid volume, as well as total glucose, fructose and protein in allantoic fluid were not affected by breed. Placental weight was greater (P less than 0.01) for LW gilts. Uterine development at Day 30 of gestation, based on total length and weight of uterine horns, width of uterine horns, total endometrial surface area and total endometrial weight was greater (P less than 0.01) for LW gilts.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of pregnancy and exogenous ovarian steroids on endometrial prolactin receptor ontogeny and uterine secretory response in pigs

During gestation, pigs have constant circulating levels of prolactin (PRL), and lack decidual PRL and placental lactogens. Effects of PRL on uterine physiology in pigs may be due to changes in endometrial PRL receptors. In this study, effects of the conceptus and cyclic hormonal environment on endometrial PRL receptors were investigated. Endometrial PRL receptor numbers were similar between Days 8 and 15 for cyclic gilts. In contrast, endometrial PRL receptor numbers for pregnant gilts were similar between Days 8 and 11, then increased (p less than 0.05) on Day 12 and remained elevated between Days 14 and 30. This day-by-status interaction approached significance (p less than 0.06) and overall receptor numbers were higher (p less than 0.01) for pregnant than for cyclic gilts. Pig conceptuses secrete estrogen between Days 11 and 12; therefore, regulation of endometrial PRL receptors by acute administration of estradiol was investigated. Uterine flushings and endometrium were collected from one uterine horn of cyclic gilts on Day 11; then, at 1, 6, 12, and 24 h following a single injection of estradiol valerate (EV; 5 mg, into adipose tissue), uterine flushings and endometrium were collected from the second uterine horn. Endometrial PRL receptor numbers were higher (p less than 0.05) at both 1 h and 6 h after treatment with EV and then decreased (p less than 0.02) by 12 h to below pretreatment values. In uterine flushings, total recoverable protein (p less than 0.05), uteroferrin (p less than 0.01), leucine aminopeptidase (p less than 0.05), calcium (p less than 0.03), sodium (p less than 0.01), and potassium (p less than 0.05) increased between 12 and 24 h following EV treatment.(ABSTRACT TRUNCATED AT 250 WORDS)     

Regulation of prostacyclin synthase expression and prostacyclin content in the pig endometrium

Prostaglandins (PGs) are critical regulators of a number of reproductive processes, including embryo development and implantation. In the present study, prostacyclin (PGI₂) synthase (PGIS) mRNA and protein expression, as well as 6-keto PGF_1α (a PGI₂ metabolite) concentration, were investigated in the pig uterus. Endometrial tissue and uterine luminal flushings were obtained on Days 4 to 18 of the estrous cycle and pregnancy. Additionally, conceptuses were collected and examined for PGIS mRNA expression and 6-keto PGF_1α concentration. Regulation of PGI₂ synthesis in the porcine endometrium by steroids, conceptus products, and cytokines was studied in vitro and/or in vivo. Endometrial PGIS protein level increased on Days 12 and 16 in pregnant but not in cyclic gilts. Moreover, higher PGIS protein expression on Day 12 of pregnancy was accompanied by a greater content of 6-keto PGF_1α in the endometrium. The concentration of 6-keto PGF_1α in uterine luminal flushings increased substantially on Days 16 and 18 in pregnant gilts and was higher than in cyclic animals. Greater PGIS mRNA expression and PGI₂ metabolite concentration were detected in Day 12 and 14 conceptuses, respectively. Incubation of endometrial explants with conceptus-conditioned medium resulted in upregulation of PGIS protein expression and increased PGI₂ secretion. Moreover, PGIS mRNA and protein expression were upregulated in the endometrium collected from gravid uterine horn on Day 14 of pregnancy. In summary, PGIS is differentially expressed in the endometrium of cyclic and pregnant gilts resulting in higher PGI₂ synthesis in pregnant animals. Porcine conceptuses are important regulators of endometrial PGIS expression and PGI₂ release during the implantation period.     

Changes in ovine conceptus and endometrial function following asynchronous embryo transfer or administration of progesterone

The secretory protein profile from conceptuses collected from naturally mated ewes on Days 10, 12, 14, and 16 was characterized by two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) and fluorography. The presence of the anti-luteolysin ovine trophoblast protein-1 (oTP-1) in culture medium from Day 10 conceptuses was confirmed by fluorography, Western blotting, and radioimmunoassay (RIA). On each of the days studied, oTP-1 was the dominant secretory protein, and was secreted in increasing quantities as pregnancy progressed. In a second experiment, Day 6 embryos were transferred to either Day 6 (SR) or Day 4 (AR) recipients. Three mated ewes (P) received daily injections of 50 mg progesterone on Days 4-9. Controls consisted of 2 groups of pregnant ewes (D8 and D10). Conceptuses and ipsilateral endometrium were collected 4 days following transfer of conceptuses to SR and AR ewes, on Day 10 in P and D10 ewes, and on Day 8 in D8 ewes. Conceptus volume was estimated upon recovery from the uterus. Tissues were cultured with 35S-methionine, and the medium was analyzed for total and trichloroacetic acid-precipitable radiolabeled proteins. Levels of specific endometrial secretory proteins were determined after protein separation by 2D-PAGE and estimation of the radioactivity associated with discrete radiolabeled proteins on fluorographs. The concentration of oTP-1 in conceptus culture medium was estimated by RIA. Thirty endometrial proteins were investigated. All 30 proteins were present in endometrial cultures from SR, AR, D10, and P ewes, but 13 proteins were absent from D8 ewes. Levels of three proteins were higher in AR compared to D8 (p less than 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)     

Porcine conceptus proteins: antiviral activity and effect on 2',5'-oligoadenylate synthetase.

Interferon-like proteins synthesized by conceptuses of domestic ruminants inhibit luteolysis during early pregnancy. Although pig conceptuses secrete trophoblast interferons during the period of CL maintenance, estrogen is involved with maintenance of the CL. The principal purposes of this work were to confirm production of trophoblast interferons by porcine conceptuses and to compare the effect of trophoblast interferons on endometrium of pigs and cattle. When measured using Madin-Darby bovine kidney (MDBK) cells challenged with vesicular stomatitis virus, antiviral activity in uterine flushings from cyclic gilts was not detectable throughout the estrous cycle; however, in pregnant gilts, antiviral activity increased from undetectable amounts to 4-11 x 10(3) U on Days 14, 16, and 18. Porcine embryos in culture produced 1,100 U/embryo/ml/24 h. Porcine conceptus secretory proteins induced 2',5'-oligo(A) synthetase in MDBK cells and in endometrial explants of cows but had no measurable effect on 2',5'-oligo(A) synthetase activity of endometrial explants of pigs. Similarly, endometrial 2',5'-oligo(A) synthetase of pregnant pigs was unaffected in vivo during the period of maximal synthesis of conceptus secretory proteins. Porcine conceptus secretory proteins produced no detectable increase in serum antiviral activity or 2',5'-oligo(A) synthetase activity of blood mononuclear leukocytes in utero-ovarian venous blood. These results suggest that conceptus interferons of pigs play different roles in the establishment of pregnancy compared to their roles in ruminants.     

Effect of guinea-pig conceptus and its secretions on endometrial prostaglandin output

The co-culture of Day-15 guinea-pig conceptuses or Day-15 pregnant guinea-pig endometrium with Day-15 non-pregnant guinea-pig endometrium had no inhibitory effect on PGF2 alpha output from the non-pregnant endometrium. Unpurified proteins secreted by the Day-15 guinea-pig conceptuses, or these proteins purified by Blue Sepharose CL-6B and ion-exchange column chromatography also had no inhibitory effect on PGF2 alpha output from Day-15 non-pregnant guinea-pig endometrium cultured in vitro. However, following the further purification of guinea-pig conceptus secreted proteins on Sephadex G-75SF, the proteins present in fraction F3:4 inhibited PGF2 alpha output from the Day-15 non-pregnant guinea-pig endometrium during the first 6 h of culture. The major protein present in F3:4 had a molecular weight of 38.2 kDa on SDS-PAGE. Proteins present in F3:4 formed only a minor proportion of the total proteins secreted. Nevertheless, the anti-luteolytic factor secreted by the guinea-pig conceptus may be this 38.2 kDa protein, but further study is required.