Biomedical subjects

F W Bazer

Publications and source records attributed to F W Bazer.

At least 73 records · Page 4Linked to original sources

PubMed1998

Maternal dietary protein deficiency decreases nitric oxide synthase and ornithine decarboxylase activities in placenta and endometrium of pigs during early gestation.

Little is known about the mechanism responsible for retarded placental and fetal growth induced by maternal dietary protein malnutrition. On the basis of the recent finding that nitric oxide (NO) and polyamines (products of L-arginine) play an important role in embryonic and placental development, the present study was designed to determine whether protein deficiency decreases placental and endometrial activities of NO synthase (NOS) and ornithine decarboxylase (ODC) (the first and key regulatory enzyme in polyamine synthesis). Primiparous gilts selected genetically for low or high plasma total cholesterol concentrations (low line and high line, respectively) were mated and then fed 1.8 kg/d of isocaloric diets containing 13% or 0.5% crude protein. At d 40 or 60 of gestation, they were hysterectomized, and placenta and endometrium were obtained for incubations, NOS and ODC assays, and measurements of free amino acids and polyamines. Maternal dietary protein restriction decreased arginine and ornithine concentrations, constitutive and inducible NOS activities and NO production, as well as ODC activity and polyamine concentrations in placenta and endometrium of both lines of gilts. Placental NO synthase activity and NO generation were lower in high line gilts than in low line gilts. ODC activities and polyamine concentrations in placenta and endometrium were decreased at d 60 compared with d 40 of gestation. These changes in placental and endometrial synthesis of NO and polyamines during early gestation may be a mechanism responsible for reduced placental and fetal growth in protein-deficient gilts and for altered conceptus development in high line gilts.

Amino Acids↗

PubMed1998

Maternal dietary protein deficiency decreases amino acid concentrations in fetal plasma and allantoic fluid of pigs.

This study was conducted to test the hypothesis that maternal dietary protein deficiency decreases amino acid availability to the fetus, thereby contributing to retarded fetal growth. Primiparous gilts selected genetically for low or high plasma total cholesterol concentrations (low line and high line, respectively) were mated, and then fed 1.8 kg/d of isocaloric diets containing 13% or 0.5% crude protein. At d 40 or 60 of gestation, they were hysterectomized, and maternal and fetal blood samples as well as amniotic and allantoic fluids were obtained for analyses of amino acids, ammonia and urea. Dietary protein restriction decreased (P < 0.05) the following: 1) maternal plasma concentrations of urea at d 40 and 60 of gestation; 2) fetal plasma concentrations of alanine, arginine, branched-chain amino acids (BCAA), glutamine, glycine, lysine, ornithine, proline, taurine, threonine and urea at d 60 of gestation; 3) amniotic and allantoic fluid concentrations of urea at d 40 and 60 of gestation; and 4) allantoic fluid concentrations of alanine, arginine, BCAA, citrulline, cystine, glycine, histidine, methionine, proline, serine, taurine, threonine and tyrosine at d 40 of gestation, in gilts of both genetic lines. At d 60 of gestation, protein deficiency decreased (P < 0.05) allantoic fluid concentrations of arginine, cystine, glycine, taurine and tyrosine in low line gilts and of cystine, glutamine, ornithine, serine, taurine and tyrosine in high line gilts. Low line and high line gilts also differed remarkably in allantoic fluid concentrations of arginine, glutamine, ornithine and ammonia at d 40 and 60 of gestation. Our results suggest the following: 1) protein-deficient gilts maintain maternal plasma concentrations of amino acids by mobilizing maternal protein stores and decreasing oxidation of amino acids during the first half of gestation; 2) protein deficiency may impair placental transport of amino acids from the maternal to the fetal blood; and 3) low line and high line gilts differ in fetal amino acid metabolism. Decreases in concentrations of the essential and nonessential amino acids in the fetus may be a mechanism whereby maternal dietary protein restriction results in fetal growth retardation.

Allantois↗

PubMed1998

Expression of interferon regulatory factors one and two in the ovine endometrium: effects of pregnancy and ovine interferon tau.

Availàble evidence suggests that interferon tau (IFNtau), the signal for pregnancy recognition in ruminants, suppresses transcription of the estrogen receptor (ER) gene in the endometrial lumenal epithelium (LE) and superficial glandular epithelium (sGE) to prevent oxytocin receptor (OTR) expression and pulsatile release of luteolytic prostaglandin F2alpha. The IFN regulatory factors one (IRF-1) and two (IRF-2) are transcription factors induced by type I IFNs that activate and silence gene expression, respectively. The objectives of these studies were to determine effects of pregnancy and IFNtau on expression of immunoreactive IRF-1 and IRF-2 proteins in the ovine endometrium. In study one, IRF-1 and IRF-2 were not detected in the LE or sGE of cyclic ewes. In pregnant ewes, IRF-1 expression was detected transiently in the LE and sGE only on Days 11 and 13, and IRF-2 was detected in these same epithelia on Days 13, 15, 17, and 20. In study two, 36 ewes were fitted with uterine catheters on Day 5 of the estrous cycle, and one uterine horn was double-ligated at the base. Uterine horns of each ewe received twice-daily injections of either recombinant ovine IFNtau or control proteins beginning on Day 11 until hysterectomy at 1, 3, 6, 12, 24, 48, 72, 96, or 120 h after initial injection. The IRF-1 was detected transiently in the endometrial LE and sGE only at 12 and 24 h in the uterine horn receiving IFNtau but not in those tissues receiving control proteins. The IRF-2 was expressed in the LE and sGE at 24 h and thereafter in the IFNtau-treated, but not control uterine horns. In control uterine horns, ER and OTR were first detected in the LE at 48 h and 72 h, respectively, and remained abundant thereafter. In horns receiving IFNtau, ER and OTR expression was not detected in the endometrial LE and sGE. Results suggest that IFNtau acts directly on the LE and sGE during pregnancy to sequentially induce IRF-1 and then IRF-2 gene expression, which is correlated temporally with an absence of ER and OTR. The ovine ER gene may contain an IFNtau-responsive element(s) that binds negative-acting, IFNtau-inducible transcription factors, such as IRF-2, which silences transcription of the ER gene in the endometrial epithelium during maternal recognition of pregnancy.

Animals↗

PubMed1998

Generation of transgenic porcine chimeras using primordial germ cell-derived colonies.

In mice, two pluripotent cell lines, embryonic stem (ES) cells and embryonic germ (EG) cells, have been identified. We present here results indicating that porcine EG cell lines can be isolated, genetically transformed, and utilized to make transgenic chimeras. Briefly, primordial germ cells (PGCs) were isolated from Day 25-27 fetuses and plated on STO feeder cells in Dulbecco's modified Eagle's medium:Ham's F-10 medium supplemented with 0.01 mM nonessential amino acids, 2 mM glutamine, 15% fetal bovine serum, 0.1 mM 2-mercaptoethanol, 40 ng/ml human stem cell factor, 20 ng/ml human basic fibroblast growth factor, and 20 ng/ml human leukemia inhibitory factor. For genetic transformation, cells were electroporated with a construct containing the green fluorescent protein under control of the cytomegalovirus promoter. After electroporation, cells were plated and later examined under fluorescein isothiocyanate excitation. Fluorescent colonies were selected for chimera generation. Blastocysts collected from gilts on Day 5 were injected with 10-15 transgenic PGC-derived cells and transferred into recipient gilts. Gilts were hysterectomized on Day 25, and fetal tissues were analyzed by Southern blotting. Three chimeras out of 20 fetuses analyzed were transgenic. Additionally, when one recipient gilt was allowed to go to term, one piglet with transgenic contribution was identified.

Animals↗

PubMed1998

Effects of the estrous cycle and early pregnancy on uterine expression of Mx protein in sheep (Ovis aries).

Conceptuses of ruminant ungulates produce large amounts of a type I interferon, interferon-tau (IFNtau), which is the signal for maternal recognition of pregnancy. Induction of cellular Mx proteins is an important component of the response to type I interferon in the immune system, but Mx regulation and function have not been studied in the uterus. This study examined temporal and spatial alterations in ovine uterine Mx expression during the cycle and early pregnancy using immunohistochemistry, in situ hybridization, and Northern and slot-blot analysis. Sheep uterine endometrium expressed a single approximately 2.5-kilobase Mx mRNA transcript that was detectable at all stages of the estrous cycle and early pregnancy examined. In cyclic ewes, mRNA abundance in endometrium increased from Day 1 to peak levels at Day 13 and then declined to Day 15. In pregnant ewes, steady-state levels of Mx mRNA were first detected above the level in cyclic ewes at Day 13 postmating, were greater than 10-fold higher at Day 15, and remained elevated at Day 19. Expression of Mx mRNA in the myometrium did not change during the estrous cycle but increased approximately 23-fold between Days 11 and 15 of pregnancy. Immunohistochemical and in situ hybridization analysis revealed a similar temporal pattern of Mx expression. In cyclic ewes, Mx protein and mRNA were initially localized to the luminal epithelium at Days 1 and 3, increased from Days 5 to 13, especially in the shallow uterine glands, and then declined at Day 15. Pregnancy resulted in up-regulation of Mx expression in the luminal and glandular epithelium, stroma, and myometrium. Punctate Mx immunostaining and Mx mRNA concentrations were greatest when progesterone production was maximal during the estrous cycle and were strongly up-regulated by the conceptus across the entire uterine wall. It is suggested that a cascade of induction of Mx gene expression proceeds from the luminal epithelium to the outer longitudinal myometrium and that transcriptional activation of the promoter may involve both soluble cytokines (i.e., IFNtau) and steroid hormones (i.e., progesterone).

Animals↗

PubMed1998

Beta transforming growth factors (TGFss) at the porcine conceptus-maternal interface. Part I: expression of TGFbeta1, TGFbeta2, and TGFbeta3 messenger ribonucleic acids.

Spatial and temporal mRNA expression of beta transforming growth factors (TGFbeta1, TGFbeta2, and TGFbeta3) in porcine uterus and conceptuses was determined during the peri-implantation period (Days 10-14 of gestation). Northern blotting identified a major 3. 5-kilobase (kb) and a minor 2.5-kb transcript for TGFbeta1 mRNA. TGFbeta2 transcripts were 6.2 kb, 5.4 kb, and 2.7 kb, and a single 3. 5-kb transcript was detected for TGFbeta3. With semiquantitative in situ hybridization analysis, progressive increases were detected in TGFbetas 1, 2, and 3 mRNA expression in uterine luminal epithelium (ULE), uterine glands (UGs), and underlying stroma (stroma spongiosum, SS) from Days 10 through 14 of gestation (p < 0.05). In myometrium, TGFbeta mRNA expression did not differ between Days 10 through 14 of gestation. In porcine conceptuses, TGFbetas 1, 2, and 3 mRNA expression was detected in trophectoderm, endoderm, embryonic ectoderm, and mesoderm. For the three TGFbeta isoforms examined, mRNA expression increased 2- to 4-fold in trophectoderm and endoderm between Days 10 and 14 of gestation. TGFbeta1 mRNA levels increase significantly in embryonic ectoderm, but not mesoderm, between Days 12 and 14 of gestation; during that same time, TGFbeta2 mRNA levels increased, but no change was detected in TGFbeta3 mRNA levels, in embryonic ectoderm and mesoderm. Progressive increases in TGFbeta mRNA expression in conceptus trophectoderm, endoderm, ULE, UGs, and SS suggest important roles for these growth factors in porcine conceptus development during the peri-implantation period.

Animals↗

PubMed1998

Beta transforming growth factors (TGFbeta) at the porcine conceptus-maternal interface. Part II: uterine TGFbeta bioactivity and expression of immunoreactive TGFbetas (TGFbeta1, TGFbeta2, and TGFbeta3) and their receptors (type I and type II).

Porcine uterine tissues were collected from Days 10 to 14 of gestation (peri-implantation period) or corresponding days of the estrous cycle. Results indicated a marked increase in beta transforming growth factors (TGFbeta1, TGFbeta2, and TGFbeta3) and TGFbeta receptor (type I and type II) immunostaining in uterine luminal epithelium (ULE) between Days 10 and 14 of gestation, but there was no increase in ULE immunostaining on the corresponding days of the estrous cycle. Uterine glands and stroma were intensely immunopositive in pregnant gilts for TGFbeta isoforms and their receptors, but immunostaining was weak to undetectable in cycling gilts. No differences were detected in myometrium, in which immunostaining was moderate in both cycling and pregnant gilts. Additionally, TGFbeta2 and TGFbeta receptor (type I and type II) immunostaining was detected in uterine monocyte/macrophage-like cells. Western blotting detected the presence of all three TGFbeta isoforms in uterine luminal flushings. The CCL64 cell TGFbeta bioassay detected bioactive TGFbetas++ in uterine luminal flushings on Days 12, 13, an 14 of gestation. These results strongly indicate that uterine expression of TGFbetas and their receptors is pregnancy specific and that bioactive TGFbetas are present at the conceptus-maternal interface in the peri-implantation period in pigs. Thus TGFbetas are likely to be involved in autocrine-paracrine interactions between the maternal uterus and the conceptus.

Animals↗

PubMed1998

Mechanisms regulating prostaglandin F2 alpha secretion from the bovine endometrium.

The mechanism that regulates luteolytic PGF2 alpha secretion as stimulated by oxytocin is thought to involve induction of the inositol (1,4,5)-trisphosphate-diacylglycerol second messenger system, which mobilizes intracellular calcium and activates protein kinase C. In Experiment 1, endometrial explants taken from heifers on d 18.5 to 19.5 postestrus had increased PGF2 alpha secretion after treatment with 1 microM calcium ionophore A23187 to increase intracellular calcium, 100 nM phorbol 12-myristate 13-acetate to activate protein kinase C, and 100 nM oxytocin. The stimulatory effects of oxytocin and calcium ionophore A23187 plus phorbol 12-myristate 13-acetate did not differ from each other. In Experiment 2, endometrial explants taken from cows on d 18.5 to 19.5 postestrus had increased PGF2 alpha secretion after treatment with 0.2 and 2 microM thapsigargin to mobilize intracellular calcium that was sensitive to inositol (1,4,5)-trisphosphate. Secretion of PGF2 alpha was also increased by 100 nM oxytocin and was influenced by the interaction of thapsigargin and oxytocin such that 100 nM oxytocin did not further increase the secretion of PGF2 alpha in the presence of 2 microM thapsigargin. In Experiment 3, 100 nM oxytocin stimulated greater production of inositol trisphosphate and total inositol phosphates in the endometrium of cyclic cows than in the endometrium of pregnant cows on d 16.5 to 17.0 postestrus, although luteolysis was not yet initiated in the cyclic cows. These results are consistent with the hypothesis that the activation of the inositol (1,4,5)-trisphosphate-diacylglycerol second messenger system by oxytocin is involved in the stimulation of PGF2 alpha secretion from the endometrium during late diestrus in cows.

Animals↗

PubMed1997

Potent anti-feline immunodeficiency virus and anti-human immunodeficiency virus effect of IFN-tau.

Ovine IFN-tau is a newly described protein related to IFN-alpha that is responsible for maternal recognition of pregnancy in sheep. It has been shown to exhibit potent antiviral and antiproliferative activity. To determine its antiviral activity against feline immunodeficiency virus (FIV) and HIV, the activity of the RNA-dependent DNA polymerase, reverse transcriptase, was assayed in FIV- and HIV-infected feline and human PBL treated with IFN-tau. Significant dose-dependent inhibition of reverse transcriptase activity by IFN-tau was detected by day 6 of culture and was maintained through the peak of virus replication. In addition, production of the FIV core protein, p25, was blocked by IFN-tau. Both the amino- and carboxyl-terminal regions of IFN-tau, as identified by synthetic peptides, appear to be involved in its antiretroviral activity. Comparison of the anti-HIV activities of IFN-tau and recombinant human IFN-alpha2 (rHuIFN-alpha2) indicated that while rHuIFN-alpha2 was toxic to cells at 10,000 U/ml, IFN-tau antiretroviral activity was not associated with a decrease in either cell viability or immunologic reactivity. Thus, IFN-tau displayed potent anti-FIV and anti-HIV activity without the cytotoxicity associated with high concentrations of rHuIFN-alpha2.

Animals↗

PubMed1997

Mucosal morphogenesis and cytodifferentiation in fetal porcine small intestine.

BACKGROUND: Development of the small intestine is essential for proper nutrition of the fetus and the neonate. This investigation examines the morphogenesis and cytodifferentiation of developing fetal porcine small intestinal mucosa. METHODS: Fetuses were collected from gilts after hysterectomy. Small intestinal segments were removed and processed for light and electron microscopy. RESULTS: Fetal porcine small intestine developed from a simple tube of stratified epithelium to a tube containing villus and intervillus regions of simple columnar epithelium. This development occurred in a proximal to distal direction. By Day 40 of gestation, cytodifferentiation was evident with the presence of goblet cells and enteroendocrine cells in the duodenum. As development progressed, microvilli lengthened and components of the apical endocytic complex (AEC) were observed. By Day 110 of gestation, tubular and vesicular components of the AEC were confined to the jejunum, whereas large lysosomal vacuoles were observed in the distal jejunum. Duodenal epithelium at Day 110 was similar to postnatal epithelium. CONCLUSIONS: The pattern of fetal porcine small intestinal development is similar to that reported for fetal human small intestine. Villus development and cytodifferentiation occur at similar relative times in gestation when compared to the human. These observations support the use of the fetal pig as a model for investigations of human small intestinal development.

Animals↗

PubMed1997

Myelosuppression in the pig (Sus scrofa): uteroferrin reduces the myelosuppressive effects of 5-fluorouracil in young pigs.

The present study investigated the ability of uteroferrin to modulate the myelosuppressive effects of 5-fluorouracil (5-FU) in young pigs (Sus scrofa). Pigs (28-35 days of age; n = 6 per treatment) were infused with equal amounts of 5-FU on days 0 and 1 of the experimental period (37.5 mg/kg cumulative dose). Uteroferrin (100 micrograms/kg in 0.9% NaCl) or control (equivalent volume of 0.9% NaCl) was administered to pigs as intramuscular injections twice daily (08:00 and 20:00 hr) on days 1 through 21. Peripheral blood cell number, composition and progenitor cells were determined over 28 days. Treatment of pigs with 5-FU resulted in a rapid dose-dependent (P < 0.05) leukocytopenia. Concurrent treatment of pigs with uteroferrin reduced (P < 0.05) the rate of 5-FU-induced leukocytopenia (44 vs 77 +/- 7% decline from baseline on day 3) and enhanced (P < 0.05), the recovery from 5-FU on days 10 and 12 postinfusion. The positive effect of uteroferrin on leukocytes resulted primarily from a protection and/or enhanced recovery of neutrophils and monocytes. In addition, uteroferrin attenuated (P < 0.05) the suppression of red blood cell numbers after 5-FU administration (6.9 vs 6.1 +/- 0.2 x 10(6) cells/microliter on day 3), an affect reflected in increased hematocrit and hemoglobin concentrations. The effects of uteroferrin appeared to result from enhancement of the proliferation and/or differentiation of primitive pluripotent stem cells resistant to 5-FU, as concurrent treatment of pigs with uteroferrin resulted in a protection and/or enhanced recovery (P < 0.05) of CFU-GEMM, CFU-GM and BFU-E progenitor cells in the peripheral blood. These results are the first to demonstrate that uteroferrin can reduce the myelosuppressive effects of 5-FU in the pig and suggest that uteroferrin has hematopoietic growth factor activity in vivo.

Acid Phosphatase↗

PubMed1997

Uteroferrin and recombinant bovine GM-CSF modulate the myelosuppressive effects of 5-fluorouracil in young female pigs (Sus scrofa).

The present study investigated the ability of uteroferrin and recombinant bovine granulocyte monocyte/macrophage-colony stimulating factor (rbGM-CSF) to modulate the myelosuppressive effects of 5-fluorouracil (5-FU) in young female pigs (Sus scrofa). Pigs (N = 3/treatment) were infused with 5-FU (32.5 mg/kg) on days 0 and 1 of the experimental period. Uteroferrin (100 micrograms/kg in 0.9% NaCl), rbGM-CSF (10 micrograms/kg in 0.9% NaCl), uteroferrin + rbGM-CSF (as above) or control (0.9% NaCl) were administered as intramuscular injections twice daily (0800 and 2000 hr). Peripheral blood cell number, composition, and progenitor cells were determined over 28 days. Treatment of pigs with 5-FU resulted in a rapid leukocytopenia and thrombocytopenia (nadirs on days 5 and 7, respectively) and a modest decrease (P < 0.05) in red blood cell (RBC) number (nadir on day 14). Although nor affecting RBC and thrombocytes, treatment of pigs with uteroferrin had an initial protective effect (P < 0.05) on the 5-FU-induced leukocytopenia (63 and 64 vs 48 and 39 +/- 6% of baseline on days 3 and 5, respectively). In contrast, rbGM-CSF enhanced (P < 0.05) the rate of the leukocytopenia and had only minor effects on thrombocyte numbers relative to controls. These effects appeared to be additive, as pigs treated with uteroferrin + rbGM-CSF had a reduced rate of leukocytopenia compared to pigs treated with rbGM-CSF alone. Uteroferrin + rbGM-CSF also attenuated (P < 0.05) the suppression and enhanced (P < 0.05) recovery of RBC and thrombocyte numbers following 5-FU treatment. In control pigs, a modest rebound leukocytosis (122 +/- 6% of baseline) and thrombocytosis (141 +/- 9% of baseline) was evident. Uteroferrin enhanced (P < 0.05) the rebound leukocytosis (135 +/- 6% of baseline), but attenuated (P < 0.05) the thrombocytosis. In contrast, rbGM-CSF enhanced (P < 0.05) the duration of the leukocytosis during the recovery phase, an effect augmented by the combination of uteroferrin + rbGM-CSF. In addition, treatment with uteroferrin + rbGM-CSF resulted in a sustained thrombocytosis (days 12 to 21). As indicated by changes in CFU-GM, BFU-E, and CFU-GEMM progenitor cells in peripheral blood, the effects of uteroferrin and rbGM-CSF appeared to reflect their ability to enhance the proliferation and/or differentiation of both similar and distinct hematopoietic progenitor cells.

Acid Phosphatase↗

PubMed1997

The effect of uteroferrin and recombinant GM-CSF on hematopoietic parameters in normal female pigs (Sus scrofa).

The present study investigated the effect of uteroferrin and recombinant bovine granulocyte-monocyte/macrophage colony stimulating factor (rbGM-CSF) on hematopoiesis in young female pigs. Uteroferrin (100 micrograms/kg in 0.9% NaCl), rbGM-CSF (10 micrograms/kg in 0.9% NaCl), uteroferrin + rbGM-CSF (as above), or control (0.9% NaCl) were administered as intramuscular injections twice daily (0800 and 2000 hr). Peripheral blood cell number, composition, and progenitor cells were determined over 28 days. Uteroferrin had minimal effects on white blood cell (WBC) number, while rbGM-CSF caused both a rapid (days 2-7; maximum 122 +/- 8% of baseline) and late (days 16-28; maximum 133 +/- 8% of baseline) increase in WBC. Combination treatment with uteroferrin + rbGM-CSF abolished the initial increase in WBC number,but resulted in a prolonged increase in WBC number (days 14-28) relative to control. The rbGM-CSF-induced increase in WBC number resulted from rapid increases (P < 0.05) in monocytes and neutrophils. The addition of uteroferrin + rbGM-CSF enhanced (P < 0.05) the initial increase in the monocyte population and augmented the neutrophilia. In addition, uteroferrin + rbGM-CSF resulted in a dramatic eosinophilia (days 2-28), which was not detected in either the uteroferrin or rbGM-CSF treatments. Although not substantially affected by uteroferrin alone, rbGM-CSF caused an increase (P < 0.05) in thrombocyte numbers from days 1 through 9 (maximum 133 +/- 11% of baseline), an effect augmented by cotreatment with uteroferrin. The ability of these cytokines to modulate blood cell number and composition appeared to result from their effects on hematopoietic progenitor cells. Treatment of pigs with uteroferrin increased (P < 0.05) CFU-GEMM, CFU-GM, and BFU-E progenitor cells in peripheral blood, while rbGM-CSF caused increases (P < 0.05) relative to control in CFU-GM and CFU-GEMM. These effects were additive, as uteroferrin + GM-CSF augmented the increases in CFU-GM, BFU-E, and CFU-GEMM. Collectively, these results indicate that uteroferrin and rbGM-CSF can modulate hematopoiesis in young pigs. These effects were both additive and, in the case of neutrophils and eosinophils, synergistic. Hence, the mechanism(s) by which uteroferrin and rbGM-CSF modulate hematopoiesis appear to be different.

Acid Phosphatase↗

PubMed1997

Lack of effect of granulocyte-macrophage colony-stimulating factor on secretion of interferon-tau, other proteins, and prostaglandin E2 by the bovine and ovine conceptus.

Three experiments tested the effects of recombinant bovine granulocyte-macrophage colony-stimulating factor (rbGM-CSF) on the preimplantation bovine and ovine conceptus. There was no effect of rbGM-CSF on the secretion of total radiolabeled protein in conditioned medium, immunoreactive interferon-tau (IFN tau), antiviral activity, or prostaglandin E2 from Day 16-18 bovine conceptuses cultured for 24 hr with, [3H]leucine and +/- 10 ng/ml rbGM-CSF. Similarly, there was no effect of 1 ng/ml rbGM-CSF on the secretion of total radiolabeled protein. IFN tau, or antiviral activity from Day 17 ovine conceptuses. There was also no beneficial effect of 1 or 10 ng/ml rbGM-CSF on the presence of immunoreactive IFN tau in conditioned medium from in vitro-produced bovine blastocysts at Day 7-8 after fertilization. Results indicate that IFN tau secretion from bovine and ovine conceptuses are unresponsive to rbGM-CSF at the concentrations tested.

Animals↗

PubMed1997

Recombinant ovine and bovine interferons tau regulate prostaglandin production and oxytocin response in cultured bovine endometrial cells.

At the time of recognition of pregnancy, the bovine conceptus (embryo and associated membranes) must produce a signal that will prevent luteolysis otherwise induced by pulsatile release of prostaglandin (PG) F2alpha from the uterus in response to oxytocin (OT). In ruminants, trophoblastic interferon tau (IFN-tau) released by the conceptus appears to be the most likely candidate to trigger the establishment of pregnancy. We have compared the effect of recombinant (r) ovine (o) and bovine (b) IFN-tau on PG production, using a fully characterized model of cultured endometrial cells. In uterine epithelial cells, the production of PGF2alpha was stimulated 7.1-fold (p < 0.0001) and that of PGE2 89.0-fold (p < 0.0001) by rbIFN-tau, and 3.6-fold and 29-fold, respectively, by roIFN-tau. The stimulation resulted in a net increase in the PGE2:PGF2alpha ratio of 7.7 with rbIFN-tau and 5.1 with roIFN-tau at the optimal concentration of 1 microg/ml (p < 0.0001). By contrast, addition of OT (100 nM) alone resulted in a decrease of the PGE2:PGF2alpha ratio. The level of stimulation of PGE2 by IFN-tau was reduced in the presence of OT, showing that there was some interaction between OT and IFN-tau at the cellular level in the regulation of PG production. In uterine stromal cells, roIFN-tau and rbIFN-tau stimulated PGE2 and PGF2alpha production 12-fold (p < 0.0001). The ratio of PGE2:PGF2alpha was not affected in a dose-dependent manner, but was increased (p < 0.001) at a single dose of rbIFN-tau (0.001 microg/ml) and roIFN-tau (0.01 microg/ml). The results indicate that 1) bovine endometrial cells are more responsive to rbIFN-tau than to roIFN-tau, 2) rIFN-tau regulates PGs by stimulating PGE2 preferentially, and 3) rIFN-tau transforms the response to OT from stimulation of PGF2alpha to stimulation of PGE2.

Animals↗

PubMed1997

Spatial and temporal analyses of integrin and Muc-1 expression in porcine uterine epithelium and trophectoderm in vitro.

The expression of the glycoproteins Muc-1 and integrin subunits (alpha1, alpha3, alpha4, alpha5, alpha(v), beta1, and beta3), all of which may be involved in the control of uterine receptivity, was examined in cultured porcine uterine epithelial (pUE) cells. Integrin subunit expression was also determined in trophoblastic vesicles generated from Day 12 and Day 15 pig conceptuses. Immunocytochemistry was performed on pUE cells cultured on several substrates (glass, serum, fibronectin, or Matrigel) in DME/F12 medium with 5% charcoal/dextran-stripped fetal bovine serum either containing no steroids or supplemented with estrogen, progesterone, or estrogen plus progesterone, or in medium with 5% complete fetal bovine serum. In addition, pUE cells were grown on Matrigel-coated Millicell filter inserts to facilitate development of structurally and functionally distinct apical and basal domains (i.e., polarized pUE cells) and were maintained in the same medium with supplements. The major findings in this investigation are that 1) Muc-1 and alpha1, alpha3, alpha4, alpha5, alpha(v), beta1, and beta3 integrin subunits are expressed on pUE cells in vitro; 2) Muc-1 and several integrin subunits are modulated by steroid hormones if culture conditions are provided that permit development of cell polarity; 3) the expression and steroid-induced modulation of Muc-1 and several integrin subunits in polarized pUE cells are similar to those detected in vivo; and 4) the expression of alpha1, alpha3, alpha4, alpha5, alpha(v), beta1, and beta3 integrin subunits by trophoblastic vesicles is identical to their expression on trophectoderm of intact conceptuses in vivo. These results suggest that the properties of polarized pUE cells and trophoblastic vesicles are comparable to those of their counterparts in vivo and are, therefore, useful as in vitro model systems to study conceptus-endometrial interactions during the periimplantation period and the control of uterine receptivity to implantation.

Animals↗

PubMed1997

Immunolocalization of acidic and basic fibroblast growth factors in porcine uterine and conceptus tissues.

Immunohistochemical techniques were used to localize acidic and basic fibroblast growth factor (FGF) polypeptides in porcine uterine and conceptus tissues collected on Days 10 through 14 of gestation, which is the peri-implantation period, and in uterine tissues collected on the same days of the estrous cycle. Our results demonstrate differential expression of acidic and basic FGF (aFGF, bFGF) in porcine uterine and conceptus tissues. Localization of these peptides in the uterus of cycling and pregnant pigs was different from that reported for other species, suggesting species-specific roles for FGFs in early pregnancy. Increases in both cytoplasmic and nuclear bFGF immunostaining were detected in uterine luminal and glandular epithelial cells from Days 10 to 14 of gestation but not in the uterine epithelium of cycling pigs. Acidic FGF immunostaining was not detected in luminal or glandular epithelium of either cycling or pregnant uterine tissue; however, differential stromal staining was observed. Uterine tissues collected from pigs on Days 10-14 of the estrous cycle had diffuse aFGF immunostaining throughout the stroma. During early pregnancy, however, intense aFGF immunostaining was concentrated around the glandular epithelial and below the luminal epithelial cells as gestation progressed. Basic FGF, but not aFGF, was detected in porcine conceptuses collected during the peri-implantation period (Days 10-14 of gestation). Although both acidic and basic FGF belong to the same family of proteins, results of the present study indicate that they likely play different roles in uterine function and conceptus development in pigs.

Animals↗

PubMed1997

Effects of exogenous recombinant ovine interferon tau on circulating concentrations of progesterone, cortisol, luteinizing hormone, and antiviral activity; interestrous interval; rectal temperature; and uterine response to oxytocin in cyclic ewes.

Interferon tau (IFN tau) is the conceptus-produced antiluteolytic signal in ruminants. Three experiments examined the effects of s.c. administration of recombinant ovine (ro)IFN tau on interestrous interval (IEI), oxytocin (OT)-induced uterine prostaglandin F2alpha metabolite (PGFM) production, rectal temperature (RT), respiration rate (RR), and plasma concentrations of progesterone, cortisol, LH, and antiviral activity (AVA) in plasma and uterine flushings. In experiment I, 20 ewes were treated s.c. with either 0, 1, 2, or 4 mg/day roIFN tau (0.7 x 10(8) U/mg; 5 ewes/dosage) from Days 11 to 15 of the estrous cycle (estrus = Day 0) and were challenged with OT (30 IU) on Day 15. Jugular blood samples were collected at -10, 0, 10, 20, 30, 40, 50, and 60 min relative to the OT challenge and assayed for PGFM. Recombinant oIFN tau increased IEI (16.7, 18.7, and 22.6 +/- 0.6 days for 0, 2, and 4 mg roIFN tau, respectively, p < 0.01). Recombinant oIFN tau did not affect peak PGFM response to OT (2309 +/- 172 pg/ml; p > 0.1). However, the 4 mg/day dosage delayed the time to peak PGFM (32.4 vs. 47.5 +/- 3.4 min; p < 0.01, 0 vs. 4 mg) and resulted in approximately 200% higher concentrations of PGFM at 60 min post-OT (0 vs. 4 mg/day, p < 0.07). Experiment II was similar to experiment I, except that only the 0- and 4-mg/day dosages of roIFN tau were administered. Ewes were hysterectomized on Day 16, and assay of uterine flushes detected no AVA from ewes treated with either 0 or 4 mg/day roIFN tau. In experiment III, 20 ewes were treated s.c. with either 0, 2, 4, or 6 mg roIFN tau on Day 12. Blood samples, RT, and RR were obtained at frequent intervals for 24 h, and plasma was assayed for progesterone, cortisol, LH, and AVA. Plasma AVA, which increased in a dose-dependent manner, was detectable within 60 min and remained elevated at 24 h compared to control values. RT (elevated 0.5-1.0 degrees C), RR, and cortisol increased in response to all dosages of roIFN tau, with peak values occurring 150-180 min postinjection. For all dosages of roIFN tau, plasma progesterone declined from 120 to 360 min posttreatment and then returned to pretreatment values by 24 h (p < 0.01) as compared to controls. Overall, exogenous roIFN tau altered uterine PGFM response to OT from a pulse to a gradual and sustained elevation and extended IEI with only a transient decline in progesterone and mild hyperthermia, effects that are not expected to compromise pregnancy.

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