Sheep Placenta Research Articles

MORPHOLOGICAL STUDY ON THE PERICYTES OF GOAT'S PLACENTOMAL CAPILLARIES

Pcricytcs arc connective tissue cells associated abluminally with capillary and postcapillary venulcs, They share a common basal lamina with endothelial cells (Majno, 1965: Schor et al., 1995). Such rnicrovcssels arc involved with the exchangeof material and extracellular space (Nehls and Drenckhahn, 1993). Perleytes were first described in the nictitating membrane of the frog by Rouget (1873). and have been seen surrounding capillaries in a wide variety of tissues(Zimmerman, 1923: Majno, 1965: Lunam and Roger, 1981: Sims, 1986: Hirschi and D'Amore, 1996; Sundberg et al., 1996; Wisse et al., 1996). The most probable function for pcricytcs is the regulation of capillary flow by their contraction (Epling, 1966; Rhodin, 1968: Wieble, 1974: Tilton et al., 1979). A possible functional role of the pericytes is the control of material exchange through the vessel wall (lmayama and Urab, 1984) and its participation in angiogenesis has been proposed (Crocker et at, 1970; Schor et at, 1995). A further role that has been attributed to pericytcs is to stabilize the capillary wall (Cohen et al., 1980; Frank et al., 1990: Jeon et at., 1996). In rat uterus. the increase in pericytes coincides with the time of implantation (Lunam and Roger, 1983). Steven and Samuel (1979) have suggested that there arc few pericytes associated with the maternal capillaries in sheep placenta to perform an important regulatory function on the transport of material from the mother to the fetus. However, Yasear et HI. (1987) found a considerably largenumber of pcricytcs in association with the maternal microvessels in sheep placenta throughout pregnancy. In the gOHt'S placenta. the materno-Ictal interface is characterized by the presence of syncytial tissue covering the maternal side of placentomes. The plcentomes arc ruminant-specific fetomatcrnal contact zones characterized by the presence ofmicrovillar junction between syncytium and fetal trophoblastic layer (Steven, 1975; Wooding and Flint, 1994). Due to the lack of information on the distribution of pericytes in the goat's placenta the present study was initiated to evaluatethe distribution of the pericytes surrounding the materna! capillaries which are located close to the syncytium, i.e. those most closely involved in the materno-fetal exchanges throughout pregnancy.

Ontogeny and regulation of ovine placental prostaglandin E2 synthase.

Recent evidence suggests that ovine placental output of prostaglandin (PG) E2 rises through late gestation partly because of a direct effect of cortisol on PGH2 synthase 2 (PGHS-2) expression and activity within trophoblast tissue. Synthesis of PGE2 is also dependent, however, on PGE2 synthase (PGES), which converts PGH2 to PGE2. We hypothesized that PGES is expressed in the ovine placenta, and that, similar to PGHS-2, expression increases through gestation and is regulated positively by cortisol. Placental tissues from pregnant ewes in mid and late gestation, at term, and during early and active labor were analyzed to determine the gestational profile of PGES. The regulation of PGES expression was assessed in placental tissues from pregnant ewes in which intrafetal cortisol infusion was administered in late gestation, in the presence or absence of an aromatase inhibitor, to block the cortisol-stimulated rise in estradiol. Expression of PGES was analyzed by in situ hybridization, Western blot analysis, and immunohistochemistry. In the placentome, PGES localized to fetal trophoblast cells and endothelial cells in maternal blood vessels, consistent with its contribution to the rise in placental PGE2 output toward the onset of labor and with a role of PGE2 in the local regulation of uteroplacental blood flow, respectively. Expression of PGES mRNA and protein increased with gestation. However, there was no significant further change with labor or during cortisol infusion in the presence or absence of a rise in fetal plasma estradiol, in contrast to reported changes in PGHS-2. These results suggest that PGES is not coregulated with PGHS-2 in the sheep placenta at term. The progressive increase in PGES, however, likely contributes to the rise in circulating PGE2 in the fetus in late pregnancy.

Clearance and disposition of indometacin in chronically instrumented fetal lambs following a 3-day continuous intravenous infusion.

Indometacin is used in pregnancy for the treatment of premature labour, but there are limited data on the disposition of the drug in the fetus. In order to elucidate fetal indometacin pharmacokinetics at plasma levels and duration comparable with those occurring with use of the drug for tocolysis in humans, indometacin was administered at doses of 1.9 (low dose, LD; n = 5) or 7.5 (high dose, HD; n = 9) microg min(-1) to steady state over a 3-day period in chronically instrumented fetal lambs. Indometacin concentrations in biological fluid samples were analysed by a sensitive capillary gas chromatography-electron capture detection method. The mean steady-state fetal arterial plasma indometacin concentrations were 68.6+/-16.5 ng mL(-1) in the LD infusion and 230.3+/-28.8 ng mL(-1) in the HD infusion. Indometacin concentrations in amniotic fluid were approximately 10% of those in fetal plasma, and below assay detection limits in tracheal fluid. Total body clearance (TBC) in the LD and HD infusions were not different and the overall mean was 11.3+/-1.2 mL min(-1) kg(-1). In the 11 experiments where paired fetal arterial and umbilical venous samples were collected, the extraction of indometacin across the placenta averaged only 5.2+/-1.1%, indicating low placental permeability to the drug in sheep. However, fetal placental clearance (CLpl) of indometacin (10.0+/-2.5 mL min(-1) kg(-1), n = 10) averaged 115.1+/-41.2% of TBC in these animals and the calculated value for fetal non-placental clearance (0.6+/-2.8 mL min(-1) kg(-1)) was not significantly different from zero. Fetal renal clearance of intact indometacin (3.8+/-1.1 microL min(-1) kg(-1); n = 12) was also very low. However, treatment of fetal urine with glucuronidase indicated the presence of glucuronide conjugates and these comprised 69.9+/-8.2% of the total drug concentration (i.e. intact+conjugated) in urine. Thus, the fetal lamb appears to be able to glucuronidate indometacin, but the contribution of this and other non-placental routes to overall fetal elimination of the drug appear minimal. CLpl of the drug is also low owing to the physicochemical properties of indometacin (high polarity) and the permeability characteristics of the sheep placenta.

Evaluation of gestational deficiencies in cloned sheep fetuses and placentae.

Sheep fetal development at 35 days of gestation was examined following natural mating, in vitro production (IVP) of fertilized embryos, or somatic cell nuclear transfer (NT). Five crossbred (Blackface x Black Welsh) and four purebred (Black Welsh) fetuses and their associated placentae produced by natural mating were morphologically normal and consistent with each other. From 10 ewes receiving 21 IVP embryos, 17 fetuses (81%) were recovered, and 15 of these (88%) were normal. The NT fetuses were derived from two Black Welsh fetal fibroblast cell lines (BLW1 and 6). Transfer of 21 BLW1 and 22 BLW6 NT embryos into 12 and 11 ewes, respectively, yielded 7 (33%) and 8 (36%) fetuses, respectively. Only three (43%) BLW1 and two (25%) BLW6 NT fetuses were normal, with the rest being developmentally retarded. The NT fetal and placental deficiencies included liver enlargement, dermal hemorrhaging, and lack of placental vascular development reflected by reduced or absent cotyledonary structures. Fibroblasts isolated from normal and abnormal cloned fetuses did not differ in their karyotype from sexually conceived fetuses or nuclear donor cell lines. Our results demonstrate that within the first quarter of gestation, cloned fetuses are characterized by a high incidence of developmental retardation and placental insufficiency. These deficiencies are not linked to gross defects in chromosome number.

The nature of the sheep chorionic oxytocic substance

The effects of infusions of fresh sheep placenta in normal saline and the filtrate obtained therefrom after boiling for 15 min were investigated on guinea-pig and rat uteri, other mammalian non-vascular smooth muscles and the cardiovascular system, noting their responses to the infusion/filtrate in the presence or absence of various inhibitors or agonists. Solvent partition, acute toxicity and thin layer chromatography (TLC) studies were also performed. It was found that the infusion/filtrate had oxytocic activity independent of histamine and muscarinic receptors. It had H 1 receptor activity agonist action on the guinea-pig ileum, antagonized adrenaline-induced contractions in the vas but unlike bradykinin did not relax rat duodenum. It induced vasoconstriction in the rat hindquarters, depressed cat blood pressure but had positive inotropic effect on the guinea-pig Langendorff heart. Only the eluent from the least mobile of the five TLC bands on silica gel had oxytocic activity. It was concluded that the sheep chorionic oxytocic substance is not acetylcholine, histamine, 5-hydroxytryptamine, noradrenaline, adrenaline, prostaglandins F or E but is a peptide, which is not bradykinin, vasopressin or oxytocin.

Immunohistochemical localization of prostaglandin G/H synthase 1 and 2 in sheep placenta after glucocorticoid-induced and spontaneous labour

Enhanced prostaglandin production and release by the placenta is an essential element in the normal transition to labour in many animal species. In sheep, expression of prostaglandin G/H synthase (PGHS) is the central enzyme regulating this process. In this study immunohistochemistry was used to examine the distribution of cells expressing PGHS-1 and PGHS-2 in ovine placenta in association with spontaneous parturition (n = 6) and glucocorticoid-induced labour (n = 5). Labour was induced in ewes after the intrafetal injection of betamethasone on day 131 of gestation. Animals administered an intrafetal injection of isotonic saline (n = 5) acted as non-labour controls. In placentomes collected from all groups, immunoreactive PGHS-1 was present in the mononuclear trophoblast cells of the fetal placenta. Cells in the maternal mesenchyme and epithelial syncytium were weakly immunopositive for this enzyme. PGHS-1 immunoreactivity was also demonstrated in the endothelial cells of the chorionic vessels. The PGHS-2 isozyme was localized exclusively to the trophoblast epithelial cells. Immunoreactive PGHS-2 was not detectable in the maternal epithelial syncytium or the stroma of the cotyledons. The binucleate cells of the fetal placenta were consistently immunonegative for both PGHS isozymes. These results indicate that the cellular localization of PGHS-1 and PGHS-2 in ovine placenta does not change during the last 15 days of pregnancy. Co-localization of these isozymes indicates that the source of arachidonic acid and the site of prostanoid formation are the same. Quantitation of the percentage area of positive staining for PGHS-1 and PGHS-2 using image analysis software demonstrated a significant increase in PGHS-2 in the fetal trophoblast after glucocorticoid-induced labour and spontaneous parturition. This finding indicates that increased formation of the PGHS-2 isozyme is responsible for the large increase in prostaglandin production by the ovine placenta at term labour.

Immunohistochemical localization of prostaglandin G/H synthase 1 and 2 in sheep placenta after glucocorticoid-induced and spontaneous labour

Immunohistochemical localization of prostaglandin G/H synthase 1 and 2 in sheep placenta after glucocorticoid-induced and spontaneous labour

Analysis of Prion Protein mRNA by in Situ Hybridization in Brain and Placenta of Sheep

In this study, prion protein (PrP) mRNA was focally detected in brain and placenta of pregnant sheep by Northern blot analysis. In addition, host-encoded cellular prion protein (PrPC) was observed in brain and placenta of the ruminant by Western blot analysis as well. Localization of PrP mRNA in pregnant sheep tissues was rendered possible with in situ hybridization. In sheep brain, PrP mRNA was predominantly localized within large neocortical neurons in the cerebrum, Purkinje cells and neurons of the molecular and granule cell layers in the cerebellum. In the placenta, signals were observed in the myometrium, including stratum longitudinale tunicae muscles and circular layers of muscular tunics. In the caruncle and placentome, signals were stronger by in situ hybridization. Since accumulation of the scrapie isoform PrP (PrPSc) is required to PrPC, these results suggest that brain and placenta of sheep may be important organs and sites for the conversion of PrPC to PrPSc.

Localization of prostaglandin synthase type-1 (PGHS-1) mRNA and prostaglandin synthase type-2 (PGHS-2) mRNA in ovine myometrium and endometrium throughout gestation.

Increased prostaglandin production by tissues in the sheep uterus and placenta are thought to be important for the onset of parturition. In the sheep placenta, this is most likely due to increased expression of prostaglandin synthase type-2 (PGHS-2) rather than prostaglandin synthase type-1 (PGHS-1). However, there is no information concerning expression of PGHS isoenzymes in maternal uterine tissues during pregnancy. Therefore, the purpose of the present study was to examine the expression of PGHS-1 and PGHS-2 in the sheep myometrium and endometrium during late gestation using in situ hybridization and immunohistochemistry. Using (35)S-labelled oligonucleotide probes, which give specific hybridization signals in other tissues, we localized PGHS-2 mRNA to endometrial epithelium, and apparently to other cells in both endometrium and myometrium. This artefactual signal was still present with 100-fold excess unlabelled oligonucleotide probe and with sense probes, but was resolved with the use of (33)P-oligonucleotides. Using (33)P-labelled oligonucleotide probes we could not detect either PGHS-1 or PGHS-2 mRNA in myometrium, and found expression only of PGHS-2 mRNA in endometrium. PGHS-2 mRNA localized to the endometrial epithelium and was undetectable in glandular epithelium. The level of PGHS-2 expression rose significantly between days 80 and 85 of pregnancy and term, and this corresponded to the appearance of immunoreactive PGHS-2 protein, measured by immunohistochemistry, in the endometrial epithelium. Therefore we conclude that (33)P-labelled probes are preferred for detection of mRNAs encoding PGHS-2 in ovine uterine tissues. Expression of PGHS-2 mRNA is greater than that of PGHS-1, increases during gestation, and predominates in the endometrial epithelium, consistent with the site of PGHS-2 protein localization.

Cellular Localization of Vascular Endothelial Growth Factor in Ovine Placenta and Fetal Membranes

To further understand the role of vascular endothelial growth factor (VEGF) in mediating angiogenesis and vascular permeability during development in the sheep placenta and fetal membranes, we examined the localization of VEGF mRNA and protein in placental, chorionic and amniotic tissues by in situ hybridization and immunohistochemistry in ovine fetuses at 62, 102 and 141 days gestation (term=150 days). In the placenta, VEGF mRNA expression and VEGF protein immunostaining were strong in cytotrophoblasts surrounding the villi. In addition, VEGF protein was localized in smooth muscle cells around fetal and maternal blood vessels and in the maternal epithelium. There was no apparent difference in placental VEGF mRNA or protein levels associated with advancing gestation. In the fetal membranes, VEGF mRNA was detected in the amniotic epithelium and the chorionic cytotrophoblastic cell layer. The intensity of the hybridization signals in both amnion and chorion appeared low at 62 days, moderate at 102 days and high at 141 days gestation. VEGF protein was detected in amniotic epithelium and chorionic cytotrophoblasts at all gestational ages studied. The increase in VEGF gene expression in fetal membranes as term approaches suggests that during fetal development VEGF may promote the vascularity and permeability of the microvessels which perfuse the fetal membranes, as well as permeability of the amniotic membrane itself. Thus VEGF may participate in the regulation of amniotic fluid volume.

Involvement of matrix metalloproteinases 2 and 9, tissue inhibitor of metalloproteinases and apoptosis in tissue remodelling in the sheep placenta

Placental growth and development is crucial for successful pregnancy. The aim of this study was to characterize the activity and localization of the matrix metalloproteinase 2 (MMP-2) and MMP-9, which are capable of degrading basement membrane collagen (predominantly collagen type IV), and their endogenous tissue inhibitor of matrix metalloproteinases (TIMPs), in amniotic fluid and in the developing ovine placenta. Cell deletion by apoptosis during placental development was also examined. Zymography with gelatin as substrate indicated that MMP-2 (72 kDa gelatinase A; predominantly latent form) was present in increasing amounts in amniotic fluid from day 70 of gestation to labour (days 140-145), and MMP-9 (92 kDa gelatinase B; predominantly latent form) was detectable from day 125 to labour; there was no increase in MMP-2 or -9 in labour. A broad range of TIMPs was detected in amniotic fluid; the molecular masses corresponded to TIMP-1, -2 and -3. Immunohistochemical techniques localized MMP-2, MMP-9 and TIMP-3 in the sheep placenta, predominantly in the trophoblast layer in uninucleate, but not binucleate, cells. However, MMP-2 and -9 activated proteins in placental homogenates were low throughout pregnancy. Apoptosis was identified by morphological criteria and also by TdT-mediated dUTP nick end labelling. Apoptosis was present in discrete regions in the placenta, predominantly in trophoblast cells near the tips and the basal regions of the fetomaternal interdigitations. During pregnancy the sheep placenta becomes more complex and the area of the fetomaternal interface increases. MMP-2 and -9 are likely to be involved in breaking down basement membranes to allow cell migration during this process. It is suggested that digestion of supporting extracellular matrix may trigger apoptosis and in some way increase the branching pattern in the villi.

Involvement of matrix metalloproteinases 2 and 9, tissue inhibitor of metalloproteinases and apoptosis in tissue remodelling in the sheep placenta

Placental growth and development is crucial for successful pregnancy. The aim of this study was to characterize the activity and localization of the matrix metalloproteinase 2 (MMP-2) and MMP-9, which are capable of degrading basement membrane collagen (predominantly collagen type IV), and their endogenous tissue inhibitor of matrix metalloproteinases (TIMPs), in amniotic fluid and in the developing ovine placenta. Cell deletion by apoptosis during placental development was also examined. Zymography with gelatin as substrate indicated that MMP-2 (72 kDa gelatinase A; predominantly latent form) was present in increasing amounts in amniotic fluid from day 70 of gestation to labour (days 140-145), and MMP-9 (92 kDa gelatinase B; predominantly latent form) was detectable from day 125 to labour; there was no increase in MMP-2 or -9 in labour. A broad range of TIMPs was detected in amniotic fluid; the molecular masses corresponded to TIMP-1, -2 and -3. Immunohistochemical techniques localized MMP-2, MMP-9 and TIMP-3 in the sheep placenta, predominantly in the trophoblast layer in uninucleate, but not binucleate, cells. However, MMP-2 and -9 activated proteins in placental homogenates were low throughout pregnancy. Apoptosis was identified by morphological criteria and also by TdT-mediated dUTP nick end labelling. Apoptosis was present in discrete regions in the placenta, predominantly in trophoblast cells near the tips and the basal regions of the fetomaternal interdigitations. During pregnancy the sheep placenta becomes more complex and the area of the fetomaternal interface increases. MMP-2 and -9 are likely to be involved in breaking down basement membranes to allow cell migration during this process. It is suggested that digestion of supporting extracellular matrix may trigger apoptosis and in some way increase the branching pattern in the villi.

Ontogeny of Serine Hydroxymethyltransferase Isoenzymes in Fetal Sheep Liver, Kidney, and Placenta

Serine is an amino acid that is not transported from the placenta to the ovine fetus. Thus, fetal plasma serine levels may be controlled by flux through their relevant biosynthetic pathways. This study was designed to determine, in fetal sheep tissues, the ontogeny of the three key enzymes in the biosynthetic pathway for serine, the cytosolic (c) and mitochondrial (m) isoforms of serine hydroxymethyltransferase (SHMT), phosphoglycerate dehydrogenase (PGD), and phosphoserine aminotransferase (PSAT). PGD and PSAT activity did not vary during gestation in either liver (PSAT, 9.4 ± 1.3 nmol/min/mg cytosolic protein; and PGD, 76 ± 10 mU/mg protein) or placenta (PGD, 8.0 ± 3.6 mU/mg protein). In the liver, cSHMT activity was low early in gestation (0.6 ± 0.5 nmol/min/mg protein at 45 days), rose in the last one-third of gestation, and peaked in the newborn period (25 ± 3 nmol/min/mg protein at 1 week of age). Hepatic cSHMT RNA levels parallel the activity pattern. Mitochondrial SHMT was stable throughout gestation and with low constant mSHMT RNA levels. In contrast, the kidney and placenta had high mSHMT and steady low cSHMT activity throughout gestation. These data support the possible role of SHMT in the fetal control of plasma serine levels. While cSHMT may contribute to fetal hepatic serine production, its activity pattern does not support a primary role in the control of fetal hepatic serine biosynthesis. In the placenta, mSHMT may be important for glycine production from serine.

Placental transfer of the thromboxane synthetase inhibitor ridogrel in the late-pregnant ewe

Objectives: To assess the occurrence of placental transfer of the thromboxane synthetase inhibitor ridogrel in the pregnant ewe and to determine its effect on prostanoid levels in the ewe and fetal lamb, on uterine contractility and on maternal and fetal hemodynamics. Study design: Five chronically instrumented pregnant ewes at 122 days of gestation received intravenous infusions of 5 mg/kg/3 h ridogrel and solvent. Maternal and fetal arterial samples were obtained at predetermined intervals to determine concentrations of ridogrel and prostaglandin metabolites TXB 2, 6-keto-PGF 1α, PGF 2α, and PGE 2. Maternal and fetal responses of blood flow and pressures were determined. Results: Fetal ridogrel levels were 25% of maternal concentrations. Ridogrel showed rapid and marked thromboxane synthetase inhibition and augmentation of levels of prostaglandin metabolites. There was no evidence of change in amniotic pressure, uterine blood flow, maternal and fetal blood pressure and heart rate. Conclusion: Ridogrel is a potent thromboxane synthetase inhibitor which passes the sheep placenta, does not influence maternal and fetal hemodynamics and uterine contractility, and shows similar antiplatelet activity in the ewe and the fetal lamb.

Differential effects of natural and synthetic glucocorticoids on cytochrome 17α-hydroxylase (P-450 17α) and cytochrome P-450 side-chain cleavage (P-450 scc) messenger ribonucleic acid in the sheep placenta

Objective: Our purpose was to determine differential effects of natural and synthetic glucocorticoids on cytochrome 17α-hydroxylase and cytochrome P-450 side-chain cleavage messenger ribonucleic acid in the sheep placenta and to determine whether feed-forward effects during labor are involved in further inducing placental cytochrome 17α-hydroxylase. Study Design: Sheep underwent placement of myometrial electromyogram electrodes while they were under general anesthesia at 117 days’ gestation. At 125 days’ gestation either saline solution (early control animals not in labor, n = 5), 0.48 mg betamethasone during 48 hours (n = 7), 0.48 mg dexamethasone during 48 hours (n = 7), or 55 mg cortisol during a maximum of 96 hours (n = 4) was directly administered intravenously to the fetus. Necropsies were performed at 127 to 129 days’ gestation. We also studied 6 ewes in spontaneous term labor at 143-147 days’ gestation, 6 term control animals not in labor at 140 to 147 days’ gestation, and 6 sheep in which myometrial activity was inhibited by intravenous infusion to the ewe of the selective cyclooxygenase 2 inhibitor nimesulide 9 hours after the onset of labor beginning at 147 to 148 days’ gestation. Total fetal placental ribonucleic acid was analyzed by Northern blot with complementary deoxyribonucleic acid probes for cytochrome 17α-hydroxylase, cytochrome P-450 side-chain cleavage, and 18S ribosomal ribonucleic acid to correct for loading. Results: Placental cytochrome 17α-hydroxylase messenger ribonucleic acid was detectable neither in term control animals not in labor nor in early control animals not in labor. Placental cytochrome 17α-hydroxylase messenger ribonucleic acid was induced in spontaneous term labor and all cortisol-infused sheep in labor with respect to term control animals not in labor and early control animals not in labor ( P < .01). All betamethasone-infused sheep had myometrial contraction activity; however, only 4 of 7 had detectable placental cytochrome 17α-hydroxylase messenger ribonucleic acid. Placental cytochrome 17α-hydroxylase messenger ribonucleic acid was not detected in dexamethasone-infused sheep, even the 2 that had myometrial contractions. After reversal of the progression of spontaneous labor with nimesulide placental cytochrome 17α-hydroxylase messenger ribonucleic acid was significantly lower than the spontaneous term labor group (without nimesulide treatment). The placentas from all animals expressed cytochrome P-450 side-chain cleavage messenger ribonucleic acid, but no changes were associated with either gestational age studied (130 versus > 140 days’ gestation) or glucocorticoid-induced premature labor and spontaneous term labor. Conclusions: (1) In sheep the expression of placental cytochrome 17α-hydroxylase is tightly associated with spontaneous term labor, and active synthesis of placental cytochrome 17α-hydroxylase is required during the progression of labor. (2) Cortisol is a more potent stimulator of placental cytochrome 17α-hydroxylase messenger ribonucleic acid in sheep than are synthetic glucocorticoids. (3) Betamethasone has a greater effect in inducing labor in sheep than does dexamethasone, possibly mediated through placental cytochrome 17α-hydroxylase. (Am J Obstet Gynecol 1999;180:1215-21.)

Changes in prostacyclin synthase in pregnant sheep myometrium, endometrium, and placenta at spontaneous term labor and regulation by estradiol and progesterone

Objective: Our purpose was to investigate, first, whether there were changes in the abundance of prostacyclin synthase protein in intrauterine tissues of pregnant ewes in association with spontaneous term labor. Second, we examined the effect of either estradiol or progesterone, or both, on regulation of prostacyclin synthase protein abundance in uterine tissues using an ovariectomized nonpregnant sheep model. Study Design: The abundance of prostacyclin synthase protein was quantified by Western blot analysis in the myometrium, endometrium, and placenta of pregnant ewes in spontaneous term labor (n = 6) and term control ewes not in labor (n = 6). The changes of prostacyclin synthase in the myometrium and endometrium of 20 ovariectomized nonpregnant sheep (n = 5 for each group) were evaluated after treatment with estradiol, progesterone, or both. Results: Prostacyclin synthase protein was present in pregnant and nonpregnant sheep myometrium, endometrium, and placenta at a molecular weight of about 55 kd. At spontaneous term labor the level of prostacyclin synthase decreased in endometrium (P < .05), increased in myometrium (P < .05), and remained unchanged in placenta. Estradiol and progesterone had no effect on prostacyclin synthase protein abundance in nonpregnant ovine endometrium and myometrium. Conclusions: The decrease in prostacyclin synthase in pregnant sheep endometrium during labor may indicate paracrine interactions between the endometrium, the myometrium, fetal membranes, or a combination of these. The significant increase of prostacyclin synthase in pregnant sheep myometrium at spontaneous term labor may contribute to the increased uterine sensitivity to oxytocin or stimulate vasodilatation during labor to increase myometrial blood flow. Neither estradiol nor progesterone at the dosages studied changed prostacyclin synthase expression in the nonpregnant myometrium and endometrium. The molecular mechanism or mechanisms that differentially regulate prostacyclin synthase expression in pregnant uterine tissues merit further study. (Am J Obstet Gynecol 1999;180:744-9.)

Bibliography of resources by and about André Hellegers.

Bibliography of Resources by and about André E. Hellegers* Compiled by Doris Mueller Goldstein (bio) This bibliography is derived from the holdings of the National Reference Center for Bioethics Literature and the BIOETHICSLINE© database (both of which are at the Kennedy Institute of Ethics and supported by the National Library of Medicine); the archives of Lauinger Library, Georgetown University; the Medline databases of the National Library of Medicine; the WorldCat database of OCLC, Dublin, Ohio, and selected Internet sites. Primarily comprised of writings by Dr. Hellegers, the list also includes interviews and media programs in which he participated, as well as writings about him that were written following his death. The arrangement within each section is by date. The 231 authored pieces reflect the transition from Dr. Hellegers’s early career as a researcher in fetal physiology at Johns Hopkins Medical School, where he published his first research report at the age of 29, to the early years of the Kennedy Institute, where he produced so many provocative columns on bioethical issues that influenced the growth of the field. Due to the diversity of sources and the 40-year time span covered, there are some variations in bibliographic style. Time and availability constraints precluded verifying all of the citations with the original documents. Any additions or corrections should be submitted to the compiler. Works Authored by André E. Hellegers No Date Ethical Issues in Sterilization of the Retarded. Unpublished paper. 1956 Vitamin B12 Serum Level and Pregnancy. American Journal of Clinical Nutrition 4: 440+, 1956. [With Bacon F. Chow and Kunio Okuda] [End Page 89] 1957 Vitamin B12 Absorption in Pregnancy and in the Newborn. American Journal of Clinical Nutrition 5: 327+, 1957. [With Bacon F. Chow, Robert E. L. Nesbitt, Kunio Okuda, and David W. Smith] 1958 The Effects of Acute Hypoxia on the Osmotic Pressure of the Plasma. Quarterly Journal of Experimental Physiology 43: 197+, 1958. [With Donald H. Barron, Frederick C. Battaglia, and Giacomo Meschia] Studies on the Absorption of Vitamin B12 in Human Pregnancy with Especial Reference to the Effect of D-Sorbitol. American Journal of Obstetrics and Gynecology 76: 91+, 1958. [With Bacon F. Chow, Harry Prystowsky, and Vernon Wong] 1959 The Alveolar PCO2 and PO2 in Pregnant and Nonpregnant Women at Altitude. Proceedings of the Society for Clinical Investigation 38: 1010+, 1959. [With Donald Barron, William Huckabee, Giacomo Meschia, James Metcalfe, and Harry Prystowsky] A Comparison of the Oxygen Dissociation Curves of the Bloods of Maternal and Fetal Goats at Various pHs. Quarterly Journal of Experimental Physiology 44: 215–21, July 1959. [With Donald Barron, Giacomo Meschia, Harry Prystowsky, and A. Stark Wolkoff] Fetal Blood Studies. XII. On the Relationship Between the Position of the Oxygen Dissociation Curve of Human Fetal Blood and Adult Fetal Hemoglobin. American Journal of Obstetrics and Gynecology 77: 585+, 1959. [With Paul D. Bruns, Julian Cotter, and Harry Prystowsky] Fetal Blood Studies. XIV. A Comparative Study of the Oxygen Dissociation Curve of Nonpregnant, Pregnant and Fetal Human Blood. American Journal of Obstetrics and Gynecology 78: 489–93, September 1959. [With Paul D. Bruns and Harry Prystowsky] Fetal Blood Studies. XVI. On the Changes in Total Osmotic Pressure and Sodium and Potassium Concentration of Amniotic Fluid During the Course of Human Gestation. Surgery, Gynecology and Obstetrics 109: 509–12, October 1959. [With Frederick Battaglia, Paul D. Bruns, Harry Prystowsky, and Clayton Smisson] Further Observations on the Metabolism of Vitamin B12 in Human Pregnancy. American Journal of Obstetrics and Gynecology 77: 1+, 1959. [With Bacon F. Chow, Harry Prystowsky, Bruno Ranke, and Elma Ranke] The Oxygen Dissociation Curves of the Bloods of Adult and Fetal Llamas. Federation Proceedings 18, 1959. [With Donald Barron, William Huckabee, Giacomo Meschia, James Metcalfe, and Harry Prystowsky] [End Page 90] Transfer of Oxygen Across the Sheep Placenta at High Altitude. Federation Proceedings 18, 1959. [With Donald Barron, William Huckabee, Giacomo Meschia, and Harry Prystowsky] Uterine Blood Flow and Metabolism in Pregnant Sheep at High Altitude. Federation Proceedings 18, 1959. [With Donald Barron, William Huckabee, Giacomo Meschia, James Metcalfe, and Harry Prystowsky] Uterine Blood Flow and Metabolism in Unanesthetized Pregnant Sheep. Federation Proceedings 18, 1959. [With Donald Barron, William Huckabee, Giacomo...

In situ hybridization for the detection and localization of swine Chlamydia trachomatis.

Gnotobiotic piglets were inoculated intralaryngeally with swine Chlamydia trachomatis strain R33 or orally with swine C. trachmatis strain R27. Archived formalin-fixed, paraffin-embedded tissues from piglets euthanatized 4-7 days postinoculation were examined by in situ hybridization for C. trachomatis nucleic acid using a nonradioactive digoxigenin-labeled DNA probes that targeted specific ribosomal RNA or omp1 mRNA molecules of the swine C. trachomatis strains. Positive hybridization signals were detected in bronchial epithelial cells, bronchiolar epithelial cells, pneumocytes, alveolar and interstitial macrophages, and jejunal and ileal enterocytes. Chlamydia-infected cells had a strong signal that was confined to the intracytoplasmic inclusions. Positive hybridization signals were not detected in tissue sections from an uninfected control piglet or in C. psittaci-infected sheep placenta. The morphology of host cells was preserved despite the relatively high temperature required in parts of the incubation procedure. The data indicate that in situ hybridization can be used to detect swine C. trachomatis in formalin-fixed, paraffin-embedded tissue specimens.

Characterisation of strain-specific sequences from an abortifacient strain of ovine Chlamydia psittaci using subtraction hybridisation

Enzootic abortion in ewes (EAE) is caused by strains of Chlamydia psittaci which have the ability to invade and colonise the placenta of sheep. In an attempt to improve diagnostic methods for the detection of EAE, subtraction hybridisation was used to isolate unique fragments of the genome of an abortifacient strain (S26/3) of C. psittaci. One S26/3 strain-specific sequence identified was shown to encode a putative helicase which is repeated throughout the EAE genome. The labelled strain-specific helicase gene fragment was used in a dot-blot hybridisation test for the detection of EAE DNA in ovine placental samples. We report the identification of C. psittaci S26/3 strain-specific sequences with potential as diagnostic probes for the detection of EAE.

Purification and cloning of the mitochondrial branched-chain amino acid aminotransferase from sheep placenta.

This paper presents the first purification of the mitochondrial branched-chain amino acid aminotransferase (BCATm) from sheep placenta. It is a homodimer with an apparent subunit molecular mass of 41 kDa. The enzyme differs from those of the rat and human as it appears to form at least one intermolecular disulfide bond. The sheep BCATm cDNA (1.4 kb) encodes a mature polypeptide of 366 amino acids with a calculated molecular mass of 41 329 Da and a partial mitochondrial targeting sequence of seven amino acids. The sheep BCATm sequence shares higher identity with other mammalian BCATm isoenzymes (82-85%) than with the cytosolic isoenzymes (60%). By Northern blot analysis, a message of 1.7 kb was detected in sheep placenta and skeletal muscle. Measurements of BCAT activity, mRNA and BCATm protein in sheep placenta and skeletal muscle revealed that BCATm is the sole BCAT isoenzyme expressed in placenta, whereas it contributes 57 and 71% of the BCAT activity in tensor fascia latae and masseter muscles from weaned lambs respectively. Skeletal muscle, the main site of branched-chain amino acid transamination, exhibits significantly lower BCAT activity in sheep than in rat. Our results suggest that the low BCATm mRNA level probably accounts for the low BCAT activity in sheep skeletal muscle, and that the metabolic scheme for branched-chain amino acid catabolism is specific to each species.