Pregnant Sheep Research Articles

Reproductive losses caused by the ingestion of Poincianella pyramidalis in sheep

Malformations have been observed in sheep and goats in the Brazilian semiarid region in areas where Poincianella pyramidalis is dominant. The objective of this trial was to determine whether Poincianella pyramidalis causes reproductive changes in pregnant sheep. Sixteen non-pregnant sheep were mated with two rams. After confirmation of the pregnancy by ultrasonography on the 18th day after mating, pregnant sheep were randomly divided into four groups (#1, 2, 3, and 4), with four animals each. Sheep received roughage in an amount equivalent to 2% of their body weight, mixed with 0%, 10%, 20% and 40% of dried leaves of P. pyramidalis for groups 1, 2, 3 and 4, respectively. In Groups 1 and 2, all animals lambed normally. In Group 3 (20% P. pyramidalis in the roughage), one lamb was born with arthrogryposis and three sheep gave birth to premature weak lambs within 128, 132, and 133 days of gestation. In Group 4 (40% P. pyramidalis in the roughage), one sheep lambed a normal lamb; another ewe had embryonic mortality after seven days of plant consumption, and two aborted on days 103 and 144 of pregnancy. One of the aborted fetuses was normal and the other showed arthrogryposis and prognathism. These results suggest that P. pyramidalis causes embryonic deaths, abortions, and malformations in sheep. Grazing pregnant sheep in areas where this plant is dominant should be avoided, and roughage for confined pregnant females should not contain more than 10% P. pyramidalis.

Interferon-τ induced gene expression in bovine hepatocytes during early pregnancy

Interferon-tau (IFNτ) is the conceptus derived specific early pregnancy signal in bovidae. Locally, IFNτ induces an IFNτ specific gene expression (ISG) in endometrial cells and by this it averts luteolysis of the corpus luteum (CL) by suppressing prostaglandin production. Moreover, it was shown that IFNτ also induces ISG in the liver in pregnant sheep and in liver biopsies from Holstein Friesian heifers on Day 18 of pregnancy. The objective of the present study was to confirm increased hepatic ISG in vivo on Day 18 of pregnancy and to prove if hepatocytes and not non-parenchymal cells react to IFNτ by using immunohistochemistry and primary bovine hepatocytes stimulated in vitro with recombinant bovine IFNτ. For the animal experiment, Angus heifers (n = 12) were cycle synchronized and the Day of ovulation (Day 0) was defined by ovarian ultrasonography and verified by progesterone < 0.1 ng/ml. Heifers were artificially inseminated either with sperm (n = 9) or with seminal plasma (mock control, n = 3). Early pregnancy was defined and detected by progesterone and pregnancy associated glycoprotein (PAG) concentration in blood before and after induced luteolysis by a PGF-injection on Day 21 in n = 3 inseminated heifers. A liver biopsy was taken on Day 18 for the analysis of gene (ISG 15, MX 1, MX 2 and OAS 1) and protein (OAS1) expression using qPCR and immunohistochemistry, respectively. Primary bovine hepatocytes were collected from bull liver using a two-step collagenase perfusion, cultured short-term in a monolayer and stimulated with IFNτ. Thereafter, gene expression was measured by qPCR. In liver biopsies obtained from pregnant heifers ISG was numerically higher expressed compared to biopsies from non-pregnant heifers. Furthermore, the OAS 1 protein expression was localized in hepatocytes on Day 18 of pregnancy. In vitro, primary bovine hepatocytes showed an increased mRNA expression of ISG after IFNτ stimulation. In conclusion, the findings confirm that IFNτ induces ISG in the parenchymal part of the liver in early pregnancy of cattle.

MicroRNA-210 Targets Ten-Eleven Translocation Methylcytosine Dioxygenase 1 and Suppresses Pregnancy-Mediated Adaptation of Large Conductance Ca2+-Activated K+ Channel Expression and Function in Ovine Uterine Arteries.

Gestational hypoxia inhibits large conductance Ca2+-activated K+ (BKCa) channel expression and function in uterine arterial adaptation to pregnancy. Given the findings that microRNA-210 (miR-210) is increased in hypoxia during gestation and preeclampsia, the present study sought to investigate the role of miR-210 in the regulation of BKCa channel adaptation in the uterine artery. Gestational hypoxia significantly increased uterine vascular resistance and blood pressure in pregnant sheep and upregulated miR-210 in uterine arteries. MiR-210 bound to ovine ten-eleven translocation methylcytosine dioxygenase 1 mRNA 3' untranslated region and decreased ten-eleven translocation methylcytosine dioxygenase 1 mRNA and protein abundance in uterine arteries of pregnant sheep, as well as abrogated steroid hormone-induced upregulation of ten-eleven translocation methylcytosine dioxygenase 1 expression in uterine arteries of nonpregnant animals. In accordance, miR-210 blocked pregnancy- and steroid hormone-induced upregulation of BKCa channel β1 subunit expression in uterine arteries. Functionally, miR-210 suppressed BKCa channel current density in uterine arterial myocytes of pregnant sheep and inhibited steroid hormone-induced increases in BKCa channel currents in uterine arteries of nonpregnant animals. Blockade of endogenous miR-210 inhibited hypoxia-induced suppression of BKCa channel activity. In addition, miR-210 decreased BKCa channel-mediated relaxations and increased pressure-dependent myogenic tone of uterine arteries. Together, the results demonstrate that miR-210 plays an important role in the downregulation of ten-eleven translocation methylcytosine dioxygenase 1 and repression of BKCa channel function in uterine arteries, revealing a novel mechanism of epigenetic regulation in the maladaptation of uterine hemodynamics in gestational hypoxia and preeclampsia.

Noninvasive embryo assessment technique based on buoyancy and its association with embryo survival after cryopreservation

Embryo cryopreservation offers many benefits by allowing genetic preservation, genetic screening, cost reduction, global embryo transport and single embryo transfer. However, freezing of embryos decreases embryo viability, as intracellular ice crystal formation often damages embryos. Success rates of frozen embryo transfer are expected to be 15–20% less than fresh embryo transfer. We have developed a noninvasive embryo assessment technique (NEAT) which enables us to predict embryo viability based on buoyancy. The purpose of this research was twofold. First was to determine if a NEAT, through a specific gravity device can detect embryo survival of cryopreservation. Second, it was to relate embryo buoyancy to embryo viability for establishing pregnancies in sheep. Blastocysts descent times were measured on one-hundred sixty-nine mice blastocysts before cryopreservation, according to standard protocol and post-thawing blastocysts descent times were measured again. There was a significant difference in blastocyst post-thaw descent times with NEAT in those blastocysts which demonstrated viability from those that did not (P < 0.05). This suggests NEAT is successful in determining blastocysts viability in cryopreserved mice blastocysts. At a commercial ovine facility, NEAT was performed on fourteen frozen and thawed ovine blastocysts. Blastocysts of similar descent times were paired and transferred into recipient ewes as twins. Pregnancy was later confirmed by blood test and multiple gestation outcomes were determined at lambing. Six of seven recipient ewes were pregnant and all pregnant ewes delivered lambs without complication. Four ewes delivered twin lambs and two ewes delivered singletons, which totals ten of the fourteen (71%) blastocysts surviving to term. This pregnancy rate is comparable to expected to pregnancy rates in a commercial setting. The blastocysts which did not establish pregnancy demonstrated less buoyancy versus those blastocysts which established pregnancies which survived to term (P < 0.05). These results suggest NEAT can identify which blastocysts survive cryopreservation, thus significantly reduce the transfer of non-viable embryos. Further studies on a larger scale commercial setting will evaluate the efficacy of NEAT.

The uptake of transdermal fentanyl in a pregnant sheep model

ObjectiveTo evaluate the maternal and foetal uptake of transdermal fentanyl patch applied to the groin of pregnant sheep following surgery. Study designProspective series. AnimalsA group of 16 singleton pregnant sheep underwent anaesthesia for laparotomy, hysterotomy and instrumentation of the foetus. Of these ewes 10 (101 ± 12 days of gestation) were used to evaluate the maternal uptake of transdermal fentanyl, and the efficacy of the drug in the postoperative period (n = 10). To determine the extent of transplacental transfer of fentanyl, six ewes from the group of 10, and six other ewes (92 ± 1 days’ gestation) were studied. MethodsA 75 μg hour−1 fentanyl patch was placed onto the woolless skin of the medial thigh close to the groin at the end of surgery. Maternal blood samples were collected from the cephalic or jugular vein, and pain and sedation scores were determined, prior to application of the patch (time 0) and at 3, 6, 12, 24, 36 and 48 hours after. A commercial Fentanyl ELISA kit was used to determine the concentration of fentanyl. Paired maternal and foetal blood samples were collected 48 hours after surgery. Animals were euthanized at the end of the study. Data were tested for normality and compared with Student t test or one-way anova and are expressed as mean ± standard deviation or median (range). ResultsRecovery from anaesthesia and surgery was uneventful in all ewes. The dose of fentanyl was 1.4 ± 0.2 μg kg−1 hour−1. The maximum maternal plasma concentration of fentanyl was 0.547 ng mL−1 (range, 0.349–0.738 ng mL−1) at 12 hours. After 48 hours, the concentration of fentanyl was 0.381 ng mL−1 (range, 0.211–0.487 ng mL−1; maternal) and 0.295 ng mL−1 (range, 0.185–0.377 ng mL−1; foetal; p = 0.175). The placental transfer rate of fentanyl was 77%. Conclusions and clinical relevanceThe uptake of fentanyl varied between animals. The placental transfer rate of fentanyl was 77%.

Fetal adaptations in insulin secretion result from high catecholamines during placental insufficiency.

Placental insufficiency and intrauterine growth restriction (IUGR) of the fetus affects approximately 8% of all pregnancies and is associated with short- and long-term disturbances in metabolism. In pregnant sheep, experimental models with a small, defective placenta that restricts delivery of nutrients and oxygen to the fetus result in IUGR. Low blood oxygen concentrations increase fetal plasma catecholamine concentrations, which lower fetal insulin concentrations. All of these observations in sheep models with placental insufficiency are consistent with cases of human IUGR. We propose that sustained high catecholamine concentrations observed in the IUGR fetus produce developmental adaptations in pancreatic β-cells that impair fetal insulin secretion. Experimental evidence supporting this hypothesis shows that chronic elevation in circulating catecholamines in IUGR fetuses persistently inhibits insulin concentrations and secretion. Elevated catecholamines also allow for maintenance of a normal fetal basal metabolic rate despite low fetal insulin and glucose concentrations while suppressing fetal growth. Importantly, a compensatory augmentation in insulin secretion occurs following inhibition or cessation of catecholamine signalling in IUGR fetuses. This finding has been replicated in normally grown sheep fetuses following a 7-day noradrenaline (norepinephrine) infusion. Together, these programmed effects will potentially create an imbalance between insulin secretion and insulin-stimulated glucose utilization in the neonate which probably explains the transient hyperinsulinism and hypoglycaemia in some IUGR infants.

The role of cavitation in vascular occlusion

Ed Carstensen’s papers on the effects of ultrasound on plants were amongst the first that attracted me to study ultrasound bio-effects. It was little surprise to me that, when I first started looking at HIFU, his name was on many of the seminal papers in this area. And now, his papers on cavitation and vasculature inform our current research. Ed’s influence on bio-effects research stretches far beyond the borders of the USA Vascular occlusion has the potential to treat life-threatening conditions including twin-twin transfusion syndrome (occurring in fetuses that share a placenta). Sheep placental vessels have been used to develop HIFU treatment guidance, delivery (1.66MHz) and monitoring. Ultrasound imaging guidance/flow monitoring, with simultaneous acoustic cavitation detection in 82 targets in 14 pregnant sheep resulted in flow occlusion in 90%. Adverse events included skin erythema (18/82) and burns (10/82), but all resolved over 21 days. Analysis of acoustic cavitation required low pass (software) filtering and careful choice of broadband integration range (0.1 to 1.1 MHz). 9% of successful treatments displayed detectable broadband, 32% with half-harmonic and 1% with drive-voltage fluctuations. In conclusion, treatment is possible, side effects are manageable, and acoustic cavitation appears not to be essential for successful occlusion.

Pregnant Sheep in a Farm Environment Did Not Develop Anaemia.

Simple SummaryAnaemia refers to a low red blood cell count and is common during pregnancy in women. Anaemia has been reported in sheep undergoing surgery during pregnancy for biomedical research projects. The incidence and severity of anaemia during pregnancy in sheep is unknown. Three groups of sheep were established: non-pregnant; pregnant with a single lamb; and pregnant with twin lambs. The stage of pregnancy was known as the sheep were mated on a known date and pregnancy was confirmed 50 days later by ultrasound examination. During pregnancy, blood samples were collected to measure red cells in the blood and protein levels. Samples were also collected to ensure that the sheep did not have any risk factors for anaemia, such as nutritional deficiency and internal parasites. Anaemia did not occur in any sheep during the study. Anaemia did not develop during pregnancy and red cell counts and protein levels were similar between each group. All the sheep lambed as expected, except for one that was expecting twins delivered a single lamb.The aim of this study was to document the haematological profile of pregnant ewes throughout gestation. Sheep were divided into three groups (n = 8 per group): non-pregnant, singleton, or twin pregnancy. Blood samples were collected every 14 days from day 55 of gestation for haemoglobin concentration; packed cell volume; total protein; and albumin concentration. On days 55 and 125 of gestation blood was collected for trace element estimation: soluble copper and zinc; glutathione peroxidase (GSHPx); and methylmalonic acid (MMA). Pooled faecal samples were collected on days 55, 97, and 139 of gestation. Pasture cuts were collected on days 97 and 153 of gestation. The haematology and protein concentrations were not different between groups throughout the study. Copper concentration increased in all animals during the study (p < 0.0001). Zinc concentration was lowest in the singleton and twin pregnant sheep on day 55 of gestation (p = 0.04). GSHPx was not different between groups during the study. MMA decreased in all animals during the study (p < 0.0001), but was not different between groups. Faecal samples were consistently negative for strongyle and nematode eggs, and coccidian oocysts. The pasture was good quality. Pregnant sheep in a farm environment with normal trace element status, no parasites, and an adequate diet, did not develop anaemia (PCV < 0.27).

Intergenerational impact of maternal overnutrition and obesity throughout pregnancy in sheep on metabolic syndrome in grandsons and granddaughters

We previously reported that maternal overnutrition and obesity (MO) throughout pregnancy and lactation in sheep (MOF0) decreases term fetal pancreatic β-cell numbers and increases perirenal adiposity producing hyperphagia, increased adiposity and insulin resistance in adult female offspring (MOF1) fed ad libitum. Pregnant female MOF1 exhibited increased blood glucose from mid to late gestation vs control F1 (CTRF1) though both groups ate only to NRC recommendations. MOF1 ewes delivered female offspring (F2) who like their MOF1 mothers exhibited increased abdominal adiposity and absent neonatal leptin surge. In the current work, we determined if adult MOF2 exhibited metabolic syndrome components when fed ad libitum. After weaning, MOF2 males (n = 5), MOF2 females (n = 6), CTRF2 males (n = 5), and CTRF2 females (n = 6) were fed to NRC requirements until 19 mo followed by 12-wk ad libitum feeding. Body weight and % fat increased (P < 0.01) in all F2 during this feeding trial. MOF2 males were heavier (P < 0.01) than CTRF2 males and females, and MOF2 females throughout the trial. By wk 8, baseline blood glucose concentrations increased (P < 0.001) in MOF2 females, but not other groups, remaining elevated throughout the trial. Baseline insulin was similar through wk 6, increasing (P < 0.05) at wk 8 in MOF2 females only. MOF2 female insulin returned to CTRF2 female levels during wk 10 and 12. The progressive increase of plasma glucose on wk 8 in association with increased insulin in MOF2 females but not other groups demonstrated a diet-induced increase (P < 0.001) in MOF2 female insulin resistance. The subsequent decline in insulin during wk 10 and 12 despite elevated glucose in MOF2 females is consistent with a decrease in glucose-stimulated pancreatic β-cell function. These data indicate that ad libitum feeding exceeds the pancreatic secretory response predisposing MOF2 females to hyperglycemia. Furthermore, there was a sex difference where MOF2 males increased body mass and MOF2 females displayed insulin/glucose dysregulation.

Visual ELISA for detection of pregnancy-associated glycoproteins (PAGs) in ewe serum

The aim of the present study was to evaluate the efficiency of visual ELISA-PAG for early pregnancy diagnosis based on the presence of PAG (pregnancy-associated glycoprotein) using sheep blood serum. Experiment 1: 140 ewes were divided into three groups with different stages of pregnancy. In the first group, 41 pregnant ewes were sampled at 26, 28, 30, and 32 days of pregnancy; in the second group, 65 ewes (49 pregnant with 30 days and 16 non-pregnant) were sampled; in the third group, 34 non-pregnant ewes were sampled. Experiment 2: 10 pregnant ewes were sampled weekly from day 35 of gestation until day 70 post-partum to verify the total period in which PAG can be detected in the blood serum by the test. Transrectal ultrasound was used as a gold standard. The detection or non-detection of PAG was analyzed by the logistic model PROC GENMOD of SAS; differences were detected by the chi-squared test. In group 1, there were no differences between the results from 28, 30, and 32 days of gestation, but samples from 30 days were easier to interpret in comparison to 28 days, with a sensitivity of 97.56%. In the second group, using 65 sheep, visual ELISA-PAG showed 100% sensitivity and 93.75% specificity, which indicates the diagnosis of an animal as a false positive. In the third group, 97.06% of the sheep were confirmed as negative and 2.94% as positive, again indicating the presence of a false positive. In 100% of the sheep, the PAG remained in the blood circulation throughout the antepartum period until birth and seven days post-partum, declining thereafter. Based on our results, the visual ELISA-PAG is an effective method for the early diagnosis of pregnancy in sheep and can be performed from day 30 of gestation.

Ureaplasma Species Multiple Banded Antigen (MBA) Variation Is Associated with the Severity of Inflammation In vivo and In vitro in Human Placentae.

Background: The multiple banded antigen (MBA), a surface-exposed lipoprotein, is a proposed virulence factor of Ureaplasma spp. We previously demonstrated that the number of Ureaplasma parvum MBA size variants in amniotic fluid was inversely proportional to the severity of chorioamnionitis in experimentally infected pregnant sheep. However, the effect of ureaplasma MBA size variation on inflammation in human pregnancies has not been reported.Methods: Ureaplasmas isolated from the chorioamnion of pregnant women from a previous study (n = 42) were speciated/serotyped and MBA size variation was demonstrated by PCR and western blot. Results were correlated with the severity of chorioamnionitis and cord blood cytokines. In vitro, THP-1-derived macrophages were exposed to recombinant-MBA proteins of differing sizes and NF-κB activation and cytokine responses were determined.Results: MBA size variation was identified in 21/32 (65.6%) clinical isolates (in 10 clinical isolates MBA size variation was unable to be determined). Any size variation (increase/decrease) of the MBA (regardless of Ureaplasma species or serovar) was associated with mild or absent chorioamnionitis (P = 0.023) and lower concentrations of cord blood cytokines IL-8 (P = 0.04) and G-CSF (P = 0.008). In vitro, recombinant-MBA variants elicited different cytokine responses and altered expression of NF-κB p65.Conclusion: This study demonstrates that size variation of the ureaplasma MBA protein modulates the host immune response in vivo and in vitro.

Biomechanical and morphological properties of the multiparous ovine vagina and effect of subsequent pregnancy

Pelvic floor soft tissues undergo changes during the pregnancy. However, the degree and nature of this process is not completely characterized. This study investigates the effect of subsequent pregnancy on biomechanical and structural properties of ovine vagina. Vaginal wall from virgin, pregnant (in their third pregnancy) and parous (one year after third vaginal delivery) Swifter sheep (n=5 each) was harvested. Samples for biomechanics and histology, were cut in longitudinal axis (proximal and distal regions). Outcome measurements describing Young’s modulus, ultimate stress and elongation were obtained from stress-strain curves. For histology samples were stained with Miller's Elastica staining. Collagen, elastin and muscle cells and myofibroblasts contents were estimated, using image processing techniques. Statistical analyses were performed in order to determine significant differences among experimental groups. Significant regional differences were identified. The proximal vagina was stiffer than distal, irrespective the reproductive status. During the pregnancy proximal vagina become more compliant than in parous (+47.45%) or virgin sheep (+64.35%). This coincided with lower collagen (−15 to −21%), higher elastin (+30 to +60%), and more smooth muscle cells (+17 to +37%). Vaginal tissue from parous ewes was weaker than of virgins, coinciding with lower collagen (−10%), higher elastin (+50%), more smooth muscle cells (+20%). It could be proposed that after pregnancy biomechanical properties of vagina do not recover to those of virgins. Since elastin has a significant influence on the compliance of soft tissues and collagen is the main “actor” regarding strength, histological analysis performed in this study justifies the mechanical behavior observed.

Erratum: Uterine artery leptin receptors during the ovarian cycle and pregnancy regulate angiogenesis in ovine uterine artery endothelial cells.

Leptin regulates body weight, reproductive functions, blood pressure, endothelial function, and fetoplacental angiogenesis. Compared to the luteal phase, the follicular phase and pregnancy are physiological states of elevated estrogen, angiogenesis, and uterine blood flow (UBF). Little is known concerning regulation of uterine artery (UA) angiogenesis by leptin and its receptors. We hypothesized that (1) ex vivo expression of leptin receptors (LEPR) in UA endothelium (UAendo) and UA vascular smooth muscle (UAvsm) is elevated in pregnant versus nonpregnant (Luteal and Follicular) sheep; (2) in vitro leptin treatments differentially modulate mitogenesis in uterine artery endothelial cells from pregnant (P-UAECs) more than in nonpregnant (NP-UAECs) ewes; and (3) LEPR are upregulated in P-UAECs versus NP-UAECs in association with leptin activation of phospho-STAT3 signaling. Local UA adaptations were evaluated using a unilateral pregnant sheep model where prebreeding uterine horn isolation (nongravid) restricted gravidity to one horn. Immunolocalization revealed LEPR in UAendo and UAvsm from pregnant and nonpregnant sheep. Contrary to our hypothesis, western analysis revealed that follicular UAendo and UAvsm LEPR were greater than luteal, nongravid, gravid, and control pregnant. Compared to pregnant groups, LEPR were elevated in renal artery endothelium of follicular and luteal sheep. Leptin treatment significantly increased mitogenesis in follicular phase NP-UAECs and P-UAECs, but not luteal phase NP-UAECs. Although UAEC expression of LEPR was similar between groups, leptin treatment only activated phospho-STAT3 in follicular NP-UAECs and P-UAECs. Thus, leptin may play an angiogenic role particularly in preparation for the increased UBF during the periovulatory period and subsequently to meet the demands of the growing fetus.

Human Amnion Epithelial Cells Protect against White Matter Brain Injury after Repeated Endotoxin Exposure in the Preterm Ovine Fetus

Intrauterine inflammation is a significant cause of injury to the developing fetal brain. Using a preterm fetal sheep model of in utero infection, we asked whether human amnion epithelial cells (hAECs) were able to reduce inflammation-induced fetal brain injury. Surgery was undertaken on pregnant sheep at ~105 days gestation (term is 147 days) for implantation of vascular catheters. Lipopolysaccharide (LPS; 150 ng/kg bolus) or saline was administered IV at 109, 110, and 111 days. Sixty million fluorescent-labeled hAECs were administered at 110, 111, and 112 days gestation via the brachial artery catheter. Brains were collected at 114 days for histological assessment. hAECs were observed within the cortex, white matter, and hippocampus. Compared to control lambs, LPS administration was associated with significant and widespread fetal brain inflammation and injury as evidenced by increased number of activated microglia in the periventricular white matter (p = 0.02), increased pyknosis, cell degeneration (p = 0.01), and a nonsignificant trend of fewer oligodendrocytes in the subcortical and periventricular white matter. Administration of hAECs to LPS-treated animals was associated with a significant mitigation in both inflammation and injury as evidenced by fewer activated microglia (p = 0.03) and pyknotic cells (p = 0.03), significantly more oligodendrocytes in the subcortical and periventricular white matter (p = 0.01 and 0.02, respectively), and more myelin basic protein-positive cells within the periventricular white matter (p = 0.02). hAEC administration to fetal sheep exposed to multiple doses of LPS dampens the resultant fetal inflammatory response and mitigates associated brain injury.

Thermal comfort indexes and physiological parameters of Santa Inês and crossbreed ewes in the semi-arid

The objective of this study was evaluated the thermal comfort indexes and physiological parameters in Santa Ines and crossbreed ewes in Brazilian semiarid. The research was developed at experimental station Benjamin Maranhao, twenty non pregnant female sheep were used, 10 pure and 10 crossbreed (½ Dorper + ½ Santa Ines), averaging 26.7 ± 2.1 kg and with 150 days of age at the beginning of the study and confined in a covered sheepfold. A completely randomized design in a 2 x 2 factorial scheme (two genotypes and two shifts) with four treatments and five repetitions was used. The interior and exterior thermal comfort indexes were, most of the day, above the thermal comfort zone for sheep. The rectal temperature, superficial temperature, respiratory rate and cardiac rate was more elevated in the afternoon, and the Santa Ines animals showed in the mornings low rectal temperature values, as well as superficial temperature, respiratory and cardiac rate when compared to the crossbreed.

Reduced fetal vitamin D status by maternal undernutrition during discrete gestational windows in sheep.

Placental transport of vitamin D and other nutrients (e.g. amino acids, fats and glucose) to the fetus is sensitive to maternal and fetal nutritional cues. We studied the effect of maternal calorific restriction on fetal vitamin D status and the placental expression of genes for nutrient transport [aromatic T-type amino acid transporter-1 (TAT-1); triglyceride hydrolase/lipoprotein uptake facilitator lipoprotein lipase (LPL)] and vitamin D homeostasis [CYP27B1; vitamin D receptor (VDR)], and their association with markers of fetal cardiovascular function and skeletal muscle growth. Pregnant sheep received 100% total metabolizable energy (ME) requirements (control), 40% total ME requirements peri-implantation [PI40, 1-31 days of gestation (dGA)] or 50% total ME requirements in late gestation (L, 104-127 dGA). Fetal, but not maternal, plasma 25-hydroxy-vitamin D (25OHD) concentration was lower in PI40 and L maternal undernutrition groups (P<0.01) compared with the control group at 0.86 gestation. PI40 group placental CYP27B1 messenger RNA (mRNA) levels were increased (P<0.05) compared with the control group. Across all groups, higher fetal plasma 25OHD concentration was associated with higher skeletal muscle myofibre and capillary density (P<0.05). In the placenta, higher VDR mRNA levels were associated with higher TAT-1 (P<0.05) and LPL (P<0.01) mRNA levels. In the PI40 maternal undernutrition group only, reduced fetal plasma 25OHD concentration may be mediated in part by altered placental CYP27B1. The association between placental mRNA levels of VDR and nutrient transport genes suggests a way in which the placenta may integrate nutritional cues in the face of maternal dietary challenges and alter fetal physiology.

Does maternal-fetal transfer of creatine occur in pregnant sheep?

Our aim was to determine the disposition of creatine in ovine pregnancy and whether creatine is transferred across the placenta from mother to fetus. Pregnant ewes received either 1) a continuous intravenous infusion of creatine monohydrate or saline from 122 to 131 days gestation, with maternal and fetal arterial blood and amniotic fluid samples collected daily for creatine analysis and fetal tissues collected at necropsy at 133 days for analysis of creatine content, or 2) a single systemic bolus injection of [13C]creatine monohydrate at 130 days of gestation, with maternal and fetal arterial blood, uterine vein blood, and amniotic fluid samples collected before and for 4 h after injection and analyzed for creatine, creatine isotopic enrichment, and guanidinoacetic acid (GAA; precursor of creatine) concentrations. Presence of the creatine transporter-1 (SLC6A8) and l-arginine:glycine amidinotransferase (AGAT; the enzyme synthesizing GAA) proteins were determined by Western blots of placental cotyledons. The 10-day creatine infusion increased maternal plasma creatine concentration three- to fourfold (P < 0.05) without significantly changing fetal arterial, amniotic fluid, fetal tissues, or placental creatine content. Maternal arterial 13C enrichment was increased (P < 0.05) after bolus [13C]creatine injection without change of fetal arterial 13C enrichment. SLC6A8 and AGAT proteins were identified in placental cotyledons, and GAA concentration was significantly higher in uterine vein than maternal artery plasma. Despite the presence of SLC6A8 protein in cotyledons, these results suggest that creatine is not transferred from mother to fetus in near-term sheep and that the ovine utero-placental unit releases GAA into the maternal circulation.

The fascinating story of surfactant.

The fascinating story of surfactant.

Effect of Hypoxemia on Fetal Ventricular Deformation in a Chronically Instrumented Sheep Model

We hypothesized that in near-term sheep fetuses, hypoxemia changes myocardial function as reflected in altered ventricular deformation on speckle-tracking echocardiography. Fetuses in 21 pregnant sheep were instrumented. After 4 d of recovery, fetal cardiac function was assessed by echocardiography at baseline, after 30 and 120 min of induced fetal hypoxemia and after its reversal. Left (LV) and right (RV) ventricular cardiac output and myocardial strain were measured. Baseline mean (standard deviation [SD]) LV and RV global longitudinal strains were −18.7% (3.8) and −14.3% (5.3). Baseline RV global longitudinal and circumferential deformations were less compared with those of the left ventricle (p = 0.016 and p < 0.005). LV, but not RV, global longitudinal strain was decreased (p = 0.003) compared with baseline with hypoxemia. Circumferential and radial strains did not exhibit significant changes. In the near-term sheep fetus, LV global longitudinal and circumferential strains are more negative than RV strains. Acute hypoxemia leads to LV rather than RV dysfunction as reflected by decreased deformation.

Uterine artery leptin receptors during the ovarian cycle and pregnancy regulate angiogenesis in ovine uterine artery endothelial cells†.

Leptin regulates body weight, reproductive functions, blood pressure, endothelial function, and fetoplacental angiogenesis. Compared to the luteal phase, the follicular phase and pregnancy are physiological states of elevated estrogen, angiogenesis, and uterine blood flow (UBF). Little is known concerning regulation of uterine artery (UA) angiogenesis by leptin and its receptors. We hypothesized that (1) ex vivo expression of leptin receptors (LEPR) in UA endothelium (UAendo) and UA vascular smooth muscle (UAvsm) is elevated in pregnant versus nonpregnant (Luteal and Follicular) sheep; (2) in vitro leptin treatments differentially modulate mitogenesis in uterine artery endothelial cells from pregnant (P-UAECs) more than in nonpregnant (NP-UAECs) ewes; and (3) LEPR are upregulated in P-UAECs versus NP-UAECs in association with leptin activation of phospho-STAT3 signaling. Local UA adaptations were evaluated using a unilateral pregnant sheep model where prebreeding uterine horn isolation (nongravid) restricted gravidity to one horn. Immunolocalization revealed LEPR in UAendo and UAvsm from pregnant and nonpregnant sheep. Contrary to our hypothesis, western analysis revealed that follicular UAendo and UAvsm LEPR were greater than luteal, nongravid, gravid, and control pregnant. Compared to pregnant groups, LEPR were elevated in renal artery endothelium of follicular and luteal sheep. Leptin treatment significantly increased mitogenesis in follicular phase NP-UAECs and P-UAECs, but not luteal phase NP-UAECs. Although UAEC expression of LEPR was similar between groups, leptin treatment only activated phospho-STAT3 in follicular NP-UAECs and P-UAECs. Thus, leptin may play an angiogenic role particularly in preparation for the increased UBF during the periovulatory period and subsequently to meet the demands of the growing fetus.