Low birthweight is a risk factor for neonatal mortality and adverse metabolic health, both of which are associated with inadequate prenatal adipose tissue development. In the present study, we ...investigated the impact of maternal undernutrition on the expression of genes that regulate fetal perirenal adipose tissue (PAT) development and function at gestation days 89 and 130 (term=145 days). Singleton fetuses were taken from adolescent ewes that were either fed control (C) intake to maintain adiposity throughout pregnancy or were undernourished (UN) to maintain conception weight but deplete maternal reserves (n=7/group). Fetal weight was independent of maternal intake at day 89, but by day 130, fetuses from UN dams were 17% lighter and had lower PAT mass that contained fewer unilocular adipocytes. Relative PAT expression of IGF1, IGF2, IGF2R and peroxisome proliferator-activated receptor gamma (PPARG) mRNA was lower in UN than in controls, predominantly at day 89. Independent of maternal nutrition, PAT gene expression of PPARG, glycerol-3-phosphate dehydrogenase, hormone sensitive lipase, leptin, uncoupling protein 1 and prolactin receptor increased, whereas IGF1, IGF2, IGF1R and IGF2R decreased between days 89 and 130. Fatty acid synthase and lipoprotein lipase (LPL) mRNAs were not influenced by nutrition or stage of pregnancy. Females had greater LPL and leptin mRNA than males, and LPL, leptin and PPARG mRNAs were decreased in UN at day 89 in females only. PAT gene expression correlations with PAT mass were stronger at day 89 than they were at day 130. These data suggest that the key genes that regulate adipose tissue development and function are active beginning in mid-gestation, at which point they are sensitive to maternal undernutrition: this leads to reduced fetal adiposity by late pregnancy.
To establish the basis for altered placental development and function previously observed at late gestation, fetoplacental growth and placental vascular development were measured at three stages of ...gestation in a nutritional paradigm of compromised pregnancy. Singleton pregnancies to a single sire were established and thereafter adolescent ewes were offered an optimal control (C) or a high (H) dietary intake. At day 50, the H group had elevated maternal insulin and amniotic glucose, whereas mass of the fetus and placenta were unaltered. At day 90, the H group exhibited elevated maternal insulin, IGF1 and glucose; fetal weight and glucose concentrations in H were increased relative to C, but placental weight was independent of nutrition. By day 130, total placentome weight in the H group was reduced by 46% and was associated with lower fetal glucose and a 20% reduction in fetal weight. As pregnancy progressed from day 50 to 130, the parameters of vascular development in the maternal and fetal components of the placenta increased. In the fetal cotyledon, high dietary intakes were associated with impaired vascular development at day 50 and an increase in capillary number at day 90. At day 130, all vascular indices were independent of nutrition. Thus, high dietary intakes to promote rapid maternal growth influence capillary development in the fetal portion of the placenta during early to mid-pregnancy and may underlie the subsequent reduction in placental mass and hence fetal nutrient supply observed during the final third of gestation.
Both high and low maternal dietary intakes adversely affect fetal nutrient supply in adolescent sheep pregnancies. Aims were: (a) to assess the impact of prenatal nutrition on pregnancy outcome, ...offspring growth and offspring glucose metabolism and (b) to determine whether the offspring metabolic phenotype could then be altered by modifying postnatal nutrition. Dams carrying a single fetus were offered either an optimal control (C) intake to maintain adiposity throughout pregnancy, undernourished to maintain weight at conception but deplete maternal reserves (UN), or overnourished to promote rapid maternal growth and adiposity (ON). Placental weight and gestation length were reduced in ON dams and lamb birth weights were C>UN>ON (P < 0·001). All offspring were fed ad libitum from weaning to 6 months of age. ON offspring exhibited rapid catch-up growth and had increased fasting glucose and relative glucose intolerance compared with C offspring (P < 0·05). Irrespective of prenatal diet and sex, birth weight correlated negatively with these indices of glucose metabolism. From 7 to 12 months offspring either had continued ad libitum diet (ADLIB; to induce an obesogenic state) or a decreased ration appropriate for normal growth (NORM). At 12 months, the negative relationship between birth weight and indices of glucose metabolism persisted in ADLIB females (for example, fasting glucose, r - 0·632; P < 0·03) but was absent in NORM females and in both male groups. Therefore, low-birth-weight offspring from differentially achieved prenatal malnutrition exhibit an early adverse metabolic phenotype, and this can apparently be ameliorated by postnatal nutrition in females but not in males.
Uterine artery (UtA) adenovirus (Ad) vector-mediated overexpression of vascular endothelial growth factor (VEGF) enhances uterine blood flow in normal sheep pregnancy and increases fetal growth in ...the overnourished adolescent sheep model of fetal growth restriction (FGR). Herein, we examined its impact on gestation length, neonatal survival, early postnatal growth and metabolism. Singleton-bearing ewes were evenly allocated to receive Ad.VEGF-A165 (5 × 1010 particles/ml, 10 ml, n = 17) or saline (10 ml, n = 16) injected into each UtA at laparotomy (0.6 gestation). Fetal growth was serially monitored (blind) by ultrasound until delivery. Lambs were weighed and blood was sampled weekly and a glucose tolerance test performed (68-day postnatal age). Hepatic DNA/RNA was extracted at necropsy (83-day postnatal age) to examine methylation status of eight somatotropic axis genes. IGF1 mRNA and protein expression were measured by RT-PCR and radioimmunoassay, respectively. All pregnancies remained viable following Ad.VEGF-A165 treatment. Fetal abdominal circumference and renal volume were greater in the Ad.VEGF-A165 group compared with the saline group at 21/28 days (P ≤ 0.04) postinjection. At delivery, gestation length (P = 0.07), lamb birthweight (P = 0.08), umbilical girth (P = 0.06), and plasma glucose (P = 0.09) tended to be greater in Ad.VEGF-A165-treated lambs. Levels of neonatal intervention required to ensure survival was equivalent between groups. Absolute postnatal growth rate (P = 0.02), insulin area under the curve (P = 0.04) and carcass weight at necropsy (P = 0.04) were increased by Ad.VEGF-A165 treatment. There was no impact on markers of insulin sensitivity or methylation/expression of key genes involved in somatic growth. Ad.VEGF-A165 gene therapy increased fetal growth in a sheep FGR model, and lambs continued to thrive during the neonatal and early postnatal period.
In the overnourished adolescent sheep, maternal tissue synthesis is promoted at the expense of placental growth and leads to a major decrease in lamb birth weight at term. Maternal growth hormone ...(GH) concentrations are attenuated in these pregnancies, and it was recently demonstrated that exogenous GH administration throughout the period of placental proliferation stimulates uteroplacental and fetal development by Day 81 of gestation. The present study aimed to determine whether these effects persist to term and to establish whether GH affects fetal growth and body composition by increasing placental size or by altering maternal metabolism. Adolescent recipient ewes were implanted with singleton embryos on Day 4 postestrus. Three groups of ewes offered a high dietary intake were injected twice daily with recombinant bovine GH from Days 35 to 65 of gestation (high intake plus early GH) or from Days 95 to 125 of gestation (high intake plus late GH) or remained untreated (high intake only). A fourth moderate-intake group acted as optimally nourished controls. Pregnancies were terminated at Day 130 of gestation (6 per group) or were allowed to progress to term (8-10 per group). GH administration elevated maternal plasma concentrations of GH, insulin, glucose, and nonesterified fatty acids during the defined treatment windows, while urea concentrations were decreased. At Day 130, GH treatment had reduced the maternal adiposity score, percentage of fat in the carcass, and internal fat depots and leptin concentrations, predominantly in the high-intake plus late GH group. Placental weight was lower in high-intake vs. control dams but independent of GH treatment. In contrast, fetal weight was elevated by late GH treatment, and these fetuses had higher relative carcass fat content, perirenal fat mass, and liver glycogen concentrations than all other groups. Expression of leptin mRNA in fetal perirenal fat and fetal plasma leptin concentrations were not significantly altered by maternal nutritional intake or GH. In pregnancies proceeding to term, the duration of gestation, fetal placental mass, and lamb birth weight were reduced in high-intake compared with control dams but were not significantly affected by GH treatment. In conclusion, exogenous GH has profound effects on maternal endocrinology, metabolism, and body composition when administered during early and late pregnancy. Treatment during late pregnancy has a modest effect on fetal growth independent of placental size and a profound effect on fetal adiposity, which may have implications beyond the fetal period.
Our aim was to identify which ultrasound parameters can be most accurately measured and best predict ovine fetal weight in late gestation. Singleton pregnancies were established using embryo transfer ...in 32 adolescent ewes, which were subsequently overnourished to produce fetuses of variable size (1720-6260 g). Ultrasound measurements at 126-133 days gestation were compared with fetal weight/biometry at late-gestation necropsy (n = 19) or term delivery (n = 13). Abdominal circumference (AC) and renal volume (RV) correlated best with physical measurements (r = 0.78-0.83) and necropsy/birth weight (r = 0.79-0.84). Combination of AC + RV produced an estimated fetal weight equation Log EFW = 2.115 + 0.003 AC + 0.12 RV - 0.005 RV(2) with highest adjusted R(2) (0.72) and lowest mean absolute/percentage prediction error (396-550 g/11.1%-13.2%). In conclusion, AC and RV are parameters of choice for assessment of late-gestation ovine fetal growth and can be used to estimate fetal weight with similar accuracy to human fetuses.
Intrauterine growth restriction (IUGR) is a risk factor for metabolic syndrome, notably when associated with rapid postnatal catch-up growth. A sheep paradigm was used to assess relationships between ...prenatal and early postnatal growth trajectories, metabolism and body composition. Singletons (single-sire embryo transfer from obese and control donors) were gestated and suckled by overnourished adolescent dams and categorised by birthweight as IUGR or normal (N). Gestation length was equivalent in both categories and all lambs were delivered spontaneously preterm (PT; mean (±s.e.m.) 139.8±1.7 days; term=145–147 days). The IUGR lambs were smaller at birth, but fractional growth rates (FGR) for eight anthropometry parameters were higher and independent of gender (except thorax girth; males (M)<females (F)). At Day 48, fasting glucose (IUGR>N; M>F) and first-phase insulin response (to 20min; IUGR<N; M<F) after glucose were influenced by prenatal growth and gender. Embryo donor adiposity influenced glucose tolerance only. Plasma insulin, insulin-like growth factor-1 (M>F) and leptin (M<F) were influenced by gender but not prenatal growth. At necropsy (Day 77), IUGR plus PT lambs had decreased carcass and visceral organ weights, but carcass composition was not different from N plus PT. In contrast, M were heavier, with lower internal fat mass, carcass fat percentage and perirenal fat leptin mRNA than F. Therefore, IUGR was associated with increased postnatal FGR and altered glucose handling, but, without absolute catch-up growth, gender had the predominant influence on postnatal leptinaemia and adiposity.
Ghrelin is an orexigenic hormone principally produced by the stomach, but also by numerous peripheral tissues including the placenta. Ghrelin acts via growth hormone secretagogue receptors (GHSR-1a) ...to alter food intake, fat utilization, and cellular proliferation, and has been suggested to play a role in the developmental growth of the fetoplacental unit. The placental expression of ghrelin and its role in ruminant species is not known. We tested the hypotheses that ghrelin and its functional receptor, GHSR-1a, are present in tissues of the ovine placenta, and that their expression is linked to the stage of development.
Antibodies raised against ghrelin and GHSR-1a were used in standard immunohistochemical protocols on placental tissues collected from pregnant ewes (n = 6 per gestational time point) at days 50, 80, 100, 128 and 135 of gestation (term approximately day 145). Immunostaining for ghrelin and GHSR-1a was quantified using computer-aided image analysis. Image analysis data were subjected to one-way ANOVA, with differences in immunostaining between time-points determined by Fisher's least significant difference.
Positive immunostaining for ghrelin was detected in ovine placentae at all gestational time points, with staining localized to the maternal epithelium, caruncle and trophectoderm. There was a significant effect of gestational age (p < 0.001) on the placental expression of ghrelin, with maximal levels at gestational day 80. GHSR-1a immunostaining was detected in the fetal trophectoderm at all time points. In contrast to the gestational pattern of ghrelin expression, there was no effect of gestational age on placental GHSR-1a immunoexpression.
Ghrelin and GHSR-1a are both present in the ovine placenta, and ghrelin displays a developmentally-related pattern of expression. Therefore, these data strongly suggest that the ghrelin system may have a role in feto-placental development in sheep.
Overnourishing the pregnant adolescent ewe promotes maternal tissue synthesis at the expense of placental growth and leads
to a major reduction in lamb birth weight at term. Growth hormone (GH) ...secretion is attenuated in these overnourished dams
and the maternal somatotrophic axis may play a key role in coordinating nutrient usage in the pregnant adolescent. Thus we
investigated whether increasing maternal GH during the period of rapid placental proliferation alters nutrient partitioning
between the maternal, placental, and fetal tissues as assessed at Day 81 of gestation. Adolescent recipient ewes were implanted
with singleton embryos, derived from superovulated dams and a single sire on Day 4 postestrus. Thereafter, the ewes were offered
either a high (H) or moderate intake (M) of the same complete diet. From Day 35 to 80 of gestation, ewes were either injected
twice daily (s.c. at 0800 and 1800 h) with recombinant bovine GH (bGH, 0.14 mg/kg live weight/day) or remained untreated (n
= 8 ewes per group). Maternal concentrations of GH, insulin, insulin-like growth factor (IGF-1), glucose, and non-esterified
fatty acids (NEFAs) were higher, and leptin secretion lower, in bGH-treated dams from both nutritional groups. Maternal body
weight gain was higher in H versus M groups and was independent of bGH treatment. Treatment with bGH reduced relative perirenal
and carcass fat deposition and increased carcass protein content in both H and M dams. Uteroplacental mass (uterus + placentomes
+ fetal membranes) averaged 1099, 1069, 1112, and 1754 g in M, H, M+GH, and H+GH groups. This significant increase in uteroplacental
development in the H+GH group was associated with higher fetal kidney and liver weights and elevated fetal insulin, glucose,
and lactate concentrations. Treatment with bGH also induced polyhydramnios in the H group. The transplacental glucose gradient
was increased twofold in the H+GH group but placental GLUT- 1 and GLUT-3 expression was unaffected. In conclusion, administration
of GH during the period of rapid placental proliferation alters endocrine status and thus nutrient partitioning in the overnourished
adolescent dam in favor of uteroplacental and fetal growth. It remains to be established whether these effects are due wholly
to alterations in maternal metabolism or if they also reflect an effect of bGH and/or the IGF system at the level of the uteroplacenta.
Overnourishing pregnant adolescent sheep promotes maternal growth but reduces placental mass, lamb birth weight and circulating progesterone. This study aimed to determine whether altered ...progesterone reflected transcript abundance for StAR (cholesterol transporter) and the steroidogenic enzymes (Cyp11A1, Hsd3b and Cyp17). Circulating and placental expression of ovine placental lactogen (oPL) was also investigated. Adolescent ewes with singleton pregnancies were fed high (H) or moderate (M) nutrient intake diets to restrict or support placental growth. Experiment 1: peripheral progesterone and oPL concentrations were measured in H (n=7) and M (n=6) animals across gestation (days 7-140). Experiment 2: progesterone was measured to mid- (day 81; M: n=11, H: n=13) or late gestation (day 130; M: n=21, H: n=22), placental oPL, StAR and steroidogenic enzymes were measured by qPCR and oPL protein by immunohistochemistry. Experiment 1: in H vs M animals, term placental (P<0.05), total cotyledon (P<0.01) and foetal size (P<0.05) were reduced. Circulating oPL and progesterone were reduced at mid- (P<0.001, P<0.01) and late gestation (P<0.01, P<0.05) and oPL detection was delayed (P<0.01). Experiment 2: placental oPL was not altered by nutrition. In day 81 H animals, progesterone levels were reduced (P<0.001) but not related to placental or foetal size. Moreover, placental steroidogenic enzymes were unaffected. Day 130 progesterone (P<0.001) and Cyp11A1 (P<0.05) were reduced in H animals with intrauterine growth restriction (H+IUGR). Reduced mid-gestation peripheral oPL and progesterone may reflect altered placental differentiation and/or increased hepatic clearance respectively. Restricted placental growth and reduced biosynthesis may account for reduced progesterone in day 130 H+IUGR ewes.
PDF
