Anne Jaquiery

Periconceptional Undernutrition of Ewes Impairs Glucose Tolerance in Their Adult Offspring

Maternal undernutrition throughout pregnancy can have long-term effects on the health of adult offspring. Undernutrition around the time of conception alters growth, metabolism, and endocrinology of the sheep fetus, but the impact on offspring after birth is largely unknown. We determined the effect of maternal periconceptional undernutrition in sheep on glucose tolerance in the offspring before and after puberty. Undernourished (UN) ewes were fed individually to maintain weight loss of 10–15% bodyweight from 61 d before until 30 d after mating. Offspring (24 UN, 30 control) underwent an i.v. glucose tolerance test at 4 and 10 mo of age. Glucose tolerance was similar in both groups at 4 mo. Insulin area under the curve increased by 33% between 4 and 10 mo (101 ± 8 versus 154 ± 12 ng · min · mL−1, p < 0.0001). At 10 mo, UN offspring had a 10% greater glucose area under the curve than controls (809 ± 22 versus 712 ± 20 mM · min, p < 0.01), a reduced first phase insulin response (p = 0.003) which was particularly apparent in females and in singletons, and a decreased insulin:glucose ratio (p = 0.01). We conclude that maternal undernutrition around the time of conception results in impaired glucose tolerance in postpubertal offspring.

Fetal exposure to excess glucocorticoid is unlikely to explain the effects of periconceptional undernutrition in sheep

Periconceptional undernutrition alters fetal growth, metabolism and endocrinology in late gestation. The underlying mechanisms remain uncertain, but fetal exposure to excess maternal glucocorticoids has been hypothesized. We investigated the effects of periconceptional undernutrition on maternal hypothalamic–pituitary–adrenal axis function and placental 11β‐hydroxysteroid dehydrogenase type 2 (11βHSD2) activity. Ewes received maintenance feed (N, n= 20) or decreased feed from −60 to +30 days from mating to achieve 15% weight loss after an initial 2‐day fast (UN, n= 21). Baseline plasma samples and arginine vasopressin (AVP)–corticotrophin‐releasing hormone (CRH) challenges were performed on days −61, −57, −29, −1, +29, 33, and 49 from mating (day 0). Maternal adrenal and placental tissue was collected at 50 days. Baseline plasma levels of adrenocorticotrophic hormone (ACTH) and cortisol decreased in the UN group (P < 0.0001). ACTH response to AVP–CRH was greater in UN ewes during undernutrition (P= 0.03) returning to normal levels after refeeding. Cortisol response to AVP–CRH was greater in UN ewes after the initial 2‐day fast, but thereafter decreased and was lower in UN ewes from mating until the end of the experiment (P= 0.007). ACTH receptor, StAR and p450c17 mRNA levels were down‐regulated in adrenal tissue from UN ewes. Placental 11βHSD2 activity was lower in UN than N ewes at 50 days (P= 0.014). Moderate periconceptional undernutrition results in decreased maternal plasma cortisol concentrations during undernutrition and after refeeding, and adrenal resistance to ACTH for at least 20 days after refeeding. Fetal exposure to excess maternal cortisol is unlikely during the period of undernutrition, but could occur later in gestation if maternal plasma cortisol levels return to normal while placental 11βHSD2 activity remains low.

Maternal undernutrition programs tissue-specific epigenetic changes in the glucocorticoid receptor in adult offspring

Epidemiological data indicate that an adverse maternal environment during pregnancy predisposes offspring to metabolic syndrome with increased obesity, and type 2 diabetes. The mechanisms are still unclear although epigenetic modifications are implicated and the hypothalamus is a likely target. We hypothesized that maternal undernutrition (UN) around conception in sheep would lead to epigenetic changes in hypothalamic neurons regulating energy balance in the offspring, up to 5 years after the maternal insult. We found striking evidence of decreased glucocorticoid receptor (GR) promoter methylation, decreased histone lysine 27 trimethylation, and increased histone H3 lysine 9 acetylation in hypothalami from male and female adult offspring of UN mothers. These findings are entirely compatible with the increased GR mRNA and protein observed in the hypothalami. The increased GR predicted the decreased hypothalamic proopiomelanocortin expression and increased obesity that we observed in the 5-year-old adult males. The epigenetic and expression changes in GR were specific to the hypothalamus. Hippocampal GR mRNA and protein were decreased in UN offspring, whereas pituitary GR was altered in a sex-specific manner. In peripheral polymorphonuclear leukocytes there were no changes in GR methylation or protein, indicating that this epigenetic analysis did not predict changes in the brain. Overall, these results suggest that moderate changes in maternal nutrition, around the time of conception, signal life-long and tissue-specific epigenetic alterations in a key gene regulating energy balance in the hypothalamus.

The effects of maternal nutrition around the time of conception on the health of the offspring.

The incidence of prematurity, diabetes and cardiovascular disease have been increasing in both the developed and developing world. Increasing numbers of human studies suggest that these serious health outcomes may have developmental origins originating from nutritional deficits in the periconceptional period, with maternal nutrition around the time of conception now shown to have important effects on the length of gestation, trajectory of fetal growth and on postnatal growth and health. Biomedical research using the pregnant sheep has been widely employed to gain a deeper understanding of the underlying mechanisms involved. There is growing awareness that this field of research has major implications for the livestock production industry. From our own studies on sheep we have evidence that maternal undernutrition during the periconceptional period results in altered fetal hypothalamic-pituitary-adrenal axis (HPAA) development, an increased rate of premature birth, altered fetal pancreatic function, insulin signalling and amino acid metabolism, and also alterations in maternal adaptation to pregnancy. We are currently studying the postnatal consequences of these changes. Other research groups have shown that restricted nutrition of sheep in the early part of pregnancy alters postnatal muscle development, fat deposition, cardiovascular regulation and HPAA function. One aim of this review is to illustrate how biomedical research using animals such as the sheep has been used to gain a better understanding of the consequences of reduced maternal nutrition during the periconceptional period. We suggest that there are equally important consequences of this research for the livestock production industries.

Nutritional programming of adult disease

Intrauterine and early neonatal life is a period of physiological plasticity, during which environmental influences may produce long-term effects. Both undernutrition and overnutrition during this period have been shown to change disease risk in adulthood. These effects are influenced by the type, timing and duration of inappropriate nutrition and by the previous nutritional environment and may not be reflected in changes in body size. An understanding of the interaction between nutrient imbalance and alteration of gene expression is likely to be the key to optimising future health outcomes.

Periconceptional Undernutrition in Sheep Affects Adult Phenotype Only in Males

Periconceptional undernutrition (PCUN) in sheep alters fetal growth and metabolism and postnatal growth regulation, but effects on adult body composition are unknown. We investigated the effects of PCUN on adult phenotype. Singleton lambs of ewes fed normally (N, n = 17) or undernourished before (UN-61-0 d, n = 23), before and after (UN-61-30 d, n = 19), or after (UN-2-30d, n = 17) mating (d0) were weighed at birth, 12 weeks, and intermittently to adulthood. At the age of 3-4 years, body composition was assessed by dual-emission X-ray absorptiometry followed by postmortem examination. Compared with N animals, male, but not female, offspring of all UN groups had greater % fat mass (all UN versus N: 9 ± 1 versus 2 ± 1%, P < 0.001) and perirenal fat (544 ± 36 versus 222 ± 44 g, P = 0.002), and proportionately smaller hearts (4.5 ± 0.1 versus 5.2 ± 0.2 g·kg−1), lungs (9.1 ± 0.2 versus 10.6 ± 0.5 g·kg−1), and adrenals (0.06 ± 0.002 versus 0.08 ± 0.003 g·kg−1). UN males also had larger testes (726 ± 21 versus 545 ± 32 g, P = 0.007), but UN females had smaller ovaries (2.7 ± 0.08 versus 3.4 ± 0.4 g, P = 0.01). Changes were independent of birth weight or postnatal growth velocity. Brief PCUN has sex-specific effects on adult phenotype, predominantly affecting males, which may contribute to adverse metabolic outcomes.

Size at birth and adult fat mass in twin sheep are determined in early gestation

•  Reduced size at birth and shorter gestation length are both associated with increased risks of non‐communicable diseases (NCD) in later adult life. •  Twins are born both smaller and earlier than singletons and adult twins also are reported to be at increased risk of common NCDs such as diabetes. •  The smaller size and shorter gestation length of twins has been presumed to be due to a lack of intrauterine space and/or limitations of placental nutrient supply in late gestation, but there are few data to support this. •  We show that size at birth and adult fat mass in twin sheep are determined largely in early gestation. •  Knowledge of the mechanisms underlying early pregnancy determination of fetal growth and gestation length in twins are likely to increase understanding of how early pregnancy factors influence lifelong health for offspring from all pregnancies.

Undernutrition Before Mating in Ewes Impairs the Development of Insulin Resistance During Pregnancy

OBJECTIVE: To assess the effects of periconceptional undernutrition on maternal adaptation of insulin-dependent metabolism during pregnancy. METHODS: Romney ewes were randomly assigned to receive normal nutrition (n=12) or undernutrition before (from 60 days before until mating, n=7), after (2 days before to 30 days after mating, n=6) or before and after mating (from 60 days before to 30 days after mating, n=10). Insulin sensitivity was measured by hyperinsulinaemic-euglycaemic clamp at 65 days of gestation (term=147 days). Lamb growth rate was measured in late gestation by chronically implanted growth catheters and weight at 133 days of gestation. RESULTS: Ewes undernourished before or both before and after mating failed to develop the insulin resistance of pregnancy seen in normally nourished ewes. Ewes undernourished after mating showed intermediate insulin sensitivity. This was not related to plasma concentrations of pregnancy-related hormones, but was related to insulin kinetics. There was an inverse relationship between insulin sensitivity and fetal growth, with ewes that were most insulin sensitive having smaller, more slowly growing lambs (highest compared with lowest tertile for insulin sensitivity: fetal weight 3.5±0.3 compared with 4.5±0.1 kg, P=.02; growth rate 2.0±0.2 compared with 2.6±0.2 mm.day-1, P=.05). CONCLUSION: Maternal undernutrition before conception impairs adaptation of insulin-related metabolism during pregnancy in ways that affect fetal growth. This suggests a key mechanism whereby prepregnancy nutritional status influences pregnancy outcome.

Do Alterations in Placental 11β-Hydroxysteroid Dehydrogenase (11βHSD) Activities Explain Differences in Fetal Hypothalamic-Pituitary-Adrenal (HPA) Function Following Periconceptional Undernutrition or Twinning in Sheep?

Periconceptional undernutrition (UN) in sheep accelerates fetal hypothalamic-pituitary-adrenal (HPA) axis activation, resulting in preterm birth. In contrast, twin conception suppresses fetal HPA function and delays prepartum HPA activation. We hypothesized that these dissimilar effects on fetal HPA activity result from different influences of maternal glucocorticoid (GC) on maturation of the fetal HPA axis, mediated via different activities of placental 11β-hydroxysteroid dehydrogenase (11βHSD) isozymes. We examined the effects of twinning and maternal periconceptional UN from 60 days before until 30 days after mating on the ontogeny of placental 11βHSD-1 and -2 enzyme activities. At day 85 of gestation, placental 11βHSD-2 activity was lower in UN than in normally nourished (N) fetuses (P < .05) and was higher in twins than in singletons (P < .05). Furthermore, placental 11βHSD-1 activity was not different between nutritional groups but was higher in twins than in singletons (P = .01). At day 85, fetal plasma cortisol (P < .001) and cortisone (P < .001) concentrations were lower in UN than in N fetuses, but the cortisol to cortisone ratio was higher in UN than in N fetuses (P = .01). There was no effect of fetus number on plasma cortisol or cortisone concentrations or on the ratio of cortisol to cortisone at day 85. Therefore, periconceptional UN and twinning may result in the alterations of placental 11βHSD isozyme activities at particular times during gestation. Changes in these activities during critical periods of fetal development could affect transplacental transfer or placental generation of GCs that reach the fetus, potentially influencing the timing of activation of the fetal HPA axis, fetal maturation, and hence the development and health later in life.

Periconceptional Events Perturb Postnatal Growth Regulation in Sheep

Periconceptional undernutrition and twin conception alter intrauterine growth and metabolism and are associated with later adverse metabolic outcomes. The contribution of postnatal growth to these outcomes is less well defined. We investigated whether maternal periconceptional undernutrition or twin conception altered postnatal growth regulation in ways that could lead to metabolic disease. Single and twin offspring of ewes undernourished (UN) from 61 d before until 30 d after mating, fed to achieve and maintain 10–15% weight loss (UN), were compared with offspring of maintenance-fed controls (N). At 2 h and 1, 6, and 12 wk after birth, lambs were weighed and plasma hormone and metabolite concentrations analyzed. Milk intake, measured by deuterium oxide dilution, was inversely related to birth weight only in N singles, although twins had the greatest postnatal growth velocity. Positive associations were seen between milk intake, growth velocity, and leptin concentrations in N, but not UN, offspring. We conclude that periconceptional undernutrition alters the relationships between regulators of postnatal growth, including nutrient intake and key hormonal axes, in both singles and twins without affecting size at birth or postnatal growth velocity. Dissociation of growth from its key regulators is one possible mechanism underlying adverse metabolic outcomes after periconceptional undernutrition.