A. L. Fowden

Imprinted Genes, Placental Development and Fetal Growth

In mammals, imprinted genes have an important role in feto-placental development. They affect the growth, morphology and nutrient transfer capacity of the placenta and, thereby, control the nutrient supply for fetal growth. In particular, the reciprocally imprinted Igf2–H19 gene complex has a central role in these processes and matches the placental nutrient supply to the fetal nutrient demands for growth. Comparison of Igf2P0 and complete Igf2 null mice has shown that interplay between placental and fetal Igf2 regulates both placental growth and nutrient transporter abundance. In turn, epigenetic modification of imprinted genes via changes in DNA methylation may provide a mechanism linking environmental cues to placental phenotype, with consequences for development both before and after birth. Changes in expression of imprinted genes, therefore, have major implications for developmental programming and may explain the poor prognosis of the infant born small for gestational age and the wide spectrum of adult-onset diseases that originate in utero.

Programming placental nutrient transport capacity

Many animal studies and human epidemiological findings have shown that impaired growth in utero is associated with physiological abnormalities in later life and have linked this to tissue programming during suboptimal intrauterine conditions at critical periods of development. However, few of these studies have considered the contribution of the placenta to the ensuing adult phenotype. In mammals, the major determinant of intrauterine growth is the placental nutrient supply, which, in turn, depends on the size, morphology, blood supply and transporter abundance of the placenta and on synthesis and metabolism of nutrients and hormones by the uteroplacental tissues. This review examines the regulation of placental nutrient transfer capacity and the potential programming effects of nutrition and glucocorticoid over‐exposure on placental phenotype with particular emphasis on the role of the Igf2 gene in these processes.

The Placenta and Intrauterine Programming

Intrauterine programming is the process by which the structure and function of tissues are altered permanently by insults acting during early development. In mammals, the placenta controls intrauterine development by supplying oxygen and nutrients, and by regulating the bioavailability of specific hormones involved in foetal growth and development. Consequently, the placenta is likely to have a key role in mediating the programming effects of suboptimal conditions during development. This review examines placental phenotype in different environmental conditions and places particular emphasis on regulation of placental nutrient transfer capacity and endocrine function by insults known to cause intrauterine programming. More specifically, it examines the effects of a range of environmental challenges on the size, morphology, blood flow and transporter abundance of the placenta and on its rate of consumption and production of nutrients. In addition, it considers the role of hormone synthesis and metabolism by the placenta in matching intrauterine development to the prevailing environmental conditions. The adaptive responses that the placenta can make to compensate for suboptimal conditions in utero are also assessed in relation to the strategies adopted to maximise foetal growth and viability at birth. Environmentally‐induced changes in placental phenotype may provide a mechanism for transmitting the memory of early events to the foetus later in gestation, which leads to intrauterine programming of tissue development long after the original insult.

Adaptations in placental phenotype support fetal growth during undernutrition of pregnant mice

Undernutrition during pregnancy reduces birth weight and programmes adult phenotype with consequences for life expectancy, but its effects on the phenotype of the placenta, responsible for supplying nutrients for fetal growth, remain largely unknown. Using molecular, morphological and functional analyses, placental phenotype was examined in mice during restriction of dietary intake to 80% of control from day 3 of pregnancy. At day 16, undernutrition reduced placental, but not fetal, weight in association with decreased junctional zone volume and placental expression of glucose transporter Slc2a1. At day 19, both placental and fetal weights were reduced in undernourished mice (91% and 87% of control, respectively, P < 0.01), as were the volume and surface area of the labyrinthine zone responsible for placental nutrient transfer (85% and 86%, respectively, P < 0.03). However, unidirectional materno‐fetal clearance of tracer glucose was maintained and methyl‐aminoisobutyric acid increased 166% (P < 0.005) per gram of undernourished placenta, relative to controls. This was associated with an 18% and 27% increased placental expression of glucose and system A amino acid transporters Slc2a1 and Slc38a2, respectively, at day 19 (P < 0.04). At both ages, undernutrition decreased expression of the placental specific transcript of the Igf2 gene by 35% (P < 0.01), although methylation of its promoter was unaffected. The placenta, therefore, adapts to help maintain fetal growth when its own growth is compromised by maternal undernutrition. Consequently, placental phenotype is responsive to environmental conditions and may help predict the risk of adult disease programmed in utero.

Thyroid hormones in fetal growth and prepartum maturation

The thyroid hormones, thyroxine (T4) and triiodothyronine (T3), are essential for normal growth and development of the fetus. Their bioavailability in utero depends on development of the fetal hypothalamic-pituitary-thyroid gland axis and the abundance of thyroid hormone transporters and deiodinases that influence tissue levels of bioactive hormone. Fetal T4 and T3 concentrations are also affected by gestational age, nutritional and endocrine conditions in utero, and placental permeability to maternal thyroid hormones, which varies among species with placental morphology. Thyroid hormones are required for the general accretion of fetal mass and to trigger discrete developmental events in the fetal brain and somatic tissues from early in gestation. They also promote terminal differentiation of fetal tissues closer to term and are important in mediating the prepartum maturational effects of the glucocorticoids that ensure neonatal viability. Thyroid hormones act directly through anabolic effects on fetal metabolism and the stimulation of fetal oxygen consumption. They also act indirectly by controlling the bioavailability and effectiveness of other hormones and growth factors that influence fetal development such as the catecholamines and insulin-like growth factors (IGFs). By regulating tissue accretion and differentiation near term, fetal thyroid hormones ensure activation of physiological processes essential for survival at birth such as pulmonary gas exchange, thermogenesis, hepatic glucogenesis, and cardiac adaptations. This review examines the developmental control of fetal T4 and T3 bioavailability and discusses the role of these hormones in fetal growth and development with particular emphasis on maturation of somatic tissues critical for survival immediately at birth.

The effects of birth weight and postnatal growth patterns on fat depth and plasma leptin concentrations in juvenile and adult pigs

Low birth weight is associated with altered adipose tissue deposition and regulation of leptin production. This study determined the effects of naturally occurring variations in birth weight in pigs on postnatal growth patterns, body fat depth and plasma leptin and other hormone concentrations. Low (< 1.47 kg) and high (> 1.53 kg) birth weight piglets were studied at 3 months (juvenile; n= 47) and 12 months of age (young adult; n= 17). At each age, arterial and venous catheters were inserted under general anaesthesia. Plasma leptin, cortisol, glucose, insulin and catecholamine concentrations were determined in basal blood samples. Body fat depth was measured by ultrasound at 12 months of age. Overall, adult fat depth was greater in low compared to high birth weight pigs and increased fat depth was associated with thinness at birth and poor early growth rates. These effects were strongest in females. Fat depth was related to current weight only in males. Compared to high birth weight pigs, plasma leptin concentrations were reduced in low birth weight females at 3 months and in low birth weight males at 12 months of age. This study demonstrates sex‐specific effects of low birth weight on postnatal growth and body fatness and on plasma leptin concentrations in pigs.

Imprinted genes and the epigenetic regulation of placental phenotype.

Imprinted genes are expressed in a parent-of-origin manner by epigenetic modifications that silence either the paternal or maternal allele. They are widely expressed in fetal and placental tissues and are essential for normal placental development. In general, paternally expressed genes enhance feto-placental growth while maternally expressed genes limit conceptus growth, consistent with the hypothesis that imprinting evolved in response to the conflict between parental genomes in the allocation of maternal resources to fetal growth. Using targeted deletion, uniparental duplication, loss of imprinting and transgenic approaches, imprinted genes have been shown to determine the transport capacity of the definitive mouse placenta by regulating its growth, morphology and transporter abundance. Imprinted genes in the placenta are also responsive to environmental challenges and adapt placental phenotype to the prevailing nutritional conditions, in part, by varying their epigenetic status. In addition, interplay between placental and fetal imprinted genes is important in regulating resource partitioning via the placenta both developmentally and in response to environmental factors. By balancing the opposing parental drives on resource allocation with the environmental signals of nutrient availability, imprinted genes, like the Igf2-H19 locus, may act as nutrient sensors and optimise the fetal acquisition of nutrients for growth. These genes, therefore, have a major role in the epigenetic regulation of placental phenotype with long term consequences for the developmental programming of adult health and disease.

Effects of prevailing hypoxaemia, acidaemia or hypoglycaemia upon the cardiovascular, endocrine and metabolic responses to acute hypoxaemia in the ovine fetus

Although it is established that the fetus can successfully withstand a single, acute hypoxaemic challenge during gestation, little is known about what effects prevailing adverse intrauterine conditions might have on the fetal response to acute hypoxaemia. The aims of this study were therefore: (1) to characterise the effects of prevailing and sustained hypoxaemia, acidaemia or hypoglycaemia on the fetal cardiovascular responses to an episode of acute hypoxaemia; and (2) to determine the effects of these adverse intrauterine conditions on mechanisms mediating these cardiovascular responses. Thirty‐three Welsh Mountain sheep fetuses were chronically instrumented (1–2 % halothane) between 117 and 125 days of gestation (term is ca 145 days) with amniotic and vascular catheters and with a transit‐time flow probe around a femoral artery. The animals were divided retrospectively into four groups based upon post‐surgical, sustained, basal blood oxygen (chronically hypoxaemic; Pa,O2, 17.3 ± 0.5 mmHg; n= 8), glucose (chronically hypoglycaemic; blood glucose, 0.49 ± 0.03 mmol l−1; n= 6) and acid‐base (chronically acidaemic; pHa, 7.25 ± 0.01; n= 5) status. Values for compromised fetuses were −2 s.d. from a group of control (n= 14) fetuses. At 130 ± 4 days, a 1 h episode of acute, isocapnic hypoxaemia (9 % O2 in N2, to reduce carotid Pa,O2 to 12 ± 1 mmHg) was induced in all fetuses by reducing the maternal inspired O2 fraction (FI,O2). Fetal cardiovascular variables were recorded at 1 s intervals throughout the experimental protocol and arterial blood samples taken at appropriate intervals for biophysical (blood gases, glucose, lactate) and endocrine (catecholamines, vasopressin, cortisol, ACTH) measures. During acute hypoxaemia all fetuses elicited hypertension, bradycardia and femoral vasoconstriction. However, prevailing fetal compromise altered the cardiovascular and endocrine responses to a further episode of acute hypoxaemia, including: (1) enhanced pressor and femoral vasoconstriction; (2) greater increments in plasma noradrenaline and vasopressin during hypoxaemia; and (3) basal upward resetting of hypothalamic‐pituitary‐adrenal axis function. Only chronically hypoxaemic fetuses had significantly elevated basal concentrations of noradrenaline and enhanced chemoreflex function during acute hypoxaemia. These data show that prevailing adverse intrauterine conditions alter the capacity of the fetus to respond to a subsequent episode of acute hypoxaemia; however, the partial contributions of hypoxaemia, acidaemia or hypoglycaemia to mediating these responses can vary.

Disproportional effects of Igf2 knockout on placental morphology and diffusional exchange characteristics in the mouse

Both complete knockout of the Igf2 gene (Igf2null+/−) and knockout of its placental specific transcript alone (Igf2P0+/−) lead to fetal growth restriction in mice. However, in the Igf2null+/− this growth restriction occurs concurrently in gestation with placental growth restriction, whereas, placental growth restriction precedes fetal growth restriction in the Igf2P0+/− mouse. Previous studies have shown that the Igf2P0+/− placenta has proportionate reductions in its cellular compartments and its diffusional exchange characteristics. Yet, nothing is known about the structural development or diffusional exchange characteristics of the Igf2null+/− mouse. Hence, this study compares the structural properties (using stereology) and diffusional exchange characteristics (using measurement of permeability–surface area product, P.S, of three inert hydrophilic tracers) of the Igf2null+/− and the Igf2P0+/− placenta to identify the role of Igf2 in the development of the labyrinthine exchange membrane and its functional consequences. Our data show disproportionate effects of complete Igf2 ablation on the compartments of the placenta, not seen when the placental‐specific transcript alone is deleted. Furthermore, although the theoretical diffusing capacity (calculated from the stereological data) of the Igf2null+/− placenta was reduced relative to control, there was no effect of the complete knockout on permeability surface area available for small hydrophilic tracers. This is in contrast to the Igf2P0+/− placenta, where theoretical diffusion capacity and P.S values were reduced similarly. Total ablation of the Igf2 gene from the fetoplacental unit in the mouse therefore results in a disproportionate growth of placental compartments whereas, deleting the placental specific transcript of Igf2 alone results in proportional placental growth restriction. Thus, placental phenotype depends on the degree of Igf2 gene ablation and the interplay between placental and fetal Igf2 in the mouse.

The effects of birth weight on basal cardiovascular function in pigs at 3 months of age

In man, epidemiological studies have shown that low birth weight (BW) is associated with an increased risk of cardiovascular disease in later life. In this study, the long‐term consequences of variations in natural BW on basal cardiovascular function were investigated in pigs at 3 months of postnatal age. Low (< 1.41 kg; n= 20) and high (> 1.52 kg; n= 20) BW Large White piglets were selected from a total of 12 litters for study at 3 months of age. Basal mean arterial pressure (MAP) and heart rate (HR) were recorded for ∼30 min using standard recording equipment and basal arterial blood samples were taken for hormone analyses. Concentrations of angiotensin‐converting enzyme (ACE) were also measured in kidney, lung and plasma. Basal MAP, but not HR, in 3‐month‐old pigs was significantly inversely related to BW and positively related to the ratio of head length to BW. Postnatal growth rate of low BW pigs was slower than that of high BW pigs such that low BW piglets remained significantly smaller at 3 months of age. There were no differences in basal plasma adrenaline or cortisol concentrations between low and high BW pigs. However, basal plasma noradrenaline concentrations were significantly elevated in low BW compared to high BW pigs. Renal and pulmonary ACE levels were significantly reduced in low BW compared to high BW pigs. These data show that basal MAP in 3‐month‐old pigs is negatively associated with BW and positively correlated to disproportionate size at birth. This effect was associated with an increase in basal plasma noradrenaline concentrations.