Beverly S. Muhlhausler

Increased maternal nutrition alters development of the appetite-regulating network in the brain

Individuals exposed to an increased nutrient supply before birth have a high risk of becoming obese children and adults. It has been proposed that exposure of the fetus to high maternal nutrient intake results in permanent changes within the central appetite regulatory network. No studies, however, have investigated the impact of increased maternal nutrition on the appetite regulatory network in species in which this network develops before birth, as in the human. In the present study, pregnant ewes were fed a diet which provided 100% (control, n=8) or ∼160% (well‐fed, n=8) of metabolizable energy requirements. Ewes were allowed to lamb spontaneously, and lambs were sacrificed at 30 days of postnatal age. All fat depots were dissected and weighed, and expression of the appetite‐regulating neuropeptides and the leptin receptor (OBRb) were determined by in situ hybridization. Lambs of well‐fed ewes had higher glucose (Glc) concentrations during early postnatal life (F=5.93, P<0.01) and a higher relative subcutaneous (s.c.) fat mass at 30 days of age (34.9±4.7 g/kg vs. 22.8±3.3 g/kg;P<0.05). The hypothalamic expression of proopiomelanocortin was higher in lambs of well‐fed ewes (0.48±0.09 vs. 0.28±0.04, P<0.05). In lambs of over‐nourished mothers, but not in controls, the expression of OBRb was inversely related to total relative fat mass (r2=0.50, P=0.05, n=8), and the direct relationship between the expression of the central appetite inhibitor CART and fat mass was lost. The expression of neuropeptide Y and AGRP was inversely related to total relative fat mass (NPY, r2=0.28, P<0.05;agouti‐related peptide, r2=0.39, P<0.01). These findings suggest that exposure to increased nutrition before birth alters the responses of the central appetite regulatory system to signals of increased adiposity after birth.—Muhlhausler, B. S., Adam, C. L., Findlay, P. A., Duffield, J. A., and McMillen, I. C. Increased maternal nutrition alters development of the appetite‐regulating network in the brain. FASEB J. 20, E556–E565 (2006)

Early origins of obesity: programming the appetite regulatory system

There is evidence that changes in perinatal nutrition programme the development of relative fat mass and the regulation of appetite in adult life. These studies have been primarily in the rodent utilizing maternal overnutrition or undernutrition imposed at different stages of pregnancy and beyond, mapping of neuropeptide localization and activity and appropriate null mutant models. Whilst the rodent offers significant advantages in terms of a short gestation and the availability of useful transgenic and null mutant models, there are also advantages to using an animal model more akin to the human, in which all components of the ‘fat–brain axis’ are present before birth, such as the sheep. This review summarizes recent work on the expression and localization of the ‘appetite regulatory’ peptides in the fetal rodent and sheep hypothalamus and their potential role in the early programming of postnatal appetite and obesity.

Epigenetics and human obesity

Background:Recent technological advances in epigenome profiling have led to an increasing number of studies investigating the role of the epigenome in obesity. There is also evidence that environmental exposures during early life can induce persistent alterations in the epigenome, which may lead to an increased risk of obesity later in life.Method:This paper provides a systematic review of studies investigating the association between obesity and either global, site-specific or genome-wide methylation of DNA. Studies on the impact of pre- and postnatal interventions on methylation and obesity are also reviewed. We discuss outstanding questions, and introduce EpiSCOPE, a multidisciplinary research program aimed at increasing the understanding of epigenetic changes in emergence of obesity.Results:An electronic search for relevant articles, published between September 2008 and September 2013 was performed. From the 319 articles identified, 46 studies were included and reviewed. The studies provided no consistent evidence for a relationship between global methylation and obesity. The studies did identify multiple obesity-associated differentially methylated sites, mainly in blood cells. Extensive, but small, alterations in methylation at specific sites were observed in weight loss intervention studies, and several associations between methylation marks at birth and later life obesity were found.Conclusions:Overall, significant progress has been made in the field of epigenetics and obesity and the first potential epigenetic markers for obesity that could be detected at birth have been identified. Eventually this may help in predicting an individual’s obesity risk at a young age and opens possibilities for introducing targeted prevention strategies. It has also become clear that several epigenetic marks are modifiable, by changing the exposure in utero, but also by lifestyle changes in adult life, which implies that there is the potential for interventions to be introduced in postnatal life to modify unfavourable epigenomic profiles.

Maternal “junk-food” feeding of rat dams alters food choices and development of the mesolimbic reward pathway in the offspring

Individuals exposed to high‐fat, high‐sugar diets before birth have an increased risk of obesity in later life. Recent studies have shown that these offspring exhibit increased preference for fat, leading to suggestions that perinatal exposure to high‐fat, high‐sugar foods results in permanent changes within the central reward system that increase the subsequent drive to overconsume palatable foods. The present study has determined the effect of a maternal “junk‐food” diet on the expression of key components of the mesolimbic reward pathway in the offspring of rat dams at 6 wk and 3 mo of age. We show that offspring of junk‐food‐fed (JF) dams exhibit higher fat intake from weaning until at least 3 mo of age (males: 16±0.6 vs. 11±0.8 g/kg/d; females: 19±1.3 vs. 13±0.4 g/kg/d; P<0.01). mRNA expression of μ‐opioid receptor (Mu) was 1.6‐fold higher (P<0.01) and dopamine active transporter (DAT) was 2‐fold lower (P<0.05) in JF offspring at 6 wk of age. By 3 mo, these differences were reversed, and Mu mRNA expression was 2.8‐fold lower (P<0.01) and DAT mRNA expression was 1.9‐fold higher (P<0.01) in the JF offspring. These findings suggest that perinatal exposure to high‐fat, high‐sugar diets results in altered development of the central reward system, resulting in increased fat intake and altered response of the reward system to excessive junk‐food intake in postnatal life.—Ong, Z. Y., Muhlhausler, B. S. Maternal “junk‐food” feeding of rat dams alters food choices and development of the mesolimbic reward pathway in the offspring. FASEB J. 25, 2167‐2179 (2011). www.fasebj.org

Fetal growth restriction, catch-up growth and the early origins of insulin resistance and visceral obesity.

There is an association between growing slowly before birth, accelerated growth in early postnatal life and the emergence of insulin resistance, visceral obesity and glucose intolerance in adult life. In this review we consider the pathway through which intrauterine growth restriction (IUGR) leads to the initial increase in insulin sensitivity and to catch-up growth. We also discuss the importance of the early insulin environment in determining later visceral adiposity and the intrahepatic mechanisms that may result in the emergence of glucose intolerance in a subset of IUGR infants. We present evidence that a key fetal adaptation to poor fetal nutrition is an upregulation of the abundance of the insulin receptor in the absence of an upregulation of insulin signalling in fetal skeletal muscle. After birth, however, there is an upregulation in the abundance of the insulin receptor and the insulin signalling pathway in the IUGR offspring. Thus, the origins of the accelerated postnatal growth rate experienced by IUGR infants lie in the fetal adaptations to a poor nutrient supply. We also discuss how the intracellular availability of free fatty acids and glucose within the visceral adipocyte and hepatocyte in fetal and neonatal life are critical in determining the subsequent metabolic phenotype of the IUGR offspring. It is clear that a better understanding of the relative contributions of the fetal and neonatal nutrient environment to the regulation of key insulin signalling pathways in muscle, visceral adipose tissue and the liver is required to support the development of evidence-based intervention strategies and better outcomes for the IUGR infant.

Omega-3 long chain fatty acid synthesis is regulated more by substrate levels than gene expression

The conversion of linoleic acid (LA) and alpha-linolenic acid (ALA) to long chain polyunsaturated fatty acids (LCPUFA) is known to involve desaturation and elongation steps. Although there is evidence that genes for these steps can be regulated by extremes of dietary PUFA, the degree to which there is meaningful regulation of LCPUFA levels in tissues by diet as a result of changes in expression of desaturase and elongase genes is unclear. In this study, we tested the effect of increasing ALA levels in diets of rats from 0.2% to 2.9% energy (en) against a constant LA level (1%en) on plasma and liver phospholipid LCPUFA content together with the expression of hepatic genes involved in PUFA metabolism, the desaturases FADS1 and FADS2, the elongases ELOV2 and ELOV5, and the transcription factors sterol regulatory element-binding protein-1c (SREBP-1c) and peroxisome proliferator-activated receptor alpha (PPARalpha). The levels of plasma and liver eicosapentaenoic acid (EPA) and docosapentaenoic acid (DPA) increased in proportion to dietary ALA whereas docosahexaenoic acid (DHA) increased only up to 1%en ALA. A low PUFA (0.4%en) reference diet stimulated the expression of delta 6 desaturase (FADS2) and elongase 2 (ELOVL2) when compared to higher PUFA diets. There was, however, no difference in the expression of any of the genes in rats, which were fed diets containing between 0.2%en and 2.9%en ALA and mRNA expression was unrelated to tissue/plasma LCPUFA content. These data suggest that the endogenous synthesis of n-3 LCPUFA from the precursor ALA is regulated independently of changes in the expression of the synthetic enzymes or regulatory transcription factor, and provides evidence that n-3 LCPUFA synthesis is regulated more by substrate competition for existing enzymes than by an increase in their mRNA expression.

Periconceptional undernutrition in normal and overweight ewes leads to increased adrenal growth and epigenetic changes in adrenal IGF2/H19 gene in offspring

Adverse conditions in early life result in increased activation of the hypothalamo‐pituitary‐adrenal axis and in stress responsiveness in offspring. We have developed a model in which “donor” ewes are either normally nourished or overnourished prior to a period of dietary restriction, before transfer of the embryo at 6–7 d after conception to a ewe of normal weight and nutritional history. A moderate restriction of energy intake during the periconceptional period in both normal weight and overweight ewes resulted in increased adrenal mass in male and female lambs and an increased cortisol response to stress in female lambs. The increase in adrenal weight in lambs exposed to periconceptional under‐nutrition was associated with a decrease in the adrenal mRNA expression of IGF2 and decreased methylation in the proximal CTCF‐binding site in the differentially methylated region of the IGF2/H19 gene. Thus, weight loss in both normal and overweight mothers during the periconceptional period results in epigenetic modification of IGF2 in the adrenal gland, adrenal overgrowth, and increased vulnerability to stress in offspring. Determining the appropriate approach to weight loss in the periconceptional period may therefore be important in overweight or obese women seeking to become pregnant.—Zhang, S., Rattanatray, L., MacLaughlin, S. M., Cropley, J. E., Suter, C. M., Molloy, L., Kleemann, D., Walker, S. K., Muhlhausler, B. S., Morrison, J. L., and McMillen, I. C. Periconceptional undernutrition in normal and overweight ewes leads to increased adrenal growth and epigenetic changes in adrenal IGF2/H19 gene in offspring. FASEB J. 24, 2772–2782 (2010). www.fasebj.org

Early-life origins of metabolic dysfunction: role of the adipocyte

More than 60% of adults in the US are classified as overweight, with most developing associated metabolic problems. It is increasingly clear that the origins of obesity and metabolic disease are early in life, yet the physiological basis for this is not well understood. We propose that perturbations to nutrient supply in utero affect adipocyte development, altering functional properties and promoting excess body fat accumulation after birth. We also propose that excessive body fat accumulation leads to leptin and insulin resistance in these individuals, rendering them more susceptible to further weight gain and metabolic deterioration. Finally, we propose that interventions that inhibit this early increase in fat deposition have the potential to interrupt the pathway to obesity.

The transition from fetal growth restriction to accelerated postnatal growth: a potential role for insulin signalling in skeletal muscle

A world‐wide series of epidemiological and experimental studies have demonstrated that there is an association between being small at birth, accelerated growth in early postnatal life and the emergence of insulin resistance in adult life. The aim of this study was to investigate why accelerated growth occurs in postnatal life after in utero growth restriction. Samples of quadriceps muscle were collected at ∼140 days gestation (term ∼150 days gestation) from normally grown fetal lambs (Control, n= 7) and from growth restricted fetal lambs (placentally restricted: PR, n= 8) and from Control (n= 14) and PR (n= 9) lambs at 21 days after birth. The abundance of the insulin and IGF1 receptor protein was higher in the quadriceps muscle of the PR fetus, but there was a lower abundance of the insulin signalling molecule PKCζ, and GLUT4 protein in the PR group. At 21 days of postnatal age, insulin receptor abundance remained higher in the muscle of the PR lamb, and there was also an up‐regulation of the insulin signalling molecules, PI3Kinase p85, Akt1 and Akt2 and of the GLUT4 protein in the PR group. Fetal growth restriction therefore results in an increased abundance of the insulin receptor in skeletal muscle, which persists after birth when it is associated with an upregulation of insulin signalling molecules and the glucose transporter, GLUT4. These data provide evidence that the origins of the accelerated growth experienced by the small baby after birth lie in the adaptive response of the growth restricted fetus to its low placental substrate supply.

Appetite regulatory neuropeptides are expressed in the sheep hypothalamus before birth

In the adult, a hypothalamic neural network acts to maintain energy balance in response to nutritional feedback from the periphery. Although there is an immediate requirement for this system to be functional at birth, it is unknown whether the components of this central neural network are expressed in the developing brain before birth. We therefore examined in the fetal sheep hypothalamus during late gestation gene expression for leptin receptor (OB‐Rb) and neuropeptides that regulate energy balance in the adult. Brains were collected from fetal sheep at 110 days (n = 12) and 140 days of gestation (n = 5) (term = 150 days) and gene expression was detected in all hypothalami using in situ hybridization with radiolabelled riboprobes for OB‐Rb, neuropeptide Y (NPY), agouti‐related peptide, pro‐opiomelanocortin and cocaine‐ and amphetamine‐regulated transcript (CART). All mRNAs were expressed in the arcuate nucleus of fetuses at both time points. Additional sites of mRNA expression were the dorsomedial hypothalamus (DMH) for NPY, the paraventricular nucleus (PVN), ventromedial hypothalamus (VMH) and lateral hypothalamic area for CART, and the DMH, PVN and VMH for OB‐Rb. We have therefore demonstrated that adult‐like localization of gene expression for OB‐Rb and key appetite regulatory neuropeptides is established in the ovine hypothalamus before birth. Thus, the fetus possesses a central appetite regulatory neural network with the potential to respond to changes in nutrient supply, which could impact on energy balance regulation both before and after birth.