Peter W. Nathanielsz

Developmental programming of the metabolic syndrome by maternal nutritional imbalance: how strong is the evidence from experimental models in mammals?

The incidence of the metabolic syndrome, a cluster of abnormalities focusing on insulin resistance and associated with high risk for cardiovascular disease and diabetes, is reaching epidemic proportions. Prevalent in both developed and developing countries, the metabolic syndrome has largely been attributed to altered dietary and lifestyle factors that favour the development of central obesity. However, population‐based studies have suggested that predisposition to the metabolic syndrome may be acquired very early in development through inappropriate fetal or neonatal nutrition. Further evidence for developmental programming of the metabolic syndrome has now been suggested by animal studies in which the fetal environment has been manipulated through altered maternal dietary intake or modification of uterine artery blood flow. This review examines these studies and assesses whether the metabolic syndrome can be reliably induced by the interventions made. The validity of the different species, diets, feeding regimes and end‐point measures used is also discussed.

A low maternal protein diet during pregnancy and lactation has sex- and window of exposure-specific effects on offspring growth and food intake, glucose metabolism and serum leptin in the rat

Extensive epidemiological and experimental evidence indicates that a sub‐optimal environment during fetal and neonatal development in both humans and animals may programme offspring susceptibility to later development of chronic diseases including obesity and diabetes that are the result of altered carbohydrate metabolism. We determined the effects of protein restriction during pregnancy and/or lactation on growth, serum leptin, and glucose and insulin responses to a glucose tolerance test in male and female offspring at 110 days postnatal life. We fed Wistar rats a normal control 20% casein diet (C) or a restricted diet (R) of 10% casein during pregnancy. Female but not male R pups weighed less than C at birth. After delivery, mothers received the C or R diet during lactation to provide four offspring groups: CC (first letter maternal pregnancy diet and second maternal lactation diet), RR, CR and RC. All offspring were fed ad libitum with C diet after weaning. Relative food intake correlated inversely with weight. Offspring serum leptin correlated with body weight and relative, but not absolute, food intake in both male and female pups. Serum leptin was reduced in RR female pups compared with CC and increased in RC males compared with CC at 110 days of age. Offspring underwent a glucose tolerance test (GTT) at 110 days postnatal life. Female RR and CR offspring showed a lower insulin to glucose ratio than CC. At 110 days of age male RR and CR also showed some evidence of increased insulin sensitivity. Male but not female RC offspring showed evidence of insulin resistance compared with CC. Cholesterol was similar and triglycerides (TG) higher in male compared with female CC. Cholesterol and TG were higher in males than females in RR, CR and RC (P < 0.05). Cholesterol and TG did not differ between groups in females. Cholesterol and TG were elevated in RC compared with CC males. Nutrient restriction in lactation increased relative whole protein and decreased whole lipid in both males and females. RC females showed decreased relative levels of protein and increased fat. We conclude that maternal protein restriction during either pregnancy and/or lactation alters postnatal growth, appetitive behaviour, leptin physiology, TG and cholesterol concentrations and modifies glucose metabolism and insulin resistance in a sex‐ and time window of exposure‐specific manner.

Sex differences in transgenerational alterations of growth and metabolism in progeny (F2) of female offspring (F1) of rats fed a low protein diet during pregnancy and lactation

Compelling epidemiological and experimental evidence indicates that a suboptimal environment during fetal and neonatal development in both humans and animals may programme offspring susceptibility to later development of several chronic diseases including obesity and diabetes in which altered carbohydrate metabolism plays a central role. One of the most interesting and significant features of developmental programming is the evidence from several studies that the adverse consequences of altered intrauterine environments can be passed transgenerationally from mother (F0) to daughter (F1) to second generation offspring (F2). We determined whether when F0 female rats are exposed to protein restriction during pregnancy and/or lactation their F1 female pups deliver F2 offspring with in vivo evidence of altered glucose and insulin metabolism. We fed F0 virgin Wistar rats a normal control 20% casein diet (C) or a protein restricted isocaloric diet (R) containing 10% casein during pregnancy. F1 female R pups weighed less than C at birth. After delivery, mothers received C or R diet during lactation to provide four F1 offspring groups CC (first letter pregnancy diet and second lactation diet), RR, CR and RC. All F1 female offspring were fed ad libitum with C diet after weaning and during their first pregnancy and lactation. As they grew female offspring (F1) of RR and CR mothers exhibited low body weight and food intake with increased sensitivity to insulin during a glucose tolerance test at 110 days of postnatal life. Male F2 CR offspring showed evidence of insulin resistance. In contrast RC F2 females showed evidence of insulin resistance. Sex differences were also observed in F2 offspring in resting glucose and insulin and insulin: glucose ratios. These sex differences also showed differences specific to stage of development time window. We conclude that maternal protein restriction adversely affects glucose and insulin metabolism of male and female F2 offspring in a manner specific to sex and developmental time window during their mothers (the F1) fetal and neonatal development.

Maternal Undernutrition from Early- to Mid-Gestation Leads to Growth Retardation, Cardiac Ventricular Hypertrophy, and Increased Liver Weight in the Fetal Sheep

Abstract Early gestation is critical for placentomal growth, differentiation, and vascularization, as well as fetal organogenesis. The fetal origins of adult disease hypothesis proposes that alterations in fetal nutrition and endocrine status result in developmental adaptations that permanently change structure, physiology, and metabolism, thereby predisposing individuals to cardiovascular, metabolic, and endocrine disease in adult life. Multiparous ewes were fed to 50% (nutrient restricted) or 100% (control fed) of total digestible nutrients from Days 28 to 78 of gestation. All ewes were weighed weekly and diets adjusted for individual weight loss or gain. Ewes were killed on Day 78 of gestation and gravid uteri recovered. Fetal body and organ weights were determined, and numbers, morphologies, diameters, and weights of all placentomes were obtained. From Day 28 to Day 78, restricted ewes lost 7.4% of body weight, while control ewes gained 7.5%. Maternal and fetal blood glucose concentrations were reduced in restricted versus control pregnancies. Fetuses were markedly smaller in the restricted group than in the control group. Further, restricted fetuses exhibited greater right- and left-ventricular and liver weights per unit fetal weight than control fetuses. No treatment differences were observed in any gross placentomal measurement. However, caruncular vascularity was enhanced in conceptuses from nutrient-restricted ewes but only in twin pregnancies. While these alterations in fetal/placental development may be beneficial to early fetal survival in the face of a nutrient restriction, their effects later in gestation as well as in postnatal life need further investigation.

Maternal nutrient restriction affects properties of skeletal muscle in offspring.

Maternal nutrient restriction (NR) affects fetal development with long‐term consequences on postnatal health of offspring, including predisposition to obesity and diabetes. Most studies have been conducted in fetuses in late gestation, and little information is available on the persistent impact of NR from early to mid‐gestation on properties of offspring skeletal muscle, which was the aim of this study. Pregnant ewes were subjected to 50% NR from day 28–78 of gestation and allowed to deliver. The longissimus dorsi muscle was sampled from 8‐month‐old offspring. Maternal NR during early to mid‐gestation decreased the number of myofibres in the offspring and increased the ratio of myosin IIb to other isoforms by 17.6 ± 4.9% (P < 0.05) compared with offspring of ad libitum fed ewes. Activity of carnitine palmitoyltransferase‐1, a key enzyme controlling fatty acid oxidation, was reduced by 24.7 ± 4.5% (P < 0.05) in skeletal muscle of offspring of NR ewes and would contribute to increased fat accumulation observed in offspring of NR ewes. Intramuscular triglyceride content (IMTG) was increased in skeletal muscle of NR lambs, a finding which may be linked to predisposition to diabetes in offspring of NR mothers, since enhanced IMTG predisposes to insulin resistance in skeletal muscle. Proteomic analysis by two‐dimensional gel electrophoresis demonstrated downregulation of several catabolic enzymes in 8‐month‐old offspring of NR ewes. These data demonstrate that the early to mid‐gestation period is important for skeletal muscle development. Impaired muscle development during this stage of gestation affects the number and composition of fibres in offspring which may lead to long‐term physiological consequences, including predisposition to obesity and diabetes.

The influence of long chain polyunsaturate supplementation on docosahexaenoic acid and arachidonic acid in baboon neonate central nervous system.

BackgroundDocosahexaenoic acid (DHA) and arachidonic acid (ARA) are major components of the cerebral cortex and visual system, where they play a critical role in neural development. We quantitatively mapped fatty acids in 26 regions of the four-week-old breastfed baboon CNS, and studied the influence of dietary DHA and ARA supplementation and prematurity on CNS DHA and ARA concentrations.MethodsBaboons were randomized into a breastfed (B) and four formula-fed groups: term, no DHA/ARA (T-); term, DHA/ARA supplemented (T+); preterm, no DHA/ARA (P-); preterm and DHA/ARA supplemented (P+). At four weeks adjusted age, brains were dissected and total fatty acids analyzed by gas chromatography and mass spectrometry.ResultsDHA and ARA are rich in many more structures than previously reported. They are most concentrated in structures local to the brain stem and diencephalon, particularly the basal ganglia, limbic regions, thalamus and midbrain, and comparatively lower in white matter. Dietary supplementation increased DHA in all structures but had little influence on ARA concentrations. Supplementation restored DHA concentrations to levels of breastfed neonates in all regions except the cerebral cortex and cerebellum. Prematurity per se did not exert a strong influence on DHA or ARA concentrations.Conclusion1) DHA and ARA are found in high concentration throughout the primate CNS, particularly in gray matter such as basal ganglia; 2) DHA concentrations drop across most CNS structures in neonates consuming formulas with no DHA, but ARA levels are relatively immune to ARA in the diet; 3) supplementation of infant formula is effective at restoring DHA concentration in structures other than the cerebral cortex. These results will be useful as a guide to future investigations of CNS function in the absence of dietary DHA and ARA.

Fetal programming of skeletal muscle development in ruminant animals.

Enhancing skeletal muscle growth is crucial for animal agriculture because skeletal muscle provides meat for human consumption. An increasing body of evidence shows that the level of maternal nutrition alters fetal skeletal muscle development, with long-term effects on offspring growth and performance. Fetal skeletal muscle development mainly involves myogenesis (i.e., muscle cell development), but also involves adipogenesis (i.e., adipocyte development) and fibrogenesis (i.e., fibroblast development). These tissues in fetal muscle are mainly derived from mesenchymal stem cells (MSC). Shifting the commitment of MSC from myogenesis to adipogenesis increases intramuscular fat (i.e., marbling), improving the quality grade of meats. Strong experimental evidence indicates that Wingless and Int (Wnt)/beta-catenin signaling regulates MSC differentiation. Upregulation of Wnt/beta-catenin promotes myogenesis, and downregulation enhances adipogenesis. A lack of nutrients in early to midgestation reduces the formation of secondary muscle fibers in ruminant animals. Nutrient deficiency during mid- to late gestation decreases the number of intramuscular adipocytes and muscle fiber sizes. Knowledge of this regulatory mechanism will allow the development of strategies to enhance muscle growth and marbling in offspring, especially in the setting of nutrient deficiency.

Efficacy of Dietary Arachidonic Acid Provided as Triglyceride or Phospholipid as Substrates for Brain Arachidonic Acid Accretion in Baboon Neonates

Arachidonic acid (AA) is a long-chain polyunsaturate (LCP) present in human breast milk as both triglyceride (TG) and as phospholipid (PL). There has been little attention to the metabolic consequences of lipid form of AA in infant formulas. Our objective was to investigate the efficacy of dietary TG and PL as carriers of AA for accretion in the brain and associated organs of term baboon neonates consuming a formula with LCP composition typical of human milk. TG and phosphatidylcholine (PC) with [U-13C]-AA in the sn-2 position and with unlabeled 16:0 in the remaining positions (TG-AA* or PL-AA*, respectively) were used as tracers to study the tissue AA* incorporation. Baboon neonates received a single oral dose of either TG-AA* (n = 3) or PL-AA* (n = 4) at 18–19 d of life. Tissues were obtained 10 d later (28–29 d of life) and isotopic enrichment was measured. In the brain, 4.5% of the PL-AA* dose and 2.1% of the TG-AA* dose were recovered as brain AA*, respectively, indicating that PL was about 2.1-fold more effective than TG as a substrate for brain AA accretion. Preferential incorporation of PL-derived AA* over TG source of AA* was also observed in the liver, lung, plasma, and erythrocytes. Because of the quantitative predominance of TG-AA in formula, total brain AA accretion, expressed as absolute weight, was 5.0-fold greater from TG-AA than from PL-AA. We estimate that about half of postnatal brain AA accretion is derived from dietary preformed AA in term baboon neonates consuming a formula with lipid composition similar to that of human milk.

A maternal low protein diet during pregnancy and lactation in the rat impairs male reproductive development

Nutrient restriction during pregnancy and lactation impairs growth and development. Recent studies demonstrate long‐term programming of function of specific organ systems resulting from suboptimal environments during fetal life and development up to weaning. We determined effects of maternal protein restriction (50% control protein intake) during fetal development and/or lactation in rats on the reproductive system of male progeny. Rats were fed either a control 20% casein diet (C) or a restricted diet (R) of 10% casein during pregnancy. After delivery mothers received either C or R diet until weaning to provide four groups: CC, RR, CR and RC. We report findings in male offspring only. Maternal protein restriction increased maternal serum corticosterone, oestradiol and testosterone (T) concentrations at 19 days gestation. Pup birth weight was unchanged but ano‐genital distance was increased by maternal protein restriction (P < 0.05). Testicular descent was delayed 4.4 days in RR, 2.1 days in CR and 2.2 days in RC and was not related to body weight. Body weight and testis weight were reduced in RR and CR groups at all ages with the exception of CR testis weight at 270 days postnatal age (PN). At 70 days PN luteinizing hormone and T concentrations were reduced in RR, CR and RC. mRNA for P450 side chain cleavage (P450scc) was reduced in RR and CR at 21 days PN but was unchanged at 70 days PN. Fertility rate was reduced at 270 days PN in RC and sperm count in RR and RC. We conclude that maternal protein delays sexual maturation in male rats and that some effects only emerge in later life.

Effect of Maternal Nutrient Restriction in Sheep on the Development of Fetal Skeletal Muscle

Abstract The effect of maternal nutrient restriction on mTOR (mammalian target of rapamyosin) signaling and the ubiquitin system as well as their possible relation to growth of fetal muscle was determined. Ewes were fed to 50% (nutrient-restricted) or 100% (control-fed) of total digestible nutrients (National Research Council requirement) from Days 28 to 78 of gestation. Ewes were killed at Day 78 of gestation, and the fetal longissimus dorsi muscle was sampled for the measurement of mTOR, ribosomal protein S6, AMP-activated protein kinase (AMPK), calpastatin, and protein ubiquitylation. No difference was observed in the content of mTOR and ribosomal protein S6, but the phosphorylation of mTOR at Ser2448 and ribosomal protein S6 at Ser235/336 were reduced (P < 0.05) in muscle from nutrient-restricted fetuses. Because phosphorylation of mTOR and ribosomal protein S6 up-regulates protein translation, these results show that nutrient restriction down-regulates protein synthesis in fetal muscle. No difference in AMPK activity was detected. The lack of difference in calpastatin and ubiquitylized protein content shows that nutrient restriction did not affect degradation of myofibrillar proteins in fetal muscle. Fetuses of nutrient-restricted ewes showed retarded development of muscles and skeleton. Muscle from nutrient-restricted fetuses contained fewer secondary myofibers than muscle from control fetuses, and the average area of fasciculi was smaller (P < 0.05). The decreased number of secondary myofibers in nutrient-restricted fetuses may result from the decreased mTOR signaling. Lower activation of mTOR signaling in nutrient-restricted fetuses may reduce the proliferation of myoblasts and, thus, reduce the formation of secondary myofibers. This decrease in secondary myofibers in fetuses may predispose fetuses to metabolic diseases, such as diabetes and obesity, in their postnatal lives.