Dysregulation of Glucocorticoid Receptor Homeostasis and Glucocorticoid-Associated Genes in Umbilical Cord Endothelial Cells of Diet-Induced Obese Pregnant Sheep.

Maternal obesity (MO) is associated with offspring cardiometabolic diseases that are hypothesized to be partly mediated by glucocorticoids. Therefore, we aimed to study fetal endothelial glucocorticoid sensitivity in an ovine model of MO. Rambouillet/Columbia ewes were fed either 100% (control) or 150% (MO) National Research Council recommendations from 60 d before mating until near-term (135 days gestation). Sheep umbilical vein and artery endothelial cells (ShUVECs and ShUAECs) were used to study glucocorticoid receptor (GR) expression and function in vitro. Dexamethasone dose-response studies of gene expression, activation of a glucocorticoid response element (GRE)-dependent luciferase reporter vector, and cytosolic/nuclear GR translocation were used to assess GR homeostasis. MO significantly increased basal GR protein levels in both ShUVECs and ShUAECs. Increased GR protein levels did not result in increased dexamethasone sensitivity in the regulation of key endothelial gene expression such as endothelial nitric oxide synthase, plasminogen activator inhibitor 1, vascular endothelial growth factor, or intercellular adhesion molecule 1. In ShUVECs, MO increased GRE-dependent transactivation and FKBP prolyl isomerase 5 (FKBP5) expression. ShUAECs showed generalized glucocorticoid resistance in both dietary groups. Finally, we found that ShUVECs were less sensitive to dexamethasone-induced activation of GR than human umbilical vein endothelial cells (HUVECs). These findings suggest that MO-mediated effects in the offspring endothelium could be further mediated by dysregulation of GR homeostasis in humans as compared with sheep.

Maternal hepatic adaptations during obese pregnancy encompass lobe-specific mitochondrial alterations and oxidative stress.

Maternal obesity (MO) is rising worldwide, affecting half of all gestations, constituting a possible risk-factor for some pregnancy-associated liver diseases (PALD) and hepatic diseases. PALD occur in approximately 3% of pregnancies and are characterized by maternal hepatic oxidative stress (OS) and mitochondrial dysfunction. Maternal hepatic disease increases maternal and fetal morbidity and mortality. Understanding the role of MO on liver function and pathophysiology could be crucial for better understanding the altered pathways leading to PALD and liver disease, possibly paving the way to prevention and adequate management of disease. We investigated specific hepatic metabolic alterations in mitochondria and oxidative stress during MO at late-gestation. Maternal hepatic tissue was collected at 90% gestation in Control and MO ewes (fed 150% of recommended nutrition starting 60 days before conception). Maternal hepatic redox state, mitochondrial respiratory chain (MRC), and OS markers were investigated. MO decreased MRC complex-II activity and its subunits SDHA and SDHB protein expression, increased complex-I and complex-IV activities despite reduced complex-IV subunit mtCO1 protein expression, and increased ATP synthase ATP5A subunit. Hepatic MO-metabolic remodeling was characterized by decreased adenine nucleotide translocator 1 and 2 (ANT-1/2) and voltage-dependent anion channel (VDAC) protein expression and protein kinase A (PKA) activity (P<0.01), and augmented NAD+/NADH ratio due to reduced NADH levels (P<0.01). MO showed an altered redox state with increased OS, increased lipid peroxidation (P<0.01), decreased GSH/GSSG ratio (P=0.005), increased superoxide dismutase (P=0.03) and decreased catalase (P=0.03) antioxidant enzymatic activities, lower catalase, glutathione peroxidase (GPX)-4 and glutathione reductase protein expression (P<0.05), and increased GPX-1 abundance (P=0.03). MO-related hepatic changes were more evident in the right lobe, corroborated by the integrative data analysis. Hepatic tissue from obese pregnant ewes showed alterations in the redox state, consistent with OS and MRC and metabolism remodeling. These are hallmarks of PALD and hepatic disease, supporting MO as a risk-factor and highlighting OS and mitochondrial dysfunction as mechanisms responsible for liver disease predisposition.

Maternal obesity alters offspring liver and skeletal muscle metabolism in early post-puberty despite maintaining a normal post-weaning dietary lifestyle.

Maternal obesity (MO) during pregnancy is linked to increased and premature risk of age-related metabolic diseases in the offspring. However, the underlying molecular mechanisms still remain not fully understood. Using a well-established nonhuman primate model of MO, we analyzed tissue biopsies and plasma samples obtained from post-pubertal offspring (3-6.5 y) of MO mothers (n = 19) and from control animals born to mothers fed a standard diet (CON, n = 13). All offspring ate a healthy chow diet after weaning. Using untargeted gas chromatography-mass spectrometry metabolomics analysis, we quantified a total of 351 liver, 316 skeletal muscle, and 423 plasma metabolites. We identified 58 metabolites significantly altered in the liver and 46 in the skeletal muscle of MO offspring, with 8 metabolites shared between both tissues. Several metabolites were changed in opposite directions in males and females in both liver and skeletal muscle. Several tissue-specific and 4 shared metabolic pathways were identified from these dysregulated metabolites. Interestingly, none of the tissue-specific metabolic changes were reflected in plasma. Overall, our study describes characteristic metabolic perturbations in the liver and skeletal muscle in MO offspring, indicating that metabolic programming in utero persists postnatally, and revealing potential novel mechanisms that may contribute to age-related metabolic diseases later in life.

Characterizing Early Cardiac Metabolic Programming via 30% Maternal Nutrient Reduction during Fetal Development in a Non-Human Primate Model.

Intra-uterine growth restriction (IUGR) is a common cause of fetal/neonatal morbidity and mortality and is associated with increased offspring predisposition for cardiovascular disease (CVD) development. Mitochondria are essential organelles in maintaining cardiac function, and thus, fetal cardiac mitochondria could be responsive to the IUGR environment. In this study, we investigated whether in utero fetal cardiac mitochondrial programming can be detectable in an early stage of IUGR pregnancy. Using a well-established nonhuman IUGR primate model, we induced IUGR by reducing by 30% the maternal diet (MNR), both in males (MNR-M) and in female (MNR-F) fetuses. Fetal cardiac left ventricle (LV) tissue and blood were collected at 90 days of gestation (0.5 gestation, 0.5 G). Blood biochemical parameters were determined and heart LV mitochondrial biology assessed. MNR fetus biochemical blood parameters confirm an early fetal response to MNR. In addition, we show that in utero cardiac mitochondrial MNR adaptations are already detectable at this early stage, in a sex-divergent way. MNR induced alterations in the cardiac gene expression of oxidative phosphorylation (OXPHOS) subunits (mostly for complex-I, III, and ATP synthase), along with increased protein content for complex-I, -III, and -IV subunits only for MNR-M in comparison with male controls, highlight the fetal cardiac sex-divergent response to MNR. At this fetal stage, no major alterations were detected in mitochondrial DNA copy number nor markers for oxidative stress. This study shows that in 90-day nonhuman primate fetuses, a 30% decrease in maternal nutrition generated early in utero adaptations in fetal blood biochemical parameters and sex-specific alterations in cardiac left ventricle gene and protein expression profiles, affecting predominantly OXPHOS subunits. Since the OXPHOS system is determinant for energy production in mitochondria, our findings suggest that these early IUGR-induced mitochondrial adaptations play a role in offspring's mitochondrial dysfunction and can increase predisposition to CVD in a sex-specific way.

Mechanisms linking hypoxia to phosphorylation of insulin-like growth factor binding protein-1 in baboon fetuses with intrauterine growth restriction and in cell culture.

Hypoxia increases fetal hepatic insulin-like growth factor binding protein-1 (IGFBP-1) phosphorylation mediated by mechanistic target of rapamycin (mTOR) inhibition. Whether maternal nutrient restriction (MNR) causes fetal hypoxia remains unclear. We used fetal liver from a baboon (Papio sp.) model of intrauterine growth restriction due to MNR (70% global diet of Control) and liver hepatocellular carcinoma (HepG2) cells as a model for human fetal hepatocytes and tested the hypothesis that mTOR-mediated IGFBP-1 hyperphosphorylation in response to hypoxia requires hypoxia-inducible factor-1α (HIF-1α) and regulated in development and DNA-damage responses-1 (REDD-1) signaling. Western blotting (n = 6) and immunohistochemistry (n = 3) using fetal liver indicated greater expression of HIF-1α, REDD-1 as well as erythropoietin and its receptor, and vascular endothelial growth factor at GD120 (GD185 term) in MNR versus Control. Moreover, treatment of HepG2 cells with hypoxia (1% pO2 ) (n = 3) induced REDD-1, inhibited mTOR complex-1 (mTORC1) activity and increased IGFBP-1 secretion/phosphorylation (Ser101/Ser119/Ser169). HIF-1α inhibition by echinomycin or small interfering RNA silencing prevented the hypoxia-mediated inhibition of mTORC1 and induction of IGFBP-1 secretion/phosphorylation. dimethyloxaloylglycine (DMOG) induced HIF-1α and also REDD-1 expression, inhibited mTORC1 and increased IGFBP-1 secretion/phosphorylation. Induction of HIF-1α (DMOG) and REDD-1 by Compound 3 inhibited mTORC1, increased IGFBP-1 secretion/ phosphorylation and protein kinase PKCα expression. Together, our data demonstrate that HIF-1α induction, increased REDD-1 expression and mTORC1 inhibition represent the mechanistic link between hypoxia and increased IGFBP-1 secretion/phosphorylation. We propose that maternal undernutrition limits fetal oxygen delivery, as demonstrated by increased fetal liver expression of hypoxia-responsive proteins in baboon MNR. These findings have important implications for our understanding of the pathophysiology of restricted fetal growth.

Postnatal persistence of nonhuman primate sex-dependent renal structural and molecular changes programmed by intrauterine growth restriction.

BACKGROUND: Poor nutrition during fetal development programs postnatal kidney function. Understanding postnatal consequences in nonhuman primates (NHP) is important for translation to our understanding the impact on human kidney function and disease risk. We hypothesized that intrauterine growth restriction (IUGR) in NHP persists postnatally, with potential molecular mechanisms revealed by Western-type diet challenge. METHODS: IUGR juvenile baboons were fed a 7-week Western diet, with kidney biopsies, blood, and urine collected before and after challenge. Transcriptomics and metabolomics were used to analyze biosamples. RESULTS: Pre-challenge IUGR kidney transcriptome and urine metabolome differed from controls. Post-challenge, sex and diet-specific responses in urine metabolite and renal signaling pathways were observed. Dysregulated mTOR signaling persisted postnatally in female pre-challenge. Post-challenge IUGR male response showed uncoordinated signaling suggesting proximal tubule injury. CONCLUSION: Fetal undernutrition impacts juvenile offspring kidneys at the molecular level suggesting early-onset blood pressure dysregulation.

Sexual dimorphism in liver cell cycle and senescence signalling pathways in young and old rats.

At the molecular level, cellular ageing involves changes in multiple gene pathways. Cellular senescence is both an important initiator and a consequence of natural ageing. Senescence results in changes in multiple cellular mechanisms that result in a natural decrease in cell cycle activity. Liver senescence changes impair hepatic function. Given the well-established sexual dimorphism in ageing, we hypothesized that the natural hepatic ageing process is driven by sex-dependent gene mechanisms. We studied our well-characterized normal, chow-fed rat ageing model, lifespan ∼850 days, in which we have reported ageing of metabolism, reproduction and endocrine function. We performed liver RNA-seq on males and females at 110 and 650 days to determine changes in the cell cycle and cellular senescence signalling pathways. We found that natural liver ageing shows sexual dimorphism in these pathways. RNA-seq revealed more male (3967) than female (283) differentially expressed genes between 110 and 650 days. Cell cycle pathway signalling changes in males showed decreased protein and expression of key genes (Cdk2, Cdk4, Cycd and PCNA) and increased expression ofp57 at 650 vs 110 days. In females, protein and gene expression of cell growth regulators, e.g. p15 and p21, which inhibit cell cycle G1 progression, were increased. The cell senescence pathway also showed sexual dimorphism. Igfbp3, mTOR and p62 gene and protein expression decreased in males while those ofTgfb3 increased in females. Understanding the involvement of cell cycling and cellular senescence pathways in natural ageing will advance evaluation of mechanisms associated with altered ageing and frailty trajectories. KEY POINTS: In rats RNA-seq analysis showed sexual dimorphism in gene expression across the life-course between 110 and 650 days of life. Fourteen times more liver transcriptome and six times more pathway changes were observed in males compared with females. Significant changes were observed in several signalling pathways during ageing. Bioinformatic analysis were focused on changes in genes and protein products related to cell cycle and cellular senescence pathways. Males showed decreased protein product and expression of the key genes Cdk2 and Cdk4 responsible for cell cycle progression while females increased protein product and expression of p21 and p15, key genes responsible for cell cycle arrest. In conclusion, normative rat hepatic ageing involves changes in cellular pathways that control cell cycle arrest but through changes in different genes in males and females. These findings identify mechanisms that underlie the well-established sexual dimorphism in ageing.

Rodent studies of developmental programming and ageing mechanisms: Special issue: In utero and early life programming of ageing and disease.

Compelling evidence exists indicating that developmental programming influences ageing. Programming alters life-course phenotype in multiple organs, predisposing to diseases such as diabetes, obesity and cardiovascular disease that shorten lifespan. This review describes studies in rodents, the most commonly studied species, addressing interactions of programming challenges with ageing. We first consider ageing and programming of insulin function that has been clearly shown to decrease with age. It is important to evaluate ageing in pancreatic islets isolated from other systems. Studies discussed show premature pancreatic islet ageing resulting from both maternal under- and overnutrition. New ways to determine programming of adipose tissue and effects on fat storage are explored. Oxidative stress is a major factor that regulates ageing in tissues. Oxidative stress is discussed in relation to reproductive and cardiovascular ageing. Premature ageing is associated with both low and high glucocorticoid function. Both over and undernutrition have offspring sex-specific programming effects on life-course glucocorticoid concentrations. Evidence is provided that maternal age at conception affects offspring endocrine and metabolism ageing. Finally, the importance of matching foetal nutrition and energy availability with composition and energy content in the post-weaning diet is demonstrated. This mismatch can lead to a greatly shortened lifespan. General principles are discussed throughout. For example, sexual dimorphism of age-related outcomes can be marked. Accelerated ageing occurs early in life. Improving knowledge on programming ageing interactions will improve health span as well as lifespan. Finally, there are considerable similarities in outcomes programmed by maternal undernutrition and overnutrition.

Developmental programming and ageing of male reproductive function.

Developmental programming predisposes offspring to metabolic, behavioural and reproductive dysfunction in adult life. Evidence is accumulating that ageing phenotype and longevity are in part developmentally programmed in each individual. Unfortunately, there are few studies addressing the effects of developmental programming by maternal nutrition on the rate of ageing of the male reproductive system. This review will discuss effects of foetal exposure to maternal environmental challenges on male offspring fertility and normal ageing of the male reproductive system. We focus on several key factors involved in reproductive ageing such as decreased hormone production, DNA fragmentation, oxidative stress, telomere shortening, epigenetics, maternal lifestyle and nutrition. There is compelling evidence that ageing of the male reproductive system is developmentally programmed. Both maternal over- or undernutrition accelerate ageing of male offspring reproductive function through similar mechanisms such as decreased serum testosterone levels, increase in oxidative stress biomarkers in both the testes and sperm and changes in sperm quality. Importantly, even in adult life, exercise in male offspring of obese mothers improves adverse effects of programming on reproductive function. Maternal consumption of a low-protein diet causes transgenerational effects in progeny via the paternal line. The seminal fluid has effects on the intrauterine environment. Programming by male factors may involve more than just the sperm. Improving knowledge on developmental programming ageing interactions will improve not only male health and life span but also the health of future generations by reducing programming via the paternal line.

Maternal obesity in sheep impairs foetal hepatic mitochondrial respiratory chain capacity.

BACKGROUND: Changes in the nutritional environment in utero induced by maternal obesity (MO) lead to foetal metabolic dysfunction predisposing offspring to later-life metabolic diseases. Since mitochondria play a crucial role in hepatic metabolism and function, we hypothesized that MO prior to conception and throughout pregnancy programmes foetal sheep liver mitochondrial phenotype. MATERIAL AND METHODS: Ewes ate an obesogenic diet (150% requirements; MO), or 100% requirements (CTR), from 60 days prior to conception. Foetal livers were removed at 0.9 gestation. We measured foetal liver mitochondrial DNA copy number, activity of superoxide dismutase, cathepsins B and D and selected protein content, total phospholipids and cardiolipin and activity of mitochondrial respiratory chain complexes. RESULTS: A significant decrease in activities of mitochondrial complexes I, II-III and IV, but not aconitase, was observed in MO. In the antioxidant machinery, there was a significant increase in activity of total superoxide dismutase (SOD) and SOD2 in MO. However, no differences were found regarding autophagy-related protein content (p62, beclin-I, LC3-I, LC3-II and Lamp2A) and cathepsin B and D activities. A 21.5% decrease in total mitochondrial phospholipid was observed in MO. CONCLUSIONS: The data indicate that MO impairs foetal hepatic mitochondrial oxidative capacity and affects total mitochondrial phospholipid content. In addition, MO affects the regulation of foetal liver redox pathways, indicating metabolic adaptations to the higher foetal lipid environment. Consequences of in utero programming of foetal hepatic metabolism may persist and compromise mitochondrial bioenergetics in later life, and increase susceptibility to metabolic diseases.

Sex-dependent vulnerability of fetal nonhuman primate cardiac mitochondria to moderate maternal nutrient reduction.

Poor maternal nutrition in pregnancy affects fetal development, predisposing offspring to cardiometabolic diseases. The role of mitochondria during fetal development on later-life cardiac dysfunction caused by maternal nutrient reduction (MNR) remains unexplored. We hypothesized that MNR during gestation causes fetal cardiac bioenergetic deficits, compromising cardiac mitochondrial metabolism and reserve capacity. To enable human translation, we developed a primate baboon model (Papio spp.) of moderate MNR in which mothers receive 70% of control nutrition during pregnancy, resulting in intrauterine growth restriction (IUGR) offspring and later exhibiting myocardial remodeling and heart failure at human equivalent ∼25 years. Term control and MNR baboon offspring were necropsied following cesarean-section, and left ventricle (LV) samples were collected. MNR adversely impacted fetal cardiac LV mitochondria in a sex-dependent fashion. Increased maternal plasma aspartate aminotransferase, creatine phosphokinase (CPK), and elevated cortisol levels in MNR concomitant with decreased blood insulin in male fetal MNR were measured. MNR resulted in a two-fold increase in fetal LV mitochondrial DNA (mtDNA). MNR resulted in increased transcripts for several respiratory chain (NDUFB8, UQCRC1, and cytochrome c) and adenosine triphosphate (ATP) synthase proteins. However, MNR fetal LV mitochondrial complex I and complex II/III activities were significantly decreased, possibly contributing to the 73% decreased ATP content and increased lipid peroxidation. MNR fetal LV showed mitochondria with sparse and disarranged cristae dysmorphology. Conclusion: MNR disruption of fetal cardiac mitochondrial fitness likely contributes to the documented developmental programming of adult cardiac dysfunction, indicating a programmed mitochondrial inability to deliver sufficient energy to cardiac tissues as a chronic mechanism for later-life heart failure.

Walking speed declines with age in male and female baboons (Papio sp.): Confirmation of findings with sex as a biological variable.

We measured walking speed in baboons (67 female, 36 male; 5-22 years) to develop regression formulas to predict biological age. The final model strongly predicted age from just speed and sex. Walking speed is a valuable baboon aging biomarker. We present the first male speed data in a nonhuman primate.

Placental fatty acid transport across late gestation in a baboon model of intrauterine growth restriction.

KEY POINTS: Intrauterine growth restriction (IUGR) is associated with perinatal morbidity and increased risk of lifelong disease, including neurodevelopmental impairment. Fatty acids (FA) are critical for normal brain development, although their transport across the placenta in IUGR pregnancies is poorly understood. The present study used a baboon model of IUGR (maternal nutrient restriction, MNR) to investigate placental expression of FA transport and binding proteins, and to determine gestational age-related changes in maternal and fetal plasma FA concentrations. We found MNR to be associated with increased placental expression of FA binding and transport proteins in late gestation, with fetal plasma FA concentrations that were similar to those of control animals. The present study is the first to report a profile of fetal and maternal plasma FA concentrations in a baboon model of growth restriction with data that suggest adaptation of placental transport to maintain delivery of critically needed FA. ABSTRACT: Intrauterine growth restriction (IUGR) is associated with specific changes in placental transport of amino acids, folate and ions. However, little is known about placental fatty acid (FA) transport in IUGR. We hypothesized that placental FA transport proteins (FATP) and FA binding proteins (FABP) are up-regulated and fetal plasma FA concentrations are decreased at term in a baboon model of IUGR. Pregnant baboons were fed control or maternal nutrient restricted (MNR) diet (70% of control calories) from gestation day (GD) 30 (term 184 days). Plasma and placental samples were collected at GD120 (control n = 8, MNR n = 9), GD140 (control n = 6, MNR n = 7) and GD170 (control n = 6, MNR n = 6). Placentas were homogenized, and syncytiotrophoblast microvillous plasma membrane (MVM) and basal plasma membranes (BM) were isolated. Protein expression of FABP1, 3, 4 and 5 (homogenate) and FATP2, 4, and 6 (MVM, BM) was determined by Western blotting. FA content in maternal and umbilical vein plasma was measured by gas chromatography-mass spectrometry. Placental FABP1 and FABP5 expression was increased in MNR compared to controls at GD170, as was MVM FATP2 and FATP6 expression at GD140 and FATP2 expression at GD170. BM FATP4 and FATP6 expression was increased in MNR at GD140. Fetal plasma FA concentrations were similar in controls and MNR. These data suggest the adaptation of placental transport when aiming to maintain delivery of critically needed FAs for fetal growth and brain development.

Hyperphosphorylation of fetal liver IGFBP-1 precedes slowing of fetal growth in nutrient-restricted baboons and may be a mechanism underlying IUGR.

In cultured fetal liver cells, insulin-like growth factor (IGF) binding protein (IGFBP)-1 hyperphosphorylation in response to hypoxia and amino acid deprivation is mediated by inhibition of mechanistic target of rapamycin (mTOR) and activation of amino acid response (AAR) signaling and casein kinase (CK)2. We hypothesized that fetal liver mTOR inhibition, activation of AAR and CK2, and IGFBP-1 hyperphosphorylation occur before development of intrauterine growth restriction (IUGR). Pregnant baboons were fed a control (C) or a maternal nutrient restriction (MNR; 70% calories of control) diet starting at gestational day (GD) 30 (term GD 185). Umbilical blood and fetal liver tissue were obtained at GD 120 (C, n = 7; MNR, n = 10) and 165 (C, n = 7; MNR, n = 8). Fetal weights were unchanged at GD 120 but decreased at GD 165 in the MNR group (-13%, P = 0.03). IGFBP-1 phosphorylation, as determined by parallel reaction monitoring mass spectrometry (PRM-MS), immunohistochemistry, and/or Western blot, was enhanced in MNR fetal liver and umbilical plasma at GD 120 and 165. IGF-I receptor autophosphorylationTyr1135 (-64%, P = 0.05) was reduced in MNR fetal liver at GD 120. Furthermore, fetal liver CK2 (α/α'/ß) expression, CK2ß colocalization, proximity with IGFBP-1, and CK2 autophosphorylationTyr182 were greater at GD 120 and 165 in MNR vs. C. Additionally, mTOR complex (mTORC)1 (p-P70S6KThr389, -52%, P = 0.05) and mTORC2 (p-AktSer473, -56%, P < 0.001) activity were decreased and AAR was activated (p-GCN2Thr898, +117%, P = 0.02; p-eIF2αSer51, +294%, P = 0.002; p-ERKThr202, +111%, P = 0.03) in MNR liver at GD 120. Our data suggest that fetal liver IGFBP-1 hyperphosphorylation, mediated by mTOR inhibition and both AAR and CK2 activation, is a key link between restricted nutrient and oxygen availability and the development of IUGR.

Strengths and validity of three methods for assessing rat body fat across the life course.

There are several different methods available for the determination of body fat composition. Two current methods requiring special instrumentation are magnetic resonance imaging (MRI) and dual energy x-ray absorptiometry (DXA). The use of these techniques is very limited despite desirable properties, due to their high costs. Dissection of all fat depots (DF) requires no special instrumentation and allows examination and evaluation of each fat depot in more detail. MRI, DXA, and DF each have their unique advantages and disadvantages when they are applied to animal models. Most studies have determined body fat in young animals, and few studies have been performed in aging models. The aim of this study was to compare MRI, DXA, and DF data in offspring (F1) of mothers fed with control and high-fat diet. We studied rats that varied by age, sex, and maternal diet. The relationships between the three methods were determined via linear regression methods (using log-transformed values to accommodate relativity in the relationships), incorporating when useful age, sex, or diet of the animal. We conclude that the three methods are comparable for measuring body fat, but that direct equivalence gets masked by age, sex, and sometimes dietary group. Depending on the equipment available, the budget of the laboratory, and the nature of the research questions, different approaches may often suggest themselves as the best one.

Maternal obesity impairs fetal cardiomyocyte contractile function in sheep.

Obesity is a major public health problem worldwide. In the United States, one-third of women of reproductive age are obese. Human studies show that maternal obesity (MO) predisposes offspring to cardiovascular disease. However, the underlying mechanisms remain unclear. Given the similarities between pregnancy in sheep and humans, we studied sheep to examine the impact of MO on fetal cardiomyocyte contractility at term. We observed that MO impaired cardiomyocyte contractility by reducing peak shortening and shortening/relengthening velocity, prolonging time to relengthening. MO disrupted Ca2+ homeostasis in fetal cardiomyocytes, increasing intracellular Ca2+ and inducing cellular Ca2+ insensitivity. The Ca2+-release channel was impaired, but Ca2+ uptake was unaffected by MO. The upstream kinases that phosphorylate the Ca2+-release channel-ryanodine receptor-2, PKA, and calmodulin-dependent protein kinase II-were activated in MO fetuses. Contractile dysfunction was associated with an increased ratio of myosin heavy chain (MHC)-ß to MHC-α and upregulated cardiac troponin (cTn)-T and tropomyosin, as well as cTn-I phosphorylation. In summary, this is the first characterization of the effects of MO on fetal cardiomyocyte contractility. Our findings indicate that MO impairs fetal cardiomyocyte contractility through altered intracellular Ca2+ handling, overloading fetal cardiomyocyte intracellular Ca2+ and aberrant myofilament protein composition. These mechanisms may contribute to developmental programming by MO of offspring cardiac function and predisposition to later life cardiovascular disease in the offspring.-Wang, Q., Zhu, C., Sun, M., Maimaiti, R., Ford, S. P., Nathanielsz, P. W., Ren, J., Guo, W. Maternal obesity impairs fetal cardiomyocyte contractile function in sheep.

Maternal obesity accelerates rat offspring metabolic ageing in a sex-dependent manner.

KEY POINTS: Maternal obesity predisposes to metabolic dysfunction in male and female offspring Maternal high-fat diet consumption prior to and throughout pregnancy and lactation accelerates offspring metabolic ageing in a sex-dependent manner This study provides evidence for programming-ageing interactions ABSTRACT: Human epidemiological studies show that maternal obesity (MO) shortens offspring life and health span. Life course cellular mechanisms involved in this developmental programming-ageing interaction are poorly understood. In a well-established rat MO model, female Wistar rats ate chow (controls (C)) or high energy, obesogenic diet to induce MO from weaning through pregnancy and lactation. Females were bred at postnatal day (PND) 120. Offspring (F1 ) of mothers on control diet (CF1 ) and MO diet (MOF1 ) delivered spontaneously at terms. Both CF1 and MOF1 ate C diet from weaning throughout the study. Offspring were killed at PND 36, 110, 450 and 650. We determined body and liver weights, liver and serum metabolite concentrations, hormones and oxidative stress biomarkers. Male and female CF1 body weight, total fat, adiposity index, serum leptin, insulin, insulin resistance, and liver weight, fat, triglycerides, malondialdehyde, reactive oxygen species and nitrotyrosine all rose with differing ageing trajectories. Female CF1 triglycerides were unchanged with age. Age-related increases were greater in MOF1 than CF1 in both sexes for all variables except glucose in males and females and cholesterol in males. Cholesterol fell in CF1 females but not MOF1 . Serum corticosterone levels were higher in male and female MOF1 than CF1 and declined with age. DHEA serum levels were lower in male and female MOF1 than CF1 . Liver antioxidant enzymes decreased with age (CF1 and MOF1 ). CONCLUSIONS: exposure to the developmental challenge of MO accelerates progeny ageing metabolic and endocrine profiles in a sex specific manner, providing evidence for programming-ageing interactions.

Role of pregnancy and obesity on vitamin D status, transport, and metabolism in baboons.

Human studies show that obesity is associated with vitamin D insufficiency, which contributes to obesity-related disorders. Our aim was to elucidate the regulation of vitamin D during pregnancy and obesity in a nonhuman primate species. We studied lean and obese nonpregnant and pregnant baboons. Plasma 25-hydroxy vitamin D (25-OH-D) and 1α,25-(OH)2-D metabolites were analyzed using ELISA. Vitamin D-related gene expression was studied in maternal kidney, liver, subcutaneous fat, and placental tissue using real-time PCR and immunoblotting. Pregnancy was associated with an increase in plasma bioactive vitamin D levels compared with nonpregnant baboons in both lean and obese groups. Pregnant baboons had lower renal 24-hydroxylase CYP24A1 protein and chromatin-bound vitamin D receptor (VDR) than nonpregnant baboons. In contrast, pregnancy upregulated the expression of CYP24A1 and VDR in subcutaneous adipose tissue. Obesity decreased vitamin D status in pregnant baboons (162 ± 17 vs. 235 ± 28 nM for 25-OH-D, 671 ± 12 vs. 710 ± 10 pM for 1α,25-(OH)2-D; obese vs. lean pregnant baboons, P < 0.05). Lower vitamin D status correlated with decreased maternal renal expression of the vitamin D transporter cubulin and the 1α-hydroxylase CYP27B1. Maternal obesity also induced placental downregulation of the transporter megalin (LRP2), CYP27B1, the 25-hydroxylase CYP2J2, and VDR. We conclude that baboons represent a novel species to evaluate vitamin D regulation. Both pregnancy and obesity altered vitamin D status. Obesity-induced downregulation of vitamin D transport and bioactivation genes are novel mechanisms of obesity-induced vitamin D regulation.

Effect of maternal baboon (Papio sp.) dietary mismatch in pregnancy and lactation on post-natal offspring early life phenotype.

BACKGROUND: Non-human primate models of developmental programing by maternal mismatch between pregnancy and lactation diets are needed for translation to human programing outcomes. We present baboon offspring morphometry from birth to 3 years, and blood cortisol and adrenocorticotropin (ACTH) from 2 to 24 months. METHODS: Control mothers ate chow; mismatch mothers ate 30% less than controls during pregnancy and high-fat high-energy diet through lactation. RESULTS: Mismatch mothers lost weight during pregnancy. At birth, there were trends toward lower weight in mismatch offspring of both sexes (P = 0.06). From 0-3 years, catch-up growth occurred. Mismatch offspring male and female body weight increased faster than controls (P < 0.001). Mismatch female offspring showed greater increase in BMI (P < 0.001) and abdominal circumference (P = 0.008) vs controls. ACTH and cortisol slopes from 2 to 24 months of age were similar between groups in both sexes. Cortisol and ACTH increased after weaning in all groups. CONCLUSIONS: Mismatch produces sexually dimorphic post-natal growth phenotypes.

Effect of maternal obesity on fetal and postnatal baboon (Papio species) early life phenotype.

BACKGROUND: Non-human primate models of developmental programming by maternal obesity (MO) are needed for translation to human programming outcomes. We present baboon offspring (F1) morphometry, blood cortisol, and adrenocorticotropic hormone (ACTH) from 0.9 gestation to 0-2 years. METHODS: Control mothers ate chow; MO mothers ate high-fat high-energy diet pre-pregnancy through lactation. RESULTS: Maternal obesity mothers weighed more than controls pre-pregnancy. Maternal obesity gestational weight gain was lower with no correlation with fetal or placenta weights. At 0.9 gestation, MO and control F1 morphometry and ACTH were similar. MO-F1 0.9 gestation male cortisol was lower, rising slower from 0-2 years vs control-F1. At birth, male MO-F1 and control-F1 weights were similar, but growth from 0-2 years was steeper in MO-F1; newborn female MO-F1 weighed more than control-F1 but growth from 0-2 years was similar. ACTH did not change in either sex. CONCLUSIONS: Maternal obesity produced sexually dimorphic fetal and postnatal growth and hormonal phenotypes.