The negative impact of nutritional deficits in the development of bronchopulmonary dysplasia is well recognized, yet mechanisms by which nutrition alters lung outcomes and nutritional strategies that optimize development and protect the lung remain elusive. Here, we use a rat model to assess the isolated effects of postnatal nutrition on lung structural development without concomitant lung injury. We hypothesize that postnatal growth restriction (PGR) impairs lung structure and function, critical mediators of lung development, and fatty acid profiles at postnatal day 21 in the rat. Rat pups were cross-fostered at birth to rat dams with litter sizes of 8 (control) or 16 (PGR). Lung structure and function, as well as serum and lung tissue fatty acids, and lung molecular mediators of development, were measured. Male and female PGR rat pups had thicker airspace walls, decreased lung compliance, and increased tissue damping. Male rats also had increased lung elastance, increased lung elastin protein abundance, and lysol oxidase expression, and increased elastic fiber deposition. Female rat lungs had increased conducting airway resistance and reduced levels of docosahexaenoic acid in lung tissue. We conclude that PGR impairs lung structure and function in both male and female rats, with sex-divergent changes in lung molecular mediators of development.

The placenta represents a critical node in fetal lipid acquisition, yet the mechanisms by which the placenta handles lipids under normal and pathologic conditions are incompletely understood. A key player in placental lipid handling is peroxisome proliferator–activated receptor gamma (PPARγ). PPARγ influences global gene expression via its regulation of the epigenetic modifier lysine methyltransferase 5A (KMT5A), which places a methyl group on histone 4 lysine 20 (H4K20me) of target genes. Here we test the hypothesis that KMT5A is present in both the human and rat placentas and is affected by uteroplacental insufficiency (UPI) in the rat in association with increased placental lipid accumulation. We assessed levels and localization of KMT5A, as well as lipid droplet accumulation, in human placental tissue collected from maternal donors after delivery by planned cesarean section. Using a rat model of UPI, we also evaluated the effects of UPI on lipid accumulation, PPARγ, KMT5A, and H4K20me in the rat placenta. In this study, we show for the first time the presence and activity of KMT5A, in human and in rat placentas. We also demonstrate that in the rat placenta, UPI increases hypoxia, KMT5a expression, and activity in association with increased lipid accumulation in placenta supporting male fetuses. Placental PPARγ-KMT5A axis may be an important mediator of placental lipid handling.

Obesity and high-fat (HF) diet are associated with over activation of the endocannabinoid system (ECS). We have demonstrated that maternal HF diet induces early obesity and modulates cannabinoid signaling in visceral (VIS) and subcutaneous (SUB) white adipose tissue (WAT) in weanling rat offspring. We hypothesized that perinatal maternal HF diet would program the expression of ECS in adipose tissue in a long-term way in parallel to alterations in epigenetic markers and sex hormone signaling. Progenitor female rats received control diet (C, 9% fat) or isocaloric high-fat diet (HF, 28% fat) for 8 weeks before mating, gestation, and lactation. All pups were weaned to C diet and they were euthanized at 180 days old. Maternal HF diet induced overweight and increased SUB WAT mass of male and female adult offspring. Maternal HF diet induced hypertrophy of VIS and SUB adipocytes only in female offspring associated with increased type 1 cannabinoid receptor protein (CB1) and mRNA (Cnr1) levels. These changes were associated with increased estrogen receptor α binding to Cnr1 promoter in SUB WAT of adult female offspring, which may contribute to higher expression of Cnr1. Increased CB1 signaling in adipose tissue might contribute to higher adiposity programmed by maternal HF diet because endocannabinoids stimulate the accumulation of fat in the adipose tissue. Our findings provide molecular insights into sex-specific targets for anti-obesity therapies based on the endocannabinoid system.

Maternal tobacco smoke exposure (MTS) affects fetal acquisition of long-chain polyunsaturated fatty acids (LCPUFA) and increases the risk of obesity and cardio-metabolic disease in the offspring. Alterations in fetal LCPUFA acquisition in maternal smoking are mediated by the placenta. The handling of LCPUFA by the placenta involves protein-mediated transfer and storage. Molecular mediators of placental LCPUFA handling include PPARγ and the fatty acid transport proteins. We previously demonstrated, in a rat model, that MTS results in programming of adult-onset obesity and metabolic disease in male, but not female, offspring. In this study, we test the hypothesis that in utero MTS exposure alters placental structure, placental LCPUFA handling, and fetal fatty acid levels, in a sex-divergent manner. We exposed pregnant rats to tobacco smoke from embryonic day 11 to term gestation. We measured placental and fetal fatty acid profiles, the systolic/diastolic ratio (SD ratio), placental histology, and expression of molecular mediators in the placenta. Our primary finding is that MTS alters fatty acid profiles in male, but not female fetuses and placenta, including increasing the ratio of omega-6 to omega-3 fatty acids. MTS also increased SD ratio in male, but not female placenta. In contrast, the expression of PPARγ and FATPs was upregulated in female, but not male placenta. We conclude that MTS causes sex-divergent changes in placental handling of LCPUFA in the rat. We speculate that our results demonstrate an adaptive response to MTS by the female placenta.

Maternal nutritional imbalances trigger developmental adaptations involving early epigenetic mechanisms associated with adult chronic disease. Maternal high-fat (HF) diet promotes obesity and hypothalamic leptin resistance in male rat offspring at weaning and adulthood. Leptin resistance is associated with over activation of the endocannabinoid system (ECS). The ECS mainly consists of endocannabinoids derived from n-6 fatty acids and cannabinoid receptors (CB1 coded by Cnr1 and CB2 coded by Cnr2). The CB1 activation in hypothalamus stimulates feeding and appetite for fat while CB2 activation seems to play an immunomodulatory role. We demonstrated that maternal HF diet increases hypothalamic CB1 in male offspring while increases CB2 in female offspring at birth, prior to obesity development. However, the molecular mechanisms behind these changes remain unexplored. We hypothesized that maternal HF diet would down-regulate leptin signaling and up-regulate Cnr1 mRNA levels in the hypothalamus of the offspring at birth, associated with sex-specific changes in epigenetic markers and sex steroid signaling. To test our hypothesis, we used progenitor female rats that received control diet (C, 9% fat) or isocaloric high-fat diet (HF, 28% fat) from 8 weeks before mating until delivery. Blood, hypothalamus and carcass from C and HF male and female offspring were collected for biochemical and molecular analyses at birth. Maternal HF diet down-regulated the transcriptional factor STAT3 in the hypothalamus of male and female offspring, but induced hypoleptinemia only in males and decreased phosphorylated STAT3 only in female offspring. Because leptin acts through STAT3 pathway to inhibit central ECS, our results suggest that leptin pathway impairment might contribute to increased levels of Crn1 mRNA in hypothalamus of both sex offspring. Besides, maternal HF diet increased the histone acetylation percentage of Cnr1 promoter in male offspring and increased the androgen receptor binding to the Cnr1 promoter, which can contribute to higher expression of Cnr1 in newborn HF offspring. Maternal HF diet increased plasma n6 to n3 fatty acid ratio in male offspring, which is an important risk factor to metabolic diseases and might indicate an over activation of endocannabinoid signaling. Thus, although maternal HF diet programs a similar phenotype in adult offspring of both sexes (obesity, hyperphagia and higher preference for fat), here we showed that molecular mechanisms involving leptin signaling, ECS, epigenetic markers and sex hormone signaling were modified prior to obesity development and can differ between newborn male and female offspring. These observations may provide molecular insights into sex-specific targets for anti-obesity therapies.

Fetal outcomes after uteroplacental insufficiency (UPI) include growth restriction and sex-divergent increase in neonatal and adult-onset disease. Fetal acquisition of long-chain polyunsaturated fatty acids (LCPUFA) is 1) linked to disease outcomes, 2) mediated by the placenta, and 3) impaired in pregnancies complicated by UPI. Peroxisome proliferator activated receptor gamma (PPARγ) regulates placental LCPUFA transport via direct control of gene expression and by initiating chromatin modifications through Setd8. We previously showed in our rat model that UPI results in fetal growth restriction and development of sex-divergent neonatal and adult-onset disease. We hypothesize that UPI in the rat results in sex-divergent changes in the placental PPARγ-Setd8 axis and sex-divergent changes in fetal serum LCPUFA profiles. UPI was induced by bilateral uterine artery ligation at embryonic day 19 in pregnant Sprague Dawley rats. Male and female fetal rat serum and corresponding placenta were surgically collected at term (embryonic day 21). Serum LCPUFA profiles were measured using gas chromatography-mass spectrometry. mRNA was measured using real-time RT-PCR, protein was measured using western blotting, and PPARγ occupancy at the Setd8 promoter was measured using ChIP. Data are expressed as mean ± standard deviation (SD), *p<0.05. UPI did not affect PPARγ mRNA levels and protein abundance in male placenta or female placenta compared to sex-matched control. UPI increased Setd8 mRNA levels, protein abundance, and total PPARγ at the promoter of the Setd8 compared to sex-matched control in male placenta but not in female placenta. In male placenta, UPI increased global H4K20me compared to sex-matched control. No differences were observed in female placenta. In male fetuses, UPI increased the LCPUFA palmitic acid (PAL), linoleic acid (LA), and arachidonic acid (AA), while decreasing palmitoleic acid (PA), eicosapentaenoic acid (EPA), and docosahexaenoic acid (DHA). In female fetuses, UPI increased AA with no change in other LCPUFA. In conclusion, UPI in the rat results in sex-divergent changes in the PPARγ-Setd8 axis in association with sex-divergent changes in fetal serum LCPUFA profiles. Given the role of Setd8 in chromatin maintenance, sex-divergent changes in PPARγ-driven Setd8 expression may influence UPI-induced sex-divergent changes in the placental transcriptome and subsequent placental transfer of LCPUFA.

BackgroundPreterm infants receiving respiratory support often experience extrauterine growth restriction (EUGR). EUGR in preterm infants occurs secondary to feeding intolerance and clinically indicated feeding volume restrictions. EUGR increases the severity and incidence of the chronic lung disease of infancy, bronchopulmonary dysplasia (BPD). BPD is characterized by impaired lung development, with decreased lung compliance, and increased lung resistance. Animal models demonstrate the importance of EUGR to lung development in the context of respiratory support. However, the isolated contribution of EUGR to lung development and function is unknown. We hypothesize that, in the juvenile rats, EUGR will impair alveolar development, decrease lung compliance, and increase lung resistance.MethodsWe induced EUGR in a rat model by cross fostering newborn rat pups to rat dams with litter sizes of 16 (EUGR) or 8 (control). Rat pup weight was measured every other day from birth to day of life 21 (d21). At postnatal d24, we measured alveolar wall thickness, second crest septal volume, and radial alveolar count (RAC) using ImageQuant software. At postnatal d24 we also measured lung compliance and resistance of the parenchyma (RPar) and airway (RAir) using a Flexivent System.ResultsResults are EUGR values as a percentage of control±SD (*=p<0.05). Rat pups in the EUGR group weighed significantly less (75±3%*) than control by postnatal d5 and continued to weigh less through d21 (70±5%*). Structural analysis of rat lungs revealed that EUGR rats had thicker alveolar walls than control in male (136±20%*) and female (134±32%*) rats, but no change in sepal density or RAC. EUGR decreased compliance in male (67±26%*) and female (66±25%*) rats. EUGR significantly increased RPar in both male (153±70%*) and female (157±56%*) rats and significantly increased RAir in male (111±25%*) and female (166±148%*) rats.ConclusionEUGR alters lung structure and function in juvenile rats. Given the role of elastin and elastic fiber formation in lung structure and function, we speculate that EUGR may disrupt elastin expression and fiber deposition. Ongoing studies are evaluating expression of elastin isoforms in the EUGR lung.Support or Funding InformationSpecial thanks to the American Physiological Society for the Undergraduate Summer Research Fellowship that funded this research, and to Lisa Joss‐Moore and the University of Utah for hosting me in their lab. In addition, thanks you to James Zhou and Haimei Wang for help with collection of mechanics data, as well as Katie Zuspan and Andrew Rebentisch for conducting image analysis for structural data.