Chronic maternal protein deprivation in mice is associated with overexpression of the cohesin-mediator complex in liver of their offspring

Epigenetic mechanisms may play an important role in the developmental programming of adult-onset chronic metabolic diseases resulting from suboptimal fetal nutrition, but the exact molecular mechanisms are incompletely understood. Given the central role of the liver in metabolic regulation, we investigated whether chronic maternal dietary protein restriction has long-term effects on liver gene expression in the offspring. We fed adult C57BL/6J dams ad libitum an 8% maternal lowprotein (MLP) or 20% protein control diet (C) from 4 wk prior to mating until the end of lactation. Male pups were weaned to standard nonpurified diet and singly housed at 21 d of age (d 21). Body weights were followed to 1 y of age (1 y). At d 21 and 1 y, organs were quantitatively dissected and analyzed. MLP offspring had significantly lower body weights at all ages and significantly lower serum activity of alanine aminotransferase and lactate dehydrogenase at 1 y. Gene expression profiling of liver at 1 y showed 521 overexpressed and 236 underexpressed genes in MLP compared to C offspring. The most important novel finding was the overexpression of genes found in liver that participate in organization and maintenance of higher order chromatin architecture and regulation of transcriptional activation. These included members of the cohesinmediator complex, which regulate gene expression by formingDNA loops between promoters and enhancers in a cell typespecific fashion. Thus, our findings of increased expression of these factors in liver of MLP offspring implicate a possible novel epigenetic mechanism in developmental programming. © 2011 American Society for Nutrition

Deletion of Porcn in Mice Leads to Multiple Developmental Defects and Models Human Focal Dermal Hypoplasia (Goltz Syndrome)

Focal Dermal Hypoplasia (FDH) is a genetic disorder characterized by developmental defects in skin, skeleton and ectodermal appendages. FDH is caused by dominant loss-of-function mutations in X-linked PORCN. PORCN orthologues in Drosophila and mice encode endoplasmic reticulum proteins required for secretion and function of Wnt proteins. Wnt proteins play important roles in embryo development, tissue homeostasis and stem cell maintenance. Since features of FDH overlap with those seen in mouse Wnt pathway mutants, FDH likely results from defective Wnt signaling but molecular mechanisms by which inactivation of PORCN affects Wnt signaling and manifestations of FDH remain to be elucidated.We introduced intronic loxP sites and a neomycin gene in the mouse Porcn locus for conditional inactivation. Porcn-ex3-7flox mice have no apparent developmental defects, but chimeric mice retaining the neomycin gene (Porcn-ex3-7Neo-flox) have limb, skin, and urogenital abnormalities. Conditional Porcn inactivation by EIIa-driven or Hprt-driven Cre recombinase results in increased early embryonic lethality. Mesenchyme-specific Prx-Cre-driven inactivation of Porcn produces FDH-like limb defects, while ectodermal Krt14-Cre-driven inactivation produces thin skin, alopecia, and abnormal dentition. Furthermore, cell-based assays confirm that human PORCN mutations reduce WNT3A secretion.These data indicate that Porcn inactivation in the mouse produces a model for human FDH and that phenotypic features result from defective WNT signaling in ectodermal- and mesenchymal-derived structures

Correction: Chronic Maternal Low-Protein Diet in Mice Affects Anxiety, Night-Time Energy Expenditure and Sleep Patterns, but Not Circadian Rhythm in Male Offspring.

[This corrects the article DOI: 10.1371/journal.pone.0170127.]

Chronic Maternal Low-Protein Diet in Mice Affects Anxiety, Night-Time Energy Expenditure and Sleep Patterns, but Not Circadian Rhythm in Male Offspring

<div><p>Offspring of murine dams chronically fed a protein-restricted diet have an increased risk for metabolic and neurobehavioral disorders. Previously we showed that adult offspring, developmentally exposed to a chronic maternal low-protein (MLP) diet, had lower body and hind-leg muscle weights and decreased liver enzyme serum levels. We conducted energy expenditure, neurobehavioral and circadian rhythm assays in male offspring to examine mechanisms for the body-weight phenotype and assess neurodevelopmental implications of MLP exposure. C57BL/6J dams were fed a protein restricted (8%protein, MLP) or a control protein (20% protein, C) diet from four weeks before mating until weaning of offspring. Male offspring were weaned to standard rodent diet (20% protein) and single-housed until 8–12 weeks of age. We examined body composition, food intake, energy expenditure, spontaneous rearing activity and sleep patterns and performed behavioral assays for anxiety (open field activity, elevated plus maze [EPM], light/dark exploration), depression (tail suspension and forced swim test), sociability (three-chamber), repetitive (marble burying), learning and memory (fear conditioning), and circadian behavior (wheel-running activity during light-dark and constant dark cycles). We also measured circadian gene expression in hypothalamus and liver at different Zeitgeber times (ZT). Male offspring from separate MLP exposed dams had significantly greater body fat (P = 0.03), less energy expenditure (P = 0.004), less rearing activity (P = 0.04) and a greater number of night-time rest/sleep bouts (P = 0.03) compared to control. MLP offspring displayed greater anxiety-like behavior in the EPM (P<0.01) but had no learning and memory deficit in fear-conditioning assay (P = 0.02). There was an effect of time on <i>Per1</i>, <i>Per 2</i> and <i>Clock</i> circadian gene expression in the hypothalamus but not on circadian behavior. Thus, transplacental and early developmental exposure of dams to chronic MLP reduces food intake and energy expenditure, increases anxiety like behavior and disturbs sleep patterns but not circadian rhythm in adult male offspring.</p></div

Body composition.

<p>(A): Body weight; (B) Body fat as percent of total body mass in male offspring (8–12 weeks age) from dams exposed to either control or MLP diet (n = 7 each). Bars are mean ± SEM and P<0.05 statistically significant by student t-test.</p

MLP male offspring mice display anxiety-like but not depression-like behavior.

<p>Elevated plus maze test (MLP and Control, n = 9 each); (A) Time spent in open arm, center and closed arm, (B) Latency to enter closed arm. (C) Tail suspension test (MLP, n = 31 and Control, n = 13) and (D) Forced swim test (MLP and Control, n = 20 each). Data is presented as mean ± SEM. P<0.05 was considered statistically significant by student t-test.</p

Food intake and energy expenditure in 12-wk-old MLP and Control offspring showing values over 24 hours, and the distribution between light and dark phases.

<p>Food intake and energy expenditure in 12-wk-old MLP and Control offspring showing values over 24 hours, and the distribution between light and dark phases.</p