Phenotypic and metabolic characteristics of the mice after 24 weeks on the CD and HFD, for the F1LM (CD1, OP1 and OR1) and F2OM (CD2, OP2 and OR2) mice.

<p>The values presented are the means±SEM. For weight and caloric intake, <i>n</i> = 50 to 70, for plasma hormone and metabolite determinations, <i>n</i> = 10 to 15, for FA level determination, <i>n</i> = 7, and for Piximus analysis and organ weights, <i>n</i> = 5.</p>a<p><i>p</i><0.05 for comparison with control,</p>b<p><i>p</i><0.05 for comparison between OP and OR mice,</p>c<p><i>p</i><0.05 for comparison between the F1LM and F2OM mice, assessed by Kruskal-Wallis tests followed by Dunn’s <i>post hoc</i> tests.</p

Analysis, by RT-qPCR, of the expression of genes encoding DNA methyltransferase enzymes in the liver and muscle of mice fed the HFD, born to either lean or obese/diabetic mothers (F1LM and F2OM).

<p>The values shown are the ratios between OP1 or OR1 and CD1 and between OP2 or OR2 and CD2. They are expressed as the mean±SEM, <i>n</i> = 8 per group. ^a<i>p</i><0.05 for comparison with control CD, ^b<i>p</i><0.05 for comparison between OP and OR mice, ^c<i>p</i><0.05 for comparison between the F1LM and F2OM born to lean and obese/diabetic mothers, respectively, assessed by Kruskal-Wallis tests and <i>post hoc</i> Dunn’s tests.</p

Analysis of mRNA levels for key adipogenic genes by RT-qPCR on the adipose tissue of mice fed either the CD or the HFD, born to either lean or obese/diabetic mothers (F1LM and F2OM).

<p>The values shown are the ratios between OP1 or OR1 and CD1 and between OP2 or OR2 and CD2 (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0066816#pone-0066816-g009" target="_blank">Figures 9</a> and <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0066816#pone-0066816-g010" target="_blank">10</a>). They are expressed as the mean±SEM, <i>n</i> = 6 per group. ^a<i>p</i><0.05 for comparison with control CD, ^b<i>p</i><0.05 for comparison between OP and OR mice, ^c<i>p</i><0.05 for comparison between F1LM and F2OM, assessed by Kruskal-Wallis tests followed by <i>post hoc</i> Dunn’s tests.</p

Heat map construction representing the differential expression of genes involved in metabolic function and neurotransmission.

<p>The heat map shows changes in the hepatic expression of genes encoding proteins involved in neurotransmission and genes encoding energy homeostasis-related proteins. Red, green and white squares represent upregulated, downregulated and unmodified genes, respectively.</p

Schematic diagram summarizing the transcriptional data obtained for the liver, muscle and adipose tissue and the results of epigenetic studies in OP1, OP2 and OR2 mice.

<p>Blue arrows represent potential lipid fluxes. In the hyperphagic OP1 and OP2 mice, lipid ingestion was much greater than in OR2 mice, in which caloric intake was normalized. In OP mice, excess lipids were initially stored in the adipose tissue, leading to adipocyte hypertrophy in OP1 mice and hyperplasia in OP2 mice, as a function of the level of expression of <i>Lep</i> and <i>Peg1</i>. In OR2 mice, lipids were stored in the adipose tissue without adipocyte abnormalities or ectopic storage, as in mice supplied with limited amounts of lipid. In OP1 mice, the excess lipids were stored in the liver, contributing to hepatic hypertrophy. Transcriptomic data indicated that <i>de novo</i> lipogenesis was activated in OP1 mice and that insulin signaling was greatly disturbed. In OP2 mice, genes related to insulin signaling were less affected, whereas genes involved in fatty acid oxidation were globally upregulated. Changes to hepatic metabolism, together with the probable redirection of lipids to muscle thus spared the liver from lipid accumulation. Finally, in OR2 mice, lipid metabolism as a whole was downregulated, whereas thyroid hormone signaling was upregulated. The HFD response was also associated, in OP1 mice, with an upregulation of potassium channels and serotonin receptors, subsequently reversed in both OP2 and OR2 mice. Changes in DNA methylation were observed in the livers of OP1 mice and the muscle of OP2 mice. In the liver, <i>Set7/9</i> expression was decreased by the HFD in mice born to either lean or obese/diabetic mothers (F1LM and F2OM), whether OP or OR. In the livers of OP1 mice, DNA hypomethylation was associated with an upregulation of <i>Suv39h1</i> and <i>Suv39h2</i> expression, whereas, in both OR2 and OP2 mice, normal DNA methylation was associated with a decrease in <i>Jhdm2a</i> expression and an increase in the level of <i>Dnmt2</i> mRNA. In muscle, normal DNA methylation was associated with an upregulation of <i>Suv39h1, Set7/9</i> and <i>Dnmt2</i> in OP1 and OR2 mice, contrasting with the lower level of expression of the <i>Set7/9</i> and <i>Dnmt2</i> genes in the muscle of OP2 mice presenting DNA hypomethylation.</p

Analysis, by RT-qPCR, of the expression of genes encoding histone-modifying enzymes in the liver and muscle of mice fed the HFD, for both lean and obese mothers.

<p>The values shown are the ratios between OP1 or OR1 and CD1 and between OP2 or OR2 and CD2. They are expressed as the mean±SEM, <i>n</i> = 8 per group. ^a<i>p</i><0.05 for comparison with control CD, ^b<i>p</i><0.05 for comparison between OP and OR mice, ^c<i>p</i><0.05 for comparison between the F1LM and F2OM (maternal effect: lean versus obese mother), assessed by Kruskal-Wallis tests and <i>post hoc</i> Dunn’s tests.</p

Schematic diagram summarizing the step-by-step evolution of the metabolic phenotype of OR and OP mice in the F1LM and F2OM offspring.

<p>Schematic diagram summarizing the step-by-step evolution of the metabolic phenotype of OR and OP mice in the F1LM and F2OM offspring.</p

Maternal effect (lean versus obese mother): Relevant functions and diseases, and representation of the major network of interaction identified by IPA analysis of the genes differentially expressed between OP1 and OP2 obese mice.

<p>The 35 genes differentially expressed between the obese mice born to lean and obese/diabetic mothers (F1LM and F2OM) obtained in the direct OP2/OP1 comparison were subjected to IPA analysis. The network shown contains 20 focal genes with a score of 50. The node color indicates the expression levels of genes: red, upregulated; green, downregulated.</p

Resistance to the obesogenic effects of the HFD: Major networks identified by IPA analysis of the genes involved in the response and adaptation to HFD of OR2 mice.

<p>(A) These networks were built from the 142 genes differentially expressed between OR2 and CD2 mice (direct comparison OR2/CD2). Only the first two networks are represented in (B) and (C). The node color indicates the level of expression of the genes: red, upregulated; green, downregulated.</p

Heat map construction representing differentially expressed imprinted genes.

<p>(A) The heat map shows the changes in expression of imprinted genes in the liver in response to the HFD (OP1/CD1, OP2/CD2, OR2/CD2), with the trait of susceptibility/resistance to the obesogenic effects of HFD (OR2/OP2; indirect comparisons OR2/CD2 and OP2/CD2), or with a maternal effect (lean versus obese mother) (OP1/OP2); indirect comparisons (OP1/CD1 and OP2/CD2). Red, green and white squares represent upregulated, downregulated and unmodified genes, respectively. The status of the imprinted gene and the preferred parental allele for gene expression presented in this table at this time may subsequently be modified, as knowledge in this domain increases. Updates accessible via <a href="http://www.geneimprint.com" target="_blank">www.geneimprint.com</a>). (B) Representation of the IGN network as described by Varrault <i>et. </i><i>al. </i><a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0066816#pone.0066816-Varrault1" target="_blank">[50]</a>. Red circles indicate the genes modulated in OR2 mice in response to the HFD and associated with maintenance of the lean phenotype (comparisons OR2/CD2 and OR2/OP2).</p