Effect of heme on PPARα target genes.

<p><b>A.</b> Categorization of heme-induced differentially expressed genes (q<0.01 and signal intensity>20 in at least treatment) according to GO Biological Process annotation. Figure is based on results from IJssennagger et al. <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0043260#pone.0043260-IJssennagger1" target="_blank">[5]</a> showing that lipid metabolism-related gene expression is substantially influenced by heme. Thirty percent of these heme-induced lipid metabolism-related genes are PPARα target genes <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0043260#pone.0043260-Bunger1" target="_blank">[18]</a>. Miscellaneous contains processes with broad and thus unspecific biological process terms. <b>B.</b> Expression of PPARα target genes in enterocytes is mainly upregulated. Behind the gene the fold- changes are indicated from colonic scrapings from heme fed vs. control mice from resp. the previous experiment with C57Bl6J mice, current experiment WT SV129 mice and current experiment KO SV129 mice. In bold are PPARα targets of which no significant induction is seen in the KO mice.</p

Physiological changes induced by heme in colon of WT and PPARα KO mice.

#<p>Calculated as percentage Ki67 positive cells per crypt. Data are represented as mean ± SEM. Groups indicated with ‘a’ are significantly (<i>P</i><0.05) different from ‘b’ by ANOVA with Bonferroni post-hoc testing. N = 6 per group, except for mucosa measurements where proliferation of one KO heme animal could not be determined due to poor tissue quality.</p

Gene expression of signaling molecules involved in hyperproliferation (A) and of PPARα targets Cyp4a10 and Fabp1 (B).

<p>Expression of the WT control group is set to one. Expression of all other groups is relative to WT control. P-values for main effects (D for diet, G for genotype and I for interaction) by a two-way ANOVA are indicated. A and b indicate significant different groups (p<0.05) determined by a Bonferroni post hoc-test.</p

Microarray clustering and pathway analysis of heme-induced differentially expressed genes.

<p><b>A.</b> Hierarchical clustering of the microarray data showing that the diet-effect (C = control and H = heme) is more pronounced than the genotype-effect. <b>B.</b> Venn diagram showing that 69% of the heme-induced changes (q<0.01 and signal intensity>20 in at least one of the treatments) in WT mice could also be found in KO mice. Ingenuity canonical pathway analysis shows that overlapping genes in Venn-diagram are involved in cell cycle-related processes (open arrows) and Nrf2-mediated oxidative stress response (gray arrow). There is hardly any effect of genotype on fatty acid metabolism-related processes (black arrow). WT mice show PPARα activation (black arrow in WT panel) and KO mice Wnt signaling (white arrow in KO panel). Note that pathways in overlap are much more significant than the WT or KO specific pathways.</p

Heme-induced stress response.

<p><b>A.</b> Representative pictures of H&E staining of mouse colonic mucosa after 14 days of control- versus heme diet. <b>B.</b> Representative pictures of colon tissue stained for Alkaline phosphatase activity, a marker for ROS stress. <b>C.</b> Expression of Vanin-1. <b>D.</b> Expression of genes involved in antioxidant response Nqo1, Cat, Gclc and Fosl1. Expression of the WT control group is set to one. Expression of all other groups is relative to WT control. P-values for main effects (D for diet, G for genotype and I for interaction) by a two-way ANOVA are indicated. A,b and c indicate significant different groups (p<0.05) determined by a Bonferroni post hoc-test.</p

Relative expression of Proglucagon mRNA (gray bar) and GLP-1 protein (black bar) in colonic mucosa of a random selection of control fed and FOS fed rats

mRNA and protein levels were normalized to Actin levels. Expression is shown as means ± SEM (n = 7). **p < 0.01, ***P < 0.001<p><b>Copyright information:</b></p><p>Taken from "Impaired barrier function by dietary fructo-oligosaccharides (FOS) in rats is accompanied by increased colonic mitochondrial gene expression"</p><p>http://www.biomedcentral.com/1471-2164/9/144</p><p>BMC Genomics 2008;9():144-144.</p><p>Published online 27 Mar 2008</p><p>PMCID:PMC2311291.</p><p></p

Differential effect of deoxycholate (A) and of control and heme fecal water (B) on viability of Gram-negative (G−) <i>E.coli</i> and Gram-positive (G+) <i>L. plantarum</i>.

<p>Fecal water pools from four separate, but identical, animal experiments were used in triplicate. Viability is measured relative to PBS (A) and control fecal water (B). Data are presented as mean ± SEM, (n = 4) *p<0.05.</p

Quantification of total bacteria and specific functional genes in colonic samples from control and heme-fed mice.

<p>Data are represented as mean ± SEM, (n = 8).</p>*<p>indicates significant change between control and heme (p<0.05),</p>**<p>indicates significant change between control and heme (p<0.001).</p><p>For ratio calculation, the in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0049868#pone-0049868-t002" target="_blank">Table 2</a> quantified Actinobacteria, Firmicutes and TM7 were included as Gram-positives and Bacteroidetes, Deferribacterres, Fibrobacteres, Fusobacteria, Proteobacteria and Verrucomicrobia were included as Gram-negatives.</p><p><a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0049868#s3" target="_blank">Results</a> are expressed in log10 of gene copy-nr/g fecal dry weight. Genes used for quantification are mentioned between brackets.</p

Phylogenetic fingerprints of the colonic microbiota of the control group and heme group.

<p>Gel-view figure of the intensity of 3,580 probes covered by MITChip and assigned to the phylogenetic class-like groups (level 1) depicted on the right side. Ward's minimum variance method was used to generate hierarchical clustering of the total microbiota probe profiles, whereas the distance matrix between the samples was based on the Pearson's product moment correlation.</p

Histological staining of mouse colonic mucosa after 14 days on control or heme diet.

<p>Histological staining was performed for H&E, AB-PAS, HID and Alpi. Ki67-immunohistochemistry was performed using a Ki67-specific antibody.</p