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Arterial endothelial methylome: differential DNA methylation in athero-susceptible disturbed flow regions in vivo.

Jiang YZ, Manduchi E, Stoeckert CJ, Davies PF - BMC Genomics (2015)

Bottom Line: Gender-specific DMRs associated with ciliogenesis that may be linked to defects in cilia development were also identified in AA DMRs.An endothelial methylome analysis identifies epigenetic DMR characteristics associated with transcriptional regulation in regions of atherosusceptibility in swine aorta in vivo.The data represent the first methylome blueprint for spatio-temporal analyses of lesion susceptibility predisposing to endothelial dysfunction in complex flow environments in vivo.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology & Laboratory Medicine and Institute for Medicine & Engineering, Perelman School of Medicine, University of Pennsylvania, 1010 Vagelos Building, 3340 Smith Walk, Philadelphia, PA, 19104, USA. jyizhou@mail.med.upenn.edu.

ABSTRACT

Background: Atherosclerosis is a heterogeneously distributed disease of arteries in which the endothelium plays an important central role. Spatial transcriptome profiling of endothelium in pre-lesional arteries has demonstrated differential phenotypes primed for athero-susceptibility at hemodynamic sites associated with disturbed blood flow. DNA methylation is a powerful epigenetic regulator of endothelial transcription recently associated with flow characteristics. We investigated differential DNA methylation in flow region-specific aortic endothelial cells in vivo in adult domestic male and female swine.

Results: Genome-wide DNA methylation was profiled in endothelial cells (EC) isolated from two robust locations of differing patho-susceptibility:--an athero-susceptible site located at the inner curvature of the aortic arch (AA) and an athero-protected region in the descending thoracic (DT) aorta. Complete methylated DNA immunoprecipitation sequencing (MeDIP-seq) identified over 5500 endothelial differentially methylated regions (DMRs). DMR density was significantly enriched in exons and 5'UTR sequences of annotated genes, 60 of which are linked to cardiovascular disease. The set of DMR-associated genes was enriched in transcriptional regulation, pattern specification HOX loci, oxidative stress and the ER stress adaptive pathway, all categories linked to athero-susceptible endothelium. Examination of the relationship between DMR and mRNA in HOXA genes demonstrated a significant inverse relationship between CpG island promoter methylation and gene expression. Methylation-specific PCR (MSP) confirmed differential CpG methylation of HOXA genes, the ER stress gene ATF4, inflammatory regulator microRNA-10a and ARHGAP25 that encodes a negative regulator of Rho GTPases involved in cytoskeleton remodeling. Gender-specific DMRs associated with ciliogenesis that may be linked to defects in cilia development were also identified in AA DMRs.

Conclusions: An endothelial methylome analysis identifies epigenetic DMR characteristics associated with transcriptional regulation in regions of atherosusceptibility in swine aorta in vivo. The data represent the first methylome blueprint for spatio-temporal analyses of lesion susceptibility predisposing to endothelial dysfunction in complex flow environments in vivo.

No MeSH data available.


Related in: MedlinePlus

Functional annotations of genes associated with athero-susceptible DMRs. a: Ingenuity Pathway Analysis of DMR-associated genes. The top five pathways and associated genes are listed. b: Gene ontology (GO) Biological Processes and c: Molecular Function categories enriched for DMR-associated genes. The enrichment analysis was performed using online tool DAVID (the Database for Annotation, Visualization and Integrated Discovery). The p-value for each GO term was adjusted by using Benjamini-Hochberg for multiple-testing correction. In b and c, categories are ranked according to the number of genes containing hypermethylated (black bar) and hypomethylated (grey bar) DMRs in AA
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Fig5: Functional annotations of genes associated with athero-susceptible DMRs. a: Ingenuity Pathway Analysis of DMR-associated genes. The top five pathways and associated genes are listed. b: Gene ontology (GO) Biological Processes and c: Molecular Function categories enriched for DMR-associated genes. The enrichment analysis was performed using online tool DAVID (the Database for Annotation, Visualization and Integrated Discovery). The p-value for each GO term was adjusted by using Benjamini-Hochberg for multiple-testing correction. In b and c, categories are ranked according to the number of genes containing hypermethylated (black bar) and hypomethylated (grey bar) DMRs in AA

Mentions: Additional file 2: Table S1a details athero-susceptible DMRs, including genomic coordinates, length, distance to the nearest transcripts and methylation fold change. We found 632 pig Ensembl genes that contain at least one DMR in the proximal promoter region (2 kb upstream to 1 kb downstream TSS) or distal regulatory region (up to 10kb upstream) as shown in Additional file 3: Table S1b. Of these pig genes, 431 have human homologs. 60 DMR-associated genes were found by Ingenuity Pathway Analysis (IPA) to be linked to cardiovascular disease in the top 5 Disease and Dysfunction category (Table 1). Previously we reported that chronic low level activation of ER stress, unfolded protein response and oxidative stress are associated with athero-susceptible endothelium [6, 7, 10]. Genes related to superoxide radical degradation and ER stress were over-represented among those containing athero- susceptible DMRs (Fig. 5a) supporting a role for DMRs in ROS and ER mechanisms in prelesional AA regions. Athero-susceptible DMRs were also associated with genes of the matrix metalloproteinase pathway and vitamin D receptor activation pathway, both of which have been shown to play critical roles in cardiovascular disease [34, 35].Table 1


Arterial endothelial methylome: differential DNA methylation in athero-susceptible disturbed flow regions in vivo.

Jiang YZ, Manduchi E, Stoeckert CJ, Davies PF - BMC Genomics (2015)

Functional annotations of genes associated with athero-susceptible DMRs. a: Ingenuity Pathway Analysis of DMR-associated genes. The top five pathways and associated genes are listed. b: Gene ontology (GO) Biological Processes and c: Molecular Function categories enriched for DMR-associated genes. The enrichment analysis was performed using online tool DAVID (the Database for Annotation, Visualization and Integrated Discovery). The p-value for each GO term was adjusted by using Benjamini-Hochberg for multiple-testing correction. In b and c, categories are ranked according to the number of genes containing hypermethylated (black bar) and hypomethylated (grey bar) DMRs in AA
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4492093&req=5

Fig5: Functional annotations of genes associated with athero-susceptible DMRs. a: Ingenuity Pathway Analysis of DMR-associated genes. The top five pathways and associated genes are listed. b: Gene ontology (GO) Biological Processes and c: Molecular Function categories enriched for DMR-associated genes. The enrichment analysis was performed using online tool DAVID (the Database for Annotation, Visualization and Integrated Discovery). The p-value for each GO term was adjusted by using Benjamini-Hochberg for multiple-testing correction. In b and c, categories are ranked according to the number of genes containing hypermethylated (black bar) and hypomethylated (grey bar) DMRs in AA
Mentions: Additional file 2: Table S1a details athero-susceptible DMRs, including genomic coordinates, length, distance to the nearest transcripts and methylation fold change. We found 632 pig Ensembl genes that contain at least one DMR in the proximal promoter region (2 kb upstream to 1 kb downstream TSS) or distal regulatory region (up to 10kb upstream) as shown in Additional file 3: Table S1b. Of these pig genes, 431 have human homologs. 60 DMR-associated genes were found by Ingenuity Pathway Analysis (IPA) to be linked to cardiovascular disease in the top 5 Disease and Dysfunction category (Table 1). Previously we reported that chronic low level activation of ER stress, unfolded protein response and oxidative stress are associated with athero-susceptible endothelium [6, 7, 10]. Genes related to superoxide radical degradation and ER stress were over-represented among those containing athero- susceptible DMRs (Fig. 5a) supporting a role for DMRs in ROS and ER mechanisms in prelesional AA regions. Athero-susceptible DMRs were also associated with genes of the matrix metalloproteinase pathway and vitamin D receptor activation pathway, both of which have been shown to play critical roles in cardiovascular disease [34, 35].Table 1

Bottom Line: Gender-specific DMRs associated with ciliogenesis that may be linked to defects in cilia development were also identified in AA DMRs.An endothelial methylome analysis identifies epigenetic DMR characteristics associated with transcriptional regulation in regions of atherosusceptibility in swine aorta in vivo.The data represent the first methylome blueprint for spatio-temporal analyses of lesion susceptibility predisposing to endothelial dysfunction in complex flow environments in vivo.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology & Laboratory Medicine and Institute for Medicine & Engineering, Perelman School of Medicine, University of Pennsylvania, 1010 Vagelos Building, 3340 Smith Walk, Philadelphia, PA, 19104, USA. jyizhou@mail.med.upenn.edu.

ABSTRACT

Background: Atherosclerosis is a heterogeneously distributed disease of arteries in which the endothelium plays an important central role. Spatial transcriptome profiling of endothelium in pre-lesional arteries has demonstrated differential phenotypes primed for athero-susceptibility at hemodynamic sites associated with disturbed blood flow. DNA methylation is a powerful epigenetic regulator of endothelial transcription recently associated with flow characteristics. We investigated differential DNA methylation in flow region-specific aortic endothelial cells in vivo in adult domestic male and female swine.

Results: Genome-wide DNA methylation was profiled in endothelial cells (EC) isolated from two robust locations of differing patho-susceptibility:--an athero-susceptible site located at the inner curvature of the aortic arch (AA) and an athero-protected region in the descending thoracic (DT) aorta. Complete methylated DNA immunoprecipitation sequencing (MeDIP-seq) identified over 5500 endothelial differentially methylated regions (DMRs). DMR density was significantly enriched in exons and 5'UTR sequences of annotated genes, 60 of which are linked to cardiovascular disease. The set of DMR-associated genes was enriched in transcriptional regulation, pattern specification HOX loci, oxidative stress and the ER stress adaptive pathway, all categories linked to athero-susceptible endothelium. Examination of the relationship between DMR and mRNA in HOXA genes demonstrated a significant inverse relationship between CpG island promoter methylation and gene expression. Methylation-specific PCR (MSP) confirmed differential CpG methylation of HOXA genes, the ER stress gene ATF4, inflammatory regulator microRNA-10a and ARHGAP25 that encodes a negative regulator of Rho GTPases involved in cytoskeleton remodeling. Gender-specific DMRs associated with ciliogenesis that may be linked to defects in cilia development were also identified in AA DMRs.

Conclusions: An endothelial methylome analysis identifies epigenetic DMR characteristics associated with transcriptional regulation in regions of atherosusceptibility in swine aorta in vivo. The data represent the first methylome blueprint for spatio-temporal analyses of lesion susceptibility predisposing to endothelial dysfunction in complex flow environments in vivo.

No MeSH data available.


Related in: MedlinePlus