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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.


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Site-specific disturbed blood flow in the aorta. Flow separation in the pig aortic arch (AA) defines an athero-susceptible site characterized by disturbed blood flow and oxidative stress. a: Flow velocity vectors in the aorta illustrating flow separation with reversal in the inner curvature of the AA during systole. Unidirectional flow recovers in the athero-protected descending thoracic (DT) segment. b: MRI-generated computational fluid dynamics of human systolic aortic blood flow. Low net flow in the AA separated flow region is below system detection. Unidirectional pulsatile laminar flow dominates all other regions. (From Markl et al. 2011) [61], reproduced with permission. c: Computed wall shear stress (WSS) and velocity distributions during systole in rat aorta. (Adapted from Bjorck et al. 2012) [62] with open access permission to reproduce. d: Aortic arch of apoE−/− mouse fed a high fat high cholesterol diet for 6 weeks, illustrating fatty streak atherosclerosis at the inner curvature of the AA. From Cheng et al. [63] reprinted with permission. e. Pig AA and DT endothelial harvest sites (i and ii) with their respective endothelial cell morphologies (iii and iv). An occasional smooth muscle cell or leukocyte was identified within the harvested endothelium (v). Endothelial nucleic acids were isolated for methylated DNA immunoprecipitation sequencing (MeDIP-seq) and for qRT-PCR (vi)
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Fig1: Site-specific disturbed blood flow in the aorta. Flow separation in the pig aortic arch (AA) defines an athero-susceptible site characterized by disturbed blood flow and oxidative stress. a: Flow velocity vectors in the aorta illustrating flow separation with reversal in the inner curvature of the AA during systole. Unidirectional flow recovers in the athero-protected descending thoracic (DT) segment. b: MRI-generated computational fluid dynamics of human systolic aortic blood flow. Low net flow in the AA separated flow region is below system detection. Unidirectional pulsatile laminar flow dominates all other regions. (From Markl et al. 2011) [61], reproduced with permission. c: Computed wall shear stress (WSS) and velocity distributions during systole in rat aorta. (Adapted from Bjorck et al. 2012) [62] with open access permission to reproduce. d: Aortic arch of apoE−/− mouse fed a high fat high cholesterol diet for 6 weeks, illustrating fatty streak atherosclerosis at the inner curvature of the AA. From Cheng et al. [63] reprinted with permission. e. Pig AA and DT endothelial harvest sites (i and ii) with their respective endothelial cell morphologies (iii and iv). An occasional smooth muscle cell or leukocyte was identified within the harvested endothelium (v). Endothelial nucleic acids were isolated for methylated DNA immunoprecipitation sequencing (MeDIP-seq) and for qRT-PCR (vi)

Mentions: To investigate the prevalence of flow-related site-specific DMRs in a physiological arterial setting, we mapped the DNA methylation landscape in swine endothelial cells from the inner curvature of the aortic arch (AA; disturbed flow) and from the nearby descending thoracic aorta (DT; undisturbed flow) representing athero-susceptible and athero-protected sites respectively. Figure 1 illustrates the regional flow differences (Fig. 1a,b) and wall shear stresses (Fig. 1c) in the AA and DT and the predisposition of AA to atherosclerosis (Fig. 1d).Fig. 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)

Site-specific disturbed blood flow in the aorta. Flow separation in the pig aortic arch (AA) defines an athero-susceptible site characterized by disturbed blood flow and oxidative stress. a: Flow velocity vectors in the aorta illustrating flow separation with reversal in the inner curvature of the AA during systole. Unidirectional flow recovers in the athero-protected descending thoracic (DT) segment. b: MRI-generated computational fluid dynamics of human systolic aortic blood flow. Low net flow in the AA separated flow region is below system detection. Unidirectional pulsatile laminar flow dominates all other regions. (From Markl et al. 2011) [61], reproduced with permission. c: Computed wall shear stress (WSS) and velocity distributions during systole in rat aorta. (Adapted from Bjorck et al. 2012) [62] with open access permission to reproduce. d: Aortic arch of apoE−/− mouse fed a high fat high cholesterol diet for 6 weeks, illustrating fatty streak atherosclerosis at the inner curvature of the AA. From Cheng et al. [63] reprinted with permission. e. Pig AA and DT endothelial harvest sites (i and ii) with their respective endothelial cell morphologies (iii and iv). An occasional smooth muscle cell or leukocyte was identified within the harvested endothelium (v). Endothelial nucleic acids were isolated for methylated DNA immunoprecipitation sequencing (MeDIP-seq) and for qRT-PCR (vi)
© Copyright Policy - open-access
Related In: Results  -  Collection

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

Fig1: Site-specific disturbed blood flow in the aorta. Flow separation in the pig aortic arch (AA) defines an athero-susceptible site characterized by disturbed blood flow and oxidative stress. a: Flow velocity vectors in the aorta illustrating flow separation with reversal in the inner curvature of the AA during systole. Unidirectional flow recovers in the athero-protected descending thoracic (DT) segment. b: MRI-generated computational fluid dynamics of human systolic aortic blood flow. Low net flow in the AA separated flow region is below system detection. Unidirectional pulsatile laminar flow dominates all other regions. (From Markl et al. 2011) [61], reproduced with permission. c: Computed wall shear stress (WSS) and velocity distributions during systole in rat aorta. (Adapted from Bjorck et al. 2012) [62] with open access permission to reproduce. d: Aortic arch of apoE−/− mouse fed a high fat high cholesterol diet for 6 weeks, illustrating fatty streak atherosclerosis at the inner curvature of the AA. From Cheng et al. [63] reprinted with permission. e. Pig AA and DT endothelial harvest sites (i and ii) with their respective endothelial cell morphologies (iii and iv). An occasional smooth muscle cell or leukocyte was identified within the harvested endothelium (v). Endothelial nucleic acids were isolated for methylated DNA immunoprecipitation sequencing (MeDIP-seq) and for qRT-PCR (vi)
Mentions: To investigate the prevalence of flow-related site-specific DMRs in a physiological arterial setting, we mapped the DNA methylation landscape in swine endothelial cells from the inner curvature of the aortic arch (AA; disturbed flow) and from the nearby descending thoracic aorta (DT; undisturbed flow) representing athero-susceptible and athero-protected sites respectively. Figure 1 illustrates the regional flow differences (Fig. 1a,b) and wall shear stresses (Fig. 1c) in the AA and DT and the predisposition of AA to atherosclerosis (Fig. 1d).Fig. 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