Altered DNA methylation pattern reveals epigenetic regulation of Hox genes in thoracic aortic dissection and serves as a biomarker in disease diagnosis

Abstract

Background Thoracic aortic dissection (TAD) is a severe disease with limited understandings in its pathogenesis. Altered DNA methylation has been revealed to be involved in many diseases etiology. Few studies have examined the role of DNA methylation in the development of TAD. This study explored alterations of the DNA methylation landscape in TAD and examined the potential role of cell-free DNA (cfDNA) methylation as a biomarker in TAD diagnosis. Results Ascending aortic tissues from TAD patients (Stanford type A; n = 6) and healthy controls (n = 6) were first examined via whole-genome bisulfite sequencing (WGBS). While no obvious global methylation shift was observed, numerous differentially methylated regions (DMRs) were identified, with associated genes enriched in the areas of vasculature and heart development. We further confirmed the methylation and expression changes in homeobox (Hox) clusters with 10 independent samples using bisulfite pyrosequencing and quantitative real-time PCR (qPCR). Among these, HOXA5, HOXB6 and HOXC6 were significantly down-regulated in TAD samples relative to controls. To evaluate cfDNA methylation pattern as a biomarker in TAD diagnosis, cfDNA from TAD patients (Stanford type A; n = 7) and healthy controls (n = 4) were examined by WGBS. A prediction model was built using DMRs identified previously from aortic tissues on methylation data from cfDNA. Both high sensitivity (86%) and specificity (75%) were achieved in patient classification (AUC = 0.96). Conclusions These findings showed an altered epigenetic regulation in TAD patients. This altered epigenetic regulation and subsequent altered expression of genes associated with vasculature and heart development, such as Hox family genes, may contribute to the loss of aortic integrity and TAD pathogenesis. Additionally, the cfDNA methylation in TAD was highly disease specific, which can be used as a non-invasive biomarker for disease prediction.

DNA methylation alternation in Stanford- A acute aortic dissection

“…Li N et al detected the DNA methylation in blood samples of TAD patients and found MMP2 promoter hypermethylated[ 18 ]. Liu P et al compared the differential DNA methylation in ascending aortic tissues of thoracic aortic dissection (TAD) patients[ 19 ]. They found that the DMP-associated genes were enriched in vasculature and heart development, and Hox genes may be important in TAD pathogenesis[ 19 ].…”

“…Liu P et al compared the differential DNA methylation in ascending aortic tissues of thoracic aortic dissection (TAD) patients[ 19 ]. They found that the DMP-associated genes were enriched in vasculature and heart development, and Hox genes may be important in TAD pathogenesis[ 19 ]. The DNA methylation signature of plasma cell-free DNA in AAD was also detected[ 19 ].…”

“…They found that the DMP-associated genes were enriched in vasculature and heart development, and Hox genes may be important in TAD pathogenesis[ 19 ]. The DNA methylation signature of plasma cell-free DNA in AAD was also detected[ 19 ]. Sun P et al used 450k BeadChip for DNA methylation exploration and found that AAD was correlated with inflammatory vascular remodeling process, which was possibly related to environmental risk factors such as smoking[ 20 ].…”

CRISPR-based targeted haplotype-resolved assembly of a megabase region

“…The bisulfite-sequencing libraries were prepared based on a method described previously 50 with modifications, by using EZ DNA Methylation-Gold Kit (NEB) and NEBNext® Ultra™ II DNA Library Prep Kit (NEB) from 10 ng of the targeted enriched HMW MHC DNA molecules, and sequenced on the Illumina Hiseq X Ten platform. For Illumina methylation EPIC beadchip, 500 ng of genomic DNA from GM12878 cells was used and the experiment was performed by Shanghai Genergy Co. Ltd (Shanghai, China) according to the manufacturer’s instructions.…”

Applying multi-omics techniques to the discovery of biomarkers for acute aortic dissection

“…We also found that alterations in DNA methylation in TAD subsequently altered the expression of these genes. The expression levels of HOXA5, HOXB6 and HOXC6, for example, were significantly downregulated in TAD patients compared to healthy controls, with the expression levels of HOXA5 and HOXB6 significantly correlated with their methylation levels (84,85). Moreover, Hox genes are known to play an important role not only in regulating cell proliferation, differentiation and migration, but also as regulators of phenotypic plasticity in VSMCs, which are closely associated with cardiovascular development and disease.…”