De novo DNA methylation induced by circulating extracellular vesicles from acute coronary syndrome patients

Background and aims: DNA methylation is associated with gene silencing, but its clinical role in cardiovascular diseases (CVDs) remains to be elucidated. We hypothesized that extracellular vesicles (EVs) may carry epigenetic changes, showing themselves as a potentially valuable non-invasive diagnostic liquid biopsy. We isolated and characterized circulating EVs of acute coronary syndrome (ACS) patients and assessed their role on DNA methylation in epigenetic modifications. Methods: EVs were recovered from plasma of 19 ACS patients and 50 healthy subjects (HS). Flow cytometry, qRT-PCR, and Western blot (WB) were performed to evaluate both intra-vesicular and intra-cellular signals. ShinyGO, PANTHER, and STRING tools were used to perform GO and PPI network analyses. Results: ACS-derived EVs showed increased levels of DNA methyltransferases (DNMTs) (p<0.001) and Ten-eleven translocation (TET) genes reduction. Specifically, de novo methylation transcripts, as DNMT3A and DNMT3B, were significantly increased in plasma ACS-EVs. DNA methylation analysis on PBMCs from healthy donors treated with HS- and ACS-derived EVs showed an important role of DNMTs carried by EVs. PPI network analysis evidenced that ACS-EVs induced changes in PBMC methylome. In the most enriched subnetwork, the hub gene SRC was connected to NOTCH1, FOXO3, CDC42, IKBKG, RXRA, DGKG, BAIAP2 genes that were showed to have many molecular effects on various cell types into onset of several CVDs. Modulation in gene expression after ACS-EVs treatment was confirmed for SRC, NOTCH1, FOXO3, RXRA, DGKG and BAIAP2 (p<0.05). Conclusions: Our data showed an important role for ACS-derived EVs in gene expression modulation through de novo DNA methylation signals, and modulating signalling pathways in target cells.

Automated summary

DNA methylation is associated with cardiovascular diseases (CVDs). This study found that circulating extracellular vesicles (EVs) from acute coronary syndrome (ACS) patients showed increased levels of DNA methyltransferases (DNMTs) and reduced Ten-eleven translocation (TET) genes. By treating lymphocytes from healthy individuals with EVs, it was discovered that DNMTs carried by EVs play an important role in DNA methylation. Furthermore, ACS-EVs were found to regulate signaling pathways and gene expression in target cells.