Methylome analysis of endothelial cells suggests new insights on sporadic brain arteriovenous malformation

Arteriovenous malformation of the brain (bAVM) is a vascular phenotype related to brain
defective angiogenesis. Involved vessels show impaired expression of vascular differentiation
markers resulting in the arteriolar to venule direct shunt. In order to clarify aberrant gene
expression occurring in bAVM, here we describe results obtained by methylome analysis per
formed on endothelial cells (ECs) isolated from bAVM specimens, compared to human cerebral
microvascular ECs. Results were validated by quantitative methylation-specific PCR and quan
titative realtime-PCR. Differential methylation events occur in genes already linked to bAVM
onset, as RBPJ and KRAS. However, among differentially methylated genes, we identified EPHB1
and several other loci involved in EC adhesion as well as in EC/vascular smooth muscle cell
(VSMC) crosstalk, suggesting that only endothelial dysfunction might not be sufficient to trigger
the bAVM phenotype. Moreover, aberrant methylation pattern was reported for many lncRNA
genes targeting transcription factors expressed during neurovascular development. Among these,
the YBX1 that was recently shown to target the arteridin coding gene. Finally, in addition to the
conventional CpG methylation, we further considered the role of impaired CHG methylation,
mainly occurring in brain at embryo stage. We showed as differentially CHG methylated genes are
clustered in pathways related to EC homeostasis, as well as to VSMC-EC crosstalk, suggesting as
impairment of this interaction plays a prominent role in loss of vascular differentiation, in bAVM
phenotype.