EZH2 Modulates Angiogenesis In Vitro and in a Mouse Model of Limb Ischemia

Abstract

Epigenetic mechanisms may regulate the expression of pro-angiogenic genes, thus affecting reparative angiogenesis in ischemic limbs. The enhancer of zest homolog-2 (EZH2) induces thtrimethylation of lysine 27 on histone H3 (H3K27me3), which represses gene transcription. We explored (i) if EZH2 expression is regulated by hypoxia and ischemia; (ii) the impact of EZH2 on the expression of two pro-angiogenic genes: eNOS and BDNF; (iii) the functional effect of EZH2 inhibition on cultured endothelial cells (ECs); (iv) the therapeutic potential of EZH2 inhibition in a mouse model of limb ischemia (LI). EZH2 expression was increased in cultured ECs exposed to hypoxia (control: normoxia) and in ECs extracted from mouse ischemic limb muscles (control: absence of ischemia). EZH2 increased the H3K27me3 abundance onto regulatory regions of eNOS and BDNF promoters. In vitro RNA silencing or pharmacological inhibition by 3-deazaneplanocin (DZNep) of EZH2 increased eNOS and BDNF mRNA and protein levels and enhanced functional capacities (migration, angiogenesis) of ECs under either normoxia or hypoxia. In mice with experimentally induced LI, DZNep increased angiogenesis in ischaemic muscles, the circulating levels of pro-angiogenic hematopoietic cells and blood flow recovery. Targeting EZH2 for inhibition may open new therapeutic avenues for patients with limb ischemia.

Epigenetic regulation of endothelial‐cell‐mediated vascular repair

“…Consistent with these results, Smits et al showed that decreased expression of miR-101, a repressor of EZH2 expression, in ECs results in increased blood vessel formation [142]. Consistent with these data, EZH2 was found to be bound to eNOS and BDNF promoters where it mediates H3K27 tri-methylation in normoxia and hypoxia [144]. capillary-like structure formation) it also increases other angiogenic properties such as adhesion and migration [143], suggesting that EZH2 inhibition could potentially be useful to improve the function of endothelial progenitors for stem cell therapy.…”

“…In addition, EZH2 silencing in tumor-associated ECs decreases angiogenesis through VASH1 re-expression, leading to decreased cancer growth [141]. A very recent study [144] highlighted the role of EZH2 in ischemic vascular repair. Interestingly, while EZH2 silencing in ECs reduces some of the cells' angiogenic properties (e.g.…”

“…Indeed EZH2 has been shown to repress angiogenic gene expression in non-ECs [140], non-pathological ECs [143] and in response to hypoxia and ischemic injury [144]. Indeed EZH2 has been shown to repress angiogenic gene expression in non-ECs [140], non-pathological ECs [143] and in response to hypoxia and ischemic injury [144].…”

“…Despite uncertainties in the molecular mechanism, DZNep has been used quite extensively in ECs. Interestingly, a recent report showed that treatment with DZNep increases eNOS expression in ECs, and improves EC functionality (migration, angiogenesis) in vitro [144]. Furthermore, DZNep treatment increases eNOS expression in CD34+ hematopoietic stem/progenitor cells [60].…”

Gestational Diabetes Mellitus Impairs Fetal Endothelial Cell Functions Through a Mechanism Involving MicroRNA-101 and Histone Methyltransferase Enhancer of Zester Homolog-2

“…30 We also showed that Ad-mediated EZH2-α overexpression impairs the angiogenic capacity of healthy HUVECs in vitro. 30 The results of our 2 studies can be reconciled if we speculate that hypoxia/ischemia and HG/GDM regulate different EZH2 isoforms and other RNAbinding molecules in ECs and that changes in EZH2-α and EZH2-β expression differently affect gene regulation.…”