The homeobox (Hox) genes encode transcription factors that are involved in the morphogenesis of body. Recent data showed that the HoxD transcription factors control the cardiovascular system development, by modulation of endothelial cell proliferation and differentiation. For our knowledge, the role of histone acetylation in expression of HoxD9 has not been studied to date; therefore, the aim of this study was to investigate the expression of HoxD9 in endothelial progenitor cells after treatment with valproic acid (VPA), a histone deacetylase inhibitor. Our results showed that VPA inhibits the histone deacetylases leaving chromatin in an acetylated state corresponding to a decondensate conformation. qRT-PCR and Western blot assays showed that the expression of HoxD9 in endothelial progenitor cells treated with VPA was increased at both gene and protein level, suggesting that acetylation regulates HoxD9 expression. Furthermore, flow cytometry analysis revealed that the expression of endothelial specific markers such as CD31, CD105, CD117 and VEGFR2 was decreased in the presence of acetylating agent, VPA. The capacity of endothelial progenitor cells to form vascular networks on Matrigel was also reduced in the presence of VPA. In conclusion, investigating the role of histone acetylation in the regulation of accessibility of transcription factors to genes involved in differentiation can contribute to understanding epigenetic mechanisms underlying the commitment of stem cells.