Mesenchymal stem cells (MSCs) are immunosuppressive multipotent cells under investigation for potential therapeutic applications in regenerative medicine and prevention of graft-versus-host disease. Human leukocyte antigen (HLA)-G contributes to the immunomodulatory properties of MSCs. HLA-G expression in MSCs is very low and diminishes during in vitro expansion. Epigenetic regulation activates HLA-G expression in some cancer cell lines but not in MSCs. In the present study, adipose- and bone marrow-derived MSCs were exposed to the DNA demethylating agent 5-aza-2-deoxycytidine (5-aza-dC) and histone deacetylase inhibitor valproic acid (VPA) and HLA-G mRNA levels assessed using semi-quantitative reverse-transcription PCR. Exposure to 5-aza-dC resulted in HLA-G1 and -G3 upregulation in both early and late passage MSCs. VPA treatment did not induce HLA-G expression in both bone marrow and adipose derived MSCs. Our results provide the first evidence that HLA-G3 could be expressed in MSCs and that methylation-mediated repression is partly responsible for the observed low levels of HLA-G expression in MSCs. Our findings provide insight that treatment of MSCs with specific epigenetic regulatory modulators may improve their immunoregulatory capability for therapeutic applications. (C) 2014 Elsevier Inc. All rights reserved.