Donor derived (allogeneic) bone marrow mesenchymal stem cells (MSCs) are promising candidates for cell therapy. Although, the outcome of initial clinical trials was promising; poor survival of MSCs following transplantation in ischemic heart has hampered their application as ‘off the shelf’ candidates. The major reason being postulated for the loss of transplanted cells is their switch from immunoprivileged to immunogenic state. Immunoprivilege of MSCs is maintained by an immunosuppressive soluble factor prostaglandin E2 (PGE2). We found that PGE2 and COX2 (enzymatic regulator of PGE2 synthesis) levels decreased in hypoxic MSCs, which was associated with their loss of immunoprivilege. However, overexpressing COX2 protein in MSCs restored PGE2 levels and preserved their immunoprivilege in vitro. COX2 overexpression also mitigated host immune response, prolonged MSC survival and improved cardiac function post transplantation in ischemic heart. Additionally, we also show that CSN5 maintains COX2 levels in MSCs; and CSN5 reduction leads to down regulation of COX2 and PGE2 levels. Besides hypoxic microenvironment, long term in vitro culture of allogeneic MSCs is known to be associated with changes in their physiological characteristics. Therefore, we investigated the effect of increased in-culture time of MSCs, on their immunoprivilege. We explored the effect of increase in passage number of MSCs on cell surface ox-Phosphatidylcholines, cellular bioenergetics, global proteomics, and immune as well as cell survival pathways. Our data demonstrated no significant changes in these parameters, confirming that increase in passage number within clinically relevant passages (P3-7) does not affect the immunological behavior of MSCs. Furthermore, owing to the inevitable nature of serum in MSC culture and cryopreservation, we analyzed the effect of different serum concentrations in the freezing media, on MSCs. We found that post thawing, there were no changes in the proliferation and viability of MSCs frozen in high or low serum media. Moreover, low serum containing freezing media did not affect the immune response generated by MSCs; indicating that serum concentration in the freezing media might not be a significant regulator of immunogenicity of MSCs. The results from these studies add novel insights to the present knowledge of immunological behavior of allogeneic MSCs, which will help in designing future trials using allogeneic MSCs.