Bone marrow-derived conventional, but not cloned, mesenchymal stem cells suppress lymphocyte proliferation and prevent graft-versus-host disease in rats. 
Bone marrow-derived mesenchymal stem cells (MSCs) could exert a potent immunosuppressive effect, and therefore may have a therapeutic potential in T-cell-dependent pathologies.  In the present study, we aimed to determine whether MSCs could be used to control graft-versus-host disease (GvHD), a major cause of morbidity and mortality after allogeneic hematopoietic stem cell transplantation (HSCT).  MSCs were isolated from Lewis rat bone morrow and then cultured in 10% FBS DMEM at 37°C for 4 weeks.  The enriched conventional MSCs and macrophages were purified by auto-MACS.  Cloned MSCs were obtained by cloning using the limiting dilution method and expanded up to more than 6 months.  The identity of MSCs was confirmed by their typical spindle-shaped morphology and immunophenotypic criteria, based on the absence of expression of CD45 and CD11b/c molecules.  Both types of MSCs were also tested for their ability to differentiate into adipocytes.  We showed that MSCs, like macrophages, exhibit immunomodulatory properties capable of inhibiting T-cell proliferation stimulated by alloantigens, anti-CD3e/CD28 mAbs, and ConA in a dose-dependent manner in vitro.  After performing adoptive transfer, MSCs suppressed systemic Lewis to (Lewis × DA)F1 rat GvHD.  In contrast to the immunosuppressive activities of conventional MSCs, the cloned MSCs enhanced T-cell proliferation in vitro and yielded no clinical benefit in regard to the incidence or severity of GvHD.  Therefore, these rat models have shown intriguing differences in the suppression effects of lymphocyte proliferation and GvHD prevention between short-term cultured conventional MSCs and cloned MSCs.