Immunosuppressive Actions of Embryonic Stem Cells and Mesenchymal Stem Cells

Abstract

Introduction: Embryonic stem cells (ESCs) and mesenchymal stem cells (MSCs) are known to have immunosuppressive effects. However, its immunosuppressive mechanisms still remain controversial. We therefore conducted this study to examine the immunosuppressive mechanisms of ESCs and MSCs in an in vitro environment and in an animal experimental model of the allograft transplantation of skin and pancreatic islet for in vivo environment. Methods: To analyze the in vitro immunosuppressive effect of stem cells, naïve T cells or memory T cells from C57BL/6 mice and antigen presenting cells from BALB/c mice with ESCs or MSCs were co-cultured for 3 days and thymidine uptake, ELISA (enzyme-linked immunosorbent assay), CFSE (carboxyfluorescein succinimidyl ester) and 7-AAD staining were conducted. Transwell experiment and RT-PCR (reverse transcription-polymerase chain reaction) on immunosuppressive molecules such as PDL-1, Fas-L, CD30, CD70, perforin and granzyme B for the immunosuppressive mechanism were performed. For the functional assay, blocking antibodies or inhibitors on each molecule were treated. To analyze the in vivo immunosuppressive effect of both stem cells, we performed the allogeneic skin and pancreatic transplantation by intravenous injection or co-transplantation of stem cells with anti-CD40L antibodies or mycophenolate mofetil (MMF). Results: ESCs or MSCs did not stimulate alloimmune response and synchronously suppressed the proliferation, survival, activation of naïve CD4+ T cells in an in vitro alloimmune response. They also suppressed the release of cytokines such as IL-2, IL-12, IFN-γ, TNF-α, IL-4, IL-5, IL-1β and IL-10. But there were no significant changes in the expression of TGF-β or IDO (indoleamine 2,3-dioxygenase). In addition, they also suppressed CD44+ memory T cells or T cell activation markers and induced regulatory T cells. The transwell assay revealed that the immunosuppressive effects of both types of stem cells originated from the cell-to-cell contact. Both types of stem cells strongly expressed PDL-1, Fas-L, CD30, perforin and granzyme B, but weakly expressed CD70. When blocking agents on granzyme B of ESCs, the immunosuppressive effect of ESCs was significantly reduced. Both ESCs and MSCs by intravenous injection or co-transplantation did not strongly suppress the immune rejection of the allogeneic skin tissue and pancreatic islet. Despite the concomitant use of them with immunosuppressant, there was no additional effect. Significant immunological changes of immune cells by stem cells were not shown in transplanted recipients. Conclusions: Murine ESCs or MSCs showed the low immunogenicity and the significant immunosuppressive effect such as the suppression of both proliferation and survival of naïve T cells and memory T cells, cytokine release reduction, the induction of Tregs in an in vitro environment. The main mechanism of immunosuppression was by cell-to-cell and granzyme B on ESCs played a key role. However, both stem cells did not suppress allogeneic rejection in the allogeneic skin and pancreatic islet transplantation. This discrepancy in the immunosuppressive effects of stem cells between an in vitro environment and an in vivo one showed that it has to be considered for the establishment of optimal transplantation condition and for the successful clinical application of stem cells in the field of organ transplantation.