Mesenchymal Stem Cells Overexpressing Interleukin-35 Propagate Immunosuppressive Effects in Mice

To explore generation of interleukin (IL)-35-expressing mouse adipocyte-derived mesenchymal stem cells (Ad-MSCs) using lentiviral vector and their potential immunosuppressive effects in mice. Ad-MSCs were isolated and cultured in vitro and transfected with a lentivirus vector for overexpression of the therapeutic murine IL-35 gene. IL-35 expression in transfected MSCs (IL-35-MSCs) was quantified by enzyme-linked immunosorbent assay (ELISA). The lymphocytes subsets after one-way mixed lymphocyte culture and in vivo intravenous transplantation were analysed by flow cytometry to evaluate the immunosuppressive effects of IL-35-MSCs. ELISA was performed to examine IL-10, IL-17A and IL-35 expression in lymphocyte culture. Mouse Ad-MSCs were isolated and cultured. IL-35 was expressed in the MSC supernatant and serum after IL-35 transduction into Ad-MSCs by lentiviral vector transfection in vitro and in vivo. The percentage of CD4⁺ CD25⁺ T regulatory (Treg) cells in mice treated with IL-35-MSCs significantly increased. IL-35-MSCs upregulated the CD4⁺ CD25⁺ Treg cells in the allogeneic mixed lymphocyte reaction system, and lowered the percentage of CD4⁺ T cells compared with the other two control groups (P < 0.01). IL-17A expression significantly decreased and IL-10 expression significantly increased in IL-35-MSCs and MSCs when compared by ELISA to the control groups (P < 0.01). IL-35-transduced Ad-MSCs in vivo can enhance proliferation of CD4⁺ CD25⁺ Treg cells and suppress the function of effector T cells such as T helper (Th) 1, Th2 and Th17 cells and may reduce the development of allograft rejection. Our data suggest that transduced Ad-MSCs overexpressing IL-35 may provide a useful approach for basic research on cell-based immunotolerance therapy for inducing transplantation tolerance.