CD8⁺ regulatory cells are responsible for the immunosuppression induced by MSCs. (A) C3 MSCs inhibit the proliferation of the CD8⁺ subset in a direct MLR. BALB/c total splenocytes (splenocytes) were depleted of CD8⁺ cells (CD8^- subset) using CD8⁺-coated beads (“Materials and methods”). Then 10⁵ cells of both populations and the CD8⁺ splenocytes (CD8⁺ subset) were stimulated by 10⁵ mitomycin-treated allogeneic splenocytes, with or without C3 MSCs. ^**P < .001. (B) The CD8⁺ subset suppresses the proliferative activity of murine lymphocytes stimulated by allogeneic splenocytes in a secondary MLR. BALB/c and DBA1 splenocytes were cultured in a primary MLR in the presence of C3 MSCs to obtain “conditioned” splenocytes. Then 10⁵ total splenocytes (Cond^ed splenocytes) or CD8⁺ (Cond^ed CD8⁺ subset) or CD8^- (Cond^ed CD8^- subset) cells were then used in a secondary MLR using 10⁵ proliferative allogeneic splenocytes from C3H mice. Results are expressed as the percentage of the proliferative response ± SD.