Mesenchymal stem cells suppress lymphocyte proliferation in vitro and prolong skin graft survival in vivo  Amelia Bartholomew, Cord Sturgeon, Mandy Siatskas, Karen Ferrer, Kevin McIntosh, Sheila Patil, Wayne Hardy, Steve Devine, David Ucker, Robert Deans, Annemarie Moseley, Ronald Hoffman  Experimental Hematology  Volume 30, Issue 1, Pages 42-48 (January 2002) DOI: 10.1016/S0301-472X(01)00769-X

Figure 1 Allogeneic baboon MSCs do not elicit a significant proliferative response when cultured with allogeneic PBL. Baboon 5957 or 6759 PBL were incubated with irradiated PBL from the MSC donor, 6656 (first set of bars); 6759, or 5957, respectively (second set of bars); or autologous source (third set of bars). Lymphocytes (1 × 105 cells/well) from baboons 5957 or 6759 were cultured with 105, 104, 103, or irradiated 105 MSCs/well. Proliferative activity against various doses of MSCs resembled autologous controls. Experimental Hematology 2002 30, 42-48DOI: (10.1016/S0301-472X(01)00769-X)

Figure 2 MSCs from animal 6593 diminish proliferative response in a one-way MLR, and this effect is abrogated with paraformaldehyde fixation of the MSCs. Responding baboon 6656 PBL were incubated with irradiated 6593 or 5957 PBL, and the mean of these responses observed in triplicate were deemed maximal stimulation. Viable or paraformaldehyde-fixed MSCs were added into these one-way MLRs on day 0 at the designated doses per well. Percent of maximal response was the mean of triplicates observed after addition of MSCs divided by the following maximal responses: 6656 vs 6593, 8457 ± 856 CPM and 6656 vs 5957, 8007 ± 1233 CPM. Experimental Hematology 2002 30, 42-48DOI: (10.1016/S0301-472X(01)00769-X)

Figure 3 Baboon 6593 MSCs (1 × 104/well) reduce MLR proliferative response when added to an ongoing MLR on day 3. Maximal response was the mean of an experiment performed in triplicate and assigned the value of 100%. Percent of maximal response was the mean of triplicates observed after addition of MSCs divided by the following maximal responses: 6656 vs 6593, 12,922 ± 3,984 CPM; 6,656 vs 5957, 4960 ± 766 CPM; 6593 vs 6656, 22,489 ± 4,828 CPM; 6593 vs 5957, 26,658 ± 4343 CPM. Experimental Hematology 2002 30, 42-48DOI: (10.1016/S0301-472X(01)00769-X)

Figure 4 MSCs from animal 6593 added to ConA-stimulated PBL from animal 6368 on day 0 suppress proliferative activity in a dose-dependent fashion. Bars show ConA alone, ConA + 105 MSCs, ConA + 104 MSCs, ConA + 103 MSCs, and ConA + 102 MSCs, respectively. Experimental Hematology 2002 30, 42-48DOI: (10.1016/S0301-472X(01)00769-X)

Figure 5 IL-2 antagonizes the immunosuppressive activity of MSCs. PBLs from animal 6368 were stimulated with either ConA, ConA + IL-2 (50 units/mL), or IL-2 alone (50 units/mL) on day 0. 1 × 104 MSCs from animal 6593 were added on either day 0 or day 3. Stimulation index for 6368 vs 6593 was 20.7. In the absence of IL-2, percent of maximal response, determined by mean CPM of the observed proliferative responses to ConA, was 4% and 27% for MSCs added on days 0 and 3, respectively. In contrast, percent of maximal response in the presence of IL-2 was 56% and 45% for MSCs added on days 0 and 3, respectively. Experimental Hematology 2002 30, 42-48DOI: (10.1016/S0301-472X(01)00769-X)

Figure 6 Hemotoxylin- and eosin-stained punch biopsies of skin grafts from control recipient 6532, which did not receive MSCs (A), and recipient 6659, which received MSCs from animal 6593 (B, C). Animal 6532 received a skin graft from a non-MSC donor animal, 5720. Eschar formation consistent with rejection was noted on the gross appearance of the skin graft from 6532 and day-7 biopsy revealed sloughed epidermis (A). Animal 6659 received a skin graft from MSC donor 6593 and an unrelated (third-party) skin graft from animal 6654. Biopsy taken from the 6654 skin graft revealed intact epidermis with normal dermis on day 7 (B) and dermal bullous formation, pyknotic nuclei, and scant neutrophilic infiltrate consistent with rejection on day 13 (C). Experimental Hematology 2002 30, 42-48DOI: (10.1016/S0301-472X(01)00769-X)