Experimental evidence and preliminary clinical studies have demonstrated that human mesenchymal stem cells (MSCs) display important immune modulatory function of potential relevant interest in the setting of allogeneic hematopoietic stem cell (HSC) transplantation. Effectiveness of MSCs in controlling severe GVHD seems to be related to the immune-regulatory role they play in suppressing alloantigen-specific T-cell activation. Aim of the present study was to extend the analysis of the mechanisms responsible for the immune regulatory effect of interaction between MSCs and alloantigen-specific immune response elicited in vitro in primary and in secondary mixed lymphocyte culture (MLC). At difference with most previously reported studies, we decided to employ non-irradiated MSCs, reasoning that irradiation might impair, beside the proliferative capacity, also the differentiation capability of MSCs and, consequently, alter their interaction pattern with lymphocyte subsets. MSC were added to primary MLC at different doses (MLC-responder-PBMC:MSC ratios = 1:1 and 10:1). Dendritic cell (DC) differentiation, lymphocyte proliferation, alloantigen-specific cytotoxic activity and differentiation of CD4+ T-cell subsets expressing CD25 and/or CTLA4 antigens were assessed in primary and secondary MLC, comparing the effect observed using third-party MSCs with that obtained employing autologous to the MLC-responder (autologous) MSCs. Results demonstrated that human MSCs: (1) strongly inhibit alloantigen-induced DC1 differentiation; (2) down-regulate, in a dose-dependent manner, alloantigen-induced lymphocyte expansion, especially that of CD8+ T cells and of NK lymphocytes; (3) favor the differentiation of CD4+ T cells co-expressing CD25 and/or CTLA4, a phenotype associated with regulatory/suppressive function of immune response; (4) cause a dose-dependent reduction of alloantigen-specific cytotoxic capacity mediated by either cytotoxic T lymphocytes or NK cells; (5) exert more effective suppressive activity on MLC-induced T-cell activation when they are allogeneic rather than autologous with respect to responder cells. In particular, higher percentages of CD4+ and of CD4+CD25+ T cells co-expressing CTLA4+ were detected when third-party, rather than autologous, MSCs were added to MLC. These data suggest that T-cell recognition of alloantigens expressed by MSCs may further facilitate the preferential differentiation of activated CD4+ T cells expressing CTLA4, a glycoprotein, known to deliver an inhibitory signal to T cells and to mediate apoptosis of previously activated T lymphocytes. Several studies previously demonstrated that MSCs exert inhibitory effect on lymphocyte activation through the release of soluble factors. Our data suggest that the preferential differentiation of CD4+CD25+ regulatory T-cell subsets may be favored by other mechanisms of MSC-mediated inhibition of alloantigen-induced effector cell activation and expansion, and, in turn, these CD4+CD25+ cells contribute to propagate and extend suppressor activity. Altogether, our results provide immunological support to the use of MSCs for prevention of immune complications related to both HSC and solid organ transplantation and to the theory that MSCs are “universal” suppressors of immune reactivity.

We studied 459 consecutive patients with hematologic malignancies, median age 44 years (range, 15 to 71 years), who underwent transplantation with grafts from identical sibling donors (SIB; n = 176), matched unrelated donors (MUD; n = 43), mismatched unrelated donors (mmUD; n = 43), unrelated cord blood (UCB; n = 105) or HLA-haploidentical family donors (HAPLO; n = 92). Graft-versus-host disease (GVHD) prophylaxis consisted of cyclosporine and methotrexate in the SIB recipients; antithymocyte globulin for the MUD, mmUD, and UCB recipients; and post-transplantation cyclophosphamide, cyclosporine, and mycophenolate in the HAPLO recipients. Conditioning regimens were mostly myeloablative (69%). Advanced disease phase was more frequent, but not significantly so, in the HAPLO and mmUD groups (P = .08). Acute GVHD grade II-IV was significantly less frequent in the HAPLO, UCB, and MUD groups (14% to 21%) compared with the SIB (31%) and mmUD (42%) groups (P < .001), and there was a trend toward less moderate-severe chronic GVHD in the HAPLO and UCB groups (P = .053). The proportion of patients off cyclosporine at 1 year ranged from 55% for the SIB group to 81% for the HAPLO group (P < .001). Transplantation-related mortality at 2 years was lower in the HAPLO and SIB groups (18% to 24%) compared with the MUD, mmUD, and UCB groups (33% to 35%; P = .10). Relapse rate was comparable in the 5 groups (P = .80). The 4-year actuarial survival was 45% in the SIB group, 43% in the MUD group, 40% in the mmUD group, 34% in the UCB group, and 52% in the HAPLO group (P = .10). In multivariate analysis, advanced disease was a negative predictor of survival (hazard ratio [HR], 2.4; P < .0001), together with a diagnosis of acute leukemia (HR, 1.8; P = .0001); HAPLO grafts were comparable to SIB (P = .80), whereas UCB had inferior survival (P = .03). In conclusion, unmanipulated haploidentical family donor transplants are an additional option for patients lacking a matched sibling donor.

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clinicaltrials.gov Identifier: NCT00297193.

Twenty-six patients with advanced Hodgkin's disease received a related HLA haploidentical unmanipulated BMT, following a nonmyeloablative conditioning with low-dose TBI, proposed by the Baltimore group; GvHD prophylaxis consisted of high-dose posttransplantation CY (PT-CY), mycophenolate and a calcineurin inhibitor. All patients had received a previous autograft, and 65% had active disease at the time of BMT. Sustained engraftment of donor cells occurred in 25 patients (96%), with a median time to neutrophil recovery (40.5 Â 10 9 /L) and platelet recovery (420 Â 10 9 /L) of þ 18 and þ 23 days from BMT. The incidence of grade II-IV acute GVHD and of chronic GVHD was 24% and 8%, respectively. With a median follow-up of 24 months (range 18-44) 21 patients are alive, 20 disease free. The cumulative incidence of TRM and relapse was 4% and 31%, respectively. The actuarial 3-year survival is 77%, the actuarial 3-year PFS is 63%. In conclusion, we confirm that high-dose PT-CY is effective as prophylaxis of GVHD after HLA haploidentical BMT, can prevent rejection and does not appear to eliminate the allogeneic graft versus lymphoma effect.