Hematopoietic stem cells have been successfully employed for tolerance induction in a variety of rodent and large animal studies. However, clinical transplantation of fully allogeneic bone marrow or blood-borne stem cells is still associated with major obstacles, such as graft-versus-host disease or cytoreductive conditioning-related toxicity. Here we show that when rat embryonic stem cell-like cells of WKY origin are injected intraportally into fully MHC-mismatched DA rats, they engraft permanently (>150 days) without supplementary host conditioning. This deviation of a potentially alloreactive immune response sets the basis for long-term graft acceptance of second-set transplanted WKY cardiac allografts. Graft survival was strictly correlated with a state of mixed chimerism, which required functional thymic host competence. Our results provide a rationale for using preimplantation-stage stem cells as vehicles in gene therapy and for the induction of long-term graft acceptance.

Soluble HLA class I molecules (sHLAs) have been identified in the serum of patients with inflammatory diseases, allografts and autoimmune diseases and in serum of healthy individuals. The biological significance of these molecules, particularly after allogeneic organ transplantation, has been enigmatic. Here we show that primary alloreactive CD8+ T cells interact with sHLA and undergo apoptosis in the absence of a second signal. Ligation of CD28 rescued T cells from death, implying that sHLAs induce apoptosis through selective stimulation of the T-cell receptor. CD95-L was upregulated after cytotoxic T lymphocytes were incubated with sHLAs, and cell death was blocked by a neutralizing anti-CD95-L antibody, suggesting that sHLAs induce endogenous mutual killing of activated T cells. These results provide a molecular basis for the capacity of sHLAs to downregulate T-cell responses, which may be especially relevant to organ transplantation.

Embryonic stem cells (ESCs) are pluripotent and therefore able to differentiate both in vitro and in vivo into specialized tissues under appropriate conditions, a property that could be exploited for cellular therapies. However, the immunological nature of these cells in vivo has not been well understood. In vitro, mouse-derived ESCs fail to stimulate T cells, but they abrogate ongoing alloresponses by a process that requires cell-cell contact. We further show that despite a high expression of the NKG2D ligand retinoic acid early inducible-1 by mouse ESCs, they remain resistant to natural killer cell lysis. In vivo, allogeneic mouse ESCs populate the thymus, spleen, and liver of sublethally irradiated allogeneic host mice, inducing apoptosis to T cells and establishing multilineage mixed chimerism that significantly inhibits alloresponses to donor major histocompatibility complex antigens. Immunohistochemical imaging revealed a significant percentage of ESC-derived cells in the splenic marginal zones, but not in the follicles. Taken together, the data presented here reveal that nondifferentiated mouse embryonic stem cells are non-immunogenic and appear to populate lymphoid tissues in vivo, leading to T-cell deletion by apoptosis.

SUMMARYHuman leucocyte antigen (HLA) -G is expressed on trophoblast cells during pregnancy, suggesting a role in protection of the semiallogeneic fetus. Published data suggest that HLA-G protects a cell against natural killer cell lysis. It has been hypothesized that HLA-G may also protect the fetus by preventing allo-cytotoxic T lymphocyte (CTL) responses. To test this hypothesis, we assayed the effects of various concentrations of puri®ed HLA-G on CTL response in a mixed lymphocyte culture (MLC) system. We found that concentrations i 0 . 1 mg/ml of HLA-G suppressed the allo-CTL response by 30±100% over the control, but, paradoxically, concentrations of 0 . 01±0 . 05 mg/ml of HLA-G augmented the allo-CTL response by 25±50% over the control. Concentrations j 0 . 001 mg/ml HLA-G had no effect. Addition of HLA-G to preprimed allo-CTL effector cells did not affect their killing ability. Allo-CTL suppressive doses of HLA-G induced a T helper type 2 (Th2) cytokine response, whereas allo-CTL-enhancing doses of HLA-G induced a Th1-type cytokine response. HLA-G puri®ed from ®rst-trimester placenta does not affect allo-proliferative responses nor does it alter the percentage of CD4 + or CD8 + T cells in MLCs. These ®ndings support a potential role for HLA-G-mediated suppression of allo-CTL formation in normal pregnancies. In addition, the effects observed at lower concentrations of HLA-G may have interesting implications for the condition of pre-eclampsia in which concentrations of this HLA class I molecule are reduced.