Background.

An understanding of the mechanisms that suppress the human anti-pig cellular response is key for xenotransplantation. We have compared the ability of human regulatory T cells (Tregs) to suppress xenogeneic and allogeneic responses in vitro.

Methods.

Human peripheral blood mononuclear cells (PBMC), CD4+ T cells, or CD4+ CD25− T cells were stimulated with irradiated human or wild type (WT) or α1, 3-galactosyltransferase gene-knockout (GT-KO) pig PBMC in the presence or absence of human CD4+ CD25 high Tregs. In separate experiments, 5-(and 6)-carboxyfluorescein diacetate succinimidyl ester-labeled human CD4+ T cells were stimulated with human or pig PBMC. The expansion and precursor frequencies of allo-and xeno-reacitve Tregs were assessed by labeling with FoxP3 mAb and flow cytometric analysis.

Results.

The responses of human PBMC, CD4+ T cells, and CD4+ CD25− T cells to pig PBMC were stronger than to human PBMC (P< 0.05). Human anti-GT-KO responses were weaker than anti-WT responses (P< 0.05). Human CD4+ CD25 high Tregs suppressed proliferation of CD4+ CD25− T cells to both human and pig PBMC stimulator cells with the same efficiency. Alloreactive CD4+ CD25+ FoxP3 high responder T cells proliferated more than their xenoreactive counterparts (P< 0.05), although xenoreactive CD4+ CD25+ T cells proliferated more than alloreactive cells (P< 0.05). There was no difference in precursor frequency between allo-and xeno-reactive CD4+ CD25+ FoxP3 high cells.

Conclusions.

Human T-cell responses to pig cells are stronger than to allogeneic cells. The human response to GT-KO PBMC is weaker than to WT PBMC. Although human Tregs can suppress both responses, expansion of CD4+ CD25+ FoxP3 high cells against pig PBMC is weaker than against human PBMC. More human Tregs may be required to suppress the stronger xenogeneic response.