Purpose: To explore the effects of adoptive transferring T regulatory cells (T reg cells) stimulated by donor corneal antigen (Ag) to prevent corneal transplantation immune rejection in mice.

Methods: C57BL/6 mice were used as donors and BALB/c mice as recipients. Corneal Ag was harvested by homogenization and centrifugation. Bone marrow dendritic cells (DCs) from BALB/c mice were cultured with stimulation of granulocyte-macrophage colony-stimulating factor and interleukin-4 for 5 days. Then, donor corneal Ag was added to obtain donor corneal Ag-loaded DCs. The DCs were used to stimulate CD4+CD25+ and CD4+CD25+ T cells from the recipient to yield Ag-stimulated T reg cells. Penetrating keratoplasty was performed in the mice. The recipients were randomly divided into 3 groups receiving 0.1 mL of phosphate-buffered saline, 1 × 10(6) naive T reg cells, and Ag-stimulated T reg cells, respectively, given by retroorbital injection at the end of surgery. The allografts were observed, and histopathological examination was performed 15 days after surgery.

Results: : The corneal Ag mainly comprised 2 proteins with molecular weight 54 and 42 kD, respectively. Corneal Ag-loaded DCs expressed higher levels of CD11c, CD80, and CD86 than bone marrow precursor cells. Both CD4CD25 and CD4+CD25+ T cells showed vigorous proliferative responses to corneal Ag-loaded DCs. Mean survival time of the mice corneal allografts in phosphate-buffered saline, naive T reg cells, and Ag-stimulated groups was 8.1 ± 1.1, 14.3 ± 2.0, and 23.3 ± 2.6 days, respectively (P < 0.01 among groups). Histopathological examination revealed less inflammatory cells infiltration in Ag-stimulated than in naive mice.

Conclusions: : Adoptive transfer of donor corneal Ag-specific T reg cells prolonged survival time of corneal allografts in our mouse model, which might suggest a useful approach to cellular immunotherapy for corneal transplantation immune rejection.