Pretreatment of transfused donor splenocytes and allografts with mitomycin C attenuates acute rejection in heart transplantation in mice.

OBJECTIVE

The aim of this study was to investigate the effect of pretreatment of donor splenocytes and grafts with mitomycin C (MMC) on heart allograft survival, as well as to demonstrate the mechanism of function.

METHODS

Donor splenocytes from Balb/C mice were incubated with MMC (40 μg/mL) in vitro and then transfused into recipients (C57BL/6 mice). The heart allograft was perfused with MMC before harvest. Graft survival and histopathology were examined. Lymphocyte activation, regulatory T cells, and donor splenocyte apoptosis were examined with the use of flow cytometry.

RESULTS

MMC incubation in vitro induced apoptosis of donor splenocytes (15.5 ± 2.3% vs 23.2 ± 4.2%; P < .01). Either intravenous injection of MMC-treated donor splenocytes or transplantation of allograft pretreated with MMC prolonged heart allograft mean survival time from 7 ± 0.8 days to 20.5 ± 1.9 days or 10 ± 0.9 days, respectively (both P < .01). A combination of MMC-pretreated donor splenocyte transfusion and allografts showed the best effect on prolongation of graft survival (28.5 ± 1.8 days). Activation of CD4(+) T cells in spleen and peripheral lymph nodes of recipients was significantly inhibited by either MMC-splenocyte transfusion or the combination treatment. Meanwhile, the percentage of CD4(+)CD25(+)Foxp3(+) regulatory T cells in the spleen was increased in the MMC-splenocyte transfusion group (15.5 ± 1.1% vs 18.2 ± 0.9%; P < .05).

CONCLUSIONS

Both injection of MMC-conditioned donor splenocytes and MMC-conditioned allograft have effects on prolongation of heart allograft survival in mice, and MMC-conditioned donor splenocytes might play an essential role. MMC pretreatment induced regulatory T cells likely through induction of donor splenocyte apoptosis, and thus it inhibited T-cell activation.