*Purpose: It has been reported that a stem cell mobilizing therapy using a combination of AMD3100 and low dose FK506 (AF) enabled immunosuppressive-free long-term kidney allograft survival through induction of allograft chimerism in rats and swine. The aim of this study is to determine if host-derived CD133 stem cells contribute to the induction of allograft chimerism by using host lineage tracing.

*Methods: 8-12 week old CD133+/C-L mice (C57/B6) containing the Rosa26GFP reporter allele (CD133+/C-L X mTmG offspring) were used as recipients and wild type Balb/c mice were used as donors. Orthotopic left kidney transplantation was performed and both recipient kidneys were removed. Transplanted mice were randomly divided into two groups and received subcutaneous injection of saline or AF combination therapy (AMD3100: 2mg/kg; FK506: 0.1mg/kg) on day 0, 2, 4, 6 and 8 and a repeat of this dosing at 1, 2 and 3 months following transplantation. Tamoxifen-induced Cre-dependent GFP whole kidney allograft fluorescence was measured using a Xenogen imaging system. GFP positive cells were further determined in tissue sections under fluorescence microscopy. In this model, the host CD133 progeny in the allograft were GFP positive.

*Results: In the saline control group, most allografts (8/13) were rejected within 2 weeks following kidney transplantation and only 23% (3/13) of the controls survived up to 3 months. In contrast, all animals with AF combination therapy (12/12) survived and had normal serum creatinine levels at 3 months after transplantation. In the treated group, Xenogen imaging showed a significant level of host GFP expression in donor Balb/c kidney allografts at three months after being transplanted into CD133 lineage tracing C57/B6 mice. Fluorescence microscopy demonstrated GFP positive renal tubules in treated allografts, indicating host repopulation.

*Conclusions: Short term AF combination therapy with monthly repeated dosing periods led to immunosuppression-free long term survival in these treated mice that received kidney allotransplants, compared with rejection in the controls. Lineage tracing in the treated recipients demonstrated the critical role of CD133 stem cells in tubule regeneration contributing to induction of allograft chimerism. This mouse kidney transplantation model provides a powerful tool for investigating the mechanisms of tolerance due to induction of allograft chimerism.

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