A LOW DONOR ISLET MASS IS SUFFICIENT TO RESTORE NORMOGLYCEMIA IN DIABETIC NOD MICE AFTER PRETRANSPLANT INDUCTION OF A SUBSTANTIAL LEVEL OF DONOR CHIMERISM

Abstract

O200 Aims: Two or more donors are required for a successful human islet allotransplantation under the current Edmonton protocol. However, human islet autotransplant recipients after pancreatectomy need fewer islets per recipient than the islet allotransplant recipients. In this study, we determined whether the islet mass required to reverse diabetes could be significantly reduced in spontaneously diabetic NOD mice after establishing a substantial level of donor lymphocyte chimerism, as compared with the islet mass required for successful allogeneic islet transplantation in immunosuppressed diabetic NOD mice. Methods: Fludarabine phosphate and cyclophosphamide were given at day -1. Donor BALB/c (H-2d) bone marrow cells (4x107) were transplanted into spontaneously diabetic NOD mice (H-2g7) at day 0. Anti-CD154 mAb were given from day 0 to day 14. Donor chimerism was measured by flow cytometric analysis before islet transplantation. Different numbers of BALB/c islets were transplanted under left kidney capsule of diabetic NOD mice with >40% donor lymphocyte chimerism at 3 weeks post-bone marrow transplantation. We also transplanted different numbers of islets into spontaneously diabetic NOD mice under immunosuppression comparable to the Edmonton protocol. Diabetic NOD mice were treated with ALS (0.3 ml) on day –1 and 0, and rapamycin (1 mg/kg/day) and low-dose tacrolimus (0.5 mg/kg/day) beginning on day –2. Results: At 3 weeks post-islet transplantation, diabetes was not reversed in any diabetic NOD mice that received 100 islets and immunosuppressants ALS, ramamycin, and tacrolimus (N=3). When 200 islets were transplanted into diabetic NOD mice under immunosuppressants ALS, ramamycin, and tacrolimus, only 16 % of recipient mice achieved normoglycemia (N=12). When the number of transplanted islets was further increased to 300 and 400 in the diabetic NOD mice under immunosuppression with ALS, ramamycin, and tacrolimus; the percentage of mice that achieved normoglycemia was increased to 50% (N=12) and 75% (N=8). However, in diabetic NOD mice with donor lymphocyte chimerism >40% before islet transplantation, 83% of mice reversed diabetes 3 weeks after receiving 100 donor islets (N=6). When the number of transplanted islets was increased to 200 (N=9), 300 (N=8), and 400 (N=8), all recipient mice achieved normoglycemia. Conclusions: Our results demonstrate that a high number of islets are required to reverse diabetes in spontaneously diabetic NOD mice under immunosuppression. However, a low islet number is sufficient to restore normoglycemia in diabetic NOD mice with a substantial level of donor chimerism. These data suggest that a small number of transplanted islets are sufficient to reverse diabetes in those diabetic NOD mice whose alloimmune response and autoimmune response were abrogated by pretransplant induction of a substantial level of donor chimerism.