Type 1 diabetes is the most common chronic disease affecting children. Islet cell or pancreas transplantation offer the most physiologic approach to glucose homostasis. However the underlying autoimmunity destroys the newly grafted organ and is not controlled by immunosuppression. Therefore, induction of tolerance is an important goal. Several autoimmune diseases, including diabetes, can be treated using hematopoietic stem cell transplant (HSCT). However, the toxicity of myeloablative conditioning has until now prevented the widespread clinical application of this approach. It has been shown that NOD mice are more resistant to engraftment and doses of TBI ≤ 500cGY with or without T cell depleting antibodies results in engraftment failure. In the present study, we have applied a non-myeloablative conditioning protocol currently in use to induce tolerance to transplanted kidneys to NOD mice to minimize the toxicity associated with conditioning.

With an aim to establish a non-myeloablative conditioning approach with clinical applicability, we used low-dose total body irradiation (200cGy) on D-1, fludarabine (30mg/kg) on D -4, -3 and -2 and cyclophosphamide (50 mg/kg) on D-2, +2 and +3. Mycophenolate mofetil (MMF; 40mg/kg) and cyclosporine A (20mg/kg) were used as maintenance immunosuppressive drugs (Day -2 to Day +9). 30×10⁶ unmodified BM cells from non-autoimmune B6 mice were transplanted into 6-8 week old NOD mice on Day 0. Engraftment for donor cells was determined four weeks after transplantation by flow cytometry. Mice were monitored for disease development. More than 90% of the mice showed various levels of durable donor chimerism ranging from 9% to 78 % at 8 weeks post-transplant, with production of multiple lineages. All engrafted mice remained disease-free irrespective of the level of chimerism whereas 70 % of untreated mice showed high blood glucose levels at 16wk of age. Diabetes-prevention was not observed in mice that received conditioning without bone marrow transplant. To determine if chimerism established tolerance, we performed donor or third party skin grafting. The chimeric mice rejected the third party skin within 14 days, whereas donor skin transplants were accepted.Our data suggest that donor-specific tolerance can be induced in the NOD mice with low-dose conditioning, which can protect transplanted islets or pancreas, theoretically providing exogenous-insulin free cure for life.

CITATION INFORMATION: Chhabra A., Huang Y., Xu H., Merchak A., Ildstad S. Low Intensity Conditioning Protocol for Inducing Chimerism-Based Tolerance in Non-Obese Diabetic Mice Am J Transplant. 2017;17 (suppl 3).

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