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Abstracts Session 3

Tuesday May 02, 2023 - 09:20 to 10:30

Room: Grand Georgian

9.1 Depletion of Donor Macrophages Permits Post-Transplant Tolerance Induction

Award Winner

Miriam Dilts, United States has been granted the AST-COTS Scientific Congress Award

Miriam Dilts, United States

PhD Student
Duke University
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Miriam Dilts is a PhD student at Duke University in the department of Pathology. She studies in the lab of Dr. Xunrong Luo. Prior to Duke, she worked in a developmental biology lab studying the effect of oxidants on neural development. At Duke, she studies the ability of donor passenger macrophages to impact tolerization of pancreatic islet grafts.


Depletion of Donor Macrophages Permits Post-Transplant Tolerance Induction

Miriam Dilts1, Anil Dangi1, Xunrong Luo1.

1School of Medicine, Duke University, Durham, NC, United States

Introduction: Using donor splenocytes fixed with cross-linker ethylcarbodiimide (ECDI-SPs), our lab has induced tolerance in a variety of murine and non-human primate models when one dose of ECDI-SPs is administered before transplant and one dose after. We aimed to induce tolerance post-transplant in a murine model by depleting donor macrophages prior to transplant. Once successful, we investigated the roles of extracellular vesicles (EVs) released by donor macrophages in alloimmunity.

Methods: We transplanted BALB/c pancreatic islets under the kidney capsule of C57BL/6J mice made diabetic with streptozotocin. Prior to islet harvest, donors were treated with either anti-CSF1R antibody to deplete macrophages or an isotype control. Recipients received two doses of ECDI-SPs following transplant. Graft rejection was defined as hyperglycemia for two consecutive days. For in vitro studies, EVs were generated by stimulation of RAW 264.7 macrophages of BALB/c origin. These EVs were fed to C57BL/6J dendritic cells, which were then cultured with 4C or TCR75 T cells, specific to direct and indirect presentation of BALB/c MHC respectively. T cells were analyzed for proliferation via CFSE dilution on flow cytometry.

Results: Depletion of donor macrophages followed by treating the recipient with two doses of ECDI-SPs resulted in indefinite graft survival in 90% of recipients, while non-depleted donors resulted in only 30% graft survival. We found that in vitro, C57BL/6J dendritic cells fed EVs released by BALB/c macrophages were able to stimulate 4C and TCR75 T cells to proliferate. This demonstrates the ability of DCs to present MHC from allogeneic EVs both semi-directly and indirectly. DCs fed BALB/c lysate stimulated TCR75, but not 4C, T cells, suggesting that intact EVs are needed for this semi-direct presentation. T cells did not respond when cultured with EVs alone, demonstrating the necessity for dendritic cells.

Conclusions: Depleting donor macrophages permitted tolerization of islet grafts with post-transplant treatments. In vitro results suggest that EVs released by donor macrophages when stimulated are capable of contributing to both semi-direct and indirect stimulation of recipient T cells. Dendritic cells play a crucial role in this phenomenon, likely contributing costimulation signals for T cell proliferation.

National Institutes of Health (NIH), R01DK132889.

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