1281: EXTRACELLULAR MITOCHONDRIA EXACERBATE GRAFT-VERSUS-HOST DISEASE MORTALITY IN A MOUSE MODEL

Abstract

Introduction: Graft-versus-host disease (GVHD) is a major complication of hematopoietic stem cell transplantation (HSCT). Previous studies have shown an association between pre-transplant conditioning mediated tissue damage and incidence of GVHD, albeit the pathobiology is not clearly delineated. Recent studies have shown that tissue damage mediates release of extracellular mitochondria (exMito) and associated byproducts, which have immunomodulatory functions due to their archaebacterial origin. However, the role of exMito in the pathogenesis of GVHD has not been explored. Methods: Intestinal epithelial cells (Caco-2) were subjected to doses of irradiation used as pre-transplant conditioning. Subsequently, changes in intracellular and extracellular mitochondrial quantity and function were evaluated. BALB/c mice were subjected to total body irradiation (TBI) and plasma was analyzed to evaluate the quantity and function of circulating exMito. Finally, BALB/c mice undergoing a MHC-mismatched allogeneic HSCT were co-transplanted with exogenous isolated syngeneic mitochondria to study the impact of exMito on GVHD mortality. Results: Seahorse XF flux assays revealed that irradiation impaired intracellular mitochondrial function with a decrease in basal, ATP-linked and maximal respiratory capacity in Caco-2 cells. Analyses of the cell-culture supernatant showed elevated exMito and increased oxidative mitochondrial protein modification (carbonylation), following irradiation. BALB/c mice which received TBI demonstrated morphological signs of mitochondrial fission and damage in intestinal epithelium along with an increase in the amount of exMito in the plasma. In addition, exMito isolated from the plasma of irradiated animals had a markedly diminished response to pyruvate/malate-dependent-induction of mitochondrial membrane potential suggestive of damage. Finally, recipient mice co-transplanted with damaged cell-free mitochondria in a MHC-mismatched allogeneic HSCT demonstrated dose-dependent increased GVHD-induced mortality. Conclusions: The results indicate that conditioning regimen-mediated extracellular release of damaged mitochondria plays a role in GVHD mortality. Future studies targeting therapies to preserve mitochondrial function and decrease extracellular mitochondrial load for GVHD management is warranted.