In Vivo T Regulatory Cell Kinetics Are Altered in a Pre-Clinical Model of Chronic Graft-Versus-Host Disease

Chronic graft-versus-host disease (cGVHD) is the main cause of late morbidity and non-relapse mortality after allogeneic hematopoietic stem cell transplantation (AHSCT).  T cells are known modulators of cGVHD, while their in vivo kinetics, as defined by cell division, cell death and trafficking, remain largely uncharacterized, and current therapies for cGVHD target T cell proliferation.  T regulatory cells (T regs) are of particular interest in cGVHD as adoptive transfer or in vivo expansion of this cell type has resulted in amelioration of cGVHD severity in pre-clinical models and in patients with cGVHD.  We have developed a method to measure T reg and non-T reg cell kinetics using deuterated water labeling/de-labeling combined with triple quadrupole GC/MS detection of deuteurium enrichment in DNA of dividing/dying cells.  Using a minor antigen mismatch murine model of cGVHD we show that in AHSCT recipients, donor CD4+ non-T regs in the spleen have a proliferative advantage over T regs, while in the syngeneic (genetically matched donor-recipient pairs) control animals cell gain rates for these populations are similar.  The net result of the differential cell kinetics in the cohort with cGVHD was fewer T regs in the spleens of allogeneic versus syngeneic marrow recipients, with a suggestion for decreased trafficking out of the spleen.  Evaluation of T reg and non-T reg CD4+ T cells within lymphoid compartments (lymph nodes and peripheral blood) and target organs (liver, intestines and skin) allowed assessment of T cell trafficking.  Preliminary results show distinct organ-specific patterns of T cell kinetics after AHSCT.  These data allow mathematical modeling of T cell behavior post AHSCT in a pre-clinical model of cGVHD, enabling interpretation of in vivo T cell kinetics in peripheral blood of patients undergoing AHSCT with and without cGVHD.