Increased Regulatory T Cell Function in a Mouse Model of Beta-Thalassemia.

Abstract 638 The development of red blood cell (RBC) alloantibodies complicates transfusion therapy for chronically transfused patients with thalassemias. Understanding the regulation of immune responses to transfused RBCs may help in future design of therapeutic interventions to prevent RBC alloimmunization in this patient population. We have previously reported in a mouse model that key immune response regulators, CD4+CD25− regulatory T cells (Tregs) expressing Foxp3, control the rate and frequency of RBC alloimmunization in wildtype recipient mice. As an initial step to study and characterize immune regulation in thalassemias, we studied the Treg status of Hbb(th1/th1) mouse model of beta-thalassemia intermedia, known to exhibit mild anemia, reticulocytosis and splenomegaly. Compared to wildtype littermate controls (WT) (n=11), 3 month old thalassemic (THAL) mice (n=12) had significantly higher frequency of CD25+Foxp3+ Tregs in CD4+ population in peripheral blood (5.5±0.5% versus 4.1±0.3%, p=0.03) but not in the spleen (11.4%±0.9% versus 9.5±0.5%, p=0.1). Sorted splenic Tregs from THAL and WT mice were equally anergic to stimulation through the T cell receptor and Tregs from THAL mice secreted the expected low background levels of IL-2 and IFN-gamma in stimulated culture supernatants, similar to Tregs from WT mice. Surprisingly, however, the THAL stimulated Tregs, but not the WT counterparts, secreted the proinflammatory cytokine IL-17. To determine whether THAL Tregs were functionally active, we performed in vitro Treg suppressive assay. We found that thalassemic Tregs were in fact more suppressive than Tregs from their WT counterparts in their ability to suppress proliferation of CD4+CD25–T cells (at 1:4 ratio of Tregs : CD4+CD25–cells 91±3% thalassemic versus 66±3% WT , p=0.03; at 1:8 ratio, 84±4% % versus 63±7%, p=0.02 and at 1:16 ratio, 76±4% % versus 28±14%, p=0.03). We next performed red cell transfusion studies to determine if the increased in vitro THAL Treg suppressive activity is associated with decrease in frequency of alloimmunization in THAL mice. A cohort of age- and sex-matched THAL and WT mice were transfused on a weekly basis for 4 weeks with red cells from transgenic mice expressing human glycophorin A. We found that considerably lower (2/9) alloimmunization rates in THAL compared to WT mice (8/11). Altogether, our data indicate that qualitative differences exist between Tregs from THAL mice and their littermate controls and raise the interesting possibility that THAL Tregs may represent a novel population of Tregs. Specifically, only THAL Tregs secrete the proinflammatory cytokine IL-17 normally associated with Th17 subset and yet they are functionally more suppressive. The mechanisms responsible for these differences are under further study. Disclosures: No relevant conflicts of interest to declare.