Introduction: Fms-like tyrosine kinase 3 ligand (Flt3L) is essential for myeloid cell homeostasis, particularly dendritic cells (DC), and acts through STAT3 to promote DC expansion. Myeloid-derived suppressor cells (MDSC) are a heterogeneous population of immunoregulatory CD11b⁺Gr1⁺ immature myeloid cells that suppress T cell responses and are required for experimental tolerance induction after heart transplantation. However, the influence of Flt3L in vivo on the expansion and activation of MDSC has not been examined. Due to the ability of Flt3L to augment expansion of multiple myeloid cell populations, we hypothesized that Flt3L administration would expand immunosuppressive MDSC able to promote heart transplant survival.

Methods: BALB/c mice were given recombinant human Flt3L (10 Μg/d i.p.) daily for 10 d, with or without STAT3 inhibitor S31-201 (5 mg/kg i.p.). Splenic CD11c⁺ and Gr1⁺ cells were isolated by immunomagnetic bead selection and tested as stimulators or suppressors, respectively, in mixed leukocyte reactions with CD3⁺ T cell responders. BALB/c recipients were given syngeneic MDSC (5×10⁶) one day prior to heterotopic transplantation of C57BL/6 hearts. Survival was monitored by abdominal palpation and rejection confirmed histologically.

Results: Flt3L increased the absolute number and frequency of splenic DC (CD11b⁺CD11c⁺) and MDSC (CD11b⁺Gr1⁺). Although DC from Flt3L-treated mice stimulated allogeneic T cell proliferation more than control DC, MDSC from these mice were more potent suppressors of T cell proliferation than control MDSC. Flt3L activated MDSC, which required IDO and HO-1 for T cell suppression. Although STAT3 is thought to be critical for MDSC expansion and activation, STAT3 inhibition blocked DC expansion but augmented Flt3L-mediated MDSC expansion. Notably, STAT3 inhibition did not alter Flt3L-expanded MDSC suppressive function. Syngeneic Flt3L-expanded MDSC, but not control MDSC, administered one day before transplantation of fully MHC-mismatched heart grafts prolonged survival significantly in the absence of immunosuppression.

Conclusions: These data provide the first demonstration that transfer of Flt3L-expanded MDSC prolongs cardiac allograft survival. Furthermore, we identify a novel immunomodulatory capacity of Flt3L for expansion of functional MDSC in a STAT3-independent manner.

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