Ligation of TLR9 induced on human IL-10-secreting Tregs by 1alpha,25-dihydroxyvitamin D3 abrogates regulatory function

Signaling through the TLR family of molecular pattern recognition receptors has been implicated in the induction of innate and adaptive immune responses. A role for TLR signaling in the maintenance and/or regulation of Treg function has been proposed, however its functional relevance remains unclear. Here we have shown that TLR9 is highly expressed by human Treg secreting the antiinflammatory cytokine IL-10 induced following stimulation of blood and tissue CD3(+) T cells in the presence of 1 alpha,25-dihydroxyvitamin D3 (1 alpha 25VitD3), the active form of Vitamin D, with or without the glucocorticoid. dexamethasone. By contrast, TLR9 was not highly expressed by naturally occurring CD4(+)CD25(+) Treg or by Th1 and Th2 effector cells. Induction of TLR9, but not other TLRs, was IL-10 dependent and primarily regulated by 1a25VitD3 in vitro. Furthermore, ingestion of calcitriol (1 alpha 25VitD3) by human volunteers led to an increase of both IL-10 and TLR9 expression by CD3(+)CD4(+) T cells analyzed directly ex vivo. Stimulation of 1 alpha 25VitD3-induced IL-10-secreting Treg with TLR9 agonists, CpG oligonucleotides, resulted in decreased IL-10 and IFN-gamma synthesis and a concurrent loss of regulatory function, but, unexpectedly, increased IL-4 synthesis. We therefore suggest that TLR9 could be used to monitor and potentially modulate the function of 1 alpha 25VitD34nduced IL-10-secreting Treg in vivo, and that this has implications in cancer therapy and vaccine design.