FOXP3-expressing T regulatory cells (Tregs) can be divided into two distinct subsets: naturally occurring Tregs (nTregs) that develop in the thymus, and induced Tregs (iTregs) that differentiate in peripheral tissues upon exposure to Ag in a tolerogenic environment. Recently it has been proposed that expression of Helios, an Ikaros family transcription factor, may specifically identify nTregs, allowing specific tracking of Tregs from different origins in health and disease. Surprisingly, we found that Helios- cells can be readily identified within naive (CD45RA+CD31+CCR7+CD62L+) FOXP3+ Tregs, a finding inconsistent with the notion that lack of Helios expression identifies Ag-experienced iTregs that should express memory markers. To investigate the phenotype and function of naive Helios+ and Helios− Tregs within the nTreg population, we isolated single-cell clones from each subset. We found that both Helios+ and Helios− nTreg clones have a similar suppressive capacity, as well as expression of FOXP3 and cell surface proteins, including CD39 and CTLA-4. Helios− nTregs, however, produced significantly more CCL3 and IFN-γ compared with Helios+ nTregs. Despite increased cytokine/chemokine production, Helios− FOXP3+ nTreg clones were demethylated at the FOXP3 Treg-specific demethylated region, indicative of Treg lineage stability. When cultured under Th1-polarizing conditions, Helios+ and Helios− nTreg clones had an equal ability to produce IFN-γ. Collectively, these data show that a lack of Helios expression does not exclusively identify human iTregs, and, to our knowledge, the data provide the first evidence for the coexistence of Helios+ and Helios− nTregs in human peripheral blood.

T-regulatory cells (Tregs) have a fundamental role in the establishment and maintenance of peripheral tolerance. There is now compelling evidence that deficits in the numbers and/or function of different types of Tregs can lead to autoimmunity, allergy, and graft rejection, whereas an over-abundance of Tregs can inhibit anti-tumor and anti-pathogen immunity. Experimental models in mice have demonstrated that manipulating the numbers and/or function of Tregs can decrease pathology in a wide range of contexts, including transplantation, autoimmunity, and cancer, and it is widely assumed that similar approaches will be possible in humans. Research into how Tregs can be manipulated therapeutically in humans is most advanced for two main types of CD4(+) Tregs: forkhead box protein 3 (FOXP3)(+) Tregs and interleukin-10-producing type 1 Tregs (Tr1 cells). The aim of this review is to highlight current information on the characteristics of human FOXP3(+) Tregs and Tr1 cells that make them an attractive therapeutic target. We discuss the progress and limitations that must be overcome to develop methods to enhance Tregs in vivo, expand or induce them in vitro for adoptive transfer, and/or inhibit their function in vivo. Although many technical and theoretical challenges remain, the next decade will see the first clinical trials testing whether Treg-based therapies are effective in humans.