Human CD4+CD25+Foxp3+ Regulatory T Cells Do Not Constitutively Express IL-35

EBV-induced gene 3 (EB13) can associate with p28 to form the heterodimeric cytokine IL-27, or with the p35 subunit of IL-12 to form the EB13/p35 heterodimer, recently named IL-35. In mice, IL-35 has been shown to be constitutively expressed by CD4(+)CD25(+)Foxp3(+) regulatory T cells (Treg cells) and suggested to contribute to their suppressive activity. In this study, we investigated whether human Treg cells express ILL-35. Double-staining analysis of human thymuses showed that neither Foxp3(+) nor CD25(+) cells coexpressed EB13. Similarly, Foxp3(+) cells present in human lymph nodes, tonsils, spleens, and intestines did not express EB13. Consistent with these in situ observations, Treg cells purified from blood or tonsils were negative for EB13 by immunoblotting. Other human T cell subsets, including effector T cells, naive and memory CD4(+) T cells, CD8(+) and gamma delta T cells also did not constitutively express EB13, which contrasts with IL-35 expression observed in murine CD8(+) and gamma delta T cells. Furthermore, although CD3/CD28 stimulation consistently induced low levels of EB13 in various CD4(+) T cell subsets, no EB13 could be detected in CD3/CD28-stimulated Treg cells. RT-PCR analysis showed that, whereas p35 transcripts were detected in both Teff and Treg cells, EB13 transcripts were detected only in activated Teff cells, but not in resting or activated Treg cells. Thus, in contrast to their murine counterpart, human Treg cells do not express detectable amounts of IL-35. The Journal of Immunology, 2008, 181: 6898-6905.