Sanne Hoeks

Defective neonatal TH17 development and reduced RORC2 mRNA 35 3 INTRODUCTION The neonatal immune system shows a different response on activation when compared with the adult system. 1 Several mechanisms contribute to this phenomenon, which involves both functioning of the innate system 2 and adaptive mechanisms of the immune system 3 of which the T H 2 bias has been investigated intensively. We previously showed an important role for human regulatory T (Treg) cells 4 in modulation of immune activation in the neonatal setting, which is induced by the interaction between antigen-presenting cells (APCs) and naive T cells. Especially programmed death-1 ligation by programmed death-L1 contributes significantly to the induction of Treg cells from human cord blood (CB)-derived precursors. 4 T H 17 and Treg cells have a reciprocal development pathway. 5 Both cells play an important role in the induction and perpetuation of inflammatory responses during infection, and also in the pathogenesis of autoimmune and allergy-related diseases. On T-cell activation in the presence of TGF-β, both signature transcription factors for T H 17 and Treg-cell lineage are upregulated: retinoid acid–related orphan receptor (ROR) γT for T H 17 cells and forkhead box (FOXP) 3 for Treg cells. In the absence of proinflammatory cytokines, FOXP3 dominates RORγT function and prevents T H 17 development. Because of the propensity of Treg-cell induction in CB, we hypothesized there to be a regulatory mechanism in CB that inhibits T H 17 cell development in neonatal T cells derived from CB cells as compared with adult peripheral blood (APB) cells. Activation of CD4 + CD25 − CD45RO − T cells in the presence of APCs (see this article’s Methods section) shows increased numbers of FOXP3 + cells in the CB (shown to be functional Treg cells earlier 4 ) than in the APB (Fig 1, A and D ). IL-17 production, however, was not observed in the CB (Fig 1, A and B ). Replacement of APCs by anti-CD28 antibody prevented T H 17 induction in the APB (see Fig E1, A , in the Online Repository). The lack of IL-17, IL-21, IL-22, and GM-CSF (Fig 1, C ) in the supernatants of CB cell cultures confirms that T H 17 phenotype is not induced in the CB after TCR stimulation. Other key cytokines, including IFN-γ and IL-13, were also reduced in CB cell cultures than in APB cells (Fig E1, B ), confirming earlier work. 6 IL-21 and IL-22 in the APB were only partly derived from IL-17–producing cells (Fig E1, C ). T H 17 cells may derive from any naive T cell when activated in a conductive cytokine milieu, or from a defined precursor cell. In humans, CD161 + CD4 + T cells are thought to be such precursors to the T H 17 cell lineage. 7 It has also been suggested that T H 17 cells more readily develop from CD45RO + memory T cells. 8 Therefore, we tested whether memory cell contamination within our naive cells at the start of culture accounts for the increased ability of APB to yield T H 17 cells. Fluorescence-activated cell sorting–sorted populations of CD25 − CD45RO − CD161 − cells from the APB can still develop into T H 17 cells, whereas the same population in the CB develops into Treg cells (Fig E1, D ). Thus, in our culture system with viable APCs present, CB-naive T cells cannot differentiate into T H 17.