Interleukin 17 (IL-17) has been linked to autoimmune diseases, although its regulation and function have remained unclear. Here we have evaluated in vitro and in vivo the requirements for the differentiation of naive CD4 T cells into effector T helper cells that produce IL-17. This process required the costimulatory molecules CD28 and ICOS but was independent of the cytokines and transcription factors required for T helper type 1 or type 2 differentiation. Furthermore, both IL-4 and interferon-gamma negatively regulated T helper cell production of IL-17 in the effector phase. In vivo, antibody to IL-17 inhibited chemokine expression in the brain during experimental autoimmune encephalomyelitis, whereas overexpression of IL-17 in lung epithelium caused chemokine production and leukocyte infiltration. Thus, IL-17 expression characterizes a unique T helper lineage that regulates tissue inflammation.

A fundamental function of CD4 + helper T (T H ) cells is the regulation of B cell-mediated humoral immunity. Development of T follicular helper (T FH ) cells that provide help to B cells is mediated by the cytokines interleukin-6 and interleukin-21 but is independent of T H 1, T H 2, and T H 17 effector cell lineages. Here, we characterize the function of Bcl6, a transcription factor selectively expressed in T FH cells. Bcl6 expression is regulated by interleukin-6 and interleukin-21. Bcl6 overexpression induced T FH -related gene expression and inhibited other T H lineage cell differentiation in a DNA binding-dependent manner. Moreover, Bcl6 deficiency in T cells resulted in impaired T FH cell development and germinal center reactions, and altered production of other effector T cell subsets. Our data thus illustrate that Bcl6 is required for programming of T FH cell generation.

T cell functional differentiation is mediated by lineage-specific transcription factors. T helper 17 (Th17) has been recently identified as a distinct Th lineage mediating tissue inflammation. Retinoic acid receptor-related orphan receptor gamma (ROR gamma) was shown to regulate Th17 differentiation; ROR gamma deficiency, however, did not completely abolish Th17 cytokine expression. Here, we report Th17 cells highly expressed another related nuclear receptor, ROR alpha, induced by transforming growth factor-beta and interleukin-6 (IL-6), which is dependent on signal transducer and activator of transcription 3. Overexpression of ROR alpha promoted Th17 differentiation, possibly through the conserved noncoding sequence 2 in Il17-Il17f locus. ROR alpha deficiency resulted in reduced IL-17 expression in vitro and in vivo. Furthermore, ROR alpha and ROR gamma coexpression synergistically led to greater Th17 differentiation. Double deficiencies in ROR alpha and ROR gamma globally impaired Th17 generation and completely protected mice against experimental autoimmune encephalomyelitis. Therefore, Th17 differentiation is directed by two lineage-specific nuclear receptors, ROR alpha and ROR gamma.

After activation, CD4+ helper T (T(H)) cells differentiate into distinct effector subsets that are characterized by their unique cytokine expression and immunoregulatory function. During this differentiation, T(H)1 and T(H)2 cells produce interferon-gamma and interleukin (IL)-4, respectively, as autocrine factors necessary for selective lineage commitment. A distinct T(H) subset, termed T(HIL-17), T(H)17 or inflammatory T(H) (T(H)i), has been recently identified as a distinct T(H) lineage mediating tissue inflammation. T(H)17 differentiation is initiated by transforming growth factor-beta and IL-6 (refs 5-7) and reinforced by IL-23 (ref. 8), in which signal transduction and activators of transcription (STAT)3 and retinoic acid receptor-related orphan receptor (ROR)-gamma mediate the lineage specification. T(H)17 cells produce IL-17, IL-17F and IL-22, all of which regulate inflammatory responses by tissue cells but have no importance in T(H)17 differentiation. Here we show that IL-21 is another cytokine highly expressed by mouse T(H)17 cells. IL-21 is induced by IL-6 in activated T cells, a process that is dependent on STAT3 but not ROR-gamma. IL-21 potently induces T(H)17 differentiation and suppresses Foxp3 expression, which requires STAT3 and ROR-gamma, which is encoded by Rorc. IL-21 deficiency impairs the generation of T(H)17 cells and results in protection against experimental autoimmune encephalomyelitis. IL-21 is therefore an autocrine cytokine that is sufficient and necessary for T(H)17 differentiation, and serves as a target for treating inflammatory diseases.

Recently, a third subset of TH cells, named as TH IL-17 , TH17, or inflammatory TH (THi), which produce IL-17, was identified by us as well as other investigators to mediate a pathogenic inflammatory response (5-7). THi cells were also found to produce IL-17F and IL-22 (5, 7-9). IL-23, sharing a p40 unit with IL-12, has been first found to regulate IL-17 expression and the development or expansion of THi cells in vitro (5-7). More recently, several groups showed that TGF-␤ in the context of IL-6 and other inflammatory cytokines supports THi differentiation in vitro, independent of IL-23 (10 -12), possibly at least in part by regulating the chromatin remodeling of the IL-17-IL-17F locus (13). IL-1␤ and TNF-␣ may also be involved in promoting THi development or in regulating expression of IL-17 at the effector phase (11,14).The downstream signaling pathways, such as STAT, that selectively mediate THi generation are unclear. STAT1 appears to negatively regulate THi differentiation (7), whereas STAT4 or STAT6 were not involved (6). Recently, Socs3-deficient T cells were found to exhibit enhanced IL-17 expression; this effect was associated with enhanced activity of STAT3 in response to IL-23 that could bind to . STAT3 has critical functions in the immune system, including control of dendritic cell production, inhibition of macrophage inflammatory signaling, and regulation of steady state and emergency granulopoiesis (16 -18). However, the precise physiological function of STAT3 in THi lineage differentiation has not been directly addressed. Whether STAT5, another STAT protein that has been shown activated by , has any function in THi differentiation is also unclear.In this study, we show that IL-6 up-regulates expression of IL-23R and that IL-23 synergizes with IL-6 in promoting THi differentiation. Retroviral expression of a hyperactive STAT3 enhances THi cell development. STAT3 deficiency in CD4 T cells results in impaired THi development and a deficiency in ROR␥t, a THi-specific transcription factor recently identified (20). These data indicate that STAT3 is a cytokine-activated essential regulator in THi development.