IL-12p70 induced IFN-γ is required to control Mycobacterium tuberculosis growth; however, in the absence of IL-12p70, an IL-12p40-dependent pathway mediates induction of IFN-γ and initial bacteriostatic activity. IL-23 is an IL-12p40-dependent cytokine containing an IL-12p40 subunit covalently bound to a p19 subunit that is implicated in the induction of CD4 T cells associated with autoimmunity and inflammation. We show that in IL-23 p19-deficient mice, mycobacterial growth is controlled, and there is no diminution in either the number of IFN-γ-producing Ag-specific CD4 T cells or local IFN-γ mRNA expression. Conversely, there is an almost total loss of both IL-17-producing Ag-specific CD4 T cells and local production of IL-17 mRNA in these mice. The absence of IL-17 does not alter expression of the antimycobacterial genes, NO synthase 2 and LRG-47, and the absence of IL-23 or IL-17, both of which are implicated in mediating inflammation, fails to substantially affect the granulomatous response to M. tuberculosis infection of the lung. Despite this redundancy, IL-23 is required to provide a moderate level of protection in the absence of IL-12p70, and this protection correlates with a requirement for IL-23 in the IL-12p70-independent induction of Ag-specific, IFN-γ-producing CD4 T cells. We also show that IL-23 is required for the induction of an IL-17-producing Ag-specific phenotype in naive CD4 T cells in vitro and that absence of IL-12p70 promotes an increase in the number of IL-17-producing Ag-specific CD4 T cells both in vitro and in vivo.

The kinetics of presentation of influenza virus–derived antigens (Ags), resulting in CD4 T cell effector and memory generation, remains undefined. Naive influenza-specific CD4 T cells were transferred into mice at various times after influenza infection to determine the duration and impact of virus-derived Ag presentation. Ag-specific T cell responses were generated even when the donor T cells were transferred 3–4 wk after viral clearance. Transfer of naive CD4 T cells during early phases of infection resulted in a robust expansion of highly differentiated effectors, which then contracted to a small number of memory T cells. Importantly, T cell transfer during later phases of infection resulted in a modest expansion of effectors with intermediate phenotypes, which were capable of persisting as memory with high efficiency. Thus, distinct stages of pathogen-derived Ag presentation may provide a mechanism by which T cell heterogeneity is generated and diverse memory subsets are maintained.

Migration of dendritic cells (DCs) to the draining lymph node (DLN) is required for the activation of naive T cells. We show here that migration of DCs from the lung to the DLN after Mycobacterium tuberculosis (Mtb) exposure is defective in mice lacking interleukin (IL)-12p40. This defect compromises the ability of IL-12p40–deficient DCs to activate naive T cells in vivo; however, DCs that express IL-12p40 alone can activate naive T cells. Treatment of IL-12p40–deficient DCs with IL-12p40 homodimer (IL-12(p40)2) restores Mtb-induced DC migration and the ability of IL-12p40–deficient DCs to activate naive T cells. These data define a novel and fundamental role for IL-12p40 in the pathogen-induced activation of pulmonary DCs.

SummaryWe have outlined the carefully orchestrated process of CD4 + T-cell differentiation from naïve to effector and from effector to memory cells with a focus on how these processes can be studied in vivo in responses to pathogen infection. We emphasize that the regulatory factors that determine the quality and quantity of the effector and memory cells generated include (i) the antigen dose during the initial T-cell interaction with antigen-presenting cells; (ii) the dose and duration of repeated interactions; and (iii) the milieu of inflammatory and growth cytokines that responding CD4 + T cells encounter. We suggest that heterogeneity in these regulatory factors leads to the generation of a spectrum of effectors with different functional attributes. Furthermore, we suggest that it is the presence of effectors at different stages along a pathway of progressive linear differentiation that leads to a related spectrum of memory cells. Our studies particularly highlight the multi-faceted roles of CD4 + effector and memory T cells in protective responses to influenza infection and support the concept that efficient priming of CD4 + T cells that react to shared influenza proteins could contribute greatly to vaccine strategies for influenza. Overview and historyOver the past decade, others and we have concluded that naïve precursor T cells must undergo many steps of division and differentiation before they acquire the effector functions necessary for their many regulatory activities (1). One of these activities is 'help' for B cells, which promotes B-cell isotype switching, somatic mutation, and differentiation in germinal centers to plasma cells and memory cells (2-4). Another key regulatory activity carried out by CD4 + T cells involves help for naïve CD8 + T cells to promote their optimum differentiation into cytotoxic effectors and memory cells and to support their maintenance (5-7). In addition, there are a host of other regulatory effects of CD4 + effectors on macrophages as well as other antigenpresenting cells (APCs). These CD4 + T-cell functions are mediated by surface coreceptors on the effector cells, including CD40L, CD28, cytotoxic T-lymphocyte antigen-4, etc., that interact with receptors on B cells, dendritic cells, macrophages, or other APCs, and by potent cytokines secreted by the CD4 + effectors upon recognition of antigen on APCs.CD4 + T-cell effectors represent a collection of distinct subsets characterized in part by their abilities to produce different patterns of cytokines. The two best characterized subsets are designated T-helper 1 (Th1), producing interferon-γ (IFN-γ), and Th2, producing interleukin-4 (IL-4), IL-5, and IL-13 as 'signature' cytokines. Recently, evidence has accumulated for a third . Most probably the APCs that stimulate the naïve CD4 + T cells are also the initial source of cytokines that imprint these subsets in situ (11). It is also increasingly accepted that the polarizing cytokines secreted by the APCs are dictated by the context of the antigen, be it from a pathogen or...

Efficient peptide presentation by professional APC to naive and effector CD4 T cells in vitro is limited to the first 1-2 days of culture, but is nonetheless optimum for effector expansion and cytokine production. In fact, prolonging Ag presentation leads to high levels of T cell death, decreased effector expansion, and decreased cytokine production by recovered effectors. Despite the absence of Ag presentation beyond day 2, T cell division continues at a constant rate throughout the 4-day culture. The Ag-independent later stage depends on the presence of IL-2, and we conclude optimum effector generation depends on an initial 2 days of TCR stimulation followed by an additional 2 days of Ag-independent, cytokine driven T cell expansion and differentiation.