NK1.1 Expression Defines a Population of CD4+ Effector T Cells Displaying Th1 and Tfh Cell Properties That Support Early Antibody Production During Plasmodium yoelii Infection

Early plasmablast induction is a hallmark of Plasmodium infection and is thought to contribute to the control of acute parasite burden. Although long understood to be a T-cell dependent phenomenon, regulation of early plasmablast differentiation, however, is poorly understood. Here, we identify a population of CD4+ T cells that express the innate NK cell marker NK1.1 as an important source of T cell help for early plasmablast and parasite-specific Ab production. Interestingly, NK1.1+ CD4+ T cells arise from conventional, naive NK1.1− CD4+ T cells, and their generation is independent of CD1d but critically reliant on MHC-II. CD4+ T cells that express NK1.1 early after activation produce IFN-γ and IL-21, and express the follicular helper T (Tfh) cell markers ICOS, PD-1 and CXCR5 more frequently than NK1.1− CD4+ T cells. Further analysis of this population revealed that NK1.1+ Tfh-like cells were more regularly complexed with plasmablasts than NK1.1− Tfh-like cells. Ultimately, depletion of NK1.1+ cells impaired class-switched parasite-specific antibody production during early Plasmodium yoelii infection. Together, these data suggest that expression of NK1.1 defines a population of rapidly expanding effector CD4+ T cells that specifically promote plasmablast induction during Plasmodium infection and represent a subset of T cells whose modulation could promote effective vaccine design

IFN-γ-producing CD4+ T cells promote experimental cerebral malaria by modulating CD8+ T cell accumulation within the brain.

It is well established that IFN-γ is required for the development of experimental cerebral malaria (ECM) during Plasmodium berghei ANKA infection of C57BL/6 mice. However, the temporal and tissue-specific cellular sources of IFN-γ during P. berghei ANKA infection have not been investigated, and it is not known whether IFN-γ production by a single cell type in isolation can induce cerebral pathology. In this study, using IFN-γ reporter mice, we show that NK cells dominate the IFN-γ response during the early stages of infection in the brain, but not in the spleen, before being replaced by CD4(+) and CD8(+) T cells. Importantly, we demonstrate that IFN-γ-producing CD4(+) T cells, but not innate or CD8(+) T cells, can promote the development of ECM in normally resistant IFN-γ(-/-) mice infected with P. berghei ANKA. Adoptively transferred wild-type CD4(+) T cells accumulate within the spleen, lung, and brain of IFN-γ(-/-) mice and induce ECM through active IFN-γ secretion, which increases the accumulation of endogenous IFN-γ(-/-) CD8(+) T cells within the brain. Depletion of endogenous IFN-γ(-/-) CD8(+) T cells abrogates the ability of wild-type CD4(+) T cells to promote ECM. Finally, we show that IFN-γ production, specifically by CD4(+) T cells, is sufficient to induce expression of CXCL9 and CXCL10 within the brain, providing a mechanistic basis for the enhanced CD8(+) T cell accumulation. To our knowledge, these observations demonstrate, for the first time, the importance of and pathways by which IFN-γ-producing CD4(+) T cells promote the development of ECM during P. berghei ANKA infection

Helper T cell IL-2 production is limited by negative feedback and STAT-dependent cytokine signals

Although required for many fundamental immune processes, ranging from self-tolerance to pathogen immunity, interleukin (IL)-2 production is transient, and the mechanisms underlying this brevity remain unclear. These studies reveal that helper T cell IL-2 production is limited by a classic negative feedback loop that functions autonomously or in collaboration with other common γ chain (IL-4 and IL-7) and IL-6/IL-12 family cytokines (IL-12 and IL-27). Consistent with this model for cytokine-dependent regulation, they also demonstrate that the inhibitory effect can be mediated by several signal transducer and activator of transcription (STAT) family transcription factors, namely STAT5, STAT4, and STAT6. Collectively, these findings establish that IL-2 production is limited by a network of autocrine and paracrine signals that are readily available during acute inflammatory responses and, thus, provide a cellular and molecular basis for its transient pattern of expression

ICOS Co-Stimulation: Friend or Foe?

Over the last 15 years, the inducible T cell co-stimulator (ICOS) has been implicated in a wide variety of immune outcomes, including the induction and regulation of Th1, Th2, and Th17 immunity. In addition to its role in directing effector T cell differentiation, ICOS has also been consistently linked with the induction of thymus-dependent Ab responses and the germinal center reaction. ICOS co-stimulation, therefore, appears to play a complex role in dictating the course of adaptive immunity. In this article, we summarize the initial characterization of ICOS and its relationship with the related co-stimulatory molecule CD28. We then address the contribution of ICOS in directing an effector T cell response, and ultimately disease outcome, against various bacterial, viral, and parasitic infections. Next, we assess ICOS in the context of TD Ab responses, connecting ICOS signaling to Tfh cell differentiation and its role in the germinal center reaction. Finally, we address the link between ICOS and human autoimmune disorders, and evaluate potential therapies aiming to mitigate disease progression by modulating ICOS signaling

IL-21 is required for optimal antibody production and T cell responses during chronic Toxoplasma gondii infection.

Previous studies have indicated that Il21r (-/-) mice chronically infected with Toxoplasma gondii display a defect in serum IgG; however, the basis for this antibody defect was not defined and questions remain about the role of IL-21 in promoting the production of IL-10, which is required to limit infection-induced pathology during toxoplasmosis. Therefore, Il21 (-/-) mice were challenged with T. gondii to determine whether IL-21 impacts the parasite-specific CD8(+) T cell response, its contribution to thymus-dependent antibody production after infection, and balance between protective and pathogenic responses. Whereas IL-21 has been implicated in the differentiation of IL-10 producing CD4(+) T cells no immune-mediated pathology was evident in Il21 (-/-) mice during the acute response, nor was there a defect in the development of this population in chronically infected Il21 (-/-) mice. However, Il21 (-/-) mice displayed a defect in IgG production after infection that correlated with a decrease in GC B cell numbers, the CD4(+) and CD8(+) T cell numbers in the brain were reduced over the course of the chronic infection leading to a decrease in total IFN-γ production and an increase in parasite numbers associated with susceptibility to toxoplasmic encephalitis. Together, these results identify a key role for IL-21 in shaping the humoral and cellular response to T. gondii, but indicate that IL-21 has a limited role in regulating immunopathology

ICOS signaling promotes a secondary humoral response after re-challenge with Plasmodium chabaudi chabaudi AS.

The co-stimulatory molecule ICOS is associated with the induction and regulation of T helper cell responses, including the differentiation of follicular helper T (Tfh) cells and the formation and maintenance of memory T cells. However, the role of ICOS signaling in secondary immune responses is largely unexplored. Here we show that memory T cell formation and maintenance are influenced by persistent infection with P. chabaudi chabaudi AS infection, as memory T cell numbers decline in wild-type and Icos-/- mice after drug-clearance. Following drug-clearance Icos-/- mice display a relapsing parasitemia that occurs more frequently and with higher peaks compared to wild-type mice after re-challenge. The secondary immune response in Icos-/- mice is characterized by significant impairment in the expansion of effector cells with a Tfh-like phenotype, which is associated with a diminished and delayed parasite-specific Ab response and the absence of germinal centers. Similarly, the administration of an anti-ICOSL antagonizing antibody to wild-type mice before and after reinfection with P. c. chabaudi AS leads to an early defect in Tfh cell expansion and parasite-specific antibody production, confirming a need for ICOS-ICOSL interactions to promote memory B cell responses. Furthermore, adoptive transfer of central memory T (TCM) cells from wild-type and Icos-/- mice into tcrb-/- mice to directly evaluate the ability of TCM cells to give rise to Tfh cells revealed that TCM cells from wild-type mice acquire a mixed Th1- and Tfh-like phenotype after P. c. chabaudi AS infection. While TCM cells from Icos-/- mice expand and display markers of activation to a similar degree as their WT counterparts, they displayed a reduced capacity to upregulate markers indicative of a Tfh cell phenotype, resulting in a diminished humoral response. Together these findings verify that ICOS signaling in memory T cells plays an integral role in promoting T cell effector responses during secondary infection with P. c. chabaudi AS

Essential role for IL-27 receptor signaling in prevention of Th1-mediated immunopathology during malaria infection.

Successful resolution of malaria infection requires induction of proinflammatory immune responses that facilitate parasite clearance; however, failure to regulate this inflammation leads to immune-mediated pathology. The pathways that maintain this immunological balance during malaria infection remain poorly defined. In this study, we demonstrate that IL-27R-deficient (WSX-1(-/-)) mice are highly susceptible to Plasmodium berghei NK65 infection, developing exacerbated Th1-mediated immune responses, which, despite highly efficient parasite clearance, lead directly to severe liver pathology. Depletion of CD4(+) T cells---but not CD8(+) T cells---prevented liver pathology in infected WSX-1(-/-) mice. Although WSX-1 signaling was required for optimal IL-10 production by CD4(+) T cells, administration of rIL-10 failed to ameliorate liver damage in WSX-1(-/-) mice, indicating that additional, IL-10-independent, protective pathways are modulated by IL-27R signaling during malaria infection. These data are the first to demonstrate the essential role of IL-27R signaling in regulating effector T cell function during malaria infection and reveal a novel pathway that might be amenable to manipulation by drugs or vaccines

IL-21 is required for resistance to oral infection with <i>T. gondii</i>.

<p>A) Survival of <i>Il21</i>^−/− mice (<i>n</i> = 11) and their wild-type littermates (WT; <i>n</i> = 7) infected orally, or intraperitoneally (<i>Il21</i>^−/−<i>n</i> = 6 and WT <i>n</i> = 6) with 20 cysts from the ME49 strain of <i>T. gondii.</i> B) Quantitative real-time PCR of parasite DNA isolated from the small intestine of WT and <i>Il21</i>^−/− mice infected orally with <i>T. gondii</i> for seven days. Results are representative of two experiments with three or four mice per group. Error bars represent SEM. C) Histopathology of the small intestine of WT and <i>Il21</i>^−/− mice infected orally for twelve days, analyzed by staining with haematoxylin and eosin. Original magnification, ×20. D) Intracellular staining for IFN-γ⁺ CD4⁺ and CD8⁺ T cells from the spleen of WT and <i>Il21</i>^−/− mice infected orally for seven days, after stimulation for 4 h <i>ex vivo</i> with PMA and ionomycin in the presence of brefeldin A. Numbers outside the boxed areas indicate percent IFN-γ⁺ CD4⁺ (top row) or CD8⁺ T cells (bottom row). Data are representative of two independent experiments with similar results.</p

<i>Il21</i>^−/− mice have reduced inflammatory cell numbers in the brain during chronic <i>T. gondii</i> infection.

<p>A) The percentage of CD4⁺ (left) and CD8⁺ (right) T cells expressing the cytokine IL-10 in the brain of individual WT and <i>Il21</i>^−/− mice infected for 35 days. Data are representative of two or three independent experiments with three to four mice per group. B) Enumeration of total cell numbers recovered from the brain of WT and <i>Il21</i>^−/− mice infected for 35 days. Data are representative of three experiments with similar results. C) Total CD4⁺ T cells, CD8⁺ T cells, macrophages and microglia in each BMNC preparation from (B), calculated by the percentages determined by flow cytometry. D) Total IL-10⁺ CD4⁺ and CD8⁺ T cells in each BMNC preparation from (A), calculated by the percentages determined by flow cytometry. E) Flow cytometry of CD4⁺ T cells isolated from WT and <i>Il21</i>^−/− mice, and activated with anti-CD3 and anti-CD28 in cRPMI (left) or IMDM (right) media in the presence of IL-27 for 4 days. Cells were then stimulated for 4 h with PMA and ionomycin in the presence of brefeldin A before staining for intracellular IL-10. Numbers in boxed areas indicate percent IL-10⁺CD4⁺ T cells. Results are representative of three independent experiments with similar results. Significance was determined by a two-tailed unpaired Student’s <i>t</i> test. Error bars represent SEM. *<i>P</i><0.05, **<i>P</i><0.01.</p

T cells from the brain of <i>Il21</i>^−/− mice produce less IFN-γ during chronic toxoplasmic encephalitis.

<p>A) Flow cytometry of BMNCs from WT and <i>Il21</i>^−/− mice infected for 35 days; cells were stimulated <i>ex vivo</i> with PMA and ionomycin in the presence of brefeldin A for 4 h and then stained intracellularly for IFN-γ. Numbers in boxed areas represent percent IFN-γ⁺ CD4⁺ (left) or CD8⁺ (right) T cells, while bold numbers indicate MFI. B) Total IFN-γ⁺CD4⁺ or CD8⁺ T cells isolated from BMNC preparations of WT (black bars) and <i>Il21</i>^−/− (white bars) mice infected for 35 days, calculated from the percentages determined by flow cytometry. Data are representative of four independent experiments with similar results. Error bars represent SEM. Significance was determined by a two-tailed unpaired Student’s <i>t</i> test. **<i>P</i><0.01.</p