Amy S. McKee

CD25+CD4+ Cells Contribute to Th2 Polarization during Helminth Infection by Suppressing Th1 Response Development

Mice infected with Schistosoma mansoni develop polarized Th2 responses in which Th1 responses are prevented by IL-10-mediated suppression of IL-12 production. We show that dendritic cells from infected mice are primed to make IL-12 in response to CD40 ligation, and that IL-10 acts by inhibiting this process. In infected mice, two subpopulations of CD4+ cells, separable by their expression of CD25, make IL-10. CD25+CD4+ cells expressed forkhead box P3, inhibited proliferation of CD4+ T cells, and made IL-10, but little IL-5. In contrast, CD25−CD4+ cells failed to express forkhead box P3 or to inhibit proliferation and accounted for all the IL-5, IL-6, and IL-13 produced by unseparated splenic populations. Thus, CD25+ and CD25− subpopulations could be characterized as regulatory T cells (Treg cells) and Th2 cells, respectively. Consistent with their ability to make IL-10, both CD25+ and CD25−CD4+ T cells from infected mice were able, when stimulated with egg Ag, to suppress IL-12 production by CD40 agonist-stimulated dendritic cells. Additionally, in adoptive transfer experiments, both CD4+ subpopulations of cells were able to partially inhibit the development of Th1 responses in egg-immunized IL-10−/− mice. The relationship of Treg cells in infected mice to natural Treg cells was strongly suggested by the ability of CD25+CD4+ cells from naive mice to inhibit Th1 response development when transferred into egg-immunized or infected IL-10−/− mice. The data suggest that natural Treg cells and, to a lesser extent, Th2 cells play roles in suppressing Th1 responses and ensuring Th2 polarization during schistosomiasis.

Helminth antigens modulate TLR-initiated dendritic cell activation.

There is increasing awareness that helminth infections can ameliorate proinflammatory conditions. In part, this is due to their inherent ability to induce Th2 and, perhaps, regulatory T cell responses. However, recent evidence indicates that helminths also have direct anti-inflammatory effects on innate immune responses. In this study, we address this issue and show that soluble molecules from the eggs of the helminth parasite Schistosoma mansoni (SEA) suppress LPS-induced activation of immature murine dendritic cells, including MHC class II, costimulatory molecule expression, and IL-12 production. SEA-augmented LPS-induced production of IL-10 is in part responsible for the observed reduction in LPS-induced IL-12 production. However, analyses of IL-10−/− DC revealed distinct IL-10-independent suppressive effects of SEA. IL-10-independent mechanisms are evident in the suppression of TLR ligand-induced MAPK and NF-κB signaling pathways. Microarray analyses demonstrate that SEA alone uniquely alters the expression of a small subset of genes that are not up-regulated during conventional TLR-induced DC maturation. In contrast, the effects of SEA on TLR ligand-induced DC activation were striking: when mixed with LPS, SEA significantly affects the expression of >100 LPS-regulated genes. These findings indicate that SEA exerts potent anti-inflammatory effects by directly regulating the ability of DC to respond to TLR ligands.

Th2 response polarization during infection with the helminth parasite Schistosoma mansoni.

Summary:  T‐helper 2 (Th2) cell responses play a critical role in protection against helminth infections. In the case of Schistosoma mansoni, an important helminth parasite of man, data from a mouse model of human disease have shown that Th2 responses are essential to allow host survival. In this infection, parasite eggs are the primary stimulus for Th2 response development. Recent work has shown that egg molecules exert multiple levels of control over the development of host interferon‐γ‐associated inflammatory responses. Soluble egg antigen inhibits the ability of dendritic cells to make interleukin‐12 and induces Th2‐polarized adaptive immune responses that in combination with regulatory T‐cell responses effectively limit Th1 response development. In this article, we discuss the factors influencing Th2 response polarization during infection with S. mansoni.

Functional Inactivation of Immature Dendritic Cells by the Intracellular Parasite Toxoplasma gondii

Despite its noted ability to induce strong cellular immunity, and its known susceptibility to IFN-γ-dependent immune effector mechanisms, the protozoan Toxoplasma gondii is a highly successful parasite, able to replicate, disseminate, and either kill the host or, more commonly, establish resistant encysted life forms before the emergence of protective immune responses. We sought to understand how the parasite gains the advantage. Using transgenic clonal parasite lines engineered to express fluorescent markers in combination with dendritic cells (DC) grown from the bone marrow of wild-type mice or transgenic mice expressing fluorescent protein-tagged MHC class II molecules, we used flow cytometry and fluorescence microscopy to analyze the responses of infected DC to both invasion by the parasite and subsequent DC maturation signals. We found that T. gondii preferentially invades immature dendritic cells but fails to activate them in the process, and renders them resistant to subsequent activation by TLR ligands or the immune-system-intrinsic maturation signal CD40L. The functional consequences of T. gondii-mediated suppression of DC activation are manifested in a relative inability of infected immature DC to activate naive CD4+ Th lymphocytes, or to secrete cytokines, such IL-12 and TNF-α, that play important roles in innate and/or adaptive immunity. The findings reveal that T. gondii suppresses the ability of immature DC to participate in innate immunity and to induce adaptive immune responses. The ability of T. gondii to temporarily evade recognition could provide a selective advantage that permits dissemination and establishment before adaptive immune response initiation.

Memory CD4 T Cells That Express CXCR5 Provide Accelerated Help to B Cells

CD4 T cell help for B cells is critical for effective Ab responses. Although many of the molecules involved in helper functions of naive CD4 T cells have been characterized, much less is known about the helper capabilities of memory CD4 T cells, an important consideration for the design of vaccines that aim to prime protective memory CD4 T cells. In this study, we demonstrate that memory CD4 T cells enable B cells to expand more rapidly and class switch earlier than do primary responding CD4 T cells. This accelerated response does not require large numbers of memory cells, and similar numbers of primary responding cells provide less effective help than do memory cells. However, only memory CD4 T cells that express the B cell follicle homing molecule, CXCR5, are able to accelerate the response, suggesting that the rapidity of the Ab response depends on the ability of CD4 memory T cells to migrate quickly toward B cells.

Vaccine adjuvants aluminum and monophosphoryl lipid A provide distinct signals to generate protective cytotoxic memory CD8 T cells

Vaccines can greatly reduce the spread of and deaths from many infectious diseases. However, many infections have no successful vaccines. Better understanding of the generation of protective CD8 memory T cells by vaccination is essential for the rational design of new vaccines that aim to prime cellular immune responses. Here we demonstrate that the combination of two adjuvants that are currently licensed for use in humans can be used to prime long-lived memory CD8 T cells that protect mice from viral challenge. The universally used adjuvant, aluminum salts, primed long-lived memory CD8 T cells; however, effective cytotoxic T-cell differentiation occurred only in the presence of an additional adjuvant, monophosphoryl lipid A (MPL). MPL-induced IL-6 was required for cytotoxic differentiation. The IL-6 acted by inducing granzyme B production and reducing expression of inhibitory molecule PD1 on the surface of the primed CD8 T cells. CD8 memory T cells generated by antigen delivered with both aluminum salts and MPL provided significant protection from influenza A challenge. These adjuvants could be used in human vaccines to prime protective memory CD8 T cells.

Host DNA released in response to aluminum adjuvant enhances MHC class II-mediated antigen presentation and prolongs CD4 T-cell interactions with dendritic cells.

Significance Alum has been used to improve the efficacy of vaccines since the 1930s. Here we show that alum acts in part via host DNA to increase the interaction time between T cells and APCs. Many vaccines include aluminum salts (alum) as adjuvants despite little knowledge of alum’s functions. Host DNA rapidly coats injected alum. Here, we further investigated the mechanism of alum and DNA’s adjuvant function. Our data show that DNase coinjection reduces CD4 T-cell priming by i.m. injected antigen + alum. This effect is partially replicated in mice lacking stimulator of IFN genes, a mediator of cellular responses to cytoplasmic DNA. Others have shown that DNase treatment impairs dendritic cell (DC) migration from the peritoneal cavity to the draining lymph node in mice immunized i.p. with alum. However, our data show that DNase does not affect accumulation of, or expression of costimulatory proteins on, antigen-loaded DCs in lymph nodes draining injected muscles, the site by which most human vaccines are administered. DNase does inhibit prolonged T-cell–DC conjugate formation and antigen presentation between antigen-positive DCs and antigen-specific CD4 T cells following i.m. injection. Thus, from the muscle, an immunization site that does not require host DNA to promote migration of inflammatory DCs, alum acts as an adjuvant by introducing host DNA into the cytoplasm of antigen-bearing DCs, where it engages receptors that promote MHC class II presentation and better DC–T-cell interactions.

Immune mechanisms of protection: can adjuvants rise to the challenge?

For many diseases vaccines are lacking or only partly effective. Research on protective immunity and adjuvants that generate vigorous immune responses may help generate effective vaccines against such pathogens.

Aluminum adjuvants elicit fibrin-dependent extracellular traps in vivo

It has been recognized for nearly 80 years that insoluble aluminum salts are good immunologic adjuvants and that they form long-lived nodules in vivo. Nodule formation has long been presumed to be central for adjuvant activity by providing an antigen depot, but the composition and function of these nodules is poorly understood. We show here that aluminum salt nodules formed within hours of injection and contained the clotting protein fibrinogen. Fibrinogen was critical for nodule formation and required processing to insoluble fibrin by thrombin. DNase treatment partially disrupted the nodules, and the nodules contained histone H3 and citrullinated H3, features consistent with extracellular traps. Although neutrophils were not essential for nodule formation, CD11b(+) cells were implicated. Vaccination of fibrinogen-deficient mice resulted in normal CD4 T-cell and antibody responses and enhanced CD8 T-cell responses, indicating that nodules are not required for aluminums adjuvant effect. Moreover, the ability of aluminum salts to retain antigen in the body, the well-known depot effect, was unaffected by the absence of nodules. We conclude that aluminum adjuvants form fibrin-dependent nodules in vivo, that these nodules have properties of extracellular traps, and the nodules are not required for aluminum salts to act as adjuvants.

CD4 memory T cells divide poorly in response to antigen because of their cytokine profile

Immunological memory is a hallmark of adaptive immunity, and understanding T cell memory will be central to the development of effective cell-mediated vaccines. The characteristics and functions of CD4 memory cells have not been well defined. Here we demonstrate that the increased size of the secondary response is solely a consequence of the increased antigen-specific precursor frequency within the memory pool. Memory cells proliferated less than primary responding cells, even within the same host. By analyzing the entry of primary and memory cells into the cell cycle, we found that the two populations proliferated similarly until day 5; after this time, fewer of the reactivated memory cells proliferated. At this time, fewer of the reactivated memory cells made IL-2 than primary responding cells, but more made IFNγ. Both these factors affected the low proliferation of the memory cells, because either exogenous IL-2 or inhibition of IFNγ increased the proliferation of the memory cells.