Showing papers by "David Voehringer published in 2014"

Basophil-mediated protection against gastrointestinal helminths requires IgE-induced cytokine secretion

Abstract: Basophils orchestrate protection against reinfections with gastrointestinal helminths and ticks, but the underlying mechanisms remain elusive. We investigated the role of Fc receptors on basophils, the antibody isotypes IgG1 and IgE, and basophil-derived IL-4/IL-13 during challenge infections with Heligmosomoides polygyrus and Nippostrongylus brasiliensis. Using mixed bone marrow chimeras, we found that activating Fc receptors on basophils were required for protective immunity but not for regulation of basophil homeostasis. Furthermore, rapid worm expulsion was impaired in IgE-deficient but not in IgG1-deficient mice. Basophils promoted the recruitment of other effector cells into the small intestine and induced expression of the antihelminthic proteins resistin-like molecule β and mucin 5ac. Selective deletion of IL-4/IL-13 in basophils resulted in impaired worm expulsion. Collectively, our results indicate that IgE-mediated activation of basophils and the release of basophil-derived IL-4/IL-13 are critical steps in protective immunity against helminths. Therefore, development of effective vaccines against helminths should consider boosting the IL-4/IgE/basophil axis of the immune system.

B-cell-intrinsic STAT6 signaling controls germinal center formation

Abstract: Infection with helminths and exposure to antigens induce a strong type 2 immune response resulting in the secretion of the cytokines IL-4 and IL-13 by CD4(+) T cells and several innate cell types. IL-4 and IL-13 promote class switch recombination to IgG1 and IgE while their role for germinal center (GC) formation is poorly understood. We found a dramatic reduction in the numbers of GC B cells when investigating different type 2 immune responses in IL-4/IL-13-deficient mice. IL-4/IL-13 from T cells located outside B-cell follicles was sufficient for GC formation. We further revealed that IL-4/IL-13 acts directly on B cells for the formation of a robust GC response. The frequency of apoptotic GC B cells was not altered in the absence of IL-4/IL-13 and proliferation was even enhanced. However, deficiency of signal transducer and activator of transcription 6 signaling in B cells resulted in failure to downregulate the chemotactic receptor Gpr183 (Ebi2) and downregulation of this receptor has been shown to be essential for proper GC B-cell differentiation. Thus, T-cell-derived extrafollicular IL-4/IL-13 and signal transducer and activator of transcription 6-regulated genes in B cells play a critical role for orchestration of the GC response in type 2 immunity.

T Cell–Derived IL-4/IL-13 Protects Mice against Fatal Schistosoma mansoni Infection Independently of Basophils

Abstract: Schistosomiasis is a severe and chronic disease caused by the parasitic trematode Schistosoma mansoni after deposition of eggs in the liver and intestines. The immune response to S. mansoni eggs is characterized by increased Th2 cells, eosinophilia, and high serum IgE levels. Granulomas are formed around the eggs to protect the organs against tissue damage caused by toxic products that are secreted from the eggs. Egg-derived components have further been shown to activate the IgE-mediated release of IL-4 and IL-13 from basophils, suggesting that basophils could be involved in protection against a fatal course of infection. Using T cell-specific IL-4/IL-13-deficient mice and basophil-deficient Mcpt8Cre mice, we determined the contribution of Th2 cells and basophils for protective immunity against S. mansoni egg-induced pathology during the patent stage of infection. Our results demonstrate that T cell-derived IL-4/IL-13 was essential for granuloma formation, IgE production, basophilia, differentiation of alternatively activated macrophages, and protection against fatal infection. Although basophils were recruited into liver granulomas, they appeared to be dispensable as a source of IL-4/IL-13 both for differentiation of Th2 cells and for prevention of weight loss and mortality.

CreER(T2) expression from within the c-Kit gene locus allows efficient inducible gene targeting in and ablation of mast cells.

Abstract: Mast cells are abundantly situated at contact sites between the body and its environment, such as the skin and, especially during certain immune responses, at mucosal surfaces. They mediate allergic reactions and degrade toxins as well as venoms. However, their roles during innate and adaptive immune responses remain controversial and it is likely that major functions remain to be discovered. Recent developments in mast cell-specific conditional gene targeting in the mouse promise to enhance our understanding of these fascinating cells. To complete the genetic toolbox to study mast cell development, homeostasis and function, it is imperative to inducibly manipulate their gene expression. Here, we report the generation of a novel knock-in mouse line expressing a tamoxifen-inducible version of the Cre recombinase from within the endogenous c-Kit locus. We demonstrate highly efficient and specific inducible expression of a fluorescent reporter protein in mast cells both in vivo and in vitro. Furthermore, induction of diphtheria toxin A expression allowed selective and efficient ablation of mast cells at various anatomical locations, while other hematopoietic cells remain unaffected. This novel mouse strain will hence be very valuable to study mast cell homeostasis and how specific genes influence their functions in physiology and pathology.

Swiprosin‐1/EFhd2 limits germinal center responses and humoral type 2 immunity

Abstract: Activated B cells are selected for in germinal centers by regulation of their apoptosis. The Ca2+ -binding cytoskeletal adaptor protein Swiprosin-1/EFhd2 (EFhd2) can promote apoptosis in activated B cells. We therefore hypothesized that EFhd2 might limit humoral immunity by repressing both the germinal center reaction and the expected enhancement of immune responses in the absence of EFhd2. Here, we established EFhd2(-/-) mice on a C57BL/6 background, which revealed normal B- and T-cell development, basal Ab levels, and T-cell independent type 1, and T-cell independent type 2 responses. However, T cell-dependent immunization with sheep red blood cells and infection with the helminth Nippostrongylus brasiliensis (N.b) increased production of antibodies of multiple isotypes, as well as germinal center formation in EFhd2(-/-) mice. In addition, serum IgE levels and numbers of IgE+ plasma cells were strongly increased in EFhd2(-/-) mice, both after primary as well as after secondary N.b infection. Finally, mixed bone marrow chimeras unraveled an EFhd2-dependent B cell-intrinsic contribution to increased IgE plasma cell numbers in N.b-infected mice. Hence, we established a role for EFhd2 as a negative regulator of germinal center-dependent humoral type 2 immunity, with implications for the generation of IgE.

Identification of murine basophils by flow cytometry and histology.

Abstract: Here we describe how murine basophils can be detected in vivo by flow cytometry and immunofluorescence staining. Basophils constitute a homogeneous population of CD4(-)CD19(-)CD49b(+)IgE(+) cells in flow cytometric analysis. When IgE levels are low one can also use anti-FceRI or anti-CD200R3 antibodies instead of anti-IgE. For immunofluorescence staining we use an anti-Mcpt8 antibody since Mcpt8 is a specific marker for murine basophils. We describe how to prepare the tissue to cut cryo-sections and how to perform the staining using a tyramide-based amplification kit.

A6.4 Induction of TH2 cells and eosinophil by infection with nippostrongylus brasiliensis protects against rheumatoid arthritis

Abstract: Background and Objective Infection with helminth triggers strong Th2 immune response, which modulates the development of systemic autoimmune diseases. The aim of this study was to investigate whether infection with Nippostrongylus brasiliensis influence rheumatoid arthritis development as well as the molecular mechanism from Th2 and eosinophil cells in this effect. Materials and Methods 5 to 7 weeks old human TNF transgenic (hTNFtg) mice were infected with Nippostrongylus brasiliensis subcutaneously. Moreover, wild-type, IL-4 -/- IL-13 -/- and CD4-Cre IL-4 -/- IL-13 -/ mice were infected with Nippostrongylus brasiliensis subcutaneously 6 days before induction of experimental arthritis through injection of 150 µl K/BxN serum intraperitoneally. Clinical arthritis score and histomorphometric analysis in the inflammatory hind paw were evaluated. Results Infection with Nippostrongylus brasiliensis alleviated spontaneous chronic arthritis in hTNFtg mice accompanied with a reduction of the systemic and local bone loss. Moreover, wild-type and CD4cre IL-4 -/- IL-13 -/ mice infected with Nippostrongylus brasiliensis have a reduction of the arthritis score whereas arthritis in IL-4 -/- IL-13 -/- mice with infection remained comparable to the non-infected control mice. Histomorphometric analyses revealed that inflammation area and bone erosion were decreased in wild-type and CD4cre IL-4 -/- IL-13 -/ mice with infection compared with IL-4 -/- IL-13 -/- or noninfected wild-type control. Consistently, osteoclast marker such as Trap, RANK and CathepsinK mRNA expression levels in the inflammatory joint were lower in wild-type mice with infection than control. In order to define the role of eosinophil, induction of K/BxN serum transfer arthritis in delta-dblGATA mice was performed. Interestingly genetically lack of eosinophil showed increased inflammation and bone erosion than wild-type control. Conclusions Infection with Nippostrongylus brasiliensis significantly alleviated bone erosion in experimental arthritis models, which is dependent of IL-4 and IL-13. In addition, eosinophils might play a crucial role in arthritis protection induced by Nippostrongylus brasiliensis infection.