Basophils induce Th2 immunity

Abstract

Despite the fact that basophils represent less than 0.5% of circulating leukocytes, recent studies have begun to unveil their potent immunoregulatory functions, i.e., induction of Th2 immunity. It is believed that basophils are capable of doing so primarily by secreting key Th2-inducing cytokines, namely IL-4 and Thymic Stromal Lymphopoietin (TSLP), and by functioning as professional antigen presenting cells. However, we have recently demonstrated that Th2 immunity can develop in the absence of basophils or IL-4 during helminth infection. In this review, how basophils may (and may not) contribute to the development of Th2 immunity in vivo is discussed.

Upon activation naïve CD4 T cells undertake a differentiation process through which they express effector cytokines and acquire ‘help and/or effector’ functions that are distinctive based on the differentiation program.¹ Among the well-studied factors controlling the process are cytokines provided from activated T cell themselves or from accessory cells such as antigen presenting cells (APC). Whereas Th1 or newly rising Th17 effector differentiation pathway has been extensively studied, the process of Th2 differentiation has always been a pathway of uncertainty.² IL-4 is the major effector cytokine produced by Th2 effector cells; yet, it is also necessary for naïve T cells to choose the Th2-lineage pathway, by which the IL-4 induces the ‘Th2-signature’ genes via key transcription factors, STAT6 and GATA3.¹ The question has always been the initial source of the IL-4. A new idea that basophils are important IL-4-providing Th2 inducers has been proposed and challenged by multiple studies.^3–5

Using a reporter animal that expresses the green fluorescence protein (GFP) under the Il4 promoter,⁶ we reported that basophils are the major IL-4 producing cells during the infection of helminth, Nippostrongylus brasiliensis (Nb).⁷ The findings that the level of IL-4 production in basophils is substantially greater than that in Th2 cells and that basophils even in naïve animals express preformed IL-4 mRNA⁷ raised an exciting hypothesis that basophils may serve as the initial source of IL-4 that drives the development of Th2 immune responses in vivo.⁸ During the infection, basophilia as well as the accumulation of basophils in the liver was observed. Interestingly, these basophil responses were not induced in Rag^-/- mice infected with Nb and the adoptive transfer of CD4 T cells into these mice was sufficient to increase the basophil levels similar to that of Nb infected WT mice.⁷ These findings strongly suggest that T cell activation plays a critical role in inducing basophil responses. In support of this, transfer of IL-3-deficient CD4 T cells into Nb infected Rag^-/- mice failed to induce basophil responses, further indicating that IL-3 produced by activated CD4 T cells is primarily responsible for inducing basophil generation in the bone marrow and the subsequent accumulation in the liver of infected mice.⁹ Therefore, Nb specific CD4 T cells produce IL-3, which then stimulates the basophil progenitors present in the bone marrow and possibly in the spleen¹⁰ to differentiate into basophils and that IL-4 production by basophils is further enhanced.¹¹ Nonetheless, it remains unclear how circulating basophils support Th2 differentiation that primarily occurs in the secondary lymphoid tissues where parasite antigens are presented to activated naïve CD4 T cells.

A clue for this conundrum came from a study by Medzhitov and colleagues that showed a transient migration of circulating basophils into the draining lymph nodes following protease antigen, papain immunization.¹² Recruited basophils were mainly found in the T-cell zones where they appear to locate in close proximity to T cells, probably providing both IL-4 and TSLP to activated naïve T cells.¹² In support of this, three independent studies have recently demonstrated that basophils can prime naïve CD4 T cells both in vitro and in vivo by directly presenting antigenic peptides via surface expression of MHC II molecules.^13–15 Of course, IL-4 production by basophils mainly drives the Th2 differentiation. Most importantly, CD11c⁺ dendritic cells were relatively dispensable for the Th2 immunity to develop, further strengthening the idea that basophils are primary antigen presenting cells to generate antigen specific Th2 CD4 T cells in vivo.

A major challenge against this hypothesis came from our new finding that basophils are not essential for Th2 immune responses to Nb infection. Interestingly, IL-3 plays a key role in recruiting circulating basophils into the draining LN after Nb infection.¹⁶ Similar to the papain-induced basophil recruitment, circulating basophils enter the draining mediastinal LN between days 3 and 4 post infection and the mesenteric LN around day 10 post infection, the kinetics of which closely matches the migration pattern of the parasites in vivo; thus the recruitment seems driven by antigen-mediated T-cell activation. Such recruitment is completely abolished in Nb infected mice deficient in IL-3 or IL-3 receptor.¹⁶ In vitro, IL-3 has been shown to upregulate expression of surface adhesion molecules and of chemokines in endothelial cells, both of which contribute to basophil adhesion as well as transendothelial migration.¹⁷ In vivo, however, the target cells of IL-3 that mediates basophil LN entry were of bone marrow origin, arguing against the in vitro observation.¹⁵ Since Nb infected IL-3-deficient mice have no basophils recruited into the draining LN, it is expected that Nb specific Th2 immune responses would fail to develop. However, Nb infected IL-3-deficient mice still mount Th2 immune responses (both Th2-producing CD4 T cells and IgE production) that are equivalent to WT mice, suggesting that basophil LN entry and possibly basophil-mediated antigen presentation may be dispensable for the induction of Th2 immunity in Nb infected mice. In support of this finding, basophil depletion in Nb infected mice did not impair the immune responses.

So, how can we reconcile the discrepancy between these studies? I would argue that the type of immune responses (or of antigens) plays dominant roles in determining basophil dependency. Schistosoma egg antigen (SEA) was shown to suppress DC functions, resulting the emergence of IL-4-producing CD4 T cells.¹⁸ On the other hand, Nb-secreted proteins could induce selective maturation of DC, thus promote Th2 differentiation.^19,20 These results suggest that parasite-associate antigens are capable of bypassing the need of basophils (or of IL-4) to induce Th2 immunity. In a T. muris infection study, worm burden was significantly elevated following basophil depletion.¹⁴ It should be noted that the contribution of basophils to the development of T. muris specific Th2 type CD4 T cells during the infection has not been examined, although basophils were still capable of inducing Th2 differentiation in vitro.¹⁴ Moreover, basophils were not required when IFNγ function was blocked.¹⁴ Therefore, basophils may regulate the balance of Th1/Th2 immunity and/or function as effector cells rather than as Th2-inducer cells during parasite infection, while they may play nonredundant roles in Th2 differentiation mediated by ‘non-parasite’ antigens with a capacity to directly activate basophils.¹²

So, what's next? First, because basophil antigen presenting functions are critical to induce antigen specific Th2 immunity under certain circumstances, it is important to understand how basophils obtain antigenic peptides (or proteins) to present to T cells in vivo. Second, a recent study reported that DC-induced reactive oxygen species induces CCL7 production by DCs, which mediates IL-4⁺ basophil recruitment into the draining LN, proposing a novel possibility of basophil-DC cooperation during Th2 immunity.²¹ Whether this type of cellular interaction occurs during helminth infection should be examined. Third, it was recently reported that Nb infection can induce Th2 type immune responses in an IL-4/STAT6-independent manner.²² Although IL-4-independent Th2 immunity has been demonstrated in many different experimental models,^23–25 it will be important to reexamine if basophils are involved in these processes. Last, basophil LN entry is a key step for the immunoregulatory roles of basophils to occur in vivo. How IL-3 or possibly dendritic cell-derived CCL7, attracts circulating basophils into the lymphoid tissues should be an important subject for future study. Because basophils themselves express IL-3 receptors,²⁶ it is possible that IL-3 may directly act on the basophils. IL-3 produced by activated T cells may upregulate adhesion molecules on the endothelial cells to which circulating basophils adhere. IL-3 may then signal through the receptor on the basophils to induce transmigration. Because IL-3 receptor expression on the endothelial cells was unnecessary in this process, it will be important to understand how IL-3 is presented to the circulating basophils. As always, more questions arise as we learn something new.

Addendum to: Kim S, Prout M, Ramshaw H, Lopez AF, LeGros G, Min B. Cutting edge: basophils are transiently recruited into the draining lymph nodes during helminth infection via IL-3, but infection-induced Th2 immunity can develop without basophil lymph node recruitment or IL-3. J Immunol. 2010;184:1143–1147. doi: 10.4049/jimmunol.0902447.