Basophils and mast cells play critical roles for leukocyte recruitment in IgE-mediated cutaneous reverse passive Arthus reaction.

BACKGROUND: The pathogenic role of IgE has been implicated in a variety of allergic and inflammatory diseases. We have previously established an IgE-mediated cutaneous reverse passive Arthus model in which eosinophil infiltration is a prominent feature. This uniquely provides a model of type III hypersensitivity in which Fc classes of Ig that forms immune complex differentially determine the disease manifestation. OBJECTIVE: To investigate the mechanisms of how mast cells and basophils regulate this IgE-mediated Arthus reaction. METHODS: IgE-mediated cutaneous reverse passive Arthus reaction was induced in wild-type C57BL/6 or WBB6F1-+/+ mice and mast-cell-deficient WBB6F1-W/W(v) mice by intradermal injection of IgE anti-trinitrophenyl antibodies followed immediately by intravenous administration of trinitrophenyl bovine serum albumin. Basophils were depleted in vivo using anti-CD200R3 monoclonal antibody prior to the IC challenge. RESULTS: Hemorrhage and infiltration of eosinophils, neutrophils, and basophils were significantly reduced but were not completely abrogated in WBB6F1-W/W(v) mice compared with those in wild-type WBB6F1-+/+ mice. Wild-type C57BL/6 mice treated by basophil-depleting mAb also showed significantly decreased hemorrhage and inflammatory cell infiltration, especially that of eosinophils, compared with control mice. Furthermore, basophil depletion in WBB6F1-W/W(v) mice led to nearly complete inhibition of eosinophil recruitment. By contrast, basophil depletion did not further decrease neutrophil infiltration in WBB6F1-W/W(v) mice. CONCLUSION: While mast cells play a central role, basophils also have an important function, especially for eosinophil recruitment, in IgE-mediated cutaneous reverse passive Arthus reaction.

Basophil-derived mouse mast cell protease 11 induces microvascular leakage and tissue edema in a mast cell-independent manner.

Mouse mast cell protease 11 (mMCP-11) is the most recently identified member of the mouse mast cell tryptase family. This tryptase is preferentially produced by basophils in contrast to other members that are expressed by mast cells but not basophils. Although blood-circulating basophils have long been considered as minor and redundant relatives of tissue-resident mast cells, recent studies illustrated that basophils and mast cells play distinct roles in vivo. To explore the in vivo role of basophil-derived mMCP-11, here we prepared recombinant mMCP-11 and its protease-dead mutant. Subcutaneous injection of the wild-type mMCP-11 but not the mutant induced edematous skin swelling with increased microvascular permeability in a dose-dependent manner. No apparent infiltration of proinflammatory cells including neutrophils and eosinophils was detected in the skin lesions. The cutaneous swelling was abolished by the pretreatment of mice with indomethacin, a cyclooxygenase inhibitor, suggesting the major contribution of prostaglandins to the microvascular leakage. Of note, the cutaneous swelling was elicited even in mast cell-deficient mice, indicating that mast cells are dispensable for the mMCP-11-induced cutaneous swelling. Thus, basophil-derived mMCP-11 can induce microvascular leakage via prostaglandins in a mast cell-independent manner, and may contribute to the development of basophil-mediated inflammatory responses.

Emerging roles of basophils in protective immunity against parasites.

Basophils, the least common type of granulocyte, have long been considered as minor effector cells in allergic responses because of their ability to release allergy-inducing chemical mediators such as histamine and leukotriene C4. However, it is unlikely that many animal species evolutionarily conserve basophils to only elicit allergic responses without any host-beneficial function. The study of basophils has been hampered by their rarity and difficult identification, as well as the lack of suitable animal models. Recent studies using novel analytical tools, including basophil-depleting antibodies and genetically engineered mice deficient only in basophils, have illuminated the crucial and nonredundant roles for basophils in protective immunity against both ecto- and endoparasites.

Nonredundant roles of basophils in immunity.

Basophils are the rarest granulocytes and represent less than 1% of peripheral blood leukocytes. They are evolutionarily conserved in many animal species, but their functional significance remained an enigma long after their discovery by Paul Ehrlich in 1879. Studies of basophils were hindered by their rarity, by difficulties in identifying them, and by the paucity of useful analytical tools. Because basophils display several characteristics shared by tissue-resident mast cells, they were often considered minor and possibly redundant relatives of mast cells or even blood-circulating precursors of mast cells. However, newly developed tools for their functional analysis, including basophil-depleting antibodies and genetically engineered mice deficient only in basophils, have fueled basophil research and defined previously unrecognized functions of basophils. We now appreciate that basophils play nonredundant roles in acquired immunity regulation, protective immunity to pathogens, and immunological disorders such as allergy and autoimmunity.

IgG-mediated systemic anaphylaxis to protein antigen can be induced even under conditions of limited amounts of antibody and antigen.

Systemic anaphylaxis is an acute, severe, and potentially fatal allergic reaction. Two classes of antibodies, IgE and IgG, contribute to the development of anaphylaxis in mice, through different mechanisms with distinct usage of effector cells and chemical mediators. Larger quantities of antibody and antigen are reportedly required to induce IgG-mediated anaphylaxis than IgE-mediated one, suggesting that the former may not happen as frequently as the latter in real life. To readdress this issue, we established in the present study a novel mouse model of passive IgG-mediated systemic anaphylaxis to a native protein antigen, ovalbumin (OVA), rather than artificially haptenated protein antigens used in previous studies. Passive sensitization of mice with a cocktail of but not individual IgG1 mAbs specific to distinct OVA epitopes elicited systemic anaphylaxis in response to OVA challenge. Importantly, much smaller doses of antibody and antigen than previously reported were sufficient for the induction of IgG-mediated systemic anaphylaxis. Moreover, a relatively small dose of antigen could induce severe anaphylaxis through both IgE- and IgG-mediated mechanisms when mice had been passively sensitized with antigen-specific IgE and IgG. These results strongly suggest that IgG-mediated systemic anaphylaxis is not rare among antibody-mediated systemic anaphylaxis, in contrast to previous thought, and significantly contributes to active systemic anaphylaxis in real life, at least in mice.

Selective ablation of basophils in mice reveals their nonredundant role in acquired immunity against ticks.

Ticks are ectoparasitic arthropods that can transmit a variety of microorganisms to humans and animals during blood feeding, causing serious infectious disorders, including Lyme disease. Acaricides are pharmacologic agents that kill ticks. The emergence of acaricide-resistant ticks calls for alternative control strategies for ticks and tick-borne diseases. Many animals develop resistance to ticks after repeated infestations, but the nature of this acquired anti-tick immunity remains poorly understood. Here we investigated the cellular and molecular mechanisms underlying acquired resistance to Haemaphysalis longicornis ticks in mice and found that antibodies were required, as was IgFc receptor expression on basophils but not on mast cells. The infiltration of basophils at tick-feeding sites occurred during the second, but not the first, tick infestation. To assess the requirement for basophil infiltration to acquired tick resistance, mice expressing the human diphtheria toxin receptor under the control of the mast cell protease 8 (Mcpt8) promoter were generated. Diphtheria toxin administration to these mice selectively ablated basophils. Diphtheria toxin-mediated basophil depletion before the second tick infestation resulted in loss of acquired tick resistance. These data provide the first clear evidence, to our knowledge, that basophils play an essential and nonredundant role in antibody-mediated acquired immunity against ticks, which may suggest new strategies for controlling tick-borne diseases.

Role for basophils in systemic anaphylaxis.

For more than 100 years since the discovery of basophils by Paul Ehrlich, the functional significance of this rare leukocyte as compared to mast cells has remained an enigma. Studies on basophils have long been hampered by their rarity (less than 1% of peripheral blood leukocytes) and the lack of useful analytical tools such as model animals deficient only in basophils. Recent studies have now defined previously-unrecognized roles for basophils in both allergic responses and immune regulation, and markedly changed our image of basophils, from a neglected minority to a key player in the immune system. We have recently demonstrated that basophils and mast cells play distinct roles in systemic anaphylaxis in mice. Basophils are dispensable for IgE-mediated systemic anaphylaxis unlike mast cells. Instead, basophils play the major role in IgG-mediated systemic anaphylaxis. In vivo depletion of basophils protects mice from anaphylactic death. Upon capture of IgG-allergen complexes, basophils release platelet-activating factor that increases vascular permeability, leading to anaphylactic shock. Thus, there are two major, distinct pathways to allergen-induced systemic anaphylaxis: one mediated by basophils, IgG and platelet-activating factor, and the other 'classical' pathway mediated by mast cells, IgE and histamine.

[The roles for basophils in allergy].

Basophils are the least common granulocytes, and account for less than 1% of peripheral blood leukocytes. Because of this minority status and their phenotypic similarity to mast cells, basophils have often been neglected in immunological studies or considered to have minor, redundant roles in immune responses in vivo. We have recently demonstrated that basophils play critical roles in IgG-mediated systemic anaphylaxis and IgE-mediated chronic allergic inflammation. Basophils also promote the Th2 differentiation and enhance the humoral memory response. Thus, basophils are a key player in both immune regulation and allergic responses. Basophils and their products seem to be promising therapeutic targets for allergic disorders.

Basophils preferentially express mouse Mast Cell Protease 11 among the mast cell tryptase family in contrast to mast cells.

Tryptases and chymases are the major proteins stored and secreted by mast cells, and they have various biological functions. However, the nature of proteases produced by basophils has been poorly characterized, particularly in mice. mMCP-11 is the most recently discovered mast cell tryptase in mice and was originally identified as Prss34, which is transcribed in some mast cell-like cell lines and at the early stage in the culture of BMMC with IL-3. Curiously, Prss34 is preferentially expressed in the BM and spleen among normal tissues in contrast to other mast cell tryptases. Therefore, it remains elusive what types of cells express mMCP-11 in vivo. Here, we show that mMCP-11 is highly expressed by primary basophils and to a much lesser extent, by some mast cells. Prss34 transcripts were detected abundantly in primary and cultured basophils and very weakly in peritoneal mast cells or cultured BMMC. Conversely, transcripts for mMCP-6 and mMCP-7 tryptases were preferentially expressed by cultured and peritoneal mast cells but not basophils. We established a mMCP-11-specific mAb and showed that mMCP-11 proteins are indeed expressed by primary basophils and those infiltrating the affected tissues during allergic inflammation and parasitic infections. Some primary mast cells also expressed mMCP-11 proteins, albeit at a much lower level. Thus, basophils rather than mast cells are the major source of mMCP-11. This is the first study to demonstrate that mouse basophils produce a trypsin-like protease.

New insights into the roles for basophils in acute and chronic allergy.

Basophils represent less than 1% of peripheral blood leukocytes. They are often recruited to the site of allergic inflammation, albeit in small numbers. However, it remained uncertain whether basophils play any significant role in allergic reactions or act as minor and redundant 'circulating mast cells'. We have recently demonstrated that basophils play critical roles in systemic anaphylaxis and chronic allergic inflammation, distinctively from mast cells. Basophils are one of the major players in the IgG- but not IgE-mediated systemic anaphylaxis, in contrast to mast cells. In response to the allergen-IgG immune complexes, basophils release the platelet-activating factor rather than histamine as the major chemical mediator to induce the systemic anaphylaxis. The depletion of basophils protects mice from death due to anaphylactic shock. Basophils also play a crucial role in the development of the IgE-mediated chronic allergic inflammation with massive eosinophil infiltration in the skin, independently of T cells and mast cells, even though basophils account for only approximately 2% of the infiltrates. The basophil depletion shows a therapeutic effect on on-going allergic inflammation. Accumulating evidence suggests that basophils function as initiators rather than effectors of the chronic allergic inflammation. Thus, basophils and their products seem to be promising therapeutic targets for allergic disorders.

Fc receptor gamma-chain, a constitutive component of the IL-3 receptor, is required for IL-3-induced IL-4 production in basophils.

The Fc receptor common gamma-chain (FcRgamma) is a widely expressed adaptor bearing an immunoreceptor tyrosine-based activation motif (ITAM) that transduces activation signals from various immunoreceptors. We show here that basophils lacking FcRgamma developed normally and proliferated efficiently in response to interleukin 3 (IL-3) but were very impaired in IL-3-induced production of IL-4 and in supporting T helper type 2 differentiation. Through its transmembrane portion, FcRgamma associated constitutively with the common beta-chain of the IL-3 receptor and signaled by recruiting the kinase Syk. Retrovirus-mediated complementation demonstrated the essential function of the ITAM of FcRgamma in IL-3 signal transduction. Our results identify a previously unknown mechanism whereby FcRgamma functions to 'route' selective cytokine-triggered signals into the ITAM-mediated IL-4 production pathway.

Newly discovered roles for basophils: a neglected minority gains new respect.

Basophils are the least common type of granulocyte and they account for less than 1% of peripheral blood leukocytes. Because of this minority status and a phenotype that is similar to mast cells, basophils have often been neglected in immunological studies or considered to have minor, redundant roles in immune responses in vivo. However, recent studies have now defined previously unrecognized roles for basophils in both immune regulation and allergic responses, and have shown that basophils and mast cells have distinct roles in immune responses.

Basophils play a pivotal role in immunoglobulin-G-mediated but not immunoglobulin-E-mediated systemic anaphylaxis.

Anaphylaxis is an acute, severe, and potentially fatal systemic allergic reaction. Immunoglobulin E (IgE), mast cells, and histamine have long been associated with anaphylaxis, but an alternative pathway mediated by IgG has been suggested to be more important in the elicitation of anaphylaxis. Here, we showed that basophils, the least common blood cells, were dispensable for IgE-mediated anaphylaxis but played a critical role in IgG-mediated, passive and active systemic anaphylaxis in mice. In vivo depletion of basophils but not macrophages, neutrophils, or NK cells ameliorated IgG-mediated passive anaphylaxis and rescued mice from death in active anaphylaxis. Upon capture of IgG-allergen complexes, basophils released platelet-activating factor (PAF), leading to increased vascular permeability. These results highlight a pivotal role for basophils in vivo and contrast two major, distinct pathways leading to allergen-induced systemic anaphylaxis: one mediated by basophils, IgG, and PAF and the other "classical" pathway mediated by mast cells, IgE, and histamine.

Mast cells and basophils are selectively activated in vitro and in vivo through CD200R3 in an IgE-independent manner.

Mast cells and basophils have been implicated in the host defense system against pathogens and in the development of allergic disorders. Although IgE-dependent responses via FcepsilonRI on these cells have been extensively studied, little is known about cell surface molecules that are selectively expressed by these cells and engaged in their activation via an IgE-independent mechanism. We have recently established two mAbs that reacted specifically with murine mast cells and basophils, and one of them selectively depleted basophils when administered in vivo. Biochemical and flow cytometric analyses revealed that both mAbs specifically recognized a CD200R-like protein, CD200R3, but not other CD200R family members. CD200R3 existed as a disulfide-linked dimer, unlike other CD200Rs, and was expressed on mast cells and basophils primarily in association with an ITAM-bearing adaptor DAP12. Cross-linking of CD200R3 with the mAbs induced degranulation in mast cells and production of the cytokine IL-4 in basophils in vitro. Administration of the nondepleting mAb in vivo elicited systemic and local anaphylaxis in a CD200R3-dependent manner. These results suggest that CD200R3 functions as an activating receptor on mast cells and basophils to regulate IgE-independent immune responses in cooperation with an inhibitory receptor CD200R, similar to the paired receptors expressed on NK cells.

Basophils are essential initiators of a novel type of chronic allergic inflammation.

Basophils represent less than 1% of peripheral blood leukocytes and have often been considered as minor and possibly redundant circulating mast cells. The recent finding that basophils readily generate large quantities of T helper 2 (Th2) cytokines such as IL-4 provided new insights into the possible role of basophils in allergic disorders and immunity to pathogens. However, in-depth studies on basophils, particularly their functions in vivo, have been hampered by the lack of appropriate animal models, such as mutant animals deficient only in basophils. Here, we established a mAb that reacted with mouse basophils and depleted them when administered in vivo. The mAb treatment of mice did not show any significant effect on classical allergic reactions such as passive cutaneous anaphylaxis and contact hypersensitivity. In contrast, it completely abolished the development of IgE-mediated chronic allergic dermatitis that is characterized by massive eosinophil infiltration, even though basophils accounted for only approximately 2% of the infiltrates. The treatment during the progression of the dermatitis resulted in drastic reduction in numbers of infiltrating eosinophils and neutrophils, concomitantly with elimination of basophils from the skin lesions. Thus, basophils play a pivotal role in the development of IgE-mediated chronic allergic inflammation, as an initiator rather than as an effector.