Hiroo Yokozeki

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.

Prostaglandin D2 Plays an Essential Role in Chronic Allergic Inflammation of the Skin via CRTH2 Receptor

PGD2 plays roles in allergic inflammation via specific receptors, the PGD receptor designated DP and CRTH2 (chemoattractant receptor homologous molecule expressed on Th2 cells). We generated mutant mice carrying a targeted disruption of the CRTH2 gene to investigate the functional roles of CRTH2 in cutaneous inflammatory responses. CRTH2-deficent mice were fertile and grew normally. Ear-swelling responses induced by hapten-specific IgE were less pronounced in mutant mice, giving 35–55% of the responses of normal mice. Similar results were seen in mice treated with a hemopoietic PGD synthase inhibitor, HQL-79, or a CRTH2 antagonist, ramatroban. The reduction in cutaneous responses was associated with decreased infiltration of lymphocytes, eosinophils, and basophils and decreased production of macrophage-derived chemokine and RANTES at inflammatory sites. In models of chronic contact hypersensitivity induced by repeated hapten application, CRTH2 deficiency resulted in a reduction by approximately half of skin responses and low levels (63% of control) of serum IgE production, although in vivo migration of Langerhans cells and dendritic cells to regional lymph nodes was not impaired in CRTH2-deficient mice. In contrast, delayed-type hypersensitivity to SRBC and irritation dermatitis in mutant mice were the same as in wild-type mice. These findings indicate that the PGD2-CRTH2 system plays a significant role in chronic allergic skin inflammation. CRTH2 may represent a novel therapeutic target for treatment of human allergic disorders, including atopic dermatitis.

Basophil recruitment and activation in inflammatory skin diseases

To cite this article: Ito Y, Satoh T, Takayama K, Miyagishi C, Walls AF, Yokozeki H. Basophil recruitment and activation in inflammatory skin diseases. Allergy 2011; 66: 1107–1113.

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.

Therapeutic effects of interleukin-6 blockade in a murine model of polymyositis that does not require interleukin-17A.

OBJECTIVE To explore new molecular targets in the treatment of polymyositis (PM) by examining a recently established murine model of PM, C protein-induced myositis (CIM), for involvement of an interleukin-6 (IL-6)/IL-17A pathway. METHODS CIM was induced by immunizing wild-type mice as well as IL-6-null and IL-17A-null C57BL/6 mice with recombinant mouse skeletal C protein fragments. Some mice were treated with anti-IL-6 receptor (anti-IL-6R) monoclonal antibodies or control antibodies. Muscle tissue samples were examined histologically and immunohistochemically. RESULTS The syngeneic C protein fragments successfully induced inflammation in the skeletal muscles of wild-type mice. IL-6 was expressed by mononuclear cells, especially in macrophages, infiltrating in the muscles. IL-6-null mice developed myositis with significantly lower incidence and milder severity than wild-type mice. In contrast, IL-17A-null mice were as susceptible to CIM as wild-type mice. Intraperitoneal administration of anti-IL-6R monoclonal antibodies, but not of control monoclonal antibodies, ameliorated CIM both preventively and therapeutically. CONCLUSION Our findings indicate that IL-6 is critically involved in the development of CIM. Although many other autoimmune models require IL-6 for differentiation of pathogenic T cells producing IL-17A, IL-17A was dispensable in CIM. Nevertheless, treatment with anti-IL-6R antibodies was effective. IL-6 blockade is potentially a new approach to the treatment of autoimmune myositis, via processes distinct from interference in the IL-6/IL-17A pathway.

Stable form of galectin-9, a Tim-3 ligand, inhibits contact hypersensitivity and psoriatic reactions: A potent therapeutic tool for Th1- and/or Th17-mediated skin inflammation

Tim-3 is a cell surface molecule preferentially expressed in Th1 and Th17 cells. Galectin-9 is a ligand for Tim-3 and the binding of galectin-9 to Tim-3 induces apoptosis. We recently developed a stable form of galectin-9 (sGal-9) by partial deletion of the linker peptide. In this study, we characterized the therapeutic effects of sGal-9 on inflammatory reactions in contact hypersensitivity and IL-23-induced psoriatic mouse models. In contact hypersensitivity in mice, the ear swelling response was suppressed by sGal-9. In vitro treatment with sGal-9 resulted in cell apoptosis of CD4, CD8, and hepatic NK cells. sGal-9-treated mice had decreased IFN-gamma- and IL-17-producing T cells. Similarly, sGal-9 reduced epidermal thickness and dermal cellular infiltrate levels in IL-23-induced psoriasis-like skin inflammation. This was accompanied by decreased skin lesion levels of IL-17 and IL-22. sGal-9 may be a unique and useful therapeutic tool for the treatment of Th1- and/or Th17-mediated skin inflammation.

Functional CD86 (B7-2/B70) is predominantly expressed on Langerhans cells in atopic dermatitis.

Recently, we reported the functional expression of CD86 on cultured human Langerhans cells derived from normal epidermis. In the present study, we investigated the expression and function of co‐stimulatory molecules in the pathogenesis of atopic dermatitis. In immunohistochemical analysis, CD80 and/or CD86 were detected on dendritic‐shaped cells not only in the epidermis but also in the dermis in the inflammatory lesions of atopic dermatitis (n = 12). CD80 was expressed in only five cases (42%), while CD86 was expressed in all cases (100%). These molecules were not detected in normal control subjects (n = 8). In non‐lesional skin of atopic dermatitis (n = 4). CD86 but not CD80 was detected in one case. CD86 was preferentially induced on dendritic‐shaped cells in positive patch test sites to Dermatophagoides pteronyssinus or house dust allergen in atopic dermatitis (n = 4). The CD80‐ or CD86‐positive cells were confirmed as Langerhans cells by double immunostaining using anti‐CD1a monoclonal antibody. Neither CD86 over that CD80 was detected n keratinocytes. Similar results of the stronger expression of CD86 over that of CD80 were obtained from psoriasis vulgaris (n = 11) and from contact dermatitis (n=7), although CD86 was expressed only in 57% of the contact dermatitis cases. The percentage of Langerhans cells positive for CD86 was higher than for CD80, i.e. 48% compared with 9%, respectively, in the epidermis of lesional skin of atopic dermatitis (n=8). The expression rate of these molecules on Langerhans cells increased in the dermis. To investigate the function of co‐stimulatory molecules on Langerhans cells in atopic dermatitis, we conducted an inhibition test with antibodies. Anti‐CD86 monoclonal antibody almost completely nhibited T‐cell proliferation stimulated with crude extract of D. pteronyssinus in the presence of epidermal cells as antigen‐presenting cells, whereas anti‐CD80 monoclonal antibody produced less of an inhibitory effect. These data indicate that CD86 expressed on Langerhans cells may play an important part in the pathogenesis of atopic dermatitis.for Investigative Dermatology. Washington, DC (1–5 May 1996).

CRTH2-dependent, STAT6-independent induction of cedar pollen dermatitis

Background Airborne contact dermatitis to cedar pollen is a recently identified disease that generally affects individuals with cedar pollinosis of the nasal and/or ocular symptoms, as well as some patients with atopic dermatitis.

Dendritic Cells Express Hematopoietic Prostaglandin D Synthase and Function as a Source of Prostaglandin D2 in the Skin

Prostaglandin D2 (PGD2), an arachidonic acid metabolite, has been implicated in allergic responses. A major source of PGD2 in the skin is mast cells that express hematopoietic PGD synthase (H-PGDS). In this study, we show the expression of H-PGDS in human dendritic cells (DCs) and the regulatory mechanisms by which DCs produce PGD2. We detected H-PGDS in epidermal Langerhans cells, dermal DCs, plasmacytoid DCs, and myeloid DCs. Monocyte-derived DCs rapidly secreted PGD2 when stimulated with the calcium ionophore A23187. More importantly, pretreatment of monocyte-derived DCs with PMA (phorbol 12-myrisate 13-acetate) synergistically enhanced the rapid PGD2 secretion induced by A23187, whereas PMA alone did not induce PGD2 secretion. Lipopolysaccharide (LPS) reduced H-PGDS expression, but interferon-gamma followed by LPS induced significant PGD2 production in a delayed time course at 6 hours. This effect was associated with inhibition of LPS-induced H-PGDS reduction. Interestingly, an irritant compound, SDS, also induced a rapid PGD2 release. PGD2 synergistically enhanced CCL22/macrophage-derived chemokine synthesis in interferon-gamma-treated human keratinocytes. In addition, bone marrow-derived DCs from wild-type mice stimulated lymph node cells to produce higher amounts of interleukin-17 than did DCs from mice lacking the H-PGDS gene. Thus, DCs could be an important source of skin PGD2 and may mediate or regulate skin inflammation by releasing PGD2 in response to various stimuli, contributing to the innate and/or acquired immune responses.

Hepatocyte growth factor both prevents and ameliorates the symptoms of dermal sclerosis in a mouse model of scleroderma

Systemic sclerosis (SSc) is a connective tissue disorder with an unknown etiology. There are currently no effective therapies for SSc. (In this study, working with a bleomycin(BLM)-induced scleroderma model mice, we performed two transfections of human hepatocyte growth factor (HGF) cDNA into the skeletal muscle and showed that this treatment not only helped to prevent the dermal sclerosis simultaneously injected BLM but also improved the symptoms of dermal sclerosis induced by BLM 4 weeks previously.) RT-PCR, ELISA and an immunohistochemical analysis revealed that both mRNA and protein of human HGF as well as murine HGF were enhanced in the skin, lung, muscle and the serum after two transfections of human HGF cDNA. These analyses also revealed that this treatment significantly reduced both the expression of the TGF-β1 mRNA and the production of TGF-β1 on macrophage-like cells that infiltrated the dermis and the fibroblastic cells in BLM-induced scleroderma. Furthermore, HGF-gene transfection both prevented and ameliorated the symptoms of not only dermal sclerosis but also of lung fibrosis induced by a subcutaneous BLM injection. These results indicated that gene therapy by the transfection of the human HGF cDNA may thus be a useful therapy for SSc and lung fibrosis involved with SSc.