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Allergen sources such as mites, insects, fungi, and pollen contain proteases. Airway exposure to proteases induces allergic airway inflammation and IgE/IgG1 responses via IL-33-dependent mechanisms in mice. We examined the epicutaneous sensitization of mice to a model protease allergen, papain; the effects of tape stripping, which induces epidermal barrier dysfunction; and the atopic march upon a subsequent airway challenge. Papain painting on ear skin and tape stripping cooperatively promoted dermatitis, the skin gene expression of proinflammatory cytokines and growth factors, up-regulation of serum total IgE, and papain-specific IgE/IgG1 induction. Epicutaneous sensitization induced T helper (Th) 2 cells and Th17 differentiation in draining lymph nodes. Ovalbumin and protease inhibitor-treated papain induced no or weak responses, whereas the co-administration of ovalbumin and papain promoted ovalbumin-specific IgE/IgG1 induction. Wild-type and IL-33-deficient mice showed similar responses in the epicutaneous sensitization phase. The subsequent airway papain challenge induced airway eosinophilia and maintained high papain-specific IgE levels in an IL-33-dependent manner. These results suggest that allergen source-derived protease activity and mechanical barrier damage such as that caused by scratching cooperatively promote epicutaneous sensitization and skin inflammation and that IL-33 is dispensable for epicutaneous sensitization but is crucial in the atopic march upon a subsequent airway low-dose encounter with protease allergens.

Protease activity of papain, a plant-derived occupational allergen homologous to mite major allergens, is essential to IgE/IgG1 production and lung eosinophilia induced by intranasal papain administration in mice, and IL-33 contributes to these responses. In this work, we investigate skin and Ab responses induced by s.c. papain administration into ear lobes and responses induced by subsequent airway challenge with papain. Subcutaneous papain injection induced swelling associated with increased epidermal thickness, dermal inflammation, serum IgE/IgG1 responses, and Th2 cytokine production in draining lymph node cells restimulated in vitro. These responses were markedly less upon s.c. administration of protease inhibitor-treated papain. Results obtained by using mast cell–deficient mice and reconstitution of tissue mast cells suggested the contribution of mast cells to papain-specific IgE/IgG1 responses and eosinophil infiltration. The responses were equivalent between wild-type and IL-33−/− mice. After the subsequent airway challenge, the s.c. presensitized wild-type mice showed more severe lung eosinophilia than those without the presensitization. The presensitized IL-33−/− mice showed modest lung eosinophilia, which was absent without the presensitization, but its severity and IgE boost by the airway challenge were markedly less than the presensitized wild-type mice, in which protease activity of inhaled papain contributed to the responses. The results suggest that mechanisms for the protease-dependent sensitization differ between skin and airway and that cooperation of mast cell–dependent, IL-33–independent initial sensitization via skin and protease-induced, IL-33–mediated mechanism in re-exposure via airway to protease allergens maximizes the magnitude of the transition from skin inflammation to asthma in natural history of progression of allergic diseases.

Background: Epicutaneous sensitization to allergens is important in the pathogenesis of not only skin inflammation such as atopic dermatitis but also "atopic march" in allergic diseases such as asthma and food allergies. We here examined antibody production and skin barrier dysfunction in mice epicutaneously administered papain, a plant-derived occupational allergen belonging to the same family of cysteine proteases as mite major group 1 allergens. Methods: Papain and Staphylococcus aureus V8 protease were patched on the backs of hairless mice. Tran- sepidermal water loss was measured to evaluate the skin barrier dysfunction caused by the proteases. Papain or that treated with an irreversible inhibitor specific to cysteine proteases, E64, was painted onto the ear lobes of mice of an inbred strain C57BL/6. Serum total IgE levels and papain-specific IgE and IgG antibodies were measured by ELISA. Results: Papain and V8 protease patched on the backs of hairless mice caused skin barrier dysfunction and increased serum total IgE levels, and papain induced the production of papain-specific IgG1, IgG2a, and IgG2b. Papain painted onto the ear lobes of C57BL/6 mice induced papain-specific IgE, IgG1, IgG2c, and IgG2b, whereas papain treated with E64 did not. IgG1 was the most significantly induced papain-specific IgG subclass among those measured. Conclusions: We demonstrated that the epicutaneous administration of protease not only disrupted skin barrier function, but also induced IgE and IgG responses in a manner dependent on its protease activity. These results suggest that protease activity contained in environmental sources contributes to sensitization through an epicutaneous route.