Human cystatin SN is an endogenous protease inhibitor that prevents allergic rhinitis

Background: Protease allergens disrupt epithelial barriers to exert their allergenicity. Cystatin SN (encoded by CST1) is an endogenous cysteine protease inhibitor upregulated in nasal epithelia in patients with allergic rhinitis (AR). Objective: We sought to investigate the protective effect of human cystatin SN on AR symptoms using pollen-induced AR mouse models. Methods: We performed an in vitro protease activity assay to evaluate the effect of recombinant human cystatin SN (rhCystatin SN) on Japanese cedar (JC) or ragweed proteases. A human nasal epithelial cell line, RPMI 2650, was used to examine tight junction (TJ) disruption in vitro. Mice were sensitized and nasally challenged with JC or ragweed pollens with or without rhCystatin SN to examine the effect of rhCystatin SN on AR symptoms and the epithelial barrier in vivo. Because mice lack CST1, we generated transgenic (Tg) mice expressing human CST1 under control of its genomic control region (hCST1-Tg mice) to examine the role of cystatin SN in physiologically expressed conditions. Results: rhCystatin SN inhibited JC but not ragweed protease activities and prevented JC-induced but not ragweed-induced TJ disruption in vitro. Exogenous administration of rhCystatin SN ameliorated JC-induced but not ragweed-induced sneezing and nasal TJ disruption in vivo. Furthermore, hCST1-Tg mice showed decreased JC-induced but not ragweed-induced sneezing symptoms and nasal TJ disruption compared with wild-type mice. Conclusion: Human cystatin SN suppresses AR symptoms through inhibiting allergen protease activities and protecting the nasal TJ barrier in an allergen-specific manner. We propose that upregulation of nasal endogenous protease inhibitors, including cystatin SN, is a novel therapeutic strategy for protease allergen–induced AR

Activation of group 2 innate lymphoid cells exacerbates and confers corticosteroid resistance to mouse nasal type 2 inflammation

Both Th2 cells and group 2 innate lymphoid cells (ILC2s) contribute to allergic diseases. However, their exact role and relationship in nasal allergic disorders are unclear. In this study, we investigated the cooperation of Th2 cells and ILC2s in a mouse model of nasal allergic disorder. To differentially activate Th2 cells and/or ILC2s in nasal mucosa, mice were intra-nasally administered ovalbumin (OVA) antigen, papain, an ILC2-activator, or both for 2 weeks. Epithelial thickness and number of eosinophils in the nasal mucosa were evaluated at 24 h after the final challenge. Intra-nasal administration of OVA and papain preferentially activated Th2 cells and ILC2s, respectively, in the nose. Both OVA and papain increased the nasal epithelial thickness and number of eosinophils, and their coadministration significantly enhanced the symptoms. Although T-/B-cell-deficient mice showed severely decreased nasal symptoms induced by OVA or OVA-plus-papain, the mice still showed slight papain-induced nasal symptoms. In ILC2-deficient mice, OVA-plus-papain-induced nasal symptoms were suppressed to the same level as OVA-alone. Similarly, IL-33- and ST2-deficient mice showed decreased OVA-plus-papain-induced nasal symptoms. IL-5 induced eosinophilia only, but IL-13 contributed to both nasal epithelial thickening and eosinophilia induced by OVA-plus-papain. Dexamethasone ameliorated OVA-alone-induced nasal epithelial thickening. However, OVA-plus-papain-induced nasal epithelial thickening was only partially controlled by dexamethasone. These results demonstrate that IL-33/ST2-pathway-mediated ILC2 activation exacerbated Th2-cell-induced nasal inflammation by producing IL-13. Although Th2-cell-alone-induced nasal inflammation was controlled by corticosteroid treatment, the activation of ILC2s conferred treatment resistance. Therefore, ILC2s and their activators could be therapeutic targets for treatment-refractory nasal allergic disorders

The expression and functional analysis of CST1 in intractable nasal polyps

好酸球性副鼻腔炎（ECRS）は，鼻腔内に多発性鼻茸を有し，鼻茸・末梢血中に好酸球増加を伴う難治性副鼻腔炎である。次世代シーケンサーを用いた，鼻茸のRNA-sequencingにおけるtranscriptome解析では，ECRS患者の鼻茸でCST1の発現が高い傾向にあった。我々はCST1がECRSの病態に関与していると考え，ECRSの鼻茸内でのCST1の発現や働きについて詳細な検討を行った。CRSwNP患者の鼻茸内におけるCST1の発現に関して，real-time PCRを用いたmRNAの発現・免疫組織化学を用いた解析では，non-ECRS患者群に比べて，ECRS患者群でCST1が有意に高発現していた。特にCST1はsevere ECRSの鼻茸上皮で強い発現を示していた。つまり，CST1の発現は，ECRSの難治性や再発性と関連する。ECRS由来の鼻茸上皮細胞を精製し，IL-4+dsRNA+CST1で刺激すると，IL-4+dsRNAで刺激した時に比べて，TSLPの発現が有意に上昇した。鼻茸上皮細胞へのTSLPあるいはIL-33の刺激は，CST1の発現を誘導した。また，ECRS由来の鼻茸線維芽細胞に対するCST1の刺激は，CCL11とperiostinの発現を誘導した。CST1は鼻茸内において，ECRSの鼻茸形成・増悪に関わる様々な因子と相互作用することにより，Th2/好酸球性炎症として作用し，鼻茸の重症化，難治性，再発に関わる。ECRSの鼻茸に対して，CST1をtargetとした治療戦略が有用となる可能性がある。本稿では，CST1のECRSにおける役割を中心に解説する