Introduction Serine proteases play a critical role in the augmented release and cleavage of IL-33, leading to the expansion of group 2 innate lymphoid cells (ILC2s) and T-helper 2 (Th2) airway inflammation. However, the protective regulation of protease-dependent interleukin-33 (IL-33) activation remains poorly understood. Therefore, we investigated the role of secretory leukocyte protease inhibitor (SLPI), as a serine protease inhibitor, in this protective regulation and aimed to clarify its contribution to type 2 immunity. Methods We evaluated the role of SLPI in the  extract-induced expansion of ILC2s and Th2-type airway inflammation via IL-33, using three models: SLPI-deficient mice, an in vivo SLPI knockdown model with shRNA, and an in vitro model utilizing primary human bronchial epithelial cells (HBECs) exposed to  extract under various conditions, including plasmid transfection. Results We showed that two mouse models of downregulation ofgene expression augmented  extract-induced release of IL-33 and the expansion of ILC2s, together with Th2 airway inflammation. Furthermore, two treatment models using  KO mice, administration of a serine protease inhibitor, bovine pancreatic trypsin inhibitor, or anti-IL-33 antibody, attenuated Th2 airway inflammation. In two in vitro experiments, SLPI, as a serine protease inhibitor, prevented both the release of IL-33 from HBECs and the cleavage of full-length IL-33 to shorter mature forms by neutrophil elastase. Discussion These findings suggest that SLPI functions as a key serine protease inhibitor in regulating IL-33-mediated type 2 immune responses. Low SLPI levels may contribute to the pathogenesis of asthma by promoting Th2 inflammation, highlighting SLPI as a potential therapeutic target in patients with low SLPI expression.