Allergic asthma (AS) is a chronic respiratory disorder, approximately 357.4 million people worldwide are afflicted with AS (Song et al., 2022). Airway inflammation, airway hyperresponsiveness, and airway remodeling are the characteristic manifestations of AS (Miller et al., 2021). Studies have indicated that type Ⅱ inflammation mediated by type Ⅱ cytokines is a key pathological mechanism of AS (Howell et al., 2023). Currently, the treatment of AS mainly relies on inhaled glucocorticoid drugs. However, these drugs still have recurrent attacks after discontinuation; long-term use not only has considerable side effects but also induces drug resistance (Henderson et al., 2020, Loke et al., 2002). Hence, it is extremely urgent to explore more safe and effective drugs for the treatment of AS.

It has been discovered that some monomers of traditional Chinese herbs can alleviate AS by influencing the secretion of Th2 cytokines and other mediators related to AS (Luo et al., 2019, Qiu et al., 2023). Formononetin (FM) is a kind of flavonoid compound that is widely present in leguminous plants (D. Jiang et al., 2019). Modern pharmacological studies have revealed that FM possesses extensive pharmacological actions, including anti-inflammatory, antioxidative, anti-tumor, lipid-lowering, spasm-relieving, anti-arrhythmic, and estrogen-like effects (Ma and Wang, 2022). In addition, recent studies have demonstrated that FM also modulates immune responses in various diseases. FM was reported to attenuate neuroinflammation in BV2 microglial cells by inhibiting the TLR4/NF-κB signaling pathway (Chen et al., 2024), ameliorates polycystic ovary syndrome by suppressing NLRP3 inflammasome activation (Liu et al., 2025), and alleviates ulcerative colitis through regulating M1/M2 macrophage polarization (Xiao et al., 2024). These findings suggest that FM can regulate innate immune responses in diverse inflammatory contexts. In the context of allergic asthma, FM has also shown promising effects. Sun et al. pointed out that FM might be an effective component of plants with protective effects against AS (Sun et al., 2022). Zhang et al. confirmed that FM promotes the repair of the airway epithelial barrier by inhibiting the ESR1/NLRP3/Caspase-1 pathway, thereby ameliorating airway inflammation in AS (Zhang et al., 2023). These results indicate that FM may exert protective effects against AS. However, the precise immunological mechanisms remain to be clarified.

In recent years, ILC2s have been confirmed to play a significant role in the pathogenesis of AS (Emami Fard et al., 2023). Upon stimulation by allergens, airway epithelial cells release cytokines such as IL-33, IL-25, and thymic stromal lymphopoietin (TSLP), which are known as "alarmins". Subsequently, these cytokines stimulate the activation of ILC2s, leading to the release of inflammatory mediators like IL-4, IL-5, IL-9, IL-13, and amphiregulin (AREG). This process aggravates airway inflammation and facilitates the initiation of adaptive immune responses (Kato, 2019). Despite the growing recognition of ILC2s as key mediators of type II inflammation, it remains unclear whether FM exerts its therapeutic effect by modulating ILC2 activation or migration dynamics. ILC2s can be classified into natural ILC2s (nILC2s) and inflammatory ILC2s (iILC2s). Under normal circumstances, ILC2s function as a defense and repair mechanism in the form of nILC2s; when inflammation emerges, ILC2s act as iILC2s to mediate inflammatory responses (Qin et al., 2024). Notably, iILC2s do not originate locally within the lungs but are primarily derived from the small intestine. Studies indicate that iILC2s are involved in mediating inflammation in lung tissues, yet the lung lacks resident precursors for these cells. Instead, under inflammatory stimulation such as IL-25, quiescent nILC2s residing in the small intestine are activated and differentiate into iILC2s. These activated cells exit the intestinal tissue via the lymphatic system, enter the peripheral circulation, and are subsequently recruited to inflamed lung tissue. This process, referred to as the “lung–gut axis migration,” is thought to be driven by key chemotactic signals, including the sphingosine kinase 1 (Sphk1)/sphingosine-1-phosphate receptor 1 (S1PR1) signaling axis, which facilitates the egress of ILC2s from the gut. Upon arrival in the lungs, iILC2s contribute to mucus hypersecretion and sustained airway inflammation, hallmarks of AS (Germain and Huang, 2019; M. Jiang et al., 2023). Given the central role of ILC2s in type 2 airway inflammation, recent studies have explored therapeutic strategies that directly target ILC2s or their upstream activators. For example, JAK3 inhibitor reduces ILC2 survival, proliferation, and cytokine production in vitro and ameliorates ILC2-driven asthma (Kim et al., 2023). Randomized controlled trials have recently shown the efficacy of biologics for targeting epithelial-derived cytokines, such as TSLP and IL-33, in patients with severe asthma (Corren et al., 2023, Wechsler et al., 2021). These approaches underscore the therapeutic value of disrupting ILC2 activation or function. However, most existing strategies focus on blocking cytokine signaling or ILC2 activation locally, while little is known about how to interfere with the long-range migration of ILC2s—particularly those derived from the gut. However, whether pharmacological intervention can suppress this lung–gut axis migration remains unknown. Thus, we put forward such a hypothesis: FM may ameliorate allergic asthma by inhibiting the activation and lung recruitment of intestinal iILC2s.

Therefore, this study aimed to investigate whether formononetin (FM) alleviates allergic asthma by inhibiting the activation and lung–gut axis migration of inflammatory ILC2s. To test this, we established an OVA-induced mouse model of allergic asthma and evaluated the effects of FM on airway inflammation, mucus hypersecretion, and ILC2 responses in vivo and in vitro. The findings may provide mechanistic insights and experimental support for the clinical application of FM in asthma therapy.