Introduction
Asthma is a chronic airway disease characterized by bronchial inflammation, airway hyperreactivity, and reversible airflow limitation; it is often caused by inhalation of allergens[1].Pollens are important outdoor allergens associated with allergic asthma and seasonal rhinitis[2].Humulus scandens(H.scandens , also known as Humulus japonicas or Lücao in Chinese) is a widespread weed in East Asia[3].In Korea, Humulus pollen constitutes approximately 18% of total pollen during the pollination period[4]. In China, Jia et al. first discovered that Humulus pollen is an important allergen in summer and autumn, second only to Artemisiapollen[5].Among individuals with fall hay fever, 59% were positive for H.scandens -specific serum IgE[6]. The symptoms of asthma triggered by H.scandenspollen are reportedly more serious than those induced byArtemisia pollen, although the mechanism underlying this effect has not been elucidated[7].
Subcutaneous immunotherapy(SCIT) with crudeH. scandens pollen extract (HSE) has been used in China for many years; this type of therapy can effectively reduce the symptom scores and medication scores of patients with rhinitis and asthma[8].However, the mechanisms underlying immunotherapy with HSE have remained unclear, thereby limiting the applications of this treatment method. Thus far, most patients with allergic asthma induced byH.scandenspollen are treated with inhaled and oral corticosteroids, despite the considerable side effects associated with long-term use of corticosteroids[9]. Therefore, it is necessary to establish an animal model of asthma with HSE and to study the mechanisms underlying successful SCIT.
Murine models of allergic asthma induced by Humulus pollen were previously established by Ya-li et al.[10] and Kong et al.[11]through intraperitoneal (IP) injection and nasal drip stimulation. We originally planned to study the mechanisms underlying immunotherapy with HSE by using this model. However, A fair number of mice died after two nasal drops. We repeated the experiment more than three times with different anesthesia methods that used for nasal drip stimulation, including IP injection with 1% pentobarbital, inhalation of ether, and inhalation of isoflurane. Our results were similar, in that most mice died. The lung histopathology of surviving mice showed acute lung injury (see Supplementary Material for representative experimental results). We speculated that these results may be attributed to direct stimulation of lung tissues with high concentrations of HSE during nasal drip challenge. Whether this is related to the fact that H.scandens is a kind of Chinese herbal medicine with heat-clearing and detoxifying effects[12] is unclear. Therefore, we attempted to use atomized inhalation for challenge, which is considered to be milder and has a lower fatality rate[13]. In addition, IP injection is the most commonly used sensitization route, but some studies have suggested that sensitization by subcutaneous (SC) injection is more effective for eliciting high IgE levels and Th2 cytokines and for inducing allergic airway inflammation[14]. For instance, Conejero et al.[15] established a murine model of olive pollen allergic asthma through SC sensitization.
In the present study, we compared the IP and SC sensitization routes, with respective reference to previously established sensitization methods and allergen dosing[10, 15], to develop an optimized model of allergic asthma induced by HSE that can better simulate the immunological and pathological features of human allergic asthma[16]. Using our asthma model, we then developed a model of rapid SCIT to simulate the process of clinical SCIT and explore the mechanisms ofHumulus immunotherapy.