平喘宁调节TRPM8通道相关蛋白干预寒性哮喘大鼠气道炎症的机制研究

目的 探究冷刺激对哮喘大鼠气道炎症的影响以及平喘宁对寒性哮喘大鼠气道炎症的治疗作用及机制。方法 将72只大鼠随机分成正常组，卵清蛋白（ovalbumin，OVA）组，模型组（OVA+冷刺激），平喘宁低、中、高剂量组，桂龙咳喘宁组以及地塞米松组，每组9只。除正常组外，其余各组大鼠采用OVA或OVA联合冷刺激建立哮喘及寒性哮喘大鼠模型，模型复制完成后，分别给予生理盐水及对应药液灌胃21 d。观察记录各组大鼠的一般行为学情况；采用HE染色检测大鼠肺组织病理变化；ELISA法检测支气管肺泡灌洗液（bronchoalveolar lavage fluid，BALF）中白细胞介素（interleukin，IL）-1β、IL-6和胸腺基质淋巴细胞生成素（thymic stromal lymphopoietin，TSLP）水平；Western blot法检测瞬时受体电位美拉塔素8（transient receptor potential melastatin subtype 8，TRPM8）、粒细胞—巨噬细胞集落刺激因子（granule macrophage-colony stimulating factor，GM-CSF）、TSLP蛋白表达水平。结果 与正常组比较，OVA组、模型组大鼠均出现呼吸急促、腹部收缩明显等行为学表现；病理结果显示支气管管壁增厚、肺组织及支气管周围可见大量炎症细胞浸润；BALF中IL-1β、IL-6、TSLP水平显著升高（P＜0.05）；肺组织TRPM8、GM-CSF、TSLP蛋白表达水平显著升高（P＜0.05）。与模型组比较，平喘宁可改善大鼠的行为学表现，减轻支气管管壁增厚、肺组织及支气管周围炎症细胞浸润，显著降低大鼠BALF中IL-1β、IL-6、TSLP水平（P＜0.05），显著降低肺组织TRPM8、GM-CSF、TSLP蛋白表达水平（P＜0.05）。结论 冷刺激会加重哮喘大鼠气道炎症反应，平喘宁可能通过调控寒性哮喘大鼠TRPM8及相关蛋白表达水平，减轻寒性哮喘大鼠气道炎症反应，进而达到治疗哮喘的作用。

Mechanism of Pingchuanning in Intervention of Airway Inflammation in Rats with Cold-type Asthma by Regulating the Transient Receptor Potential Melastatin Subtype 8 Channel-related Proteins

Objective To investigate the effect of cold stimulation on airway inflammation in asthmatic rats, as well as the therapeutic effect of Pingchuanning on airway inflammation in rats with cold-type asthma and its mechanism. Methods A total of 72 rats were randomly divided into normal group, ovalbumin (OVA) group, model group (OVA+cold stimulation), Guilong Kechuanning group, dexamethasone group, and low-, middle-, and high-dose Pingchuanning groups, with 9 rats in each group. All rats except those in the normal group were given OVA or OVA combined with cold stimulation to establish a model of asthma or cold-type asthma, and after modeling, the rats were given normal saline or the corresponding drug solution by gavage for 21 days. General behavioral manifestations were recorded for each group; HE staining was used to observe the pathological changes of lung tissue; ELISA was used to measure the levels of the inflammatory factors interleukin（IL）-1β, IL-6 and thymic stromal lymphopoietin (TSLP) in bronchoalveolar lavage fluid (BALF); Western blot was used to measure the expression levels of transient receptor potential melastatin subtype 8 (TRPM8), granule macrophage-colony stimulating factor (GM-CSF) and TSLP. Results Compared with the normal group, the OVA group and the model group had the behavioral manifestations such as shortness of breath and obvious abdominal contraction; pathological results showed thickening of the bronchial wall and massive inflammatory cell infiltration in lung tissue and the peribronchial area; there were significant increases in the levels of IL-1β, IL-6, and TSLP in BALF (P<0.05) and the protein expression levels of TRPM8, GM-CSF, and TSLP in lung tissue (P<0.05). Compared with the model group, Pingchuanning improved the behavioral manifestation of rats, alleviated bronchial wall thickening and inflammatory cell infiltration in lung tissue and the peribronchial area, and significantly reduced the levels of IL-1β, IL-6, and TSLP in BALF (P<0.05) and the protein expression levels of TRPM8, GM-CSF, and TSLP in lung tissue (P<0.05). Conclusion Cold stimulation aggravates airway inflammatory response in asthmatic rats, and Pingchuanning may exert a therapeutic effect on asthma by regulating the expression levels of TRPM8 and related proteins and alleviating airway inflammatory response in rats with cold-type asthma.