基于PI3K/Akt/NF-κB信号通路探讨运脾泻肺化痰方调控幼龄哮喘大鼠气道粘液高分泌的机制研究

Asthma is a global disease, children are high incidence of asthma, and asthma seriously affects the growth and quality of life of children. Airway mucus hypersecretion is an important pathological feature of asthma, were independent risk factors for the prognosis of asthma and its development. At present, the lack of effective treatment of Western medicine drugs mucus hypersecretion. The spleen and lung phlegm is the experience of diarrhea in our hospital professor Huang Jianye's clinical application in treatment of children asthma sputum for more than 50 years, remarkable effect. The previous experimental study also confirmed that this prescription could effectively relieve the lung inflammation in asthmatic mice. The project selection and airway mucus hypersecretion is closely related to the PI3K/Akt/NF- B signaling pathway as a starting point, in the young rat asthma model as the research object, using a variety of techniques to explore the molecular mechanism of Yunpi xiefei phlegm inhibiting asthmatic airway mucus hypersecretion through the regulation of PI3K/Akt/NF- B signaling pathway, to provide ideas for the key scientific problem of the traditional Chinese medicine the regulation of airway mucus hypersecretion, and provide a theoretical basis for the clinical application experience of old doctor of traditional Chinese medicine. At the same time, in-depth exploration of relationship of airway hypersecretion in PI3K/Akt signaling pathway and B signaling pathway downstream of NF-, has important scientific significance and practical application value.

哮喘是全球性疾病，儿童是哮喘高发人群，哮喘严重影响儿童的生长发育及生活质量。气道黏液高分泌是哮喘的重要病理特征，是影响哮喘病情发展及其预后的独立危险因素。目前西医缺乏有效治疗粘液高分泌的药物。运脾泻肺化痰方是我院名老中医黄建业教授的经验方，临床运用治疗儿童哮喘痰多50余年，效果显著。前期的实验研究亦证实该方可有效缓解哮喘小鼠肺部炎症。本项目选择与哮喘气道粘液高分泌密切相关的PI3K/Akt/NF-κB信号通路为切入点，以幼龄哮喘模型大鼠为研究对象，采用多种技术手段探索运脾泻肺化痰方通过调控PI3K/Akt/NF-κB信号通路抑制哮喘气道粘液高分泌的分子机制，为中药调控哮喘气道粘液高分泌的关键科学问题提供思路，为临床推广运用名老中医的经验方提供理论依据。本课题同时深入探索哮喘气道高分泌中PI3K/Akt信号通路与下游NF-κB信号通路的相互联系，具有重要科学意义和实际应用价值。

哮喘是全球性疾病，儿童是哮喘高发人群，哮喘严重影响儿童的生长发育及生活质量。气道黏液高分泌是哮喘的重要病理特征，是影响哮喘病情发展及其预后的独立危险因素。运脾泻肺化痰方是我院名老中医黄建业教授的经验方，临床运用治疗儿童哮喘痰多50余年，效果显著。前期的实验研究亦证实该方可有效缓解哮喘小鼠肺部炎症。本项目选择与哮喘气道粘液高分泌密切相关的PI3K/Akt/NF-κB信号通路为切入点，以幼龄哮喘模型大鼠为研究对象，采用多种技术手段探索运脾泻肺化痰方通过调控PI3K/Akt /NF-κB信号通路抑制哮喘气道粘液高分泌的分子机制，为中药调控哮喘气道粘液高分泌的关键科学问题提供思路，为临床推广运用名老中医的经验方提供理论依据。本课题同时深入探索哮喘气道高分泌中PI3K/Akt信号通路与下游NF-κB信号通路的相互联系，具有重要科学意义和实际应用价值。其主要内容包括：在体实验：（1）成功建立幼龄哮喘动物模型。（2）给予动物模型药物干预，检测PI3K/Akt/NF-κB 信号通路蛋白及其 mRNA 的含量，研究运脾泻肺化痰方抑制哮喘气道粘液高分泌的分子机制及该方对上皮细胞细信号转导通 PI3K/Akt/NF-κB 的调控作用。结果及数据包括：细胞计数及分类（BALF）嗜酸性粒细胞、肺脏病理 （肺组织）、酶联免疫检测血清 IL-6 、IL-8 、TNF-α、IL-4、IL-13 的含量，Western-blot 法及RT-PCR 技术检测肺组织 IκBα、IκBβ、P-50、P-65、IKK-ɑ 、IKK-β、PI3K、AKT、MUC5AC 蛋白表达及mRNA 的表达。离体实验：（1）制备不同组大鼠含药血清，培养大鼠支气管上皮细胞。（2）以大鼠含药血清作用于大鼠支气管上皮细胞，检测各项指标，从细胞水平研究运脾泻肺化痰方抑制哮喘气道粘液高分泌的分子机制及 该 方对上皮细胞细信号转导通PI3K/Akt/NF-κB 的调控作用。以明确运脾泻肺化痰方究竟是怎样通过调节调控PI3K/Akt/NF-κB信号转导通路来抑制气道粘液高分泌的这一分子生物学机制。其离体实验关键数据包括：Western-blot 法和RT-PCR 技术检测上皮细胞 IκBα、IκBβ、P-50、P-65、IKK-ɑ 、IKK-β、PI3K、AKT、MUC5AC蛋白及基因表达。