薄荷醇通过mTOR活化促进人支气管上皮细胞气道炎症相关因子的表达  

Role of mammalian target of rapamycin activation in menthol?induced expressions of airway inflammation-related factors in human bronchial epithelial cells in vitro

作　　者：陈海博 李敏超[1] CHEN Haibo;LI Minchao(Department of Respiratory Medicine,Second Affiliated Hospital of Chongqing Medical University,Chongqing 400010,China)

机构地区：[1]重庆医科大学附属第二医院呼吸内科

出　　处：《南方医科大学学报》2019年第11期1344-1349,共6页Journal of Southern Medical University

基　　金：国家自然科学基金（81270102）;重庆市自然科学基金（cstc2012jjA10050）;重庆市教委科学技术研究基金（KJ120301）~~

摘　　要：目的探讨哺乳动物雷帕霉素靶蛋白(mTOR)活化对薄荷醇诱导的人支气管上皮细胞(BEAS-2B)气道炎症相关因子表达的影响及其机制。方法将细胞分为4组:正常对照组、薄荷醇组、雷帕霉素组、薄荷醇+雷帕霉素组。用CCK-8法检测细胞存活率,实时荧光定量PCR检测瞬时受体电位蛋白-8(TRPM8)、肿瘤坏死因子(TNF)-α和白细胞介素(IL)-1βmRNA的表达,Western blot检测BEAS-2B中磷酸化的mTOR(p-mTOR)、TRPM8、TNF-α和IL-1β的蛋白表达。用流式细胞术检测胞内Ca2+荧光强度。结果与正常对照组相比,薄荷醇组中细胞TNF-α和IL-1βmRNA及蛋白表达升高,p-mTOR蛋白表达升高,胞内Ca2+浓度升高(P<0.05)。与正常对照组相比,雷帕霉素组中细胞TNF-α和IL-1βmRNA及蛋白表达均无差异(P>0.05),p-mTOR蛋白表达降低(P<0.05),胞内Ca2+浓度无差异(P>0.05)。与薄荷醇组相比,薄荷醇+雷帕霉素组中细胞TNF-α和IL-1βmRNA及蛋白表达降低,p-mTOR蛋白表达降低,胞内Ca2+浓度降低(P<0.05)。结论薄荷醇可能通过活化mTOR诱导胞内Ca2+浓度升高,进而促进气道炎症相关因子IL-1β和TNF-α的表达。Objective To investigate the role of mammalian target of rapamycin(mTOR) activation in menthol-induced expression of airway inflammation-related factors in human bronchial epithelial cells and explore its mechanism. Methods Cultured human bronchial epithelial cells(BEAS-2 B) were divided into normal control group, menthol group, rapamycin group, and menthol + rapamycin group with corresponding treatments. The cell viability was measured with CCK-8 method.The mRNA levels of transient receptor potential melastatin 8(TRPM8), tumor necrosis factor(TNF)-α and interleukin(IL)-1βwere detected by RT-PCR, and the protein expressions of phosphorylated mTOR(p-mTOR), TRPM8, TNF-α and IL-1β were determined using Western blotting. The intracellular Ca2+fluorescence intensity was measured by flow cytometry. Results Compared with the normal control cells, menthol-treated cells showed significantly increased TNF-α, IL-1β, and p-mTOR expression and elevated intracellular Ca2+concentration(P<0.05), and the rapamycin-treated cells exhibited significantly decreased p-mTOR expression(P<0.05). No significant difference was found in TNF-α, IL-1β or intracellular Ca2+concentration between the normal control and rapamycin-treated cells(P>0.05). Compared with the menthol-treated cells, the cells treated with both menthol and rapamycin showed significantly decreased TNF-α, IL-1β, and p-mTOR expression and obviously lowered intracellular Ca2+concentration(P<0.05). Conclusion Menthol promotes the expressions of airway inflammationrelated factors IL-1β and TNF-α possibly by activating mTOR to cause the increase of intracellular Ca2+concentration.

关 键 词：人支气管上皮细胞 薄荷醇 瞬时受体电位蛋白-8 哺乳动物雷帕霉素靶蛋白 气道炎症 

分 类 号：R73[医药卫生—肿瘤]