β_2受体下调细胞模型的构建研究  

Study on construction of β_2-adrenoceptor downregulation cell model

作　　者：刘华[1] 徐翼[2] 盛泽波[2] 章艳菊[1] 张引子[1] 倪松石[1] 

机构地区：[1]南通大学附属医院呼吸科,南通226001 [2]江苏省常熟市第一人民医院呼吸科

出　　处：《南通大学学报（医学版）》2011年第1期10-13,16,共5页Journal of Nantong University(Medical sciences)

基　　金：国家自然科学基金资助项目(30971306);南通市社会发展科技项目(S2008017;S2009023);南通大学附属医院科技项目(TDFY0914)

摘　　要：目的:通过沙丁胺醇刺激的方法构建细胞水平β2肾上腺素能受体(β2AR)下调的模型,为研究β2AR下调的分子生物学机制奠定基础。方法:取正常Balb/c小鼠气道平滑肌细胞(ASMC)进行原代培养(组织贴块法),第2～5代细胞经形态学及间接免疫荧光法进行鉴定,将第3～4代纯度在95%以上的ASMC随机分为2组:正常ASMC对照组和沙丁胺醇刺激组。采用RT-PCR和Western Blot分别检测对照组和沙丁胺醇刺激组β2AR mRNA及蛋白水平的表达。结果:(1)原代培养的ASMC呈典型的"峰谷"状生长,细胞浆内有与细胞纵轴平行的特异性平滑肌α-肌动蛋白丝的阳性表达,证实了平滑肌细胞的来源,并符合平滑肌细胞的形态学和生物学特性;(2)沙丁胺醇刺激组β2AR mRNA的表达较对照组降低(P<0.05),β2AR蛋白表达亦明显降低(P<0.05)。结论:成功构建了小鼠气道平滑肌β2AR下调的细胞模型,为进一步研究哮喘患者β2AR减敏的机制奠定了良好的基础。Objective：To establish cell model of β2-adrenergic receptor（β2AR） downregulation of murine airway smooth muscle induced by salbutamol.So,and on the basis of this platform to elucidate the molecular and biological mechanism of β2AR downregulation in detail.Methods：Airway smooth muscle cell（ASMC） from Balb/c mice was primarily cultured.Two to five generations of cells were identified by cell morphology and indirect immunofluorescence.They were divided into two groups at random：control group and salbutamol-treated group,whose purity was more than 95% from third to fourth generations.The level of β2AR mRNA and protein expression of control group and salbutamol-treated group were detected by RT-PCR and Western Blot.Results：Primarily cultured cells demonstrated typical characteristic of ＂peak and valley＂-like growth trend.There was positive expression of specific smooth muscle α-actin filament paralleling to cell longitudinal axis in the cytoplasm.The origin of smooth muscle cell had been validated which was consistent with its morphology and biological characteristics.The expression level of β2AR mRNA in salbutamol-treated group was lower than that in the control group（P0.05）.The expression level of β2AR protein was also markedly lower than that in the control group（P0.05）.Conclusion：Cell model of airway smooth muscle cell related to β2AR downregulation had been successfully established.It may lay a good foundation for further studying the mechanism of β2AR downregulation in asthmatic patients.

关 键 词：气道平滑肌细胞 Β2肾上腺素能受体 沙丁胺醇 细胞培养 

分 类 号：R562.25[医药卫生—呼吸系统]