基于实时无标记细胞分析系统的气道平滑肌细胞收缩舒张效应检测方法的建立  

Establishment of the Contraction and Relaxation Effect System Based on RTCA of Airway Smooth Muscle Cells

作　　者：陈采陶 史阳琳 刘佳缘 杨永清 陈艳焦 CHEN Caitao;SHI Yanglin;LIU Jiayuan;YANG Yongqing;CHEN Yanjiao(Shanghai Research Institute of Acupuncture and Meridian,University of Traditional Chinese Medicine,Shanghai 200030,China)

机构地区：[1]上海中医药大学上海市针灸经络研究所,上海200030

出　　处：《中国细胞生物学学报》2020年第7期1187-1193,共7页Chinese Journal of Cell Biology

基　　金：国家自然科学基金(批准号:81973951);上海市卫生健康委员会(批准号:20194Y0164)资助的课题。

摘　　要：该研究利用实时无标记细胞分析系统(xCELLigence real-time cell analysis system,RTCA)建立气道平滑肌细胞(airway smooth muscle cells,ASMCs)收缩舒张效应检测方法。筛选适宜细胞接种密度,接种于RTCA检测板中监测细胞动态生长曲线;利用组胺刺激细胞收缩,并计算组胺半数抑制浓度(IC50);在适宜组胺刺激浓度的基础上,利用特布他林模拟细胞舒张效应,并计算半数有效浓度(EC50)。结果显示,ASMCs为4000个/孔接种密度培养于(16±4)h区间内,符合后续实验需求;组胺作用ASMCs 2 h后的IC50为8.07 mmol/L,故选择8 mmol/L组胺刺激ASMCs可见较为明显的收缩效应;特布他林作用ASMCs 2 h后的EC50为8.08×10–8 mol/L,故浓度为10μmol/L和20μmol/L的特布他林可检测到明显舒张效应(P<0.05)。利用RTCA可以稳定、有效、可靠地检测ASMCs收缩舒张效应,为以ASMCs为主要研究对象的呼吸系统疾病提供参考数据和准确的检测方法。In this study,RTCA(xCELLigence real-time cell analysis system)was used to establish a method to detect the contraction and relaxation effect of ASMCs(airway smooth muscle cells).ASMCs were inoculated into the E-plate of RTCA detection plate with different densities to monitor the dynamic growth curve,and the appropriate cell density was selected.Histamine was used to stimulate cell contraction and the IC50(median inhibition concentration)of histamine was calculated.Based on appropriate histamine stimulation concentration,terbutaline was used to simulate the relaxation effect of cells,and the EC50(median effective concentration)was calculated.The results showed that the density of ASMCs was 4000 cells/well[the cell culture was within(16±4)h],which was in line with the requirements of subsequent experiments.The IC50 of histamine on ASMCs for 2 h was 8.07 mmol/L,so the contractile effect of ASMCs stimulated by histamine of 8 mmol/L was obvious.The EC50 of terbutaline on ASMCs 2 h was 8.08×10–8 mol/L.Based on the cell contraction model,significant relaxation effect of terbutaline was detected at the concentration of 10μmol/L and 20μmol/L(P<0.05).ASMCs contraction and relaxation effect can be effectively detected by RTCA,providing reference data and accurate detection method for respiratory diseases with ASMCs as the main research object.

关 键 词：气道平滑肌 实时无标记细胞分析系统 收缩舒张效应 组胺 特布他林 

分 类 号：R562.25[医药卫生—呼吸系统]