中文摘要：

目的：用两种不同的方法构建β3肾上腺素能受体（β3-AR）过表达的 SD 乳鼠心肌成纤维细胞（CFBs）模型，并对两种方法的效果进行比较，为今后在细胞水平观察β3-AR 对心功能，尤其是对慢性心力衰竭（CHF）的影响提供新的实验方法。方法2015年3—6月，选取出生1~3 d 的 SFP 级 SD 乳鼠8只。分离 SD 乳鼠CFBs 并体外培养，随机分为3个感染复数（MOI）组，分别为 MOI 50组、MOI 100组、MOI 200组，采用β3-AR 基因过表达的慢病毒载体感染 CFBs，利用流式细胞仪检测感染率，确定最佳 MOI。将 SD 乳鼠 CFBs 随机分为4个β3-AR 激动剂干预组，分别为正常对照组、10-6 mol/ L 组、10-5 mol/ L 组、10-4 mol/ L 组，采用不同浓度β3-AR 激动剂干预CFBs，利用 Western blotting 法检测β3-AR 表达水平，挑选出β3-AR 激动剂干预 CFBs 的最适浓度。结果普通光学显微镜下观察 CFBs，无搏动性，大多数呈梭形，其细胞核较大，胞质透明。倒置荧光显微镜下观察感染24、48、72、96 h 时 CFBs 状态及荧光表达情况，感染72 h 时，各 MOI 组 CFBs 生长状态较好，细胞大多数呈梭形，无搏动性，细胞核较大，且肉眼观察荧光表达最佳。MOI 100组、MOI 200组感染率高于 MOI 50组（P ＜0.05）；MOI 200组感染率低于MOI 100组（P ＜0.05）。10-6 mol/ L 组、10-5 mol/ L 组、10-4 mol/ L 组β3-AR 表达水平高于正常对照组（ P ＜0.05）；10-5 mol/ L组β3-AR 表达水平高于10-6 mol/ L 组、10-4 mol/ L 组（P ＜0.05）。10-5 mol/ L 组、MOI 100组β3-AR 表达水平高于正常对照组（P ＜0.05）；MOI 100组β3-AR 表达水平高于10-5 mol/ L 组（P ＜0.05）。结论使用β3-AR 过表达的慢病毒载体感染 CFBs 与β3-AR 激动剂干预 CFBs 均可使其β3-AR 过表达，且前者效果优于后者。

英文摘要：

Objective To build neonatal SD rat cardiac fibroblasts（CFBs）models with β3-AR overexpression by two different methods and compare the two methods so as to provide new experimental method for exploring the effect of β3-AR on heart function,especially on chronic cardiac failure. Methods From March to June in 2015,8 SPF neonatal SD rats born 1 to 3 days ago were selected. CFBs of the rats were cultured in vitro and were randomly divided into three MOI groups which were MOI 50 group,MOI 100 group and MOI 200 group. These CFBs were infected by lentiviral vector with β3-AR overexpression,and infection rate was detected by flow cytometry to determine the optimal MOI. CFBs were randomly divided into four β3-AR agonist intervention groups which were normal control group,10 - 6 mol/ L group,10 - 5 mol/ L group,and 10 - 4 mol/ L group. Intervention was conducted on these CFBs by different concentrations of β3-AR agonist,and β3-AR expression level was detected by Western blotting method to find out the optimal concentration of β3-AR agonist in the intervention on CFBs. Results Observed under bspnormal optics microscope,CFBs showed no pulsation and mostly took on fusiform shape,with large cell nucleus and transparent cytoplasm. The status of CFBs and fluorescent expression were observed under inverted fluorescence microscope at 24,48,72 and 96 h after infection. At 72 h,CFBs of each MOI group were in good growth status,they were mostly in fusiform shape and had no pulsation,with large cell nucleus,and the fluorescent expression observed by naked eye was optimal. MOI 100 group and MOI 200 group were higher than MOI 50 group in infection rate（P 〈 0. 05）;MOI 200 group was lower than MOI 100 group in infection rate（P 〈 0. 05）. 10 - 6 mol/ L group,10 - 5 mol/ L group and 10 - 4 mol/ L group were higher than normal control group in the expression level of β3-AR（P 〈 0. 05）;10 - 5 mol/ L group was higher than 10 - 6 mol/ L group and 10 - 4 mol/ L group in the expression level of β3-AR（P 〈 0. 05）.