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心肌梗死是一种严重的心血管疾病,由于冠状动脉血供急剧减少或中断,使心肌持续性缺血缺氧以致坏死,损害心功能且可能导致心律失常、休克以及心力衰竭等严重后果,已成为威胁人类生命健康的重大疾病之一[1-2]。近几十年来,随着医疗技术的进步,再灌注心肌治疗可显著改善心梗患者的生存率,但由于心梗发生发展及转归过程极其复杂,使其在临床治疗中仍然面临许多挑战[3],因此构建合适的动物模型对探究人心肌梗死的发病机制和病理过程、评价药物疗效以及探索新的治疗方法至关重要。通过结扎冠状动脉模拟心肌缺血过程来建立心肌梗死的动物模型是目前广泛应用的较为成熟的方法[4-5]。以往更倾向于选择较大动物构建模型,随着基因工程技术的发展,基因工程小鼠成为炙手可热的研究工具,因此建立简便有效的小鼠心梗模型对心肌梗死疾病的深入研究有重要意义。但目前国内关于小鼠心梗模型构建方法的报道较少,缺乏一种比较便捷的模型制作方法和无创评价手段。本文基于Gao等[6]报道的心梗模型构建方法结合实际操作总结了一些经验。
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小鼠心梗模型总体成功率为79.3%(23/29)。其中,冠脉结扎术中死亡率为6.8%(2/29),1只由于穿刺过深导致心脏破裂出血,1只由于胸腔暴露时间过长导致气胸。术后早期(<4 h)死亡率为10.3%(3/29);2只解剖后发现胸腔内有大量积血,1只疑为发生不可逆转的致死型心律失常。此外,1只造模失败未发生心梗,术后24 h TTC染色未见明显梗死区,且术后即刻和术后4 h心电图均无ST段改变(表1)。
表 1 小鼠心梗模型失败原因分析及解决方法
失败时段 比例(%) 原因分析 只数 解决方法 术中死亡 6.8 术中心脏破裂致死 1 控制穿刺深度和宽度 术中气胸致死 1 穿刺结扎迅速,快速闭合胸腔 术后死亡 10.3 术后胸腔出血致死 2 保证视野开阔,用纱布保护胸壁及肺,避免刺破大血管和肺脏 术后致死型心律失常致死 1 结扎动作迅速轻柔,减少对心脏的刺激 其他 3.4 术后未发生心梗 1 (造模失败) -
造模小鼠均施行术后即刻和术后4 h两次心电图检查,发现经24 h TTC染色后证实有心肌梗死发生的23只造模成功的小鼠中,两次心电图中均呈现出明显的ST段抬高以及变异T波(图1A),而造模失败的小鼠心电图则没有明显异常(图1B)。
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术后24 h打开小鼠胸腔,肉眼可见左心室前下壁靠近心尖处呈灰白色(图2A)。取心脏标本经常规TTC染色后,可观察到梗死区域呈白色,而非梗死区域呈红色(图2B)。根据前文提到心梗区大小的计算方法,本实验中,心梗模型梗死区大小最大可达40%,几乎累及整个左心室;最小为17%,主要集中在心尖部位;梗死区平均大小为(28±6)%(见表2)。
表 2 23只小鼠心肌梗死区大小情况
项目 体重/g 全心室重量(M1)/mg 梗死心肌重量(M2)/mg 梗死大小(M2/M1)/(%) 总计($ {\rm{\bar X}} $±s) 29.65±5.35 76±13 21.91±7.06 28±6
Establishment of mouse myocardial infarction model and early electrocardio- gram assessment
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摘要:
目的 构建并优化小鼠心肌梗死模型,联合使用冠脉结扎术后即刻和术后4 h的两次肢体导联心电图对心梗发生情况进行早期评价。 方法 C57BL/6J雄性小鼠29只,异氟烷吸入麻醉后,经左侧第3/4肋间进入胸腔,结扎冠状动脉左前降支建立模型,施行术后即刻和术后4 h肢体导联心电图评价心梗发生情况。术后24 h打开胸腔观察梗死情况,留取心脏标本进行TTC染色确定梗死区域并计算梗死面积。 结果 小鼠行冠状动脉结扎术,术中死亡率为6.8%(2/29),术后早期(<4 h)死亡率为10.3%(3/29),24 h存活率为82.8%(24/29);术后24 h TTC染色明确梗死发生,则心梗模型建立,造模成功率为79.3%(23/29),平均梗死区域大小(梗死心肌重量/全心室重量)为(28±6)%;成功建立模型的小鼠在术后4 h心电图可见明显ST-T改变,提示心梗已发生。 结论 成功建立小鼠心肌梗死模型,且联合使用术后即刻和术后4 h两次心电图可以作为小鼠心肌梗死模型快速无创的评价方法。 Abstract:Objective To establish and optimize a mouse myocardial infarction (MI) model, and to use twice limb lead ECGs immediately after coronary ligation and 4 h after surgery to evaluate the occurrence of myocardial infarction. Methods Twenty-nine male C57BL/6J mice were anesthetized with isoflurane. then a myocardial infarction model was established by ligating the left anterior descending (LAD) coronary artery through the third/fourth intercostal space of left anterior chest. Immediate and 4 h postoperative limb lead ECGs were performed. Twenty-four hours after surgery, the chest was opened and the occurrence of myocardial infarction was evaluated. The heart samples were taken for TTC staining to determine the infarct area and calculate the infarct area. Results During the mice underwent coronary artery ligation the intraoperative mortality was 6.8% (2/29), and the early postoperative (<4 h) mortality was 10.3% (3/29). The 24 h survival rate was 82.8% (24/29). 24 hours after TTC staining confirmed the occurrence of infarction, the myocardial infarction model was established. The success rate of the model was 79.3% (23/29), and the average infarct size (infarcted myocardial weight / whole ventricular weight) was (28 ± 6)%; The mice successfully established by the model showed obvious ST-T changes in the ECG at 4 hours after surgery, suggesting that a myocardial infarction has occurred. Conclusions The mouse myocardial infarction model was successfully established. The combined use of ECG immediately after surgery and 4 h after surgery could be used as a rapid and non-invasive evaluation method for mouse myocardial infarction. -
Key words:
- myocardial infarction /
- mouse /
- animal model /
- ECG assessment /
- TTC staining
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表 1 小鼠心梗模型失败原因分析及解决方法
失败时段 比例(%) 原因分析 只数 解决方法 术中死亡 6.8 术中心脏破裂致死 1 控制穿刺深度和宽度 术中气胸致死 1 穿刺结扎迅速,快速闭合胸腔 术后死亡 10.3 术后胸腔出血致死 2 保证视野开阔,用纱布保护胸壁及肺,避免刺破大血管和肺脏 术后致死型心律失常致死 1 结扎动作迅速轻柔,减少对心脏的刺激 其他 3.4 术后未发生心梗 1 (造模失败) 表 2 23只小鼠心肌梗死区大小情况
项目 体重/g 全心室重量(M1)/mg 梗死心肌重量(M2)/mg 梗死大小(M2/M1)/(%) 总计( $ {\rm{\bar X}} $ ±s)29.65±5.35 76±13 21.91±7.06 28±6 -
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