这项工作得到了瑞士心血管基金会的赠款 (编号 32016) 的支持。

该协议中使用了 雄性 C57BL/6J 鼠 (10 周, 25-30g, n=60)。根据法国农业部和高等教育和研究部制定的准则, 动物接受人道护理, 所有程序都按照欧洲共同体理事会1986年11月24日的指示执行 (86/609/欧洲经济共同体) 和法国法律。该议定书得到了 #34 的批准; 动物实验 CREMEAS 和 #34 区域伦理委员会; (#2016092816207606). 
 1. 手术准备 <ol> <li> 在到达动物设施后一周内, 在标准的笼子里, 用12小时/12 小时的光照/黑暗周期, 用食物 (详见材料表) 和可用的水来维护小鼠。随意 . </li>
	<li> 在手术当天, 将小鼠放在单独的笼子里, 在麻醉诱导前几分钟, 以避免对动物造成任何额外的压力。手术前一天对所有手术器械进行消毒. </li>
	<li> 注射 intra-peritoneally 单剂量氯胺酮 (51.4 毫克/千克) 和嗪 (3.3 毫克/千克) 稀释盐水溶液 (0.9% NaCl). </li>
	<li> 在没有脚趾后退反射的情况下确保麻醉的深度. </li>
	<li> 用市面上可用的剃须刀剃掉动物的颈部和胸部, 用70% 酒精消毒剃须的区域. </li>
	<li> 将动物仰卧在干净的软木工作台上, 并用胶带固定爪子. </li>
</ol> 2。外科手术 <ol> <li> 无菌外科技术在整个过程中使用。在一个自发呼吸的动物, 执行纵向中线颈部切口超过10毫米与11刀片刀从上部胸骨切口到 mid-chest 为了暴露胸骨 ( 图 1 ). </li>
	<li> 通过一个4/0 单丝聚丙烯保持缝线与一个克瑞理木针持有人, 并将其贴在工作台上, 以收回甲状腺. </li>
	<li> 将 pre-tracheal 肌肉与微外科钳分开, 以发现气管. </li>
	<li> 在气管和胸骨后的闭合颚上轻轻滑动平滑倾斜的微外科钳. </li>
	<li> 通过仔细地打开和关闭光滑倾斜的弯曲显微外科钳的颌骨, 在 pre-tracheal 肌肉和胸骨后进行钝性解剖, 将胸膜移开. </li>
	<li> 抓住右锁骨上的肌肉与光滑的直微外科钳, 并轻轻拉起动物的胸部. </li>
	<li> 在胸骨下滑动骨钳的下颚, 并执行 3-4 mm 上部部分胸骨 ( 图 2 )。将胸骨的下部稍稍朝左. </li>
	<li> 通过7/0 单丝聚丙烯保持缝线从内到外通过胸骨两侧的第二个肋间空间使用微外科针持有人。靠近肋骨-胸骨角, 避免肋间和胸廓内血管或胸膜损伤. </li>
	<li> 在每一侧使用7/0 单丝聚丙烯保持缝线将胸骨边缘展开, 并用胶带将其固定在工作台上. </li>
	<li> 在低功耗放大 (2-3X) ( 图 3 ) 的条件下, 使用平滑的弯曲显微外科钳将 pre-tracheal 肌、纵隔脂肪和胸腺的部位轻轻移开, 以可视化主动脉弓。注意不要接触或损伤壁胸膜, 以防气胸的发展. </li>
	<li> 通过结扎钳 ( 图 4 A ) 将主动脉弓下的软组织露出, 轻轻地张开它的颚。在软组织中, 通过轻轻地打开和闭合颌骨, 在主动脉弓下的软组织下准备一条隧道. </li>
	<li> 通过结扎辅助的眼睛 ( 图 4 B ) 在主拱下的左侧传递6/0 丝线结扎的片段, 并通过在右手之间的绑钳进行检索。右无名和左颈共同动脉 ( 图 5 ). </li>
	<li> 将一根27口径的针切割成5毫米的长度, 然后用克瑞理针钳将其两端钝。将钝的27口径针放在主动脉弓 ( 图 6 ) 旁边, 用平滑的直微手术钳将缝合线紧贴在针和主动脉之间, 右无名和左颈共同使用两个结扎钳的动脉 ( 图 7 )。为了紧贴缝合, 执行一个初始的双结, 后跟四额外的结。确保所有节点都是平坦的. </li>
	<li> 结扎后, 快速移除针头, 以达到0.4 毫米直径缩小和可重复的横向65-70% 主动脉收缩. </li>
	<li> 检查腹主动脉弓周围软组织的止血, 胸骨边缘和 pre-tracheal 肌。在观察血液渗出的地方, 放可止血纱布。取下7/0 单丝聚丙烯保持缝合线用于胸骨边缘的传播. </li>
	<li> 通过一个简单的6/0 单丝聚丙烯缝合线与微外科针持有人从外部到左第二肋间空间, 然后从内到外的右第二肋间空间。靠近肋骨-胸骨角, 避免肋间和胸廓内血管或胸膜损伤. </li>
	<li> 将6/0 单丝聚丙烯缝合线与克瑞理木针夹合在一起, 将胸骨边缘拼接在一起. </li>
	<li> 用5/0 单丝聚丙烯纤维在一层中用克瑞理-木针夹紧皮肤. </li>
	<li> 执行与收缩操作相同的假程序, 但不在主动脉周围绑缝合. </li>
</ol> 3。术后恢复 <ol> <li> 非常密切地监视动物. #160; 将鼠标转移到单个的笼子, 并将其置于俯卧位置. </li>
	<li> 允许鼠标在变暖光下恢复, 直到完全清醒 (在诱导麻醉后不到1小时). </li>
	<li> 为术后镇痛, 注入0.1 毫克/千克丁丙诺啡腹腔. 和 #160; 重复皮下注射0.1 毫克/千克丁丙诺啡每8小时, 在前三天表明. </li>
	<li> 将操作的小鼠放在标准笼子里 (每个笼子最多3只老鼠, 每个笼子至少2只老鼠). </li>
</ol> 4。心脏收获 <ol> <li> 在分析的当天, 安乐用氯胺酮300毫克/千克和嗪20毫克/千克的溶液进行腹腔注射. </li>
	<li> 首先从下腔静脉采集血液, 然后通过同一条线在盐水中注入5毫升的2.6 毫米 EDTA 溶液. </li>
	<li> 收获心脏, 去除心房和重量心脏 (左和右心室没有心房). </li>
	<li> 将左侧与右侧 ventr 分开icle 与左心室部分残余的隔膜。称量两种组织标本, 并将其冷冻在液氮中. </li>
</ol>

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作者没有利益冲突的披露。

本议定书的目的是提出一个 step-by 的外科技术, 微侵袭横动脉狭窄小鼠。对小鼠横主动脉狭窄的详细技术描述已由其他作者2,8报告。然而, 这些研究者在动物插管和通气后进行手术。使用额外的一步插管通气增加了整个过程的复杂性和持续时间和全球压力的动物接触。因此, 微创横脉狭窄的概念得到了一定的重视。小鼠的微创横动脉狭窄用于诱发左心室肥厚及心力衰竭的进展9,10,11。这些研究的重点是在左心室肥厚和心力衰竭的成因, 而不是对手术技术的描述9,10,11。
在本协议中, 我们详细报道了小鼠微创 TAC 的简化和重现技术。熟练的外科医生可以在15分钟内进行20分钟的收缩手术和假手术 (无缝合栓)。在我们最初的技术证明中, 我们发现, 引进一个关键的仪器, 结扎的援助, 使一个非常低的手术死亡率1.7%。这与人 et 报告的4% 的手术死亡率相比比较有利。al。5, 3.7% 由廖 et al。13和2.7% 由塔拉斯坦费尔德 et al14。此外, 观察期长达28天, 术后晚期死亡率也很低, 为1.7%。再次, 这与人 et al (10%)5、廖 et al (19%)13或塔拉斯坦费尔德 et al (2.6%)14所报告的晚期死亡率比较好。
主动脉弓下的通道是整个手术过程中最关键的一步。这一步骤的重现性没有描述的胡和同事谁使用了自制的电线与一个圈套在其年底通过主动脉下的权利无名的来源和左颈总动脉9, 也没有由 Tarnavski 谁定位在上升主动脉下内侧侧的弯曲钳, 在另一侧抓住7/0 丝缝合线, 并将其移动到主动脉2下方。在我们的技术使用的结扎援助, 使一个标准化和重复性的机动, 低风险的主动脉撕裂。
该程序的另一个决定性步骤是张力施加在27口径针的领带上, 以有效降低主动脉弓的腔内 homogenously。首先, 我们使用栓钳, 这有助于应用均匀和重现张力的缝合周围的主动脉弓。在心脏和主动脉弓的收获过程中, 对缝线的适当放置进行了验证。安徒生和工友通过评估颈动脉的多普勒信号证实了乐队的适当的安置在主动脉带的安置之前和之后11。在他们的报告中, 当多普勒流速比从右至左颈动脉的一倍11时, 接受了适当的带状。在我们的技术, 我们选择测量的效率, TAC 的程度诱导左心室肥厚与假动物相比, 为了验证的程序, 因为它不需要任何延长的程序或动物的补充麻醉。在我们的技术, 在实验结束时, 检查左心室肥厚程度和适当的带状位置。我们的技术对左心室肥厚的程度与其他研究人员在 TAC 后3周报告的结果相比, 在小鼠15。此外, 在我们的带状动物中观察到的心脏与体重比值的低变化证明了在领带上施加的张力的低波动。
总之, 通过避免在本协议中提出的插管通气, 我们的小鼠微创技术提供了一个可靠的和可重复的模型。这一模型减少了对动物的全球应变, 是时间和成本节省比 TAC 使用插管通风的动物。该方法手术和后期死亡率低, 使该技术成为一种诱导小鼠左心室肥厚的选择手段。

在过去的几年中, 心力衰竭的研究已经在可行的动物模型中进行了1。与大型动物模型的心力衰竭相比, 小动物模型有许多潜在的优势。除了低成本的住房和维修, 小动物模型可以访问更多的研究人员, 由于不太复杂的设施需要2。
小鼠心力衰竭模型提供了许多相同的优势, 与老鼠模型。除了减少的住房成本3, 鼠标模型受益于相关的转基因和淘汰赛 (KO) 菌株的可用性。细胞类型特异、诱导性 KO 或转基因策略的可能性使小鼠成为研究心力衰竭发病机制的宝贵工具, 并尝试识别新的治疗方案3。
在目前使用的心力衰竭的小鼠模型中4中, 首次用人5描述的横主动脉收缩 (TAC) 是产生压力超负荷引起的左心室肥厚 (肥厚)1的首选模型。,3. 该模型的最大优点是能够允许左心室肥厚的分层, 尽管在不同的小鼠品系中, 对 TAC 的反应是可变的.特别是, C57BL/6 小鼠发展快速 LV 扩张后, TAC 可能不会发生其他菌株4,6,7。
在2周内, 以 TAC 引起的高血压的突然发作使 LV 肿块增加约 50%, 从而可以快速检查旨在调节左心室肥厚4发育的药理或分子干预活动。急性高血压的 TAC 不完全再现左心室肥厚和重构观察在主动脉狭窄或动脉高血压的临床设置。然而, 该模型被许多研究者用来识别和修改心力衰竭的新治疗指标4。
执行 TAC 在小鼠需要更大的手术经验比其他技术所需的诱导肥厚和随后心力衰竭2。大多数作者执行此过程的插管和通风的动物2,8, 这使得这个过程更苛刻和耗时, 并增加了对动物的外科负担。只有很少的调查人员在他们的研究中使用了微创的 TAC, 简要地参考了手术过程9,10,11。
本议定书的目的是描述 step-by 一步简化和 user-friendly 的小鼠横主动脉狭窄的技术, 突出了程序的关键阶段。通过遵循这些关键步骤, 可以轻松地执行此项技术。

<table><tbody><tr><td>Surgical microscope</td><td>Olympus</td><td>SZX2-TR30</td><td></td></tr><tr><td>Razor</td><td>Rowenta</td><td>Nomad TN3650FO</td><td></td></tr><tr><td>&lt;strong&gt;Sutures:&lt;/strong&gt;</td><td></td><td></td><td></td></tr><tr><td>Polypropylene 7/0</td><td>Ethicon</td><td>BV-1X</td><td></td></tr><tr><td>Polypropylene 6/0</td><td>BBraun</td><td>C0862061</td><td></td></tr><tr><td>Silk 6/0 ligature&amp;nbsp;</td><td>FST</td><td>18020-60</td><td></td></tr><tr><td>Polypropylene 4/0</td><td>Ethicon</td><td>8683</td><td></td></tr><tr><td>Polypropylene 5/0</td><td>Ethicon</td><td>Z303</td><td></td></tr><tr><td>&lt;strong&gt;Drugs:&lt;/strong&gt;</td><td></td><td></td><td></td></tr><tr><td>Ketamin</td><td>Merial</td><td>Imalg&amp;egrave;ne 1000, LBM154AD</td><td></td></tr><tr><td>Xylazine</td><td>Bayer</td><td>Rompun 2%, KP09PPC</td><td></td></tr><tr><td>Buprenorphine</td><td>Ceva</td><td>Vetergesic, 072013</td><td></td></tr><tr><td>&lt;strong&gt;Instruments:&amp;nbsp;&lt;/strong&gt;</td><td></td><td></td><td></td></tr><tr><td>Bone nippers</td><td>Fine Surgical Tools</td><td>16101-10</td><td></td></tr><tr><td>Ligation aid</td><td>Fine Surgical Tools</td><td>18062-12</td><td></td></tr><tr><td>Tying forceps</td><td>Fine Surgical Tools</td><td>18026-10</td><td></td></tr><tr><td>Needle holder Crile-Wood</td><td>Fine Surgical Tools</td><td>12003-15</td><td></td></tr><tr><td>Microsurgery forceps&amp;nbsp;</td><td>Fine Surgical Tools</td><td>11003-12</td><td></td></tr><tr><td>Microsurgery forceps&amp;nbsp;</td><td>Fine Surgical Tools</td><td>11002-12</td><td></td></tr><tr><td>Tissue forceps</td><td>Fine Surgical Tools</td><td>11021-12</td><td></td></tr><tr><td>Microsurgery needle holder</td><td>Fine Surgical Tools</td><td>12076-12</td><td></td></tr><tr><td>Microsurgery scissors</td><td>Fine Surgical Tools</td><td>91501-09</td><td></td></tr><tr><td>Mayo scissors</td><td>Fine Surgical Tools</td><td>14511-15</td><td></td></tr><tr><td>11-blade knife</td><td>Fine Surgical Tools</td><td>10011-00</td><td></td></tr><tr><td>&lt;strong&gt;RNA extraction and qPCR:&lt;/strong&gt;</td><td></td><td></td><td></td></tr><tr><td>TriReagent</td><td>Euromedex</td><td>TR-118-200</td><td></td></tr><tr><td>Rneasy Mini kit</td><td>Qiagen</td><td>74704</td><td></td></tr><tr><td>Qubit Fluorimetric RNA assay</td><td>Fisher Scientific</td><td>10034622</td><td></td></tr><tr><td>RNA 6000 Nano kit</td><td>Agilent</td><td>5067-1511</td><td></td></tr><tr><td>High Capacity cDNA kit</td><td>Fisher Scientific</td><td>10400745</td><td></td></tr><tr><td>Taqman Master Mix</td><td>Fisher Scientific</td><td>10157154</td><td></td></tr><tr><td>Taqman BNP primers</td><td>Fisher Scientific</td><td>Mm01255770_g1</td><td></td></tr><tr><td>Taqman ANP primers</td><td>Fisher Scientific</td><td>Mm01255747_g1&amp;nbsp;</td><td></td></tr><tr><td>Taqman ACE primers</td><td>Fisher Scientific</td><td>Mm00802048_m1</td><td></td></tr><tr><td>Taqman Col1a1 primers&amp;nbsp;</td><td>Fisher Scientific</td><td>Mm00801666_g1</td><td></td></tr><tr><td>Taqman TGFb primers</td><td>Fisher Scientific</td><td>Mm01178820_m1</td><td></td></tr><tr><td>Taqman Gapdh primers</td><td>Fisher Scientific</td><td>Mm99999915_g1</td><td></td></tr><tr><td>ABIPrism&amp;nbsp; Thermocycler</td><td>Applied Biosystems</td><td>7000</td><td></td></tr><tr><td>&lt;strong&gt;Software:&lt;/strong&gt;</td><td></td><td></td><td></td></tr><tr><td>GraphPad Prism</td><td>GraphPad</td><td>Prism 7</td><td></td></tr><tr><td>&lt;strong&gt;Animal food&lt;/strong&gt;</td><td></td><td></td><td></td></tr><tr><td>Complete diet for adult rats/mice</td><td>Safe</td><td>UB220610R</td><td></td></tr></tbody></table>

technique of minimally invasive transverse aortic constriction in mice for induction of left ventricular hypertrophy

小鼠横主动脉狭窄 (TAC) 是压力超负荷所致左心室肥厚的实验研究和心力衰竭进展的最常用的外科技术之一。在大多数报告的调查中, 这一程序是用插管和通气的动物进行的, 这使得它要求和耗时, 并增加了对动物的外科负担。本协议的目的是描述一个简单的技术, 无插管和通气小鼠微创 TAC。强调了该技术的关键步骤, 以达到低死亡率和高效率的诱导肥厚。
雄性 C57BL/6 小鼠 (10 周大, 25-30 克, n=60) 被麻醉, 单腹腔注射氯胺酮和嗪的混合物。在一个自发呼吸的动物在3-4 毫米上部部份胸骨之后, 6/0 丝缝线穿线通过结扎援助的眼睛通过在主动脉弓之下和栓在一条钝的27测量针。假手术的动物进行了相同的外科准备, 但没有主动脉收缩。该程序在诱导肥厚中的疗效证实了心脏/体重比的显著增加。这个比率在 3, 7, 14 和28在手术以后获得 (n = 6-10 在每个小组和每个时间点)。运用我们的技术, 在 TAC 中观察到7天28的假动物的肥厚。手术和晚 (超过28天) 死亡率都非常低, 在1.7%。
总之, 我们的小鼠微创 cost-effective 技术具有非常低的手术和术后死亡率, 是高效的诱导肥厚。它简化了手术过程, 减少了对动物的压力。通过遵循本协议中描述的关键步骤, 可以很容易地执行该操作。

手术和晚期生存 手术的生存是非常高的, 98.3% (59 出 60) 为整个系列 (TAC 和假操作的动物)。唯一的手术死亡是由于一个小鼠的出血并发症, 计划为假手术。术后存活28天的观察期也很好, 98.3% (58 出 59)。唯一的晚术后死亡发生在一天 (D) 16, 可能是心脏起源的 TAC 鼠标。
技术验证 所提出的技术是非常可靠和重现性。在所有接受 TAC 的动物中, 正确的无名和左颈总动脉之间的缝合位置得到了确认。
通过测定3、7、14、28天的心脏重量/体重比值 (硬件/BW、镁/克), 验证了该技术对诱发左心室肥厚的后。硬件是左、右心室的重量, 没有心房。与术后 D7 (4.9±0.2 vs 4.1±0.05 毫克/克, p & #60; 0.01) 上的假组相比, 在条纹上的硬件/BW 比值显著增加, 并保持显著高于 D28 (5.8±0.3 vs 4.1±0.1 毫克/克, p & #60; 0.0001)后 (图 8)。观察到的增加的硬件/BW 比是完全由于左心室/体重比 (图 9a) 的上升, 因为右心室/体重比仍然是 TAC 和假操作动物在整个观察中的比较期间 (图 9B)。
此外, 我们在左心室组织中测量了心肌肥大的标志物的 mRNA 表达, 如前所述12。在 D14, 脑利钠蛋白 (BNP)、心房利钠蛋白 (ANP)、血管紧张素转换酶 (ACE)、胶原 1a1 (Col1a1) 和转化生长因子？ (TGFß) 的 mRNA 表达明显高于假操作的动物 (图 10)。因此, 观察到的左心室肥厚验证了我们的 TAC 技术的效率。
用 one-way 方差分析和 Bonferroni 的事后试验对配对数据进行比较, 比较 TAC 和假组均值的平均值和标准误差。
<img alt="Figure 1" class="xfigimg" src="/files/ftp_upload/56231/56231fig1.jpg"> 
图 1: 切口. 皮肤被切割超过10毫米从胸骨切口到 mid-sternum 和甲状腺被缩回与逗留缝合。<a href="//ecsource.jove.com/files/ftp_upload/56231/56231fig1large.jpg" target="_blank">请单击此处查看此图的较大版本.</a>
<img alt="Figure 2" class="xfigimg" src="/files/ftp_upload/56231/56231fig2.jpg"> 
图 2: 骨钳. 该仪器允许在骨头上短而精确地切割3-4 毫米上部偏高级胸骨。<a href="//ecsource.jove.com/files/ftp_upload/56231/56231fig2large.jpg" target="_blank">请单击此处查看此图的较大版本.</a>
<img alt="Figure 3" class="xfigimg" src="/files/ftp_upload/56231/56231fig3.jpg"> 
图 3: 曝光. 在胸骨边缘的缩回与7/0 中止缝合之后, 主动脉弓, 右无名和左颈总动脉与气管一起被暴露。<a href="//ecsource.jove.com/files/ftp_upload/56231/56231fig3large.jpg" target="_blank">请单击此处查看此图的较大版本.</a>
<img alt="Figure 4" class="xfigimg" src="/files/ftp_upload/56231/56231fig4.jpg"> 
图 4: 结扎钳.这些钳是必要的, 以执行一个温和的和钝性解剖后, 胸骨和周围的主动脉弓。B. 结扎辅助.这是一个关键的工具, 以实现一个微妙的和创伤的通道下主动脉弓在 TAC 和假手术的小鼠。<a href="//ecsource.jove.com/files/ftp_upload/56231/56231fig4large.jpg" target="_blank">请单击此处查看此图的较大版本.</a>
<img alt="Figure 5" class="xfigimg" src="/files/ftp_upload/56231/56231fig5.jpg"> 
图 5: 主动脉弓下的通道. 6/0 丝结扎的一部分是通过在主动脉弓下使用结扎援助, 并放置在右无名和左颈总动脉。<a href="//ecsource.jove.com/files/ftp_upload/56231/56231fig5large.jpg" target="_blank">请单击此处查看此图的较大版本.</a>
<img alt="Figure 6" class="xfigimg" src="/files/ftp_upload/56231/56231fig6.jpg"> 
图 6: 结扎的准备. 短段2-3 毫米的钝27口径针被放置在主动脉弓上。<a href="//ecsource.jove.com/files/ftp_upload/56231/56231fig6large.jpg" target="_blank">请单击此处查看此图的较大版本.</a>
<img alt="Figure 7" class="xfigimg" src="/files/ftp_upload/56231/56231fig7.jpg"> 
图 7: 横向主动脉收缩. 用结扎钳将丝线缝合在右侧无名和左颈总动脉之间的针和主动脉弓上。而不是聚丙烯缝合线是首选的主动脉结扎, 因为结将更好地举行。<a href="//ecsource.jove.com/files/ftp_upload/56231/56231fig7large.jpg" target="_blank">请单击此处查看此图的较大版本.</a>
<img alt="Figure 8" class="xfigimg" src="/files/ftp_upload/56231/56231fig8.jpg"> 
图 8: 横向主动脉狭窄的验证. 与假手术 (白棒) 小鼠相比, 带状 (黑条) 的心脏重量/体重比明显增加, 通过微创的横向主动脉狭窄诱导心肌肥大。心脏肥大已经出现在 D7 术后, 并逐步增加, 随着时间的推移, 以 D28 (n=6-10 每组. ** 和 #60; 0.01, *** 和 #60; 0.001, *** 和 #60; 0.0001)。数据显示为平均± SEM (误差线)。<a href="//ecsource.jove.com/files/ftp_upload/56231/56231fig8large.jpg" target="_blank">请求</a>se 点击这里查看更大版本的这个数字。
<img alt="Figure 9" class="xfigimg" src="/files/ftp_upload/56231/56231fig9v2.jpg"> 
图 9: 左 (a) 和右 (B) 心室/体重比. 在观察期间, 左心室/体重比增加, 而右心室/体重比在 TAC (黑酒吧) 与假手术 (白酒吧) 动物的比例保持相似。这证实了左心室肥厚, 在右心室没有改变, 并加强了我们的技术验证 (n=6-10 每组. ** 和 #60; 0.01, *** 和 #60; 0.001, *** 和 #60; 0.0001)。数据显示为平均± SEM (误差线)。<a href="//ecsource.jove.com/files/ftp_upload/56231/56231fig9v2large.jpg" target="_blank">请单击此处查看此图的较大版本.</a>
<img alt="Figure 10" class="xfigimg" src="/files/ftp_upload/56231/56231fig10.jpg"> 
图 10: BNP-mRNA 表达式. 脑钠蛋白 (BNP)、心房利钠蛋白 (ANP)、血管紧张素转换酶 (ACE)、胶原 1a1 (Col1a1) 和转化生长因子？ (TGFß) 的 mRNA 表达, 主动脉带状心肌肥厚的阳性对照研究 (黑酒吧)vs 假动物 (白吧) (每组 n=6) 在 D14。表达式被计算为 2(-ΔCt) , 其中校准器是 Gapdh 参考基因的 mRNA 水平。数据显示为平均± SEM (误差线)。* p & #60; 0.05, *** 和 #60; 0.01, ***p 和 #60; 0.001 与假组比较 (t-测试)。<a href="//ecsource.jove.com/files/ftp_upload/56231/56231fig10large.jpg" target="_blank">请单击此处查看此图的较大版本.</a>

Watch this Scientific Journal Video about Technique of Minimally Invasive Transverse Aortic Constriction in Mice for Induction of Left Ventricular Hypertrophy at JoVE.com

Technique of Minimally Invasive Transverse Aortic Constriction in Mice for Induction of Left Ventricular Hypertrophy

本议定书的目的是描述 step-by 微创的横向主动脉狭窄 (TAC) 技术的小鼠。通过消除气管插管和通气是常用的标准程序的强制性, 微创 TAC 简化了手术程序, 减少了对动物的压力。

小鼠横主动脉狭窄诱导左心室肥厚的技术研究

Transverse aortic constriction (TAC) in mice is one of the most commonly used surgical techniques for experimental investigation of pressure ...

technique-minimally-invasive-transverse-aortic-constriction-mice-for

Universidad San Sebastian

Research

JoVE Journal

Medicine

24.2K 次观看.  University of Zurich. 本议定书的目的是描述 step-by 微创的横向主动脉狭窄 (TAC) 技术的小鼠。通过消除气管插管和通气是常用的标准程序的强制性, 微创 TAC 简化了手术程序, 减少了对动物的压力。

视频: Technique of Minimally Invasive Transverse Aortic Constriction in Mice for Induction of Left Ventricular Hypertrophy

Ascending Aortic Constriction in Rats for Creation of Pressure Overload Cardiac Hypertrophy Model

Ascending aortic constriction is the most common and successful surgical model for creating pressure overload induced cardiac hypertrophy and heart failure. Here, we describe a detailed surgical procedure for creating pressure overload and cardiac hypertrophy in rats by constriction of the ascending aorta using a small metallic clip. After anesthesia, the trachea is intubated by inserting a cannula through a half way incision made between two cartilage rings of trachea. Then a skin incision is made at the level of the second intercostal space on the left chest wall and muscle layers are cleared to locate the ascending portion of aorta. The ascending aorta is constricted to 50&#8211;60% of its original diameter by application of a small sized titanium clip. Following aortic constriction, the second and third ribs are approximated with prolene sutures. The tracheal cannula is removed once spontaneous breathing was re-established. The animal is allowed to recover on the heating pad by gradually lowering anesthesia. The intensity of pressure overload created by constriction of the ascending aorta is determined by recording the pressure gradient using trans-thoracic two dimensional Doppler-echocardiography. Overall this protocol is useful to study the remodeling events and contractile properties of the heart during the gradual onset and progression from compensated cardiac hypertrophy to heart failure stage.

We describe a stepwise procedure for creating pressure overload and left ventricular hypertrophy in Wistar rats by constriction of the ascending aorta using a small metallic clip. This model is extensively used for studying remodeling changes during cardiac hypertrophy and for identifying and evaluating strategies for regression of such changes.

大鼠建立压力超负荷心肌肥厚模型在升主动脉缩窄

Ascending aortic constriction is the most common and successful surgical model for creating pressure overload induced cardiac hypertrophy and heart ...

科研

医学

Transverse Aortic Constriction in Mice

Transverse aortic constriction (TAC) in the mouse is a commonly used experimental model for pressure overload-induced cardiac hypertrophy and heart failure.1 TAC initially leads to compensated hypertrophy of the heart, which often is associated with a temporary enhancement of cardiac contractility. Over time, however, the response to the chronic hemodynamic overload becomes maladaptive, resulting in cardiac dilatation and heart failure.2 The murine TAC model was first validated by Rockman et al.1, and has since been extensively used as a valuable tool to mimic human cardiovascular diseases and elucidate fundamental signaling processes involved in the cardiac hypertrophic response and heart failure development. When compared to other experimental models of heart failure, such as complete occlusion of the left anterior descending (LAD) coronary artery, TAC provides a more reproducible model of cardiac hypertrophy and a more gradual time course in the development of heart failure. Here, we describe a step-by-step procedure to perform surgical TAC in mice. To determine the level of pressure overload produced by the aortic ligation, a high frequency Doppler probe is used to measure the ratio between blood flow velocities in the right and left carotid arteries.3, 4 With surgical survival rates of 80-90%, transverse aortic banding is an effective technique of inducing left ventricular hypertrophy and heart failure in mice.

Transverse aortic constriction (TAC) in the mouse is a commonly used experimental model to study mechanisms underlying cardiac hypertrophy and heart failure development. Here, we describe procedures to constrict the aorta to create a reproducible degree of cardiac hypertrophy in mice.

在小鼠体内的横向主动脉缩窄

Transverse aortic constriction (TAC) in the mouse is a commonly used experimental model for pressure overload-induced cardiac hypertrophy and heart ...

生物学

Assessment of Cardiac Morphological and Functional Changes in Mouse Model of Transverse Aortic Constriction by Echocardiographic Imaging

Transverse aortic constriction (TAC) in mice has been used as a valuable model to study mechanisms of cardiac hypertrophy and heart failure1. A reliable noninvasive method is essential to assess real-time cardiac morphological and functional changes in animal models of heart disease. Transthoracic echocardiography represents an important tool for noninvasive assessment of cardiac structure and function2. Here we used a high-resolution ultrasound imaging system to monitor myocardial remodeling and heart failure progression over time in a mouse model of TAC. B-mode, M-mode, and Doppler imaging were used to precisely assess cardiac hypertrophy, ventricular dilatation, and functional deterioration in mice following TAC. Color and pulse wave (PW) Doppler imaging was used to noninvasively measure pressure gradient across the aortic constriction created by TAC and to assess transmitral blood flow in mice. Thus transthoracic echocardiographic imaging provides comprehensive noninvasive measurements of cardiac dimensions and function in mouse models of heart disease.

The goal of this protocol is to noninvasively assess cardiac structural and functional changes in a mouse model of heart disease created by transverse aortic constriction, using B- and M-mode echocardiography and color/pulse wave Doppler imaging.

在横向主动脉缩窄的小鼠模型和心脏形态学评估功能改变通过超声心动图成像

Transverse aortic constriction (TAC) in mice has been used as a valuable model to study mechanisms of cardiac hypertrophy and heart failure1. A reliable ...

Minimally Invasive Transverse Aortic Constriction in Mice

Minimally invasive transverse aortic constriction (MTAC) is a more desirable method for the constriction of the transverse aorta in mice than standard open-chest transverse aortic constriction (TAC). Although transverse aortic constriction is a highly functional method for the induction of high pressure in the left ventricle, it is a more difficult and lengthy procedure due to its use of artificial ventilation with tracheal intubation. TAC is oftentimes also less survivable, as the newer method, MTAC, neither requires the cutting of the ribs and intercostal muscles nor tracheal intubation with a ventilation setup. In MTAC, as opposed to a thoracotomy to access to the chest cavity, the aortic arch is reached through a midline incision in the anterior neck. The thyroid is pulled back to reveal the sternal notch. The sternum is subsequently cut down to the second rib level, and the aortic arch is reached simply by separating the connective tissues and thymus. From there, a suture can be wrapped around the arch and tied with a spacer, and then the sternal cut and skin can be closed. MTAC is a much faster and less invasive way to induce left ventricular hypertension and enables the possibility for high-throughput studies. The success of the constriction can be verified using high-frequency trans-thoracic echocardiography, particularly color Doppler and pulsed-wave Doppler, to determine the flow velocities of the aortic arch and left and right carotid arteries, the dimension of the blood vessels, and the left ventricular function and morphology. A successful constriction will also trigger significant histopathological changes, such as cardiac muscle cell hypertrophy with interstitial and perivascular fibrosis. Here, the procedure of MTAC is described, demonstrating how the resulting flow changes in the carotid arteries can be examined with echocardiography, gross morphology, and histopathological changes in the heart.

Minimally invasive transverse aortic constriction (MTAC) conserves the essentials of regular transverse aortic constriction (TAC) while eliminating the use of a ventilator with tracheal intubation. It proves to be a highly desirable method for high-throughput studies on left ventricular overload, particularly in translational studies.

微创横向主动脉缩窄小鼠

Minimally invasive transverse aortic constriction (MTAC) is a more desirable method for the constriction of the transverse aorta in mice than standard ...

A Closed-chest Model to Induce Transverse Aortic Constriction in Mice

Research on cardiac hypertrophy and heart failure is frequently based on pressure overload mouse models induced by TAC. The standard procedure is to perform a partial thoracotomy to visualize the transverse aortic arch. However, the surgical trauma caused by the thoracotomy in open-chest models changes the respiratory physiology as the ribs are dissected and left unattached after chest closure. To prevent this, we established a minimally invasive, closed chest approach via lateral thoracotomy. Herein we approach the aortic arch via the 2nd intercostal space without entering the chest cavities, leaving the mouse with a less traumatic injury to recover from. We perform this operation using standard laboratory settings for open chest TAC procedures with equal survival rates. Apart from maintaining physiological breathing patterns due to the closed chest approach, the mice seem to benefit by showing rapid recovery, as the less invasive technique appears to facilitate a fast healing process and to reduce immune response after trauma.

Here, we present a protocol of Transverse Aortic constriction (TAC) via a lateral thoracotomy. This technique is a minimally invasive, closed chest surgical procedure aiming to simulate pressure overload and heart failure in mice utilizing standard TAC laboratory settings.

诱导小鼠横向主动脉狭窄的闭合胸模型

Research on cardiac hypertrophy and heart failure is frequently based on pressure overload mouse models induced by TAC. The standard procedure is to ...

Studying Left Ventricular Reverse Remodeling by Aortic Debanding in Rodents

To better understand the left ventricular (LV) reverse remodeling (RR), we describe a rodent model wherein, after aortic banding-induced LV remodeling, mice undergo RR upon removal of the aortic constriction. In this paper, we describe a step-by-step procedure to perform a minimally invasive surgical aortic debanding in mice. Echocardiography was subsequently used to assess the degree of cardiac hypertrophy and dysfunction during LV remodeling and RR and to determine the best timing for aortic debanding. At the end of the protocol, terminal hemodynamic evaluation of the cardiac function was conducted, and samples were collected for histological studies. We showed that debanding is associated with surgical survival rates of 70-80%. Moreover, two weeks after debanding, the significant reduction of ventricular afterload triggers the regression of ventricular hypertrophy (~20%) and fibrosis (~26%), recovery of diastolic dysfunction as assessed by the normalization of left ventricular filling and end-diastolic pressures (E/e&#39;&#160;and LVEDP). Aortic debanding is a useful experimental model to study LV RR in rodents. The extent of myocardial recovery is variable between subjects, therefore, mimicking the diversity of RR that occurs in the clinical context, such as aortic valve replacement. We conclude that the aortic banding/debanding model represents a valuable tool to unravel novel insights into the mechanisms of RR, namely the regression of cardiac hypertrophy and the recovery of diastolic dysfunction.

Here we describe a step-by-step protocol of surgical aorta debanding in the well-established mice model of aortic-constriction. This procedure not only allows studying the mechanisms underlying the left ventricular reverse remodeling and regression of hypertrophy but also to test novel therapeutic options that might accelerate myocardial recovery.

研究左心室反向改造由主动脉脱体在啮齿动物

To better understand the left ventricular (LV) reverse remodeling (RR), we describe a rodent model wherein, after aortic banding-induced LV remodeling, ...

Operating Transverse Aortic Constriction with Absorbable Suture to Obtain Transient Myocardial Hypertrophy

Based on twice transverse aortic constrictions (TACs) in mice, it is proved that myocardial hypertrophic preconditioning (MHP) could attenuate cardiomyocyte hypertrophy and slow down progression to heart failure. For novices, however, the MHP model is usually quite difficult to establish because of the technical obstacles in ventilator operation, opening the chest repeatedly, and bleeding caused by debanding. To facilitate this model, to increase the surgical success rate and to reduce the incidence of bleeding, we switched to absorbable sutures for the first TAC combing with a ventilator-free technique. Using a 2-week absorbable suture, we demonstrated that this procedure could cause significant myocardial hypertrophy in 2 weeks; and 4 weeks after surgery, myocardial hypertrophy was almost completely regressed to the baseline. Using this protocol, the operators could master the MHP model easily with a lower operation mortality.

This protocol presents an improved method to obtain transient myocardial hypertrophy with absorbable suture, simulating left ventricular hypertrophy decrease after removing pressure overload. It could be valuable for the studies on myocardial hypertrophic preconditioning.

操作横向主动脉收缩与可吸收的缝合获得瞬时心肌萎缩

Based on twice transverse aortic constrictions (TACs) in mice, it is proved that myocardial hypertrophic preconditioning (MHP) could attenuate ...

A Modified Technique for Transverse Aortic Constriction in Mice

Transverse aortic constriction (TAC) is a frequently used surgery in research regarding heart failure and cardiac hypertrophy based on the formation of pressure overload in mouse models. The main challenge of this procedure is to clearly visualize the transverse aortic arch and precisely band the target vessel. Classical approaches perform a partial thoracotomy to expose the transverse aortic arch. However, it is an open-chest model that causes a rather large surgical trauma and requires a ventilator during the surgery. To prevent unnecessary trauma and simplify the operating procedure, the aortic arch is approached via the proximal proportion of the sternum, reaching and binding the target vessel using a small self-made retractor that contains a snare. This procedure can be conducted without entering the pleura cavity and does not need a ventilator or microsurgical operation, which leaves the mice with physiological breathing patterns, simplifies the procedure, and significantly reduces operation time. Due to the less invasive approach and less operation time, mice can undergo fewer stress reactions and recover rapidly.

The present protocol describes a modified and simplified technique with a minimally invasive transverse aortic constriction (TAC) procedure using a self-made retractor. This procedure can be conducted without a ventilator or microscope and introduces pressure overload, eventually leading to cardiac hypertrophy or heart failure.

一种改良的小鼠横主动脉收缩技术

Transverse aortic constriction (TAC) is a frequently used surgery in research regarding heart failure and cardiac hypertrophy based on the formation of ...

Transverse Aortic Arch Banding in Mouse Model: A Minimally Invasive Technique in Mice for Induction of High Pressure in the Left Ventricle

Source: Zaw, A. M. et al. <a href="https://www.jove.com/video/55293" target="_blank">Minimally Invasive Transverse Aortic Constriction in Mice</a>. J. Vis. Exp. (2017)
In this video, we demonstrate an experimental protocol for minimally invasive transverse aortic constriction in mice to examine pressure overload-induced heart failure.

小鼠横主动脉弓条带模型:小鼠诱导左心室高压的微创技术

Source: Zaw, A. M. et al. Minimally Invasive Transverse Aortic Constriction in Mice. J. Vis. Exp. (2017)
In this video, we demonstrate an experimental ...

JoVE Encyclopedia of Experiments

Rodent Models

Adult

Surgical Aortic Debanding: A Procedure to Study Left Ventricular Reverse Remodeling in Murine Model of Aortic Constriction

Source: Goncalves-Rodrigues, P. et al. <a href="https://www.jove.com/video/60036" target="_blank">Studying Left Ventricular Reverse Remodeling by Aortic Debanding in Rodents</a>. J. Vis. Exp. (2021)
This video demonstrates the technique of aortic debanding, which can lead to reverse remodeling to limit the progression towards heart failure in a murine model. Post debanding, a significant reduction of ventricular afterload triggers regression of ventricular hypertrophy and fibrosis, causing a recovery in cardiac function.