This work was supported by a National Heart, Lung, and Blood Institute grant 1RO1-HL-105296-01 (to D.E. Berkowitz) and an Australian Research Council Grant DP110101134 (to A. Avolio).

该协议已经被批准的机构动物护理和使用委员会在约翰·霍普金斯大学。 1.准备的解决方案，材料和动物<ol><li>制备300微克/毫升的溶液去氧肾上腺素（PE）和300微克/毫升溶液硝普钠（SNP）在0.9％盐水中。通过混合1毫升1000单位/毫升肝素到10ml的0.9％盐水中制备单独的肝素盐溶液。 注:药物应该是在室温下在使用前。 </li><li>使两个30国祥½"注射针头和PE 10聚乙烯管导管静脉输注药物。为了使导管，插入一个针入软管的一端。除去其它皮下注射针的针部并插入钝端插入管的另一端。附加导管以1ml注射器，并填写与肝素盐溶液导管。 </li><li>鼠标放置在麻醉诱导室Containing 2-2.5％异氟烷在100％的氧。离开鼠标的感应室，直到它反应迟钝对外界的刺激。 </li><li>从感应腔室中取出小鼠，并将其放置在加热的心电图（ECG）垫。维持动物在2％异氟醚。 </li><li>适用兽医软膏或盐溶液到该动物的眼睛，以防止在操作过程中干。 </li></ol> 2.插入导管的尾静脉<ol><li>由于尾静脉上尾部的两侧横向地位于，放置在其一侧的动物更好的访问。确保鼠标到ECG垫带。确保动物保​​持温暖，以促进尾静脉的血管扩张。 </li><li>使用一块硅橡胶管作为止血带，扎止血带绕尾的基部。扎止血带不够紧崩的静脉，但不足以切断动脉循环。经过2-3分钟，静脉应凸出而变得更加明显。 </li><li>轻轻一拉绷紧的尾巴。弯曲的尾部在用一只手的角度，并保持平行的针，以与其它的尾巴。皮尔斯其中尾部穿过皮肤进入静脉弯曲针。血液会推回到所述导管如果针插入静脉。 </li><li>放置一滴组织胶，其中所述针被插入到固定导管。取出止血带，并注入生理盐水少许阻力确认通畅。 </li></ol>通过股动脉插入3.血压导管<ol><li>将压力导管到30ml注射器充满蒸馏水和导管连接到压力控制部。泡导管在水中，接通电源后，30-45分钟期间设置和手术过程。 </li><li>将动物仰卧并用胶带它的爪子上的心电图垫。胸部和面积在股动脉申请脱毛霜。 <ol><li>等待3-5分钟，取出奶油和头发。日oroughly从胸部取出头发，防止超声波期间的文物。无论擦拭胸部和后肢地区用湿垫去除多余脱毛霜。 </li></ol></li><li>使用细剪刀，做一个切口在上述股动脉的位置的皮肤。解剖通过皮下脂肪组织，露出股动脉。股动脉被部分覆盖的腹部。用止血移动腹部而去。保持组织湿润通过用湿纱布覆盖，或替代地周期性地滴盐水超过它，以防止组织干燥。 </li><li>用细镊子，从动脉静脉丛分离的神经了。通过围绕动脉静脉丛鞘轻轻刺穿到动脉从静脉分离。通过动脉周围1缝合在近端和放置两个缝合在远端。 </li><li>牢固地结最远侧缝合停止远端的血流。用止血拉Proximal缝合暂时停止血流进入股动脉。使用microscissors做一个小切口插入股动脉。使近远端结切口。 </li><li>使用压力控制装置上的校准设置的数据采集软件校准到导管。切换压力控制单元回读换能器和平衡压力导管，以使导管输出0毫米汞柱，在装满水的注射器。 </li><li>插入导管进入股动脉。用一只手打开所述切口用细镊子和插入导管头部与另一方面的动脉。 <ol><li>结的导管导线围在中间缝合来固定导管进入动脉。放松近侧缝合并向前推进所述导管进入腹主动脉。结近侧缝合线，以进一步固定导管，并防止出血。 </li></ol></li><li>小心地移动心电图垫鼠标，压力导管和生理盐水syringe到超声成像阶段。血压导管连接到压力控制部。放置在注射器泵的盐水注射器。允许动物和导管，以平衡20分钟。 </li></ol> 4.测量主动脉内径过血压的范围<ol><li>减少异氟烷1.5％。纵向形象化升主动脉上使用长轴视图B模式。安装该换能器上的导轨系统，以使相同的视图被保持在实验的持续时间。 </li><li>在超声大型机，将M型光标置于主动脉的部分进行跟踪。跟踪随着使用M型心动周期主动脉直径变化。 </li><li>在注射器给PE溶液改变盐水并将注射器放入注射器泵。 <ol><li>记录M型在基线主动脉的压力。开始输注360微克/公斤/分钟，并注入1分钟主动脉压，达到平台期。对于25克鼠标，该做的本身相当于30微升/分钟。 </li><li>记录的M模​​式，然后停止输液，并等待2分钟，血压返回到基线。 </li></ol></li><li>低输注速率至240微克/千克/分钟。对于一个25克鼠标，该剂量相当于20微升/分钟。开始输液，注入1分钟的血压平稳，并记录M-模式。停止输液，并等待2分钟，血压返回到基线。 </li><li> 120微克/公斤，重复步骤4.4 /分钟PE（10微升/分钟的25克鼠标）。 </li><li>取代的PE用盐水和注入的盐水在用于360微克/公斤/分钟输注的速率（30微升/分钟为一个25克小鼠）。注入为2-3分钟，直到进一步输注不产生增加主动脉压和压力返回到基线。等待5分钟，使血压在基线稳定。 </li><li>与SNP更换盐水。 <ol><li>记录M型在基线主动脉的压力。开始输注在240微克/千克/分钟（20微升/分钟25克小鼠）和注入​​，持续1分钟。当主动脉压力达到一个高原，记录M-模式。停止输液，等待2分钟为血压回到基线。 </li></ol></li><li>低输注速率至120微克/千克/分钟（10微升/分钟25克鼠标）。开始输液，注入1分钟的血压平稳，并记录M-模式。停止输注并等待2分钟为血压回到基线。 </li><li> 60微克/千克，重复步骤4.8 / min的SNP（5微升/分钟25克鼠标）。 </li></ol> 5.终止实验<ol><li>安乐死的动物，增加异氟烷至4％。当呼吸速度减慢，一般在1-2分钟，通过削减用剪刀打开胸腔，暴露心脏的胸骨。 </li><li>掌握中等钳心脏和削减在升主动脉，用剪刀从体内切除它。 </li></ol>

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取直径测量在几个压力增量在宽范围的压力值是必需的压径关系精确表征。可药理学诱导的上下压力极限可通过实验组变化，但是理想的范围是约25毫米汞柱至125毫米汞柱舒张压和50毫米汞柱至200毫米汞柱收缩。 360微克/千克/分钟PE和240微克/千克的剂量/分钟的SNP通常引起的压力范围的限制。然而，剂量的PE的可提高到480微克/千克/分钟和SNP至360微克/千克/分钟，以验证该限制已经达到。聚乙烯和SNP的工作浓度可以降低到实现更精细的压力增量。随着直径将与主动脉压力变化，诱发动物和实验组之间相同的压力值是不重要的。 静脉和动脉插管可以在其他位置具有相同的OU进行tcomes。尾静脉插管可以是由于尾静脉小尺寸有挑战性。此外，尾静脉不在深色小鼠容易看见。股静脉可以插管作为替代。这条路线可能会更容易，因为股静脉是更容易获得。对于压力的​​导管插入，除了股动脉，导管可通过颈动脉插入。股动脉优选在颈动脉，但是，因为胸部区域保持完整的超声成像。股动脉插管可以更困难，因为股动脉越小。使用1.2˚F导管和引入所述导管在腹腔下方的近端股动脉将方便插管过程。放置几滴像利多卡因血管扩张剂的上股动脉或使用导管导引器还可以帮助扩大容器，以方便的导管插入。压力导管应处理，并使用根据制造商的说明。 主动脉内导管的位置并不需要是动物之间是一致的主动脉内的压力降是微不足道的。然而，将所述导管在腹主动脉可能更好，以尽量减少与胸主动脉的超声成像的干扰。一些超声大型机可以记录压力实时与M型跟踪，从而给出为对M模式测量每直径的压力的测量。不幸的是，因为那里的直径测量的位置不相同的位置，其中压力被记录时，记录在导管中的压力，并在升主动脉中的实际压力之间的滞后存在。其结果是，只有最大和最小直径的测量可用于数据分析。 该方法的主要限制是由在主动脉和移出ö移引入在测量的不确定性f显示心动周期中的超声平面。运动引入误差是通用于所有基于成像研究，包括MRI和CT。补偿策略包括使用的解剖特点来参考帧的运动18移位和数据处理过程中实现的。作为运动补偿的软件是不容易获得的，研究者必须警惕调整探针的位置来跟踪此消彼长的主动脉血压上升的位置和降低。直径测量也应当通过主动脉的中心截取。然而，确定的M型记录位置是否被通过中心可能难以判断超声波图像上，尤其是与主动脉换档位置。通过这些限制体现在分散的数据的程度， 如图6引入，如明显的不确定性。获得ascendi的纵向轴线的横截面的图像，而不是纳克主动脉可能是一个解决办法。不过，获得这一观点有时会更具挑战性，所产生的M型跟踪可以就不太清楚了。从B模式图像的横截面周长可被测量，而不是从对M模式图像的直径。然而，确定当最大和最小圆周已经实现将由B模式帧速率是有限的，可能更难以判断比在M模式。 制造的压径积多个测量和增加实验组的大小可以提高数据的准确性。的压径数据可以从沿着胸部的几个位置被收集。该协议将首先进行与探针放置在一个位置上的胸部。主动脉然后将可视化与探针放在另一个位置和反复的协议。 用来调制血压血管活性剂可能会影响主动脉平滑肌MUSC乐调，这反过来会影响刚度。然而，操纵通过静脉回流主动脉压力已经显示出产生在侵入性测量PWV作为药理操纵大鼠类似的变化。这些结果表明，输注血管活性药物作用主要是外周阻力动脉和不显著影响主动脉平滑肌音19。 该协议可以在大鼠进行了一些小的改动。之前申请脱毛霜胸口剃光。市售27国祥½"导管用于药物注射。用来调制主动脉压的药物的剂量是40次，80次和PE和40，80的120微克/千克/分钟，和120微克/千克/ SNP的分除了升主动脉，主动脉刚度区域差异可与该协议来确定。通过这种方法测得的区域刚度会比更精确了PWV的测量ARE从一个位置作为反对两个位置的PWV。然而，沿着主动脉的区域，可以使用这种技术来测量被限制为那些可以通过超声波来可视化。 弹性模量也可以从若能够获得壁厚测量通过该方法收集的数据计算出来的。准确的小鼠主动脉的体内测量是通过电流超声技术的分辨率极限的限制。超声技术的未来改进可以使体内的壁厚测量更可行。作为替代，厚度测量可以进行体外 。压力肌动描记将提供最精确的测量，因为厚度可以在每个压力增量来测量。

增加动脉硬化是心血管疾病的一个标志。老化1，吸烟2，糖尿病3，高脂血症4，和心血管疾病的其它危险因素已被证明能增加主动脉刚度。流行病学研究已经进一步证明主动脉刚度冠状动脉心脏疾病和中风发病的一个有力的独立预测因子，以及心血管事件和死亡率5-8的发生。由于增加了主动脉僵硬的临床和公共卫生意义的，目前的研究重点是了解背后血管硬化的发生和发展的机制。因此极大的兴趣存在于显影在心血管疾病的实验模型中血管硬度的准确测量。 的材料的硬度的特征可在于它的应力 - 应变关系和定量为弹性模汗国。线性弹性材料可逆变形和应力比例提高应变。主动脉和大动脉是非线性弹性体:拉伸时，在动脉的刚度不会保持恒定，但具有腹胀的程度增大。这种非线性在大动脉的机械性能是由于在承载元件，即弹性蛋白和胶原蛋白，构成容器壁的不同的刚度特性。弹性是0.6兆帕的弹性模量高可扩展性。相比较而言，胶原蛋白是非常僵硬的为1GPa 9的弹性模量。在较低的应变值显示出由主动脉初始刚度是由于弹性而高刚度显示出在高应变值是由于胶原蛋白。载荷从弹性蛋白胶原转移作为容器distends和负载的该区域转移是其中血管系统进行操作。因此，在生理压力，动脉硬度取决于这两个弹性蛋白和胶原蛋白10的贡献。 的分布和弹性蛋白和胶原蛋白的取向由动脉壁内层而有所不同。在媒体中，弹性蛋白，胶原和平滑肌细胞被捆绑成紧螺旋是层状同心。这种布置允许在动脉抵抗高负荷在圆周方向上。外膜主要是胶原蛋白与小弹性蛋白和胶原纤维被组织在一个网状方式。这些胶原纤维波浪在无应力状态，理顺随着负载的增加。刚度随着胶原纤维理顺，从而防止动脉从过度拉伸和破裂。由于胶原纤维的结构组织和不同取向的，动脉是各向异性:表现出刚度取决于如果容器被纵向或周向11拉伸体内 stiffnes。因此s是主动脉的纵向和周向刚性的复合材料。 动脉僵硬度一般是量化体内的合规性和脉搏波速度（PWV）。动脉顺应性被定义为C =ΔD/ΔP其中ΔD是变化的直径和ΔP是在压力的相应变化。达标值越低表明硬的船只。遵从动脉的压力 - 维关系计算，因此直接测量刚度。如刚度在脉管12均匀地散发非，依从应在每个受试者相同/相似的位置进行测量，以使实验组之间进行有意义的比较。 合规性和弹性模量之间的差别在于，弹性模量被归一化的材料的尺寸。因此，合规体现了结构刚度，而弹性模量裁判脉冲编码材料刚度。随着衰老，动脉壁的厚度的增加和弹性蛋白/胶原蛋白的比例降低，所以两者的结构刚度和材料刚性更大。 相比于合规性，PWV是动脉僵硬度的间接测量。 PWV是在哪一个压力脉冲行进沿着动脉的长度，并且由血管壁的性质的影响的速度。所述Moens-Korteweg等式用于PWV和弹性模量之间的关系进行建模:PWV 2 = E的h /（2ρR）其中E是增量弹性模量，h是壁厚，ρ是血液粘度，并且r是容器的半径。因此，一个更高的PWV值提出了一个更硬的容器。 合规性和弹性模量，可以对船只的切除部分实验体外测量。以确定是否符合，将容器段被安装在压力肌动描记13,14。容器内的压力增加逐步与日Ë产生变化直径使用视频显微镜跟踪。遵从压径数据确定。增量弹性系数可通过拉伸试验来测量。在这些实验中，将容器拉开分步进行和力 - 位移数据被收集，直到容器环断裂。应力和应变值可以被计算并绘制，以确定增量弹性模量。这些离体方法可用于评估的变化，影响刚度的被动特性。 在体内 ，除了墙壁内容，血管刚度由平滑肌张力和血压13,15,16动态影响。 PWV是最广泛使用的方法，用于测量在实验模型中体内主动脉刚度。 PWV可以使用无创多普勒超声或压平眼压计17来决定。压力脉冲的测量是在两个不同的位置和所需的时间遍历的距离的脉冲的脉波速度。因为PWV测量在主动脉的长度，它是刚性的平均值。大动脉是非线性弹性，所以刚度，因此PWV将随动脉压。因此，一个更高的PWV值可以从增加的僵硬或高压出现。 PWV值因此必须标准化为血压得出关于所述容器的刚性的结论。结合了血压与血管壁的被动性和血管活性介质的该改变色调会产生动脉僵硬的生理相关指数的影响的影响的测量方法。这种方法是通过测量PWV侵入使用的导管与在一个固定的距离13分开的两个压力传感器来实现。这种双重压力导管插入主动脉和血管活性药物，如苯福林或硝普钠，通过静脉内输注静脉导管，以提高和降低血压。 这个协议描述了一种方法，以确定从在小鼠模型中它的压力维关系体内主动脉刚度。这种方法有几个优点在侵入PWV测量。刚性指数，如合规性，可以从由该过程所收集的压力维数据来计算。此外，该技术允许对本地主动脉刚度测量，因为刚度是从单个位置测量。这种方法是在测量升主动脉刚度作为该区域的长度短使得PWV测量难以获得特别有用。存在的研究兴趣特别是在升主动脉，因为其机械性能影响冠脉循环的灌注和血管功能障碍的心脏反应。 以测量的主动脉体内的压径关系</e米&gt;，升主动脉被可视化和它的直径由超声成像测量。主动脉血压与一个压力导管侵入测量。血压是通过血管活性药物静脉输注增量改变。苯福林收缩血管，并用于提高主动脉的压力。硝普钠扩张血管和用于降低主动脉的压力。收缩压和舒张压主动脉直径及相应的主动脉压力测量每个压力增量。遵守可以从收集到的压径数据进行计算。

<table><tbody><tr><td>Equipment</td><td></td><td></td><td></td></tr><tr><td>High-resolution ultrasound machine</td><td>Visual Sonics</td><td>Vevo2100</td><td></td></tr><tr><td>13-24 MHz transducer</td><td>Visual Sonics</td><td>MS250</td><td>Used for imaging rats</td></tr><tr><td>22-55 MHz transducer</td><td>Visual Sonics</td><td>MS550D</td><td>Used for imaging mice</td></tr><tr><td>Imaging Station</td><td>Visual Sonics</td><td>Imagine Station 1</td><td></td></tr><tr><td>1.2F Pressure catheter</td><td>Transonic</td><td>FTH-1211B-0018</td><td></td></tr><tr><td>SP200 pressure control unit</td><td>Transonic</td><td>FFS-095-DP01</td><td></td></tr><tr><td>Standard Infusion Only Harvard Pump 11 Plus syringe pump</td><td>Harvard Apparatus</td><td>702208</td><td></td></tr><tr><td>Isoflurane vaporizer</td><td>VetEquip</td><td>911103</td><td></td></tr><tr><td>Induction chamber</td><td>VetEquip</td><td>941443</td><td></td></tr><tr><td>100% O&lt;sub&gt;2&lt;/sub&gt;</td><td>Airgas</td><td>OX USP200</td><td></td></tr><tr><td>Single Stage Brass 0-50 psi General Purpose Cylinder Regulator CGA540</td><td>Airgas</td><td>Y11215B540</td><td></td></tr><tr><td>Stereo Boom Stand Microscope</td><td>National Optical</td><td>420-BMSQ</td><td></td></tr><tr><td>Fiber optic illuminator &amp;amp; light pipe</td><td>Cole Palmer</td><td>EW-41500-50</td><td></td></tr><tr><td>Supplies</td><td></td><td></td><td></td></tr><tr><td>30G x 1/2&quot; BD PrecisionGlide Needle</td><td>BD</td><td>305106</td><td>For tail vein cannulation in mice</td></tr><tr><td>Polyethylene Tubing PE10</td><td>Becton Dickinson</td><td>427401</td><td>For tail vein cannulation in mice</td></tr><tr><td>27G x 1/2&quot; Surfloe winged infusion set</td><td>Terumo</td><td>SV*27EL</td><td>For tail vein cannulation in rats</td></tr><tr><td>Signa Gel Electrode Gel</td><td>Parker</td><td>15-25</td><td>Use for ECG recording</td></tr><tr><td>Aquasonic Clear Ultrasound Gel</td><td>Parker</td><td>03-08</td><td>Use for ultrasound</td></tr><tr><td>1ml Sub-Q Syringes, 26G x 5/8&quot;</td><td>BD</td><td>309597</td><td></td></tr><tr><td>Nair</td><td>Nair</td><td></td><td>Depilatory cream</td></tr><tr><td>Histoacryl</td><td>TissueSeal</td><td>TS1050071FP</td><td>Tissue glue</td></tr><tr><td>Braided Silk Suture 6-0</td><td>Teleflex</td><td>104-S</td><td></td></tr><tr><td>Dumostar P55 fine forceps</td><td>Roboz</td><td>RS-4984</td><td></td></tr><tr><td>Microscissors</td><td>WPI</td><td>501839</td><td></td></tr><tr><td>Fine scissors</td><td>FST</td><td>14060-11</td><td></td></tr><tr><td>Medium forceps</td><td>Ted Pella</td><td>5665</td><td></td></tr><tr><td>Hemostatic forceps</td><td>Roboz</td><td>RS-7131</td><td></td></tr><tr><td>Non-sterile cotton gauze sponge</td><td>Fisherbrand</td><td>22-362-178</td><td></td></tr><tr><td>Cotton tipped applicators</td><td>Oritan</td><td>803-WC</td><td></td></tr><tr><td>Label tape</td><td>Fisherbrand</td><td>15-901-20</td><td></td></tr><tr><td>Drugs</td><td></td><td></td><td></td></tr><tr><td>Sodium chloride</td><td>Sigma Aldrich</td><td>S7653</td><td></td></tr><tr><td>R-Phenylephrine hydrochloride</td><td>Sigma Aldrich</td><td>P6126</td><td></td></tr><tr><td>Sodium nitroprusside dihydrate</td><td>Sigma Aldrich</td><td>71778</td><td></td></tr><tr><td>Software</td><td></td><td></td><td></td></tr><tr><td>Prism</td><td>GraphPad</td><td></td><td></td></tr><tr><td>Excel</td><td>Microsoft</td><td></td><td></td></tr></tbody></table>

measuring ascending aortic stiffness in vivo in mice using ultrasound

我们提出了一个协议，用于测量体内主动脉刚度利用高分辨率超声波成像的小鼠。主动脉直径由超声测量和主动脉血压与固态压力导管侵入测量。血压升高然后通过血管活性药物苯肾上腺素和硝普钠静滴增量降低。主动脉直径测量为每个压力步骤来表征升主动脉的压径的关系。从压径关系导出硬度指数可以从收集到的数据来计算。动脉顺应性的计算本协议描述。 这种技术可以用于研究底层与心血管疾病和衰老相关的增加的主动脉僵硬的机制。该技术生产的刚度相比体外的生理相关度量方法，因为pH值主动脉刚度ysiological影响在测量被并入。该技术的主要限制是在心动周期期间从主动脉的运动引入的测量误差。这种运动可以通过调整探针与主动脉运动的位置以及制备的主动脉压径关系的多个测量结果和扩大实验组大小进行补偿。

左心室和升主动脉的纵向图像心脏期间捕获在B模式，如图1中所示。可选择地，可以得到只在主动脉的纵向图像，如在图2中 。主动脉壁的运动周期出现在M型两条白线， 如图3，主动脉内腔中的行之间的区域。主动脉压力通过输注血管活性药物调制。 PE引起的主动脉压力， 如图4A所示 ，和SNP降低压力， 如图4B所示 。 M型记录时血压高原，输注开始后1分钟。主动脉压力逐渐改变通过改变施用的药物的剂量， 如图5。药物的剂量是通过输注的速率来控制。所有药物的剂量是在微克/公斤/分钟。最大和最小直径从吨测量他M模式， 如图3所示。这些直径对应于收缩压和记录由压力导管舒张主动脉的压力。 的3个心动周期的收缩和舒张的直径和压力值在基线和在各PE和SNP剂量被测量。在一种药物剂量的直径三次测量之间的标准偏差的范围为0.01mm至0.04毫米。主动脉直径可以绘制针对其相应的主动脉压来说明压径的关系， 如图6A所示 。 这些压径值被用来计算主动脉柔顺性。动脉顺应性的计算方法 C =（D SYS - ð 直径 ）/（P SYS - P 直径 ）（1） 其中D SYS和D 直径是收缩和舒张的直径和P sys和 P 直径是收缩压和舒张压。合规性和平均主动脉压（MAP），计算在基线和在各PE和SNP剂量。是否合格，对绘制地图展示的压力依赖性刚​​度。因为主动脉的非线性弹性的行为，符合增加的MAP，如在图6B中看到的降低。 <img alt="图1" src="/files/ftp_upload/52200/52200fig1highres.jpg" /> 图1:在B模式升主动脉纵向视图直径测量是从升主动脉离开左心室的纵向图像。 LV:左心室; PA:肺动脉; AA:升主动脉。肺动脉可视取决于探针放置。主动脉直径测量远端的主动脉瓣。用于捕捉这一形象探头的频率为40 MHz的。 图2:在B模式升主动脉的备选视图升主动脉的特征更加显着，并在左心室和心脏壁是不太明显。 AA:升主动脉; LV:左心室。探头用来记录这个形象频率为40 MHz的。 <img alt="图3" src="/files/ftp_upload/52200/52200fig3highres.jpg" /> 图3:主动脉上可视化的M型主动脉直径从M模式图像进行测定。主动脉壁的运动显示为两个波浪线。在两条线之间的空间是在主动脉管腔。的3个心动周期的收缩和舒张主动脉直径从M模式中测得。在该图像记录由压力导管，ECG信号和呼吸循环主动脉压力上的M模式显示为红色，绿色和黄色。探头FREQUENCŸ用来记录这个形象是40 MHz和采集扫描速度是1200赫兹。 <img alt="图4" src="/files/ftp_upload/52200/52200fig4highres.jpg" /> 图4:更改主动脉压力增量主动脉压力由药物的剂量输注增量改变。药物剂量是由输注速率调制。所有剂量均在微克/公斤/分钟。 <img alt="图5" src="/files/ftp_upload/52200/52200fig5highres.jpg" /> 图5:调制用血管活性药物的主动脉压力主动脉压力增加与输注血管收缩剂去氧肾上腺素（PE），并与输注血管扩张剂硝普钠（SNP）的下降。主动脉压高原1分钟的药物输注开始后。主动脉直径的M型被记录在平台期。（A）示出的上升主动脉压用360微克/公斤/分钟的PE输注。（B）中示出了与240微克/公斤/分钟的SNP输注主动脉压的下降。时的输注开始和M模式被记录的时间标记上的痕迹。 &#160;主动脉压力由药物的剂量输注增量改变。药物剂量是由输注速率调制。所有剂量均在微克/公斤/分钟。 <img alt="图6" src="/files/ftp_upload/52200/52200fig6highres.jpg" /> 图6:直径与压力和法规遵从与平均动脉压地块主动脉直径能抵挡它的相应的主动脉压，显示压力直径的关系（A）绘制。遵守可以计算每个压力递增和作图的平均主动脉压（MAP），以显示主动脉刚性（B）的压力依赖性。

Watch this Scientific Journal Video about Measuring Ascending Aortic Stiffness In Vivo in Mice Using Ultrasound at JoVE.com

Measuring Ascending Aortic Stiffness In Vivo in Mice Using Ultrasound

We describe a technique for measuring aortic stiffness from its pressure-diameter relationship in vivo in mice. Aortic diameter is recorded by ultrasound and aortic pressure is measured invasively with a solid-state pressure catheter. Blood pressure is changed incrementally and the resulting diameter is measured.

测量升主动脉刚度<em&gt;在体内</em&gt;小鼠使用超声

We present a protocol for measuring in vivo aortic stiffness in mice using high-resolution ultrasound imaging. Aortic diameter is measured by ultrasound ...

measuring-ascending-aortic-stiffness-in-vivo-in-mice-using-ultrasound

Universidad de Cartagena UDEC

Research

JoVE Journal

Medicine

Measuring Ascending Aortic Stiffness In Vivo in Mice Using Ultrasound

15.9K 次观看.  Johns Hopkins University. We describe a technique for measuring aortic stiffness from its pressure-diameter relationship in vivo in mice. Aortic diameter is recorded by ultrasound and aortic pressure is measured invasively with a solid-state pressure catheter. Blood pressure is changed incrementally and the resulting diameter is measured.

视频: Measuring Ascending Aortic Stiffness In Vivo in Mice Using Ultrasound

Multi-modal Imaging of Angiogenesis in a Nude Rat Model of Breast Cancer Bone Metastasis Using Magnetic Resonance Imaging, Volumetric Computed Tomography and Ultrasound

Angiogenesis is an essential feature of cancer growth and metastasis formation. In bone metastasis, angiogenic factors are pivotal for tumor cell proliferation in the bone marrow cavity as well as for interaction of tumor and bone cells resulting in local bone destruction. Our aim was to develop a model of experimental bone metastasis that allows in vivo assessment of angiogenesis in skeletal lesions using non-invasive imaging techniques.
For this purpose, we injected 105 MDA-MB-231 human breast cancer cells into the superficial epigastric artery, which precludes the growth of metastases in body areas other than the respective hind leg1. Following 25-30 days after tumor cell inoculation, site-specific bone metastases develop, restricted to the distal femur, proximal tibia and proximal fibula1. Morphological and functional aspects of angiogenesis can be investigated longitudinally in bone metastases using magnetic resonance imaging (MRI), volumetric computed tomography (VCT) and ultrasound (US).
MRI displays morphologic information on the soft tissue part of bone metastases that is initially confined to the bone marrow cavity and subsequently exceeds cortical bone while progressing. Using dynamic contrast-enhanced MRI (DCE-MRI) functional data including regional blood volume, perfusion and vessel permeability can be obtained and quantified2-4. Bone destruction is captured in high resolution using morphological VCT imaging. Complementary to MRI findings, osteolytic lesions can be located adjacent to sites of intramedullary tumor growth. After contrast agent application, VCT angiography reveals the macrovessel architecture in bone metastases in high resolution, and DCE-VCT enables insight in the microcirculation of these lesions5,6. US is applicable to assess morphological and functional features from skeletal lesions due to local osteolysis of cortical bone. Using B-mode and Doppler techniques, structure and perfusion of the soft tissue metastases can be evaluated, respectively. DCE-US allows for real-time imaging of vascularization in bone metastases after injection of microbubbles7.
In conclusion, in a model of site-specific breast cancer bone metastases multi-modal imaging techniques including MRI, VCT and US offer complementary information on morphology and functional parameters of angiogenesis in these skeletal lesions.

In the pathogenesis of bone metastasis, angiogenesis is a crucial process and therefore represents a target for imaging and therapy. Here, we present a rat model of site-specific breast cancer bone metastasis and describe strategies to non-invasively image angiogenesis in vivo using magnetic resonance imaging, volumetric computed tomography and ultrasound.

利用磁共振成像，容积电脑断层扫描及超音波的裸鼠乳腺癌骨转移大鼠模型血管生成的多模态成像

Angiogenesis is an essential feature of cancer growth and metastasis formation. In bone metastasis, angiogenic factors are pivotal for tumor cell ...

科研

医学

Ultrasound Based Assessment of Coronary Artery Flow and Coronary Flow Reserve Using the Pressure Overload Model in Mice

Transthoracic Doppler echocardiography (TTDE) is a clinically useful, noninvasive tool for studying coronary artery flow velocity and coronary flow reserve (CFR) in humans. Reduced CFR is accompanied by marked intramyocardial and pericoronary fibrosis and is used as an indication of the severity of dysfunction. This study explores, step-by-step, the real-time changes measured in the coronary flow velocity, CFR and systolic to diastolic peak velocity (S/D) ratio in the setting of an aortic banding model in mice. By using a Doppler transthoracic imaging technique that yields reproducible and reliable data, the method assesses changes in flow in the septal coronary artery (SCA), for a period of over two weeks in mice, that previously either underwent aortic banding or thoracotomy. 
During imaging, hyperemia in all mice was induced by isoflurane, an anesthetic that increased coronary flow velocity when compared with resting flow. All images were acquired by a single imager. Two ratios, (1) CFR, the ratio between hyperemic and baseline flow velocities, and (2) systolic (S) to diastolic (D) flow were determined, using a proprietary software and by two independent observers. Importantly, the observed changes in coronary flow preceded LV dysfunction as evidenced by normal LV mass and fractional shortening (FS). 
The method was benchmarked against the current gold standard of coronary assessment, histopathology. The latter technique showed clear pathologic changes in the coronary artery in the form of peri-coronary fibrosis that correlated to the flow changes as assessed by echocardiography. 
The study underscores the value of using a non-invasive technique to monitor coronary circulation in mouse hearts. The method minimizes redundant use of research animals and demonstrates that advanced ultrasound-based indices, such as CFR and S/D ratios, can serve as viable diagnostic tools in a variety of investigational protocols including drug studies and the study of genetically modified strains.

Coronary flow reserve (CFR) is useful for assessment of myocardial oxygen demand and evaluation of cardiovascular risk. This study establishes a step-by-step transthoracic Doppler echocardiographic (TTDE) method for longitudinal monitoring of the changes in CFR, as measured from coronary artery in mice, under the experimental pressure overload of aortic banding.

冠脉流量和冠状动脉血流储备用小鼠压力超负荷基于模型的超声评估

Transthoracic Doppler echocardiography (TTDE) is a clinically useful, noninvasive tool for studying coronary artery flow velocity and coronary flow ...

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 ...

Echocardiographic Approaches and Protocols for Comprehensive Phenotypic Characterization of Valvular Heart Disease in Mice

The aim of this manuscript and accompanying video is to provide an overview of the methods and approaches used for imaging heart valve function in rodents, with detailed descriptions of the appropriate methods for anesthesia, the echocardiographic windows used, the imaging planes and probe orientations for image acquisition, the methods for data analysis, and the limitations of emerging technologies for the evaluation of cardiac and valvular function. Importantly, we also highlight several future areas of research in cardiac and heart valve imaging that may be leveraged to gain insights into the pathogenesis of valve disease in preclinical animal models. We propose that using a systematic approach to evaluating cardiac and heart valve function in mice can result in more robust and reproducible data, as well as facilitate the discovery of previously underappreciated phenotypes in genetically-altered and/or physiologically-stressed mice.

This protocol provides a detailed description of the echocardiographic approach for comprehensive phenotyping of heart and heart valve function in mice.

超声心动图方法和协议心脏瓣膜疾病的综合表型特征小鼠

The aim of this manuscript and accompanying video is to provide an overview of the methods and approaches used for imaging heart valve function in ...

Ultrasound-based Pulse Wave Velocity Evaluation in Mice

Arterial stiffness can be evaluated by calculating pulse wave velocity (PWV), i.e., the speed with which the pulse wave travels in a conduit vessel. This parameter is being increasingly investigated in small rodent models in which it is used for assessing alterations in vascular function related to particular genotypes/treatments or for characterizing cardiovascular disease progression. This protocol describes an image processing algorithm which leads to non-invasive arterial PWV measurement in mice using ultrasound (US) images only. The proposed technique has been used to assess abdominal aorta PWV in mice and evaluate its age-associated changes.
Abdominal aorta US scans are obtained from mice under gaseous anesthesia using a specific US device equipped with high-frequency US probes. B-mode and Pulse-Wave Doppler (PW-Doppler) images are analyzed in order to obtain diameter and mean velocity instantaneous values, respectively. For this purpose, edge detection and contour tracking techniques are employed. The single-beat mean diameter and velocity waveforms are time aligned and combined in order to achieve the diameter-velocity (lnD-V) loop. PWV values are obtained from the slope of the linear part of the loop, which corresponds to the early systolic phase.
With the present approach, anatomical and functional information about the mouse abdominal aorta can be non-invasively achieved. Requiring the processing of US images only, it may represent a useful tool for the non-invasive characterization of different arterial sites in the mouse in terms of elastic properties. The application of the present technique can be easily extended to other vascular districts, such as the carotid artery, thus providing the possibility to obtain a multi-site arterial stiffness assessment.

Arterial stiffness represents a key factor in cardiovascular disease and pulse wave velocity (PWV) can be considered as a surrogate index for arterial stiffness. This protocol describes an image processing algorithm for calculating PWV in mice based on ultrasound image processing that is applicable at different arterial sites.

基于超声脉搏波速度评价小鼠

Arterial stiffness can be evaluated by calculating pulse wave velocity (PWV), i.e., the speed with which the pulse wave travels in a conduit vessel. This ...

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 ...

Measurement of Pulse Propagation Velocity, Distensibility and Strain in an Abdominal Aortic Aneurysm Mouse Model

An abdominal aortic aneurysm (AAA) is defined as a localized dilation of the abdominal aorta that exceeds the maximal intraluminal diameter (MILD) by 1.5 times of its original size. Clinical and experimental studies have shown that small aneurysms may rupture, while a subpopulation of large aneurysms may remain stable. Thus, in addition to the measurement of intraluminal diameter of the aorta, knowledge of structural traits of the vessel wall may provide important information to assess the stability of the AAA. Aortic stiffening has recently emerged as a reliable tool to determine early changes in the vascular wall. Pulse propagation velocity (PPV) along with the distensibility and radial strain are highly useful ultrasound-based methods relevant for assessing aortic stiffness. The primary purpose of this protocol is to provide a comprehensive technique for the use of ultrasound imaging system to acquire images and analyze the structural and functional properties of the aorta as determined by MILD, PPV, distensibility and radial strain.

This manuscript describes a detailed protocol for using high frequency ultrasound imaging to measure luminal diameter, pulse propagation velocity, distensibility and radial strain on a mouse model of abdominal aortic aneurysm.

腹部主动脉瘤小鼠模型中脉冲传播速度、失散性和应变的测量

An abdominal aortic aneurysm (AAA) is defined as a localized dilation of the abdominal aorta that exceeds the maximal intraluminal diameter (MILD) by 1.5 ...

免疫与感染

Ultrasound Imaging of the Thoracic and Abdominal Aorta in Mice to Determine Aneurysm Dimensions

Contemporary high-resolution ultrasound instruments have sufficient resolution to facilitate the measurement of mouse aortas. These instruments have been widely used to measure aortic dimensions in mouse models of aortic aneurysms. Aortic aneurysms are defined as permanent dilations of the aorta, which occur most frequently in the ascending and abdominal regions. Sequential measurements of aortic dimensions by ultrasound are the principal approach for assessing the development and progression of aortic aneurysms in vivo. Although many reported studies used ultrasound imaging to measure aortic diameters as a primary endpoint, there are confounding factors, such as probe position and cardiac cycle, that may impact the accuracy of data acquisition, analysis, and interpretation. The purpose of this protocol is to provide a practical guide on the use of ultrasound to measure the aortic diameter in a reliable and reproducible manner. This protocol introduces the preparation of mice and instruments, the acquisition of appropriate ultrasound images, and data analysis.

Ultrasound imaging has become a common modality to determine the luminal dimensions of thoracic and abdominal aortic aneurysms in mice. This protocol describes the procedure to acquire reliable and reproducible two-dimensional ultrasound images of the ascending and abdominal aorta in mice.

小鼠胸腹主动脉超声检查确定动脉瘤大小

Contemporary high-resolution ultrasound instruments have sufficient resolution to facilitate the measurement of mouse aortas. These instruments have been ...

Measuring the Stiffness of Ex Vivo Mouse Aortas Using Atomic Force Microscopy

Arterial stiffening is a significant risk factor and biomarker for cardiovascular disease and a hallmark of aging. Atomic force microscopy (AFM) is a versatile analytical tool for characterizing viscoelastic mechanical properties for a variety of materials ranging from hard (plastic, glass, metal, etc.) surfaces to cells on any substrate. It has been widely used to measure the stiffness of cells, but less frequently used to measure the stiffness of aortas. In this paper, we will describe the procedures for using AFM in contact mode to measure the ex vivo elastic modulus of unloaded mouse arteries. We describe our procedure for isolation of mouse aortas, and then provide detailed information for the AFM analysis. This includes step-by-step instructions for alignment of the laser beam, calibration of the spring constant and deflection sensitivity of the AFM probe, and acquisition of force curves. We also provide a detailed protocol for data analysis of the force curves.

Measuring the Stiffness of Ex Vivo Mouse Aortas Using Atomic Force Microscopy

We present detailed protocols for isolation of aortas from mouse and measurement of their elastic modulus using atomic force microscopy.

测量的刚度<em&gt;离体</em&gt;鼠标主动脉用原子力显微镜

Arterial stiffening is a significant risk factor and biomarker for cardiovascular disease and a hallmark of aging. Atomic force microscopy (AFM) is a ...

生物学

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 ...

测量升主动脉刚度

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