We thank Fred Roberts for exemplary technical support. We thank Brigham Women&#8217;s Hospital Cardiovascular Physiology Core for providing with the instrumentation and the funds for this work. This work was supported in part by NHLBI grants HL093148, HL086967, and Departmental Funds from (SUNDRY).

执行从查尔斯河实验室按照美国兽医协会（AVMA）的指引购买，使用批准的机构动物护理和使用委员会（IACUC）协议雄性SD大鼠的所有程序。 1.动物的制备和外科手术<ol><li>任何实验步骤之前适应所有动物一个星期。 </li><li>使用异氟醚麻醉的动物（2-3％诱导和1.0％维持），并应用眼膏双眼，防止干燥，发炎或溃疡。 </li><li>使用＃40刀片，并根据需要脱毛霜动物的胸部取出头发。它可能不得不删除头发形式动物的回来，如果我们不能有从腹侧成像获得了良好的图像数据。 </li></ol> 2.肾毒性大鼠模型<ol><li>顺铂肾毒性模型，管理顺铂，使用协议š如前所述。15 </li><li>在基线，24小时前给予顺铂（0天）进行超声检查。 （请参阅步骤3中，成像协议） </li><li>随机鼠（n = 6）分为两组。第1天，给予顺铂动物的体重计算（10毫克/毫升）（10毫克/千克体重，单剂量肾毒性感应），注射量（1毫升/ kg体重）），腹膜内研究组和生理盐水（ NS）对照组。 </li><li>麻醉动物如步骤1.2在24，48，72，96，120，顺铂给药后144小时。 </li><li>根据动物的麻醉平稳阶段使用高分辨率超声系统（参见材料和设备表）采取的图像。继续通过完全恢复从麻醉诱导成像期间监测动物的基本生理功能。 </li><li>在成像过程监控动物的生命体征:鼠温度:35.9-37.5，呼吸频率:66-144 /分钟，心脏率:250-600 /分钟。最佳生命体征在我们的研究中提出的阅读是:温度:36.5-37.0，呼吸频率:80-100 /分钟，心脏率:450〜550 /分钟。 注意:使用静脉输液，并加热灯维持动物的正常生理条件，以减少外科手术和麻醉的效果。如有必要，在手术过程中协助机械通气机呼吸。然而，机械通气很少需要在该实验。 </li></ol> 3.成像协议注:超声机供应商提供长期成像过程加热平台。但是，我们不要用加热的平台，在我们的实验证明，因为它只需要5到15分钟。其控制对与连接到所述生理控制开关直肠温度计监测体温。 <ol><li>肾脏（B模式）的横向图像: <ol><li>使用MS 250超声与21兆赫的中心频率连接编到有源端口，应用预设设置为"通用影像"。 </li><li>与平台上的动物仰卧，定位在动物使用的导轨系统的21兆赫超声波探头，中线和隔离主动脉。在这个位置上，探针角度为90度到左侧胸骨旁线（横轴）（ 图1A，B）。 </li><li>从该位置滑动与动物的平台，使得探针现在是在肾动脉的水平（左或右，可以形象一次一个）。 </li><li>通过使用显微操作，查看的右侧或左侧肾动脉。 </li><li>由沿着探针的y轴稍微倾斜，以获得在屏幕的中心的完整肾视图调整探头角度。 </li><li>一旦适当地标（肾盂，肾动脉）， 如图1C和D，电影商店使用允许与所使用的探针的最高帧速率的图像示出被识别。 </l我&gt; </ol></li><li>肾（彩色多普勒视图）的横向图像: <ol><li>使用键盘上的彩色多普勒键，打开彩色多普勒声窗。这有助于隔离肾动脉和肾静脉（ 图1D）。 （蓝色代表动脉血流;而红色表示静脉血流）。 </li><li>确保焦点深度（在Y轴上以和黄色箭头指示）位于肾脏的中心。录制电影存储中的数据。 </li><li>确保记录在尽可能高的帧速率可能的（&gt; 200帧/秒）的数据。 </li></ol></li><li>肾（脉冲波或PW视图）的横向图像: <ol><li>点击PW关键，而在彩色多普勒模式，调出屏幕（ 图1F）上的黄色指示线（脉冲多普勒取样容积）。 </li><li>放置在肾动脉黄线，在该通过PW角度键平行通过容器的流动的方向性的角度。 </li><lI&gt;对于在右肾动脉流动评估，将PW黄色指示线沿流动方向的肾动脉（这在图1（D）蓝色和E示出）确保了多普勒角为60度以下。 </li><li>在这种模式下，accoustic窗口分割成上部和下部。 </li><li>使用电影存储捕捉到的指示在峰值收缩和舒张动脉流动的速度的波形图像。 </li></ol></li></ol> 4.动物处理成像后<ol><li>从第0天至第5天，将动物到一个干净的恢复区（带床上用品干净的纸巾）在成像后胸骨斜卧位。请注意，我们处理极为谨慎所有动物与攻击性的动物，如动物的"尾巴控股"的方法，从麻醉中恢复。 </li><li>在麻醉恢复，保持动物的体温与外部老天ŧ源和显示器动物与电探针生命体征，直到动物从麻醉中完全恢复。 </li><li>返回恢复动物设施住房房间时，他们是警觉和活跃。 </li><li>根据6日和收获肾脏机构准则安乐死所有动物（见步骤4.7）进行组织学评估，以及步4.5。 </li><li>收集附着在代谢笼肌酐测试，以检查肾功能收集管动物的尿。 </li><li>执行动物肾脏的石蜡切片，并进行HE（苏木和曙红）染色检查肾毒性（见步骤4.7查看详细）。 </li><li>牺牲动物和用0.9％NaCl溶液，接着用10％缓冲的福尔马林固定通过左心室抽血。用0.9％NaCl溶液放血后，取出大鼠肾脏组织学评估16。 <ol><li>石蜡嵌入6毫米切片观察肾吗啉术和肾毒性。在4ºC脱水30％蔗糖肾组织在磷酸盐缓冲盐水（PBS）48小时。然后于4ºC固定在10％缓冲福尔马林区段24-48小时。 </li><li>接着，嵌入石蜡肾组织，并在室温的组织石蜡块存储直到切片。进一步部使用石蜡切片机将组织块，并放置在一被涂覆载玻片的章节。 </li><li> Deparrafinize部分和再水化并用苏木染色10分钟，随后曙红3分钟。安装在一个幻灯片上的部分，并让他们通过一种啮齿动物病理学家进行评估。 </li></ol></li></ol> 5.数据计算与分析<ol><li>计算从3.2步获得的彩色多普勒超声图像肾动脉峰值速度。选择速度时间积分（VTI）工具跟踪收缩和舒张速度的峰值。 </li><li>使用方程计算阻力指数（RI）和搏动指数（PI）下面纳秒。 RI =（收缩期峰值速度，舒张末期速度）/收缩期峰值速度 PI =（收缩期峰值速度，舒张末期速度）/平均血流速度。 </li><li>执行的RI和PI结果与平均三个周期的测量标准偏差的统计分析。对于其他的标准参数，请参考手册从制造商使用专用软件进行数据分析（见材料和​​设备表）。 </li></ol>

<ol>
	<li>Bonventre, J. V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Diagnosis+of+acute+kidney+injury:+from+classic+parameters+to+new+biomarkers.">Diagnosis of acute kidney injury: from classic parameters to new biomarkers.</a> Contrib Nephrol. 156, 213-219 (2007).</li><li>Lerman, L. O., Textor, S. C., Grande, J. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mechanisms+of+tissue+injury+in+renal+artery+stenosis:+ischemia+and+beyond.">Mechanisms of tissue injury in renal artery stenosis: ischemia and beyond.</a> Progress in cardiovascular diseases. 52 (3), 196-203 (2009).</li><li>Karas, M. G., Kizer, J. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Echocardiographic+assessment+of+the+right+ventricle+and+associated+hemodynamics.">Echocardiographic assessment of the right ventricle and associated hemodynamics.</a> Progress in cardiovascular diseases. 55 (2), 144-160 (2012).</li><li>Milman, Z., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Hemodynamic+response+magnetic+resonance+imaging:+application+for+renal+hemodynamic+characterization.">Hemodynamic response magnetic resonance imaging: application for renal hemodynamic characterization.</a> Nephrol Dial Transplant. 28, 1150-1156 (2013).</li><li>Anavekar, N. S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Usefulness+of+right+ventricular+fractional+area+change+to+predict+death,+heart+failure,+and+stroke+following+myocardial+infarction+(from+the+VALIANT+ECHO+Study).">Usefulness of right ventricular fractional area change to predict death, heart failure, and stroke following myocardial infarction (from the VALIANT ECHO Study).</a> Am J Cardiol. 101 (5), 607-612 (2008).</li><li>Lindqvist, P., Calcutteea, A., Henein, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Echocardiography+in+the+assessment+of+right+heart+function.">Echocardiography in the assessment of right heart function.</a> Eur J Echocardiogr. 9 (2), 225-234 (2008).</li><li>Sullivan, J. C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Novel+use+of+ultrasound+to+examine+regional+blood+flow+in+the+mouse+kidney.">Novel use of ultrasound to examine regional blood flow in the mouse kidney.</a> American journal of physiology. Renal physiology. 297, F228-F235 (2009).</li><li>Bonnin, P., Sabaa, N., Flamant, M., Debbabi, H., Tharaux, P. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ultrasound+imaging+of+renal+vaso-occlusive+events+in+transgenic+sickle+mice+exposed+to+hypoxic+stress.">Ultrasound imaging of renal vaso-occlusive events in transgenic sickle mice exposed to hypoxic stress.</a> Ultrasound Med Biol. 34 (7), 1076-1084 (2008).</li><li>Desberg, A. L., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Renal+artery+stenosis:+evaluation+with+color+Doppler+flow+imaging.">Renal artery stenosis: evaluation with color Doppler flow imaging.</a> Radiology. 177 (3), 749-753 (1990).</li><li>Ciccone, M. M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+clinical+role+of+contrast-enhanced+ultrasound+in+the+evaluation+of+renal+artery+stenosis+and+diagnostic+superiority+as+compared+to+traditional+echo-color-Doppler+flow+imaging.">The clinical role of contrast-enhanced ultrasound in the evaluation of renal artery stenosis and diagnostic superiority as compared to traditional echo-color-Doppler flow imaging.</a> International angiology : a journal of the International Union of Angiology. 30 (2), 135-139 (2011).</li><li>Bauer, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Echocardiographic+speckle-tracking+based+strain+imaging+for+rapid+cardiovascular+phenotyping+in+mice.">Echocardiographic speckle-tracking based strain imaging for rapid cardiovascular phenotyping in mice.</a> Circ Res. 108 (8), 908-916 (2011).</li><li>Boulikas, T., Vougiouka, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Recent+clinical+trials+using+cisplatin,+carboplatin+and+their+combination+chemotherapy+drugs+(review).">Recent clinical trials using cisplatin, carboplatin and their combination chemotherapy drugs (review).</a> Oncology reports. 11 (3), 559-595 (2004).</li><li>Vaidya, V. S., Ferguson, M. A., Bonventre, J. V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Biomarkers+of+acute+kidney+injury.">Biomarkers of acute kidney injury.</a> Annu Rev Pharmacol Toxicol. 48, 463-493 (2008).</li><li>Hye Khan, M. A., Abdul Sattar, M., Abdullah, N. A., Johns, E. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cisplatin-induced+nephrotoxicity+causes+altered+renal+hemodynamics+in+Wistar+Kyoto+and+spontaneously+hypertensive+rats:+role+of+augmented+renal+alpha-adrenergic+responsiveness.">Cisplatin-induced nephrotoxicity causes altered renal hemodynamics in Wistar Kyoto and spontaneously hypertensive rats: role of augmented renal alpha-adrenergic responsiveness.</a> Exp Toxicol Pathol. 59, 253-260 (2007).</li><li>Vaidya, V. S., Bonventre, J. V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mechanistic+biomarkers+for+cytotoxic+acute+kidney+injury.">Mechanistic biomarkers for cytotoxic acute kidney injury.</a> Expert Opin Drug Metab Toxicol. 2 (5), 697-713 (2006).</li><li>Lu, T. S., Chen, H. W., Huang, M. H., Wang, S. J., Yang, R. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Heat+shock+treatment+protects+osmotic+stress-induced+dysfunction+of+the+blood-brain+barrier+through+preservation+of+tight+junction+proteins.">Heat shock treatment protects osmotic stress-induced dysfunction of the blood-brain barrier through preservation of tight junction proteins.</a> Cell stress, &amp; chaperones. 9 (4), 369-377 (2004).</li><li>Kaya, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Hemodynamics - New Diagnostic and Therapeutic Approaches. , 1-30 (2012).</li><li>Bjornerheim, R., Grogaard, H. K., Kjekshus, H., Attramadal, H., Smiseth, O. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=High+frame+rate+Doppler+echocardiography+in+the+rat:+an+evaluation+of+the+method.">High frame rate Doppler echocardiography in the rat: an evaluation of the method.</a> European journal of echocardiography : the journal of the Working Group on Echocardiography of the European Society of Cardiology. 2 (2), 78-87 (2001).</li><li>Zhang, L., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+high-frequency,+high+frame+rate+duplex+ultrasound+linear+array+imaging+system+for+small+animal+imaging.">A high-frequency, high frame rate duplex ultrasound linear array imaging system for small animal imaging.</a> IEEE transactions on ultrasonics, ferroelectrics, and frequency. 57, 1548-1557 (2010).</li><li>Frea, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Echocardiographic+evaluation+of+right+ventricular+stroke+work+index+in+advanced+heart+failure:+a+new+index?.">Echocardiographic evaluation of right ventricular stroke work index in advanced heart failure: a new index?.</a> J Card Fail. 18 (12), 886-893 (2012).</li><li>Marwick, T. H., Raman, S. V., Carrio, I., Bax, J. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Recent+developments+in+heart+failure+imaging.">Recent developments in heart failure imaging.</a> JACC Cardiovasc Imaging. 3 (4), 429-439 (2010).</li><li>Pokreisz, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Pressure+overload-induced+right+ventricular+dysfunction+and+remodelling+in+experimental+pulmonary+hypertension:+the+right+heart+revisited.">Pressure overload-induced right ventricular dysfunction and remodelling in experimental pulmonary hypertension: the right heart revisited.</a> Eur Heart J supplements. 9 (Supplement H), H75-H84 (2007).</li><li>Senechal, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+simple+Doppler+echocardiography+method+to+evaluate+pulmonary+capillary+wedge+pressure+in+patients+with+atrial+fibrillation.">A simple Doppler echocardiography method to evaluate pulmonary capillary wedge pressure in patients with atrial fibrillation.</a> Echocardiography. 25 (1), 57-63 (2008).</li><li>Souders, C. A., Borg, T. K., Banerjee, I., Baudino, T. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Pressure+overload+induces+early+morphological+changes+in+the+heart.">Pressure overload induces early morphological changes in the heart.</a> Am J Pathol. 181 (4), 1226-1235 (2012).</li><li>Tanaka, N., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Transthoracic+echocardiography+in+models+of+cardiac+disease+in+the+mouse.">Transthoracic echocardiography in models of cardiac disease in the mouse.</a> Circulation. 94 (5), 1109-1117 (1996).</li></ol>

作者什么都没有透露。

在过去的十年中，许多进步都发生在超声技术，包括高频机械探头，它提供高品质，灵敏度和准确度超声数据的发展。这些探针可在5到12毫米和高帧速率（大于200帧/秒）的穿透深度为大约50微米轴向分辨率，因此，它可以作为一个强大的工具来研究小动物如大鼠和小鼠18 19。另外，它也允许收集关于轻度镇静或清醒动物的超声图像与在生理水平的生命体征。此外，这种非侵入性的方式也提供了机会，以在不牺牲动物19执行的疾病进展过程中的结构和功能的改变纵向评估。 1959年，博士。 Rusell和刷最初描述的三个"R"规则（替换，减少和细化），以提高AWAR在研究中使用动物伦理问题eness。该协议显示了首次非侵入性的小动物超声可以利用下Nephorotoxicity研究最少痛苦，折磨或痛苦最小数量的动物。因此，它是为满足实验动物的三个"R&#39;规则的潜在有效方式。 许多超声研究都集中在心脏应用;肾功能评估，往往是从心的状态而不是肾脏20-25的直接研究测量得出。我们已经建立了一个成像方法，以可视化的肾脏实时解剖和功能的变化。我们使用了预先选择的一组互补的声学窗，灰阶/ B模式和彩色多普勒，特异于肾视图。我们所使用的RI和PI指数来评估这些指数与肾功能中的顺铂诱导的毒性模型中的变化之间的关系。 <p class="jove_content"&gt;但是，也有一些挑战和限制所提出的成像方法如下:1）麻醉剂的适当选择和麻醉的程度是心脏和呼吸稳定性是至关重要的。不一致的生理现象（包括呼吸和心脏的波动）影响肾动脉血流，成像和肾功能评估的质量。我们将使用IACUC批准注射麻醉剂，戊巴比妥（50毫克/公斤体重，IP），如我们的备份麻醉剂，以确保在成像和肾功能的评估在适当的正常生理功能; 2）脱毛是关键的一步，为胸毛存在影响了超声图像的质量; 3）当肾脏超声检查是受过训练的操作相对简单;对于一般的操作者，这是至关重要的，以适应技术动物个体的独特解剖学和作出轻微的操作; 4）如大鼠的尺寸是相当大的（大于350克），较低的频率探针（小于在这项研究中21兆赫使用）可能需要最佳成像。这可能是谨慎的成像系统的操作建议之前采取的培训课程。 在使用超声提出的方法和衍生协议检测药物肾毒性的新颖之处在于对肾损伤的情况下，血流动力学改变其早期强大的检测。结果表明，事实上，肾内血管血液动力学变化先上升血肌酐。这些数据是使用组织学分析对常规金标准基准，并证明小动物超声是一种无创，敏感和可重复的方式，其具有动物使用的最低限度的要求。因此，这是早期发现使用大鼠模型药物引起的肾毒性的有效工具。

血肌酐（SCR）一直是黄金标准指标来评估超过二十年的肾功能。最近，许多研究报告说，出现肾损伤比SCR中1的变化要早得多。不过，也有用于检测在肾损伤包括药物引起的肾毒性的进程的初期出现血流动力学改变不稳健的方法。 药物引起的急性肾功能不全的血流动力学导致的肾组织的损伤，并进一步发展为肾功能衰竭2,3。在过去的几十年里，研究表明，成像工具，如计算机断层扫描（CAT），功能性磁共振成像（fMRI）技术，并发挥超声血流动力学评估4的作用。在当前的成像工具，灰阶超声加上彩色多普勒技术，是最常用的建立和评估肾3,5,6-解剖状态。 Sullivan 等。人与BonniN 等。人最近报道说，超声是在血管收缩和缺氧应激动物模型7,8分析血流动力学改变一个有效的，强大的，非侵入性的工具。这种技术也通常用于检测动脉狭窄9,10。 在高分辨率超声成像领域最新的技术进步已经允许研究者使用高频率（25-80兆赫）和高分辨率（&lt;0.03毫米分辨率）探针来解决心血管毒性，体内 11。我们推测，使用这种高分辨率的超声研究肾脏将为早期发现肾毒性的非侵入性和灵敏的方法了前所未有的机遇。 顺铂是用于治疗睾丸癌，卵巢癌，膀胱癌，头，肺，颈部癌症的组合与其他药物12-14。顺铂已充分证明肾毒性是由于磷细胞坏死roximal小管（PT）和收集管导致上升的血液尿素氮（BUN）和SCR 15。在此，我们提供了使用非侵入性的肾脏超声表征使用药物（顺铂）的大鼠模型肾功能不全的详细的一步一步的方法诱导的肾毒性。

<table><tbody><tr><td>Depilatory cream</td><td>Miltex, Inc.</td><td>Surgi-Prep</td><td>Apply 24 hours prior to imaging</td></tr><tr><td>cis-Diamineplatinum(II) dichloride</td><td>Sigma</td><td>479306</td><td>To induce acute kidney injury at small animals.</td></tr><tr><td>Isoflurane</td><td>Baxter International Inc.</td><td>NDC 10019-773-40</td><td>2-3% for induction, and 1-1.5 % for maintenance; heart beats will be maintained at above 500 beats per minute</td></tr><tr><td>High Frequency Ultrasound</td><td>FUJIFILM VisualSonics, Inc.</td><td>Vevo 2100</td><td></td></tr><tr><td>High-frequency Mechanical Transducer</td><td>FUJIFILM VisualSonics, Inc.</td><td>MS250, MS550D, MS400</td><td></td></tr></tbody></table>

early detection of drug-induced renal hemodynamic dysfunction using sonographic technology in rats

肾功能正常，以保持血流动力学稳态和是由药物毒性损害的主要部位。药物引起的肾毒性估计危重病人导致的急性肾损伤（AKI）的所有临床病例19- 25％。 AKI检测历来依靠指标，如血清肌酐（SCR）或血液尿素氮（BUN），这是在肾功能不全的早期肾毒性的全面评估明显不足。目前，对在AKI的早期阶段正确地检测血流动力学改变，而这种改变可能实际上先于血清生物标记物水平的上升不健壮的诊断方法。这种早期发现可以帮助医生作出准确的诊断，并有助于在决策制定治疗策略。大鼠用顺铂治疗诱导AKI。肾毒性进行了评估，使用高频超声，可控硅测量六天后的组织病理学肾。使用二维和彩色多普勒血流图像评价被用来研究连续大鼠肾毒性，使用超声检查。我们的数据通过组织学检查显示，成年大鼠成功的药物性肾损伤。彩色多普勒基于AKI的超声评估显示，阻力指数（RI）和搏动指数（PI）治疗组均增加;和峰收缩速度（毫米/秒），舒张末期速度（毫米/秒）和速度 - 时间积分（VTI，毫米）在肾动脉在同一组降低。重要的是，通过超声检查评估这些血流动力学改变之前血肌酐水平的上升。因此，基于超声指数如RI或PI可在啮齿类动物肾功能下降的有用的预测指标。从在接受AKI大鼠肾脏我们基于超声观测，我们发现，这些无创血流动力学测量可以考虑在检测早期肾功能不全的准确，灵敏，可靠的方法。 ŧ他的研究还强调与研究动物的使用相关的伦理问题的重要性。

在本研究中提出的图像是由一个操作员。使用高频超声机（请参阅材料和设备表）收集的所有图像。所有成像数据是由一个单一的研究者进行分析。结果表明，顺铂处理的动物有sCr的范围从0.5到在第6天（ 图2A）2.1（正常范围&lt;1.1）。然而，组织学研究时相比生理盐水处理的动物表现出急性肾小管间质性损伤的一致的模式。 采用高分辨率的超声成像测量肾血流动力学改变，数据显示，没有与NS0天6日间治疗的动物形态的变化，而在动物在治疗Cisplain六年后一天被检测奇脉哲水蚤的形态。正常的RI和PI的上限为0.7和1.15，分别在大鼠17。使用上述指标来评估血液动力学<html>肾脏的变化，这表明，有在顺铂处理的动物在第6天的RI和PI的显著增加。 <img alt="图1" src="/files/ftp_upload/52409/52409fig1.jpg" /> 图1.大鼠探测肾脏图像超声装置的设置。动物阶段（A）和成像探头位置（B）的大鼠肾超声成像的操作期间的设定的成像系统的图解说明。采用高频，高分辨率超声系统（请参阅材料andand设备表）从大鼠肾得到的样品超声图像。 （CF）。这些数据表明在肾血管有足够的信息来进一步血流动力学参数测量和分析，明确肾脏解剖结构和血液流动。09fig1large.jpg"目标="_空白"&gt;点击此处查看该图的放大版本。 <img alt="图2" src="/files/ftp_upload/52409/52409fig2.jpg" /> 图2. 组织学和肾下顺铂治疗的大鼠的超声图像。血肌酐（SCR）和组织学研究提出车辆处理的大鼠和严重近端肾小管肾损伤（黄色箭头）正常肾组织中顺铂治疗的大鼠（A，B）。 sCr的顺铂治疗后略有增加，但仍维持在正常范围（&lt;1.1）之内。车辆和顺铂处理的大鼠（C）的彩色多普勒模式大鼠的右肾的超声图像在第0天，第3，和6;血流动力学参数，RI和PI分别显著上升，由彩色多普勒超声（D，E）进行评估。正常的RI的上限为PI 0.7和1.15。更重要的，T他上面的数据显示这些血流动力学改变之前SCR的上升。脉搏波速度测量表明顺铂治疗后缓慢和微弱的脉冲（奇脉水蚤标志，黄色圆圈）与组织学研究成果相关。这种现象说明肾动脉狭窄，阻塞，进一步肾功能不全。组织学数据显示，成功的药物诱导肾小管肾损伤和超声评估表明使用彩色多普勒技术在RI，PI和脉搏波速度显著的变化。 N = 3，* P &lt;0.05。 <a href="https://jove.unicartagenaproxy.elogim.com/files/ftp_upload/52409/52409fig2large.jpg" target="_blank">请点击此处查看该图的放大版本。</a> <img alt="表格1" src="/files/ftp_upload/52409/52409table1.jpg" /> 对于药物引起的AKI 表1. 肾血流动力学参数

Watch this Scientific Journal Video about Early Detection of Drug-Induced Renal Hemodynamic Dysfunction Using Sonographic Technology in Rats at JoVE.com

Early Detection of Drug-Induced Renal Hemodynamic Dysfunction Using Sonographic Technology in Rats

Early stage hemodynamic dysfunction is critical to the development of kidney disease. Yet, detection methodologies are limited. Recent advances in sonography provide a noninvasive, accurate option for early detection of kidney injury. This study outlines a step-by-step, sonographic methodology for detecting kidney dysfunction using a drug-induced nephrotoxicity rat model.

大鼠使用超声技术药物引起的肾血流动力学障碍的早期检测

The kidney normally functions to maintain hemodynamic homeostasis and is a major site of damage caused by drug toxicity. Drug-induced nephrotoxicity is ...

early-detection-drug-induced-renal-hemodynamic-dysfunction-using

Universidad de Cartagena UDEC

Research

JoVE Journal

Medicine

12.0K 次观看.  Brigham and Women's Hospital, Harvard Medical School. Early stage hemodynamic dysfunction is critical to the development of kidney disease. Yet, detection methodologies are limited. Recent advances in sonography provide a noninvasive, accurate option for early detection of kidney injury. This study outlines a step-by-step, sonographic methodology for detecting kidney dysfunction using a drug-induced nephrotoxicity rat model. 

视频: Early Detection of Drug-Induced Renal Hemodynamic Dysfunction Using Sonographic Technology in Rats

Transcutaneous Assessment of Renal Function in Conscious Rodents

Glomerular filtration rate (GFR) is the gold standard to assess overall kidney function. However, traditional methods to evaluate GFR are cumbersome and time-consuming. In addition, serial blood or urine samples are required, with the associated stress for the experimental animals. A recent technique significantly reduces the investment in time and resources, minimizing the invasiveness and the animal stress, but being equally valid as the traditional approaches. The method measures transcutaneously renal function. Using an optical device and the exogenous renal marker fluorescein isothiocyanate (FITC)-sinistrin, this technique is capable of measuring the elimination kinetics of the marker through the skin. With neither blood nor urine samples nor the associated laboratory assays needed, the results of the transcutaneous measurement are almost instantaneously available. The method has been already validated in different species and successfully applied in several models of renal pathology. Moreover, due to its minimally invasive characteristics, it is suitable for sequential measurements within the same animal. Here is provided a detailed protocol to carry out the transcutaneous assessment of renal function in rodents.

Determination of glomerular filtration rate (GFR) is the gold standard to assess overall kidney function. However, traditional procedures to measure this parameter are cumbersome and require a large investment of time. Here we describe a faster and minimally invasive method to determinate GFR transcutaneously.

在鼠害自觉肾功能的评估皮

Glomerular filtration rate (GFR) is the gold standard to assess overall kidney function. However, traditional methods to evaluate GFR are cumbersome and ...

科研

医学

Intravascular Delivery of Biologics to the Rat Kidney

The renal microvascular compartment plays an important role in the progression of kidney disease and hypertension, leading to the development of End Stage Renal Disease with high risk of death for cardiovascular events. Moreover, recent clinical studies have shown that renovascular structure and function may have a great impact on functional renal recovery after surgery. Here, we describe a protocol for the delivery of drugs into the renal artery of rats. This procedure offers significant advantages over the frequently used systemic administration as it may allow a more localized therapeutic effect. In addition, the use of rodents in pharmacodynamic analysis of preclinical studies may be cost effective, paving the way for the design of translational experiments in larger animal models. Using this technique, infusion of rat recombinant Vascular Endothelial Growth Factor (VEGF) protein in rats has induced activation of VEGF signaling as shown by increased expression of FLK1, pAKT/AKT, pERK/ERK. In summary, we established a protocol for the intrarenal delivery of drugs in rats, which is simple and highly reproducible.

The administration of drugs for recovery of kidney function requires control of the localization and distribution of the therapeutic compound. Here, we describe in detail a simple technique for intrarenal delivery of drugs in rats. This procedure may be easily performed with no mortality and high reproducibility.

生物血管内送货到大鼠肾脏

The renal microvascular compartment plays an important role in the progression of kidney disease and hypertension, leading to the development of End Stage ...

Ultrasound Assessment of Flow-Mediated Dilation of the Brachial and Superficial Femoral Arteries in Rats

Arterial vasodilation to increases in wall shear rate is indicative of vascular endothelial function. In humans, the non-invasive measurement of endothelial function can be achieved by employing the flow-mediated dilation technique, typically performed in the brachial or superficial femoral artery. Briefly, a blood pressure cuff placed distal to an ultrasound probe is inflated to a suprasystolic pressure, which results in limb ischemia. After 5 min of occlusion the cuff is deflated, resulting in reactive hyperemia and increases in wall shear rate that signal vasodilatory molecules to be released from the endothelium eliciting vasodilation. Despite the thousands of studies performing flow-mediated dilation in humans, surprisingly, no studies have performed this technique non-invasively in living rats. Considering the recent shift in focus to translational research, the establishment of guidelines for non-invasive measurement of flow-mediated dilation in rats and other rodents would be extremely valuable. In the following article, a protocol is presented for the non-invasive measurement of flow-mediated dilation in brachial and superficial femoral arteries of rats, as those sites are most commonly measured in humans.

Non-invasive assessment of endothelial function in humans can be determined by the flow-mediated dilation technique. Although thousands of studies have used this technique, no study has performed this technique non-invasively in rats. The following article describes non-invasive measurement of flow-mediated dilation in the brachial and superficial femoral arteries of rats.

肱血流介导的扩张和大鼠股浅动脉超声评估

Arterial vasodilation to increases in wall shear rate is indicative of vascular endothelial function. In humans, the non-invasive measurement of ...

Ultrasonography in Experimental Reproductive Investigations on Rats

With the development of assisted reproductive technology and the ethical limitations of research on humans, rat animal models have been widely used in reproductive medicine. In the past, the study of reproductive system development in rodents has been based on one-time histological examination of excised tissues. Recently, with the development of high-resolution transabdominal ultrasound, high-quality sonography can now be performed to evaluate the reproductive organs of rats, allowing a new method for studying the reproductive system. Images were obtained using a high-resolution ultrasonographic system. Gynecological ultrasonography was performed on 28 eight-week-old non-pregnant rats and 5 pregnant Sprague-Dawley rats. We describe how to recognize organs of the reproductive system and associated structures in typical views during different phases of the estrus cycle. Color flow Doppler was used to measure uterine artery blood flow and evaluate uterine blood flow pattern changes during different stages of pregnancy. We have demonstrated that ultrasound exploration is a useful method for evaluating changes in internal reproductive organs. Its use raises the possibility of conducting additional experiments, including medical or surgical procedures, and provides the ability to monitor sonographic changes to internal organs without sacrificing animals.

This manuscript describes the utility of ultrasound performed on female rats to design experimental models for reproductive and gynecological investigation. A step-by-step explanation of how to perform ultrasonographic evaluation is shown.

大鼠实验性生殖研究中的超声检查

With the development of assisted reproductive technology and the ethical limitations of research on humans, rat animal models have been widely used in ...

Noninvasive and Invasive Renal Hypoxia Monitoring in a Porcine Model of Hemorrhagic Shock

Up to 50% of patients with trauma develop acute kidney injury (AKI), in part due to poor renal perfusion after severe blood loss. AKI is currently diagnosed based on a change in serum creatinine concentration from baseline or prolonged periods of decreased urine output. Unfortunately, baseline serum creatinine concentration data is unavailable in most patients with trauma, and current estimation methods are inaccurate. In addition, serum creatinine concentration may not change until 24-48 h after the injury. Lastly, oliguria must persist for a minimum of 6 h to diagnose AKI, making it impractical for early diagnosis. AKI diagnostic approaches available today are not useful for predicting risk during the resuscitation of patients with trauma. Studies suggest that urinary partial pressure of oxygen (PuO2) may be useful for assessing renal hypoxia. A monitor that connects the urinary catheter and the urine collection bag was developed to measure PuO2 noninvasively. The device incorporates an optical oxygen sensor that estimates PuO2 based on luminescence quenching principles. In addition, the device measures urinary flow and temperature, the latter to adjust for confounding effects of temperature changes. Urinary flow is measured to compensate for the effects of oxygen ingress during periods of low urine flow. This article describes a porcine model of hemorrhagic shock to study the relationship between noninvasive PuO2, renal hypoxia, and AKI development. A key element of the model is the ultrasound-guided surgical placement in the renal medulla of an oxygen probe, which is based on an unsheathed optical microfiber. PuO2 will also be measured in the bladder and compared to the kidney and noninvasive PuO2 measurements. This model can be used to test PuO2 as an early marker of AKI and assess PuO2 as a resuscitative endpoint after hemorrhage that is indicative of end-organ rather than systemic oxygenation.

Presented here is a protocol to measure renal oxygenation in the medulla and noninvasive urine oxygen partial pressure in a hemorrhagic shock porcine model to establish urine oxygen partial pressure as an early indicator of acute kidney injury (AKI) and a novel resuscitative endpoint.

猪出血性休克模型中的无创和侵入性肾缺氧监测

Up to 50% of patients with trauma develop acute kidney injury (AKI), in part due to poor renal perfusion after severe blood loss. AKI is currently ...

生物工程

Echocardiographic Assessment of Cardiac Anatomy and Function in Adult Rats

The use of experimental animal models has become crucial in cardiovascular science. Most studies using rodent models are focused on two-dimensional imaging to study the cardiac anatomy of the left ventricle and M-mode echo to assess its dimensions. However, this could limit a comprehensive study. Herein, we describe a protocol that allows an assessment of the heart chamber size, left ventricular function (systolic and diastolic) and valvular function. A conventional medical ultrasound machine was used in this protocol and different echo views were obtained through left parasternal, apical and suprasternal windows. In the left parasternal window, the long and short axis were acquired to analyze left chamber dimensions, right ventricle and pulmonary artery dimensions, and mitral, pulmonary and aortic valve function. The apical window allows the measurement of heart chamber dimensions and evaluation of systolic and diastolic parameters. It also allows Doppler assessment with detection and quantification of heart valve disturbances (regurgitation or stenosis). Different segments and walls of the left ventricle are visualized throughout all views. Finally, the ascending aorta, aortic arch, and descending aorta can be imaged through the suprasternal window. A combination of ultrasound imaging, Doppler flow and tissue Doppler assessment have been obtained to study cardiac morphology and function. This represents an important contribution to improve the assessment of cardiac function in adult rats with impact for research using these animal models.

A non-invasive protocol for transthoracic echocardiography assessment of cardiac anatomy and function for adult rats is presented in the current study. The heart valves, all four cardiac chambers and the ascending aorta, aortic arch and descending aorta are studied in detail.

成人大鼠心脏解剖与功能的超声心动图评估

The use of experimental animal models has become crucial in cardiovascular science. Most studies using rodent models are focused on two-dimensional ...

A Large Animal Model for Acute Kidney Injury by Temporary Bilateral Renal Artery Occlusion

Acute kidney injury (AKI) is associated with higher risk for morbidity and mortality post-operatively. Ischemia-reperfusion injury (IRI) is the most common cause of AKI. To mimic this clinical scenario, this study presents a highly reproducible large animal model of renal IRI in swine using temporary percutaneous bilateral balloon-catheter occlusion of the renal arteries. The renal arteries are occluded for 60 min by introducing the balloon-catheters through the femoral and carotid artery and advancing them into the proximal portion of the arteries. Iodinated contrast is injected in the aorta to assess any opacification of the kidney vessels and confirm the success of the artery occlusion. This is furtherly confirmed by the flattening of the pulse waveform at the tip of the balloon catheters. The balloons are deflated and removed after 60 min of bilateral renal artery occlusion, and the animals are allowed to recover for 24 h. At the end of the study, plasma creatinine and blood urea nitrogen significantly increase, while eGFR and urine output significantly decrease. The need for iodinated contrast is minimal and does not affect renal function. Bilateral renal artery occlusion better mimics the clinical scenario of perioperative renal hypoperfusion, and the percutaneous approach minimizes the impact of the inflammatory response and the risk of infection seen with an open approach, such as a laparotomy. The ability to create and reproduce this clinically relevant swine model eases the clinical translation to humans.

This study presents a highly reproducible large animal model of renal ischemia-reperfusion injury in swine using temporary percutaneous bilateral balloon-catheter occlusion of the renal arteries for 60 min and reperfusion for 24 h.

临时双性肾动脉闭塞的急性肾损伤大型动物模型

Acute kidney injury (AKI) is associated with higher risk for morbidity and mortality post-operatively. Ischemia-reperfusion injury (IRI) is the most ...

Use of 3D Robotic Ultrasound for In Vivo Analysis of Mouse Kidneys

Common modalities for in vivo imaging of rodents include positron emission tomography (PET), computed tomography (CT), magnetic resonance imaging (MRI), and ultrasound (US). Each method has limitations and advantages, including availability, ease of use, cost, size, and the use of ionizing radiation or magnetic fields. This protocol describes the use of 3D robotic US for in vivo imaging of rodent kidneys and heart, subsequent data analysis, and possible research applications. Practical applications of robotic US are the quantification of total kidney volume (TKV), as well as the measurement of cysts, tumors, and vasculature. Although the resolution is not as high as other modalities, robotic US allows for more practical high throughput data collection. Furthermore, using US M-mode imaging, cardiac function may be quantified. Since the kidneys receive 20%-25% of the cardiac output, assessing cardiac function is critical to the understanding of kidney physiology and pathophysiology.

Use of 3D Robotic Ultrasound for In Vivo Analysis of Mouse Kidneys

This protocol demonstrates robotic ultrasound (US) as a practical, cost-effective, and quick alternative to traditional non-invasive image modalities.

使用3D机器人超 声进行小鼠 肾脏体内分析

Common modalities for in vivo imaging of rodents include positron emission tomography (PET), computed tomography (CT), magnetic resonance imaging (MRI), ...

Long-Term Continuous Measurement of Renal Blood Flow in Conscious Rats

The kidneys play a crucial role in maintaining the homeostasis of body fluids. The regulation of renal blood flow (RBF) is essential to the vital functions of filtration and metabolism in kidney function. Many acute studies have been carried out in anesthetized animals to measure RBF under various conditions to determine mechanisms responsible for the regulation of kidney perfusion. However, for technical reasons, it has not been possible to measure RBF continuously (24 h/day) in unrestrained unanesthetized rats over prolonged periods. These methods allow the continuous determination of RBF over many weeks while also simultaneously recording blood pressure (BP) with implanted catheters (fluid-filled or by telemetry). RBF monitoring is carried out with rats placed in a circular servo-controlled rat cage that enables the unrestrained movement of the rat throughout the study. At the same time, the tangling of cables from the flow probe and arterial catheters is prevented. Rats are first instrumented with an ultrasonic flow probe placement on the left renal artery and an arterial catheter implanted in the right femoral artery. These are routed subcutaneously to the nape of the neck, and connected to the flowmeter and pressure transducer, respectively, to measure RBF and BP. Following surgical implantation, rats are immediately placed in the cage to recover for at least one week and stabilize the ultrasonic probe recordings. Urine collection is also feasible in this system. The surgical and post-surgical procedures for continuous monitoring are demonstrated in this protocol.

The present protocol describes a long-term continuous measurement of renal blood flow in conscious rats and simultaneously recording blood pressure with implanted catheters (fluid-filled or by telemetry).

意识大鼠肾血流量的长期连续测量

The kidneys play a crucial role in maintaining the homeostasis of body fluids. The regulation of renal blood flow (RBF) is essential to the vital ...

Application of Ultrasound and Shear Wave Elastography Imaging in a Rat Model of NAFLD/NASH

Nonalcoholic Steatohepatitis (NASH) is a condition within the spectrum of Non-Alcoholic Fatty Liver Disease (NAFLD), which is characterized by liver fat accumulation (steatosis) and inflammation leading to fibrosis. Preclinical models closely recapitulating human NASH/NAFLD are essential in drug development. While liver biopsy is currently the gold standard for measuring NAFLD/NASH progression and diagnosis in the clinic, in the preclinical space, either collection of whole liver samples at multiple timepoints during a study or biopsy of liver is needed for histological analysis to assess the disease stage.
Conducting a liver biopsy mid-study is an invasive and labor-intensive procedure, and collecting liver samples to assess disease level increases the number of research animals needed for a study. Thus, there is a need for a reliable, translatable, non-invasive imaging biomarker to detect NASH/NAFLD in these preclinical models. Non-invasive ultrasound-based B-mode images and Shear Wave Elastography (SWE) can be used to measure steatosis as well as liver fibrosis. To assess the utility of SWE in preclinical rodent models of NASH, animals were placed on a pro-NASH diet&#160;and underwent non-invasive ultrasound B-mode and shear wave elastography imaging to measure hepatorenal (HR) index and liver elasticity, measuring progression of both liver fat accumulation and tissue stiffness, respectively, at multiple time points over the course of a given NAFLD/NASH study.
The HR index and elasticity numbers were compared to histological markers of steatosis and fibrosis. The results showed strong correlation between the HR index and percentage of Oil Red O (ORO) staining, as well as between elasticity and Picro-Sirius Red (PSR) staining of livers. The strong correlation between classic ex vivo methods and in vivo imaging results provides evidence that shear wave elastography/ultrasound-based imaging can be used to assess disease phenotype and progression in a preclinical model of NAFLD/NASH.

This protocol describes the use of an enhanced ultrasound technique to non-invasively observe and quantify liver tissue changes in rodent models of nonalcoholic fatty liver disease.

在 NAFLD/NASH 的鼠模型中应用超声波和剪切波弹性成像

Nonalcoholic Steatohepatitis (NASH) is a condition within the spectrum of Non-Alcoholic Fatty Liver Disease (NAFLD), which is characterized by liver fat ...

大鼠使用超声技术药物引起的肾血流动力学障碍的早期检测

摘要

探索更多视频

此视频中的章节

在鼠害自觉肾功能的评估皮

生物血管内送货到大鼠肾脏

肱血流介导的扩张和大鼠股浅动脉超声评估

大鼠实验性生殖研究中的超声检查

猪出血性休克模型中的无创和侵入性肾缺氧监测

成人大鼠心脏解剖与功能的超声心动图评估

临时双性肾动脉闭塞的急性肾损伤大型动物模型

使用3D机器人超声进行小鼠肾脏体内分析

意识大鼠肾血流量的长期连续测量

在 NAFLD/NASH 的鼠模型中应用超声波和剪切波弹性成像

大鼠使用超声技术药物引起的肾血流动力学障碍的早期检测

摘要

探索更多视频

此视频中的章节

在鼠害自觉肾功能的评估皮

生物血管内送货到大鼠肾脏

肱血流介导的扩张和大鼠股浅动脉超声评估

大鼠实验性生殖研究中的超声检查

猪出血性休克模型中的无创和侵入性肾缺氧监测

成人大鼠心脏解剖与功能的超声心动图评估

临时双性肾动脉闭塞的急性肾损伤大型动物模型

使用3D机器人超 声进行小鼠 肾脏体内分析

意识大鼠肾血流量的长期连续测量

在 NAFLD/NASH 的鼠模型中应用超声波和剪切波弹性成像

使用3D机器人超声进行小鼠肾脏体内分析