出生后右心室容积超负荷小鼠模型的建立与确认
1Department of Pediatric Intensive Care Unit, Shanghai Children’s Medical Center, School of Medicine,Shanghai Jiaotong University, 2Department of Thoracic and Cardiovascular Surgery, Shanghai Children’s Hospital, School of Medicine,Shanghai Jiaotong University, 3Department of Plastic and Reconstructive Surgery, Shanghai Ninth People’s Hospital, School of Medicine,Shanghai Jiao Tong University, 4Department of Radiology, Huangpu Branch, Shanghai Ninth People’s Hospital,Shanghai Jiao Tong University School of Medicine, 5Institute of Pediatric Translational Medicine, Shanghai Children’s Medical Center, School of Medicine,Shanghai Jiaotong University, 6Shanghai Institute for Pediatric Congenital Heart Disease, Shanghai Children’s Medical Center, School of Medicine,Shanghai Jiaotong University
Summary
该方案介绍了腹动静脉瘘（AVF）小鼠出生后右心室容积超负荷（VO）模型的建立和确认，该模型可用于研究VO如何促进出生后心脏发育。
Abstract
右心室 （RV） 容量超负荷 （VO） 在先天性心脏病患儿中很常见。鉴于不同的发育阶段，与成人相比，儿童心室心肌对VO的反应可能不同。本研究旨在使用改良的腹动静脉瘘在小鼠中建立出生后 RV VO 模型。为了确认 VO 的产生以及 RV 的以下形态学和血流动力学变化，进行了为期 3 个月的腹部超声、超声心动图和组织化学染色。结果，在出生后小鼠中进行的手术显示出可接受的存活率和瘘管成功率。在VO小鼠中，RV腔增大，游离壁增厚，术后2个月内每搏输出量增加约30%-40%。此后右心室收缩压升高，观察到相应的肺动脉瓣反流，并出现小肺动脉重塑。综上所述，改良动静脉瘘（AVF）手术在出生后小鼠中建立RV VO模型是可行的。考虑到瘘管闭合和肺动脉阻力升高的概率，在应用前必须进行腹部超声和超声心动图以确认模型状态。
Introduction
右心室 （RV） 容量超负荷 （VO） 在先天性心脏病 （CHD） 患儿中很常见，可导致病理性心肌重塑和长期预后不良 1,2,3。深入了解左心室重塑和相关的早期靶向干预对于冠心病患儿的良好预后至关重要。成人和儿童心脏的分子结构、生理功能和对刺激的反应存在一些差异1,4,5,6。例如，在压力超负荷的影响下，心肌细胞增殖是新生儿心脏的主要反应，而纤维化发生在成人心脏中5,6。此外，许多治疗成人心力衰竭的有效药物对儿童心力衰竭没有治疗作用，甚至可能造成进一步的损害7,8。因此，从成年动物中得出的结论不能直接应用于幼年动物。
几十年来，动静脉瘘 （AVF） 模型已被用于在不同物种的成年动物中诱导慢性心脏 VO 和相应的心功能障碍 9,10,11,12,13。然而，人们对出生后小鼠的模型知之甚少。在我们之前的研究中，通过创建腹部 AVF 成功生成了 VO 出生后小鼠模型。出生后心脏中RV发育轨迹的变化也被证明是14,15,16,17。
为了探索本模型的基本改良手术过程和特点，提出了详细的方案;在这项研究中，该模型进行了为期 3 个月的评估。
Protocol
本文介绍的所有程序均符合《赫尔辛基宣言》中概述的原则，并得到了上海儿童医学中心动物福利与人类研究委员会（SCMC-LAWEC-2023-003）的批准。本研究使用C57BL / 6小鼠幼崽（P7，雄性，3-4g）。这些动物是从商业来源获得的（见 材料表）。将小鼠幼崽及其哺乳母亲（幼崽：单个笼子中的母亲= 6：1）保持在无特定病原体的实验室条件下，在22±2°C下进行12小时光暗循环，自由获得水和营?…
Representative Results
3个月内的生存率和AVF通畅率VO组共有30只（75%）小鼠和假手术组19只（95%）小鼠在AVF手术中存活（图4A）。在VO组中，8只小鼠在手术后1天内因出血过多（n = 5）或自相残杀（n = 3）而死亡，而2只小鼠在1个月时因不明原因死亡。 在存活的VO小鼠（n = 30）中，超声证实术后21只小鼠成功建立了瘘管，这些小鼠在术后1周（P14）时被证明是未闭的?…
Discussion
以前，经典的 RV VO 模型是使用瓣膜反流21 创建的;然而，与AVF相比，心内直视瓣膜手术可能需要更复杂的技术，并且可能与显着更高的死亡率有关，特别是在出生后小鼠中。由于动物研究表明，AVF22 已达到与 VO 相同的效果，因此本研究使用了创伤较小的改良腹瘘手术。
在成功建立瘘管的过程中考虑了某些因素。首先，该程序是在未进行气?…
Disclosures
The authors have nothing to disclose.
Acknowledgements
这项工作得到了国家自然科学基金（第82200309项）和宁波市杰出医疗团队创新项目（第2022020405项）的支持
Materials
| 70% Ethanol | Tiandz,Chia | ||
| ACETAMINOPHEN Oral Solution | VistaPharm, Inc. Largo, FL 33771, USA | NDC 66689-054-01 | |
| Anesthesia machine | RWD Life Science,China | R550IP | |
| Anesthesia mask | RWD Life Science,China | 68680 | |
| C57BL/6 mice | Xipu’er-bikai Experimental Animal Co., Ltd (Shanghai, China) | ||
| Hair removal cream | Veet, France | VT-200 | |
| Hematoxylin and eosin Kit | Beyotime biotech | C0105M | |
| Isoflurane | RWD Life Science,China | R510-22-10 | |
| Microscope | Yuyan Instruments, China | SM-301 | |
| Surgical suture needles | NINGBO MEDICAL NEEDLE CO.,LTD, China | ||
| Thermostatic heating platform | Qingdao Juchuang Environmental Protection Group Co., Ltd, China | ||
| Ultrasound device | FUJIFILM VisualSonics, Inc. | Vevo 2100 | Image modes includes B-Mode, Color Doppler Mode and Pulsed Wave Doppler Mode |
| Ultrasound gel | Parker Laboratories,United States | REF 01-08 | |
| Ultrasound transducer | FUJIFILM VisualSonics, Inc. | MS 400 |
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Cite This Article
Sun, S., Zhu, H., Wang, S., Xu, X., Ye, L. Establishment and Confirmation of a Postnatal Right Ventricular Volume Overload Mouse Model. J. Vis. Exp. (196), e65372, doi:10.3791/65372 (2023).