β-aminopropionitrile 诱发主动脉瘤和解剖小鼠模型 Aortopathy 的定量显微 CT 分析
Summary
本文描述了一个详细的方法, 使用显影铅基硅橡胶灌注小鼠血管的主动脉直径量化的主动脉瘤和解剖模型。
Abstract
主动脉瘤和解剖与人口中严重的发病率和死亡率有关, 而且可能具有极高的致死性。虽然存在主动脉疾病的动物模型, 但在体内成像的血管已经受到限制。近年来, 微电脑断层扫描 (微 CT) 已成为一种在体内和体外成像大、小血管的首选方式. 结合血管铸型的方法, 我们成功地利用微 CT 来表征β-aminopropionitrile 治疗 C57/Bl6 小鼠主动脉病变的频率和分布。这种方法的技术局限性包括: 不良动物制剂所引入的灌注质量的变化, 适用于容器尺寸量化的适当方法, 以及本程序的不可生存性。本文详细介绍了一种以铅为基础的显影硅橡胶血管内灌注的方法, 用于 aortopathy 在动脉瘤和解剖小鼠模型中的定量表征。除了可视化的主动脉病理, 该方法可用于检查其他血管床在体内或血管床切除后的尸体。
Introduction
主动脉夹层的发生率为3例, 每10万年1。在美国, 主动脉夹层和动脉瘤性疾病每年超过1万人死亡, 占 1-2% 的死亡在西方国家2。主动脉夹层是由血管内膜层撕裂引起的, 在生理压力下通过主动脉壁层的血液传播而引发。患者脉压升高与解剖和并发症发生率增高有关。增加壁面剪应力与主动脉壁扩张有关, 导致动脉瘤形成3,4。主动脉夹层的后果包括阻塞血液流向远端器官, 包括大脑, 肾脏, 内脏和四肢, 形成慢性动脉瘤, 破裂, 或死亡5,6,7。
目前, 在主动脉瘤和解剖的启动和进展中所涉及的生物化学和细胞过程尚不清楚。可再生动物模型的主动脉瘤和解剖是了解其病理生理学的关键。β-aminopropionitrile (BAPN) 是一种赖氨酰氧化酶抑制剂, 它可以防止弹性蛋白和胶原蛋白的交联, 并已被证明大大改变了血管壁细胞外基质的结构及其生物力学完整性6, 8。用 BAPN 治疗的啮齿动物被用作主动脉瘤和解剖9、10的常见动物模型。
血管成像方法有助于识别血管病理, 确认血管通畅, 评价器官灌注。近年来, 微计算机断层扫描 (显微 CT) 已被用来研究小鼠和类似大小动物的血管。与骨不同的是, 血管的轴向成像是有限的, 因为腔内血液本身相对阴性。然而, 当与血管造影剂结合时, 微 CT 可以对动物 vasculatures 进行详细的三维重建, 用于研究宏观解剖血管病理学11。
所选的对比剂 (见材料表) 是一种含有铬酸铅和硫酸铅的显影硅橡胶。在催化剂的存在下灌注时, 它很快硬化形成一个血管的投射, 在血管的宏观解剖结构中最小的改变, 使血管高度显影与背景组织对比时,检查 x 线。这种造影剂是有利的, 因为它易于处理和避免组织退化和船舶损失由于断裂经常与血管铸型腐蚀有关。由于它治愈的最低萎缩12, 血管清除的血液仍然是专利, 并允许准确评估动物宏观血管在非生存实验。以前的工作已经成功地使用了显影硅橡胶-对比在各种动物研究。具体来说, 在可视化冠状动脉, 肾小球, 胎盘和脑循环11,12,13,14,15的适用性已经显示。本文详细介绍了以铅为基础的显影硅橡胶血管内灌注的开放性左心室穿刺方法, 以定量表征 BAPN 诱导的小鼠主动脉病变的显微 CT 表现。
Protocol
Representative Results
Discussion
微 CT 成像可用于提供高度详细和三维的血管病理重建动物模型。通过使用血管造影增强的培养基, 非增强的软组织, 如血管的流明, 可以区别于那些加强。虽然激光多普勒, microangiography, 磁共振血管造影, 组织学与共焦, 或双光子显微镜可用于评估血管床, 他们通常侧重于有限的研究领域和/或仅限于二维评估。微 CT 提供了一种成本效益高的方法来获得详细的血管结构成像, 这有助于了解潜在的血管生…
Disclosures
The authors have nothing to disclose.
Acknowledgements
我们要感谢马克. 史密斯在射线成像方面的帮助。这项工作得到了 nih T32 赠款的支持, 为心血管疾病 (蟒蛇) 的跨学科研究, 美国心脏协会 (SMC) 和 nih R35 补助金 (拿 dk)。
Materials
| Microfil | Flow Tech, Inc | MV-122 | We use yellow, a different color can be ordered as desired. Kit includes MV-Compound, MV-Diluent, and MV-Curing Agent. |
| Heparin (1000 U/mL) | Sagent Pharmaceuticals | 25021-400-10 | |
| Phosphate buffered saline | Corning | 21-031-CV | |
| Isoflurane | Vet One, MWI | 502017 | |
| 3-Aminopropionitrile fumarate salt | Sigma-Aldrich | A3134 | |
| Single syringe pump | Fisher Scientific | 14-831-200 | |
| 27-gauge scalp vein set needle | Exel Int | 26709 | 27G x 3/4", 12" tube |
| Inveon Micro-CT scanner | Siemens Medical Solutions | ||
| Osirix MD | Pimxmeo SARL | Version 8.0.2 | |
| Inveon Research Workplace | Siemens Medical Solutions | Version 4.2 | |
| Rodent Chow | Harlan Teklad | 2018sx |
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