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| 硅胶微导管套管缩窄术建立小鼠颈总动脉血管重塑模型 |
| 赵峰1,徐强1,2,胡舒婷3,郭志福1,宋晓伟1*,吴弘1* |
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(1. 海军军医大学(第二军医大学)长海医院心血管内科, 上海 200433;
2. 海军905医院心血管内科, 上海 200050;
3. 宁夏医科大学基础医学院生理学教研室, 银川 750004
*通信作者) |
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| 摘要: |
| 目的 通过硅胶微导管套管缩窄术建立一种新的小鼠颈总动脉血管重塑模型，分析血管缩窄时间与血管重塑之间的关系。方法 12只C57BL/6小鼠经异氟烷麻醉后，取颈正中切口，分离左、右侧颈总动脉。左侧颈总动脉行缩窄术，将长1 cm、内径0.3 mm的硅胶微导管纵向剖开后套入颈总动脉，固定并扎紧；右侧颈总动脉仅行假手术，不放置硅胶微导管，不进行缩窄术。在术后2周和4周时分别处死6只小鼠，3只用于组织病理学观察，3只用于RNA和蛋白提取。取小鼠两侧颈总动脉制作病理切片，采用H-E染色观察颈总动脉血管的显微结构，免疫组织化学染色检测颈总动脉组织中血管平滑肌细胞增殖细胞核抗原（PCNA）的表达；取两侧颈总动脉RNA和蛋白样品，采用qRT-PCR检测血管平滑肌表型转化标志分子钙调理蛋白1（CNN1）、平滑肌肌动蛋白α2（ACTA2）的表达，蛋白质印迹法检测细胞增殖相关分子周期蛋白A2、周期蛋白D1和PCNA的表达。结果 12只小鼠左侧颈总动脉套管缩窄手术均获得成功，套管扎紧后即刻可见血管管腔狭窄但血流未被阻断，术后2周和4周取材时小鼠均无明显异常表现。H-E染色结果显示，术后2周时小鼠左侧颈总动脉血管外膜组织明显增生，4周时小鼠左侧颈总动脉血管中膜和内膜明显增生。免疫组织化学染色结果显示，术后2周和4周时小鼠左侧颈总动脉出现大量PCNA表达，提示平滑肌细胞异常增殖。qRT-PCR检测结果显示，术后2周和4周时小鼠左侧颈总动脉收缩型平滑肌细胞标志分子CNN1、ACTA2的表达水平均降低。蛋白质印迹法检测结果显示，术后2周和4周时小鼠左侧颈总动脉细胞周期蛋白A2、周期蛋白D1和PCNA的表达水平均升高。结论 硅胶微导管套管致小鼠颈总动脉缩窄是一种简单、高效的血管重塑模型。 |
| 关键词: 血管重塑 血管平滑肌表型转化 颈总动脉缩窄 微导管 内膜增生 |
| DOI：10.16781/j.0258-879x.2021.08.0875 |
| 投稿时间：2021-04-30修订日期：2021-08-03 |
| 基金项目:国家自然科学基金（81760076，82070419），上海市优秀学术带头人（青年）计划（19XD1423600）. |
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| Establishment of a mouse common carotid artery remodeling model by silicone microcatheter cannula constriction |
| ZHAO Feng1,XU Qiang1,2,HU Shu-ting3,GUO Zhi-fu1,SONG Xiao-wei1*,WU Hong1* |
(1. Department of Cardiovasology, Changhai Hospital, Naval Medical University(Second Military Medical University), Shanghai 200433, China;
2. Department of Cardiovasology, No. 905 Hospital of PLA Navy, Shanghai 200050, China;
3. Department of Physiology, College of Basic Medical Sciences, Ningxia Medical University, Yinchuan 750004, Ningxia Hui Autonomous Region, China
*Corresponding authors) |
| Abstract: |
| Objective To establish a new mouse model of common carotid artery remodeling by silicone microcatheter cannula constriction, and to analyze the relationship between vascular constriction time and vascular remodeling. Methods Twelve C57BL/6 mice were anesthetized with isoflurane. The median cervical incision was taken to separate the left and right common carotid arteries. The left common carotid artery was constricted by a silicone microcatheter (1 cm in length and 0.3 mm in internal diameter), which was longitudinally cut and inserted into the common carotid artery, fixed and tied tightly; the right common carotid artery received sham operation without silicone microcatheter and constriction. Six mice were sacrificed at 2 and 4 weeks after operation, 3 for histopathological observation and 3 for RNA and protein extraction. Pathological sections were made from both common carotid arteries of mice. The microstructure of common carotid arteries was observed by hematoxylin-eosin (H-E) staining, and the expression of proliferating cell nuclear antigen (PCNA) in vascular smooth muscle cells of common carotid arteries was detected by immunohistochemical staining; RNA and protein samples were extracted from both common carotid arteries, the expression of molecular markers (calponin 1 [CNN1] and actin alpha 2, smooth muscle [ACTA2]) for smooth muscle cell phenotype switches was detected by quantitative real-time polymerase chain reaction (qRT-PCR), and the expression of proliferation markers (cyclinA2, cyclinD1 and PCNA) was detected by Western blotting. Results The left common carotid artery cannula constriction was successfully performed in all the 12 mice. The vascular lumen stenosis was observed immediately after the cannula was tightened, but the blood flow was not blocked. There were no obvious abnormal manifestations in the mice at 2 or 4 weeks after operation. H-E staining showed that the adventitia of the left common carotid artery proliferated significantly at 2 weeks, and the intima and media of the left common carotid artery proliferated significantly at 4 weeks. Immunochemical staining revealed abundant PCNA expression in the left common carotid artery at 2 and 4 weeks after operation, suggesting abnormal proliferation of smooth muscle cells. Additionally, qRT-PCR showed that the expression levels of contraction markers of smooth muscle cells (CNN1 and ACTA2) in the left common carotid artery were significantly decreased at 2 and 4 weeks after operation. Western blotting showed that the expression levels of the proliferation markers (cyclinA2, cyclinD1 and PCNA) were all increased in the left common carotid artery at 2 and 4 weeks after operation. Conclusion The constriction of common carotid artery with a silicone microcatheter cannula is a simple and efficient method for the construction of vascular remodeling model. |
| Key words: vascular remodeling vascular smooth muscle phenotype switch common carotid arterial constriction microcatheter neointima formation |
