中国组织工程研究

中国组织工程研究 ›› 2023, Vol. 27 ›› Issue (11): 1708-1714.doi: 10.12307/2023.183

• 组织构建实验造模 experimental modeling in tissue construction • 上一篇    下一篇

线栓法与化学损毁法造模引起模型大鼠吞咽功能损害的对比

华晓琼1，李彦杰2，李斯锦1，张淑芹1，刘昊源1，丁慧敏1，赵楠楠1   

  1. 1河南中医药大学康复医学院，河南省郑州市  450046；2河南省中医院康复科，河南省郑州市  450046
  • 收稿日期:2022-05-12 接受日期:2022-06-17 出版日期:2023-04-18 发布日期:2022-09-27
  • 通讯作者: 李彦杰，硕士，主任中医师，硕士生导师，河南省中医院康复科，河南省郑州市 450046
  • 作者简介:华晓琼，女，1997年生，河南中医药大学在读硕士，主要从事脑病、脊髓损伤的康复研究。
  • 基金资助:
    河南省科技攻关计划项目(222102310569)，项目负责人：李彦杰；河南省中医药科学研究专项课题(重点课题)(2019ZYBJ14)，项目负责人：李彦杰；河南省中医药科学研究专项课题(2022ZY1083)，项目负责人：李彦杰；河南省中医药拔尖人才培养项目(豫中医科[2018]35号)，项目负责人：李彦杰

Suture-occluded method versus chemical destruction method to induce swallowing disorders in model rats

Hua Xiaoqiong1, Li Yanjie2, Li Sijin1, Zhang Shuqin1, Liu Haoyuan1, Ding Huimin1, Zhao Nannan1   

  1. 1Rehabilitation School of Medicine, Henan University of Chinese Medicine, Zhengzhou 450046, Henan Province, China; 2Department of Rehabilitation, Henan Province Hospital of TCM, Zhengzhou 450046, Henan Province, China
  • Received:2022-05-12 Accepted:2022-06-17 Online:2023-04-18 Published:2022-09-27
  • Contact: Li Yanjie, Master, Chief physician, Master’s supervisor, Department of Rehabilitation, Henan Province Hospital of TCM, Zhengzhou 450046, Henan Province, China
  • About author:Hua Xiaoqiong, Master candidate, Rehabilitation School of Medicine, Henan University of Chinese Medicine, Zhengzhou 450046, Henan Province, China
  • Supported by:
    Henan Province Science and Technology Research Project, No. 222102310569 (to LYJ); Henan Province Traditional Chinese Medicine Scientific Research Special Project (Key Project), No. 2019ZYBJ14 (to LYJ); Henan Province Traditional Chinese Medicine Scientific Research Special Project, No. 2022ZY1083 (to LYJ); Henan Province Traditional Chinese Medicine Top-notch Talent Training Project, No. [2018]35 (to LYJ)

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摘要：

文题释义：
吞咽：是一个非常复杂的反射过程，属于多水平的调节，控制吞咽的神经机制是从传入到传出的过程。吞咽主要受3个方面的神经调控：①外周神经，主要调控支配舌体和吞咽肌群的相关活动；②脑干吞咽中枢，是吞咽的核心部位，能反射性地调节吞咽活动；③皮质吞咽中枢，起着协调自主吞咽及控制口咽的作用。脑干吞咽中枢模式发生器的完整性在吞咽正常活动中起着重要的作用。中枢模式发生器包括了3个部分，孤束核及周围的网状结构、疑核以及疑核上的延髓腹外侧区部分。
化学损毁：通过对神经元的极度兴奋作用而造成注射部位神经元胞体的死亡而不损伤其过路纤维，较电解损毁能够更准确地判断神经核团的作用。NMDA受体激动剂鹅膏蕈氨酸与内源性兴奋性氨基酸的作用基本相同，均可造成细胞过度兴奋而死亡，对不同类型神经元的敏感性差异不大，相比海人酸其毒性更低，可避免远隔损害，能使目标区域产生满意的局灶性损毁，又不会引发癫痫，所以损毁实验中其应用相对普遍，具有较好的可行性。

背景：建立动物模型是研究卒中后吞咽障碍的重要研究方法之一，线栓法是缺血性脑卒中最常用的经典模型，新近研究表明，化学损毁吞咽中枢疑核也能造成大鼠表现吞咽功能障碍，但其造模成功率、临床相似性、可调控性仍不确定。
目的：观察线栓法与化学损毁法两种不同造模方法对大鼠模型的吞咽功能改变，探究能较完整表现与人类相似的吞咽障碍临床及病理特征。
方法：将40只SD大鼠随机分为3组，正常组10只，线栓组15只，化学损毁组15只，线栓组以线栓法短暂脑缺血90 min后再灌注制备左侧卒中脑缺血模型，化学损毁组以鹅膏蕈氨酸化学损毁疑核造成大鼠吞咽障碍模型。记录各组大鼠每周存活数、体质量及24 h进食、进水量，在造模后第2，7，14，30天时采用生物信号采集器及张力换能器检测大鼠吞咽启动反应时间及吞咽次数，并采用苏木精-伊红染色观察延髓吞咽中枢疑核处病理学形态变化，ELISA法观察各组大鼠血清的白细胞介素6、白细胞介素1β、肿瘤坏死因子α的表达变化。
结果与结论：①和化学损毁组相比，线栓组大鼠死亡率在第2，7天显著降低(P < 0.05)，其余时间段无明显差异(P > 0.05)，且线栓组最终死亡率明显低于化学损毁组(40%，73%，P < 0.05)；②与正常组相比，造模后第7天化学损毁组大鼠体质量及24 h进食、进水量均减少、吞咽启动反应时间延长、吞咽次数减少，差异均有显著性意义(P < 0.05)，线栓组无明显差异(P > 0.05)；与正常组相比，第14天后2个造模组大鼠体质量及24 h进食、进水量均减少，吞咽启动反应时间均延长，吞咽次数均减少，差异均有显著性意义(P < 0.05)；线栓组和化学损毁组相比，大鼠体质量及24 h进食、进水量、吞咽启动反应时间、吞咽次数在第7天差异有显著性意义(P < 0.05)，其余时间段无明显差异(P > 0.05)；③2个造模组大鼠的延髓处均较出现明显病理性改变，延髓处炎症细胞均明显增多；线栓组和化学损毁组相比，病理性改变不明显；④提示无论是线栓法和化学损毁疑核法都可引起大鼠吞咽功能改变；线栓组的死亡率在造模后的所有时间段内均低于化学损毁组，从模型最终的成功率而言，线栓法短暂脑缺血90 min后再灌注大鼠模型更具复制意义。

https://orcid.org/0000-0001-7332-9876(华晓琼)

中国组织工程研究杂志出版内容重点：组织构建；骨细胞；软骨细胞；细胞培养；成纤维细胞；血管内皮细胞；骨质疏松；组织工程

关键词: 吞咽功能, 化学损毁, 疑核, 脑缺血, 动物模型

Abstract: BACKGROUND: The establishment of animal models is one of the important methods to study swallowing disorders after stroke. Suture-occluded method is the most commonly classic model of ischemic stroke. Recent studies have shown that the chemical destruction of the nucleus ambiguus can also cause swallowing disorders in rats. However, its modeling success rate, clinical similarity, and controllability remain uncertain.
OBJECTIVE: To observe the changes in swallowing function in the rat model established by two different modeling methods, namely, suture-occluded method and chemical destruction method, and to investigate the clinical and pathological characteristics of swallowing disorders similar to those in humans. 
METHODS: Forty Sprague-Dawley rats were randomly divided into three groups: normal group (n=10), suture-occluded group (n=15), and chemical destruction group (n=15). The left-sided ischemic stroke models were prepared using the suture-occluded method through reperfusion after 90 minutes of transient cerebral ischemia in the suture-occluded group, while rat models of swallowing disorders were established through the chemical destruction of the nucleus ambiguus by gooseberry muscarinic acid in the chemical destruction group. Survival numbers per week, body mass, and 24-hour food and water intake in each group were recorded. The rats were examined on days 2, 7, 14, and 30 after modeling using a biosignal collector and a tension transducer to detect the swallowing initiation response time and the number of swallows. Hematoxylin-eosin staining was used to observe the pathomorphological changes of the nucleus ambiguus of the medulla oblongata swallowing center. ELISA method was used to detect the levels of serum interleukin-6, interleukin-1β and tumor necrosis factor-α.
RESULTS AND CONCLUSION: The mortality rate in the suture-occluded group was significantly lower than that in the chemical destruction group on days 2 and 7 (P < 0.05), but there was no significant difference between the two groups at other observation time points (P > 0.05). The final mortality rate in the suture-occluded group was significantly lower than that in the chemical destruction group (40% vs.73%, P < 0.05). Compared with the normal group, body mass, 24-hour food and water intake, swallowing initiation response time, and the number of swallows were significantly worse in the chemical destruction group on day 7 (P < 0.05) but showed no significant changes in the suture-occluded group (P > 0.05). Compared with the normal group, the other two model groups showed a significant reduction in body mass, 24-hour food and water intake, and the number of swallows and significantly prolonged swallowing initiation response time (P < 0.05). There were significant differences in body mass, 24-hour food and water intake, swallow initiation response time, and the number of swallows between the suture-occluded group and the chemical destruction group on day 7 (P < 0.05), but no significant difference at other observation time points (P > 0.05). Rats in the two model groups showed obvious pathological changes in the medulla oblongata with a significant increase in inflammatory cells. However, there was no significant difference between the two groups (P > 0.05). To conclude, both the suture-occluded method and the chemical destruction method could cause swallowing disorders in rats. The mortality rate in the suture-occluded group was significantly lower than that in the chemical destruction group at all times after modeling. Therefore, the suture-occluded method with reperfusion following 90 minutes of transient ischemia is more replicative in terms of the final success rate.

Key words: swallowing function, chemical destruction, nucleus ambiguus, cerebral ischemia, animal model

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