Protective Effect and Mechanism of Sanhuatang and Its Modifications on Cerebral Ischemia-reperfusion Injury in Rats

Zi-han GONG; Wen-jie LI; Ming-jie SUN; Ying HUANG

doi:10.13422/j.cnki.syfjx.20220102

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Protective Effect and Mechanism of Sanhuatang and Its Modifications on Cerebral Ischemia-reperfusion Injury in Rats

更新时间：2021-12-14

- Protective Effect and Mechanism of Sanhuatang and Its Modifications on Cerebral Ischemia-reperfusion Injury in Rats
- Chinese Journal of Experimental Traditional Medical Formulae Vol. 28, Issue 2, Pages: 11-18(2022)
- 作者机构：
  
  1.中国中医科学院中医基础理论研究所，北京 100700
  2.中国中医科学院医学实验中心，中医药防治重大疾病基础研究北京市重点实验室，北京 100700
- 作者简介：
- 基金信息：
- DOI：10.13422/j.cnki.syfjx.20220102
  CLC： R2-0;R22;R285.5;R289;R33
- Published：20 January 2022，
  
  Published Online：15 November 2021，
  
  Received：25 August 2021，
- 稿件说明：
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巩子汉,李雯洁,孙明杰等.三化汤及其拆方对大鼠脑缺血再灌注损伤的保护作用及机制[J].中国实验方剂学杂志,2022,28(02):11-18.

GONG Zi-han,LI Wen-jie,SUN Ming-jie,et al.Protective Effect and Mechanism of Sanhuatang and Its Modifications on Cerebral Ischemia-reperfusion Injury in Rats[J].Chinese Journal of Experimental Traditional Medical Formulae,2022,28(02):11-18.
巩子汉,李雯洁,孙明杰等.三化汤及其拆方对大鼠脑缺血再灌注损伤的保护作用及机制[J].中国实验方剂学杂志,2022,28(02):11-18. DOI： 10.13422/j.cnki.syfjx.20220102.

GONG Zi-han,LI Wen-jie,SUN Ming-jie,et al.Protective Effect and Mechanism of Sanhuatang and Its Modifications on Cerebral Ischemia-reperfusion Injury in Rats[J].Chinese Journal of Experimental Traditional Medical Formulae,2022,28(02):11-18. DOI： 10.13422/j.cnki.syfjx.20220102.

摘要

目的

观察三化汤及其拆方对脑缺血再灌注大鼠脑组织的保护作用，探讨三化汤的作用机制及其组方配伍特点。

方法

将140只清洁级SD大鼠随机分为正常组，假手术组及模型组，造模组采用Longa线栓法制作大鼠脑缺血再灌注损伤（CIRI）动物模型，后按照随机数字表法分为模型组，尼莫地平组，三化汤去羌活组，羌活组及三化汤组，灌胃给药5 d。采用2，3，5-氯化三苯基四氮唑（TTC）染色法检测各组大鼠脑组织梗死面积，苏木素-伊红（HE）染色及电镜检测各组大鼠脑组织病理变化情况，实时荧光定量聚合酶链式反应（Real-time PCR）检测脑组织紧密连接闭合蛋白-5（Claudin-5），咬合蛋白（Occludin）及闭锁小带蛋白-1（ZO-1） mRNA表达水平，蛋白免疫印迹法（Western blot）检测脑组织Claudin-5，Occludin及ZO-1蛋白表达水平。

结果

与正常组比较，模型组大鼠脑组织梗死灶显著，病理形态及超微结构改变明显，Claudin-5，Occludin及ZO-1 mRNA及蛋白表达水平显著降低（

0.01）；与模型组比较，尼莫地平组、三化汤组大鼠脑组织梗死面积显著降低（

0.01），病理损伤减轻；三化汤去羌活组及羌活组大鼠脑组织梗死面积及病理改变有减小趋势，差异无统计学意义；与模型组比较，尼莫地平组、三化汤组、羌活组大鼠脑组织Claudin-5，Occludin，ZO-1 mRNA及蛋白表达水平明显升高（

0.05，

0.01）；与三化汤去羌活组比较，羌活组大鼠脑组织Claudin-5，ZO-1 mRNA及蛋白表达水平明显升高（

0.05，

0.01）。

结论

三化汤对大鼠脑缺血再灌注损伤具有保护作用，其机制可能与调节Claudin-5，Occludin及ZO-1表达，改善血脑屏障的功能密切相关。其中羌活可能在三化汤保护大鼠脑缺血再灌注损伤中发挥了重要作用。

Abstract

Objective

To observe the protective effect of Sanhuatang and its modifications on the brain tissue of rats exposed to cerebral ischemia-reperfusion injury （CIRI） and explore its action mechanism and compatibility characteristics.

Method

One hundred and forty SD male rats of clean grade were randomly divided into the control group， sham-operation group， and operation group. The Longa suture method was employed to establish the CIRI model. The successfully modeled CIRI rats were further divided into five groups， namely the model group， nimodipine group， Sanhuatang without Notopterygii Rhizoma et Radix group， Notopterygii Rhizoma et Radix group， and Sanhuatang group， and treated with the corresponding medicines by gavage for five days. The cerebral infarct size in each group was examined by 2，3，5-triphenyltetrazolium chloride （TTC） staining， and the pathological changes in the brain tissue were observed by hematoxylin-eosin （HE） staining and electron microscopy. The mRNA and protein expression levels of Claudin-5， Occludin， and zonula occludens-1 （ZO-1） in brain tissues were detected by real-time fluorescence quantitative polymerase chain reaction （Real-time PCR） and Western blot， respectively.

Result

Compared with the control group， the model group exhibited markedly increased infarct size， obvious changes in brain morphology and ultrastructure， and down-regulated mRNA and protein expression of Claudin-5， Occludin， and ZO-1 （

0.01）. Compared with the model group， both nimodipine and Sanhuatang significantly decreased the infarct size （

0.01） and relived the pathological changes. The infarct sizes in the Sanhuatang without Notopterygii Rhizoma et Radix group and Notopterygii Rhizoma et Radix group were reduced without exhibiting a statistically significant difference. The mRNA and protein expression levels of Claudin-5， Occludin， and ZO-1 in the nimodipine group， Sanhuatang group， and Notopterygii Rhizoma et Radix group were up-regulated significantly in comparison with those in the model group （

0.01，

0.01）. The mRNA and protein expression levels of Claudin-5 and ZO-1 were higher in the Notopterygii Rhizoma et Radix group than in the Sanhuatang without Notopterygii Rhizoma et Radix group （

0.01，

0.01）.

Conclusion

Sanhuatang exerts the protective effect against CIRI in rats possibly by regulating the expression of Claudin-5， Occludin， and ZO-1 and improving the blood-brain barrier function. Notopterygii Rhizoma et Radix in Sanhuatang may play an important role in the protection of rats from CIRI.

关键词

三化汤羌活脑缺血/再灌注血脑屏障紧密连接闭合蛋白-5（Claudin-5）咬合蛋白（Occludin）闭锁小带蛋白-1（ZO-1）

Keywords

SanhuatangNotopterygii Rhizoma et Radixcerebral ischemia-reperfusion injury （CIRI）blood-brain barrierClaudin-5Occludinzonula occludens-1 （ZO-1）

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