中国组织工程研究

中国组织工程研究 ›› 2019, Vol. 23 ›› Issue (1): 55-60.doi: 10.3969/j.issn.2095-4344.1526

• 骨髓干细胞 bone marrow stem cells • 上一篇    下一篇

补阳还五汤联合骨髓间充质干细胞移植脑缺血再灌注大鼠脑组织紧密连接蛋白的表达

张运克1，2，车志英1，李 可1   

  1. 1河南中医药大学，河南省郑州市 450046；2河南中医药大学第一附属医院，河南省郑州市 450000
  • 修回日期:2018-10-18 出版日期:2019-01-08 发布日期:2018-11-28
  • 通讯作者: 张运克，博士，教授，河南中医药大学，河南省郑州市 450046
  • 作者简介:张运克，男，1966年生，河南省平顶山市人，汉族，2002年黑龙江中医药大学毕业，博士，2007年从中国中医科学院博士后流动站出站，教授，主任医师，博士生导师，博士后合作导师，主要从事中医药防治神经系统疾病的研究。
  • 基金资助:

    国家自然科学基金课题(81072799/H2708)，项目负责人：张运克；河南省基础与前沿技术研究计划(162300410098)，项目负责人：张运克；河南省科技创新杰出人才项目(2018JR0006)，项目负责人：张运克

Effect of Buyang Huanwu Decoction combined with bone marrow mesenchymal stem cell transplantation on expression of tight junction proteins in the brain of cerebral ischemia-reperfusion rats

Zhang Yunke1, 2, Che Zhiying1, Li Ke1   

  1. 1Henan University of Chinese Medicine, Zhengzhou 450046, Henan Province, China; 2First Affiliated Hospital, Henan University of Chinese Medicine, Zhengzhou 450000, Henan Province, China
  • Revised:2018-10-18 Online:2019-01-08 Published:2018-11-28
  • Contact: Zhang Yunke, Henan University of Chinese Medicine, Zhengzhou 450046, Henan Province, China; First Affiliated Hospital, Henan University of Chinese Medicine, Zhengzhou 450000, Henan Province, China
  • About author:Zhang Yunke, MD, Professor, Chief physician, Doctoral supervisor, Henan University of Chinese Medicine, Zhengzhou 450046, Henan Province, China; First Affiliated Hospital, Henan University of Chinese Medicine, Zhengzhou 450000, Henan Province, China
  • Supported by:

    the National Natural Science Foundation of China, No. 81072799/H2708 (to ZYK); the Basic and Frontier Technology Research Program of Henan Province, No. 162300410098 (to ZYK); Henan Provincial Science and Technology Innovation Outstanding Talents Project, No. 2018JR0006 (to ZYK)

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摘要：

文章快速阅读：

文题释义：
血脑屏障的基本结构：由脑的连续毛细血管内皮及其细胞间的紧密连接、完整的基膜、周细胞以及星形胶质细胞脚板围成的神经胶质膜构成。这种结构可使脑组织少受甚至不受循环血液中有害物质的损害，从而保持脑组织内环境的基本稳定，对维持中枢神经系统正常生理状态具有重要的生物学意义。
紧密连接：存在于多种组织器官中，由跨膜蛋白、胞质附着蛋白和细胞骨架蛋白共同组成。跨膜蛋白包括Occludin、Claudin和JAM 3种完整的膜蛋白；胞质附着蛋白有3种，即ZO-1，2，3；细胞骨架蛋白主要成分为纤维状肌动蛋白F-actin。紧密连接的特殊结构使大分子物质难以通透，而只允许水分子和离子从索条衔接处的小孔透过，具有重要的生理作用，主要包括两个方面：一是发挥其渗透性调节功能，通过限制穿过组织的离子与蛋白的运动，维持脑组织内环境；二是有助于在细胞单层间形成化学与离子梯度，从而进行载体转运过程。

 

摘要
背景：急性脑缺血时内皮细胞间的紧密连接受到破坏，紧密连接蛋白转录或表达发生改变，引起血脑屏障通透性增加而形成脑水肿。
目的：旨在从紧密连接蛋白的调控方面，探讨补阳还五汤联合骨髓间充质干细胞移植对脑缺血再灌注大鼠血脑屏障修复的作用机制。
方法：80只大鼠(山东省动物实验中心提供)随机分为假手术组(n=16)，模型组(n=16)，骨髓间充质干细胞组(n=16)，补阳还五汤组(n=16)，补阳还五汤联合骨髓间充质干细胞组(联合组，n=16)。利用改良线栓法制备大脑中动脉栓塞缺血再灌注大鼠模型，造模前3 d开始，补阳还五汤组、联合组给予补阳还五汤15 mL/kg灌胃，假手术组、模型组、骨髓间充质干细胞组给予等量生理盐水灌胃；造模后1 d时，骨髓间充质干细胞组、联合组尾静脉注射含1×106个骨髓间充质干细胞悬液1 mL，其他组给予等量生理盐水。在造模后3，5 d取缺血侧脑组织，采用Western Blot法检测脑组织中Occludin、Claudin-5、JAM-A、ZO-1等紧密连接相关蛋白的表达。
结果与结论：①与假手术组相比，模型组大鼠缺血脑组织中Occludin、Claudin-5、JAM-A、ZO-1的蛋白表达显著下降(P < 0.01)；②与模型组相比，补阳还五汤组、骨髓间充质干细胞组和联合组缺血脑组织中Occludin、Claudin-5、JAM-A、ZO-1的蛋白表达量上升(P < 0.05)，以联合组上调最为明显(P < 0.01)；③结果表明，补阳还五汤联合骨髓间充质干细胞对血脑屏障的修复具有协同作用，可能是通过上调缺血脑组织中Occludin、Claudin、JAM-A、ZO-1的蛋白表达来维持血脑屏障紧密连接的完整性，以实现脑保护的作用。

中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程
ORCID: 0000-0003-1500-1535(张运克)

关键词: 大脑中动脉栓塞, 再灌注损伤, 补阳还五汤, 骨髓间充质干细胞, Occludin, Claudin, JAM-A, ZO-1, 动物模型, 国家自然科学基金, 干细胞

Abstract:

BACKGROUND: In acute cerebral ischemia, the tight junction between endothelial cells is damaged, and variation in the transcription or expression of tight junction proteins increases the blood-brain barrier permeability eventually triggering the formation of brain edema. 
OBJECTIVE: To explore the effect mechanism by which Buyang Huanwu Decoction combined with bone marrow mesenchymal stem cell (BMSCs) transplantation treats a cerebral ischemia-reperfusion rat model via the blood brain barrier. 
METHODS: Eighty rats provided by the Animal Experimental Central of Shandong Province, China, were randomized into sham operation group (n=16), model group (n=16), transplantation group (n=16), Buyang Huanwu Decoction group (n=16), and Buyang Huanwu Decoction+transplantation group (combination group, n=16). Ischemia-reperfusion rat model was established via middle cerebral artery occlusion using improved Zea Longa’s suture-occlusion method. In the Buyang Huanwu Decoction group and combination group, the rats were gavaged with 15 ml/kg Buyang Huanwu Decoction 3 days before modeling, while in the sham operation group, model group and transplantation group, the rats were gavaged with the equal volume of normal saline. The transplantation group and combination group were injected with bone marrow mesenchymal stem cell suspension (1x106 mL, 1 mL) 1 day after modeling, and the other groups were gavaged with the equal volume of normal saline. The expression of Occludin, Claudin-5, Jam-A, and ZO-1 in ischemic brain tissues was detected by western blot assay at 3 and 5 days after modeling. 
RESULTS AND CONCLUSION: Compared with the sham operation group, the protein expression of Occludin, Claudin-5, Jam-A, and ZO-1 in the ischemic brain tissue of rats in the model group decreased significantly (P < 0.01). Compared with the model group, the protein expression levels of Occludin, Claudin-5, Jam-A, and ZO-1 in the rat ischemic brain tissue in the Buyang Huanwu Decoction group, transplantation group and combination group were up-regulated significantly (P < 0.05), with the most significant increase in the combination group (P < 0.01). Buyang Huanwu Decoction combined with bone marrow mesenchymal stem cell transplantation exerts a synergistic effect in blood-brain barrier repair, and keeps the tight junction with the blood-brain barrier through the upregulation of ischemic brain tissue Occludin, Claudin-5, Jam-A and ZO-1protein expression, to achieve the brain protection.

中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程

Key words: Infarction, Middle Cerebral Artery, Reperfusion Injury, Bone Marrow, Mesenchymal Stem Cell Transplantation, Claudins, Tissue Engineering

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