中国神经再生研究(英文版)

中国神经再生研究(英文版) ›› 2019, Vol. 14 ›› Issue (3): 470-479.doi: 10.4103/1673-5374.245471

• 原著:脑损伤修复保护与再生 • 上一篇    下一篇

中药小续命汤提取物可调节慢性脑低灌注海马组织蛋白的表达

  

  • 出版日期:2019-03-15 发布日期:2019-03-15
  • 基金资助:

    中国国家自然科学基金项目（81473383），国家科技重大专项重大新药创制（2018ZX09711001-003-019），北京协和医学院研究生创新基金项目（2017-1007-02）

Xiao-Xu-Ming decoction extract regulates differentially expressed proteins in the hippocampus after chronic cerebral hypoperfusion

Yue-Hua Wang 1, 2 , Ying-Lin Yang 2 , Xiao Cheng 2 , Jun Zhang 2 , Wan Li 1, 2 , Guan-Hua Du 1, 2   

  1. 1 State Key Laboratory of Bioactive Substance and Function of Natural Medicines, Institute of Materia Medica, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing, China 2 Beijing Key Laboratory of Drug Target Identification and Drug Screening, Institute of Materia Medica, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing, China
  • Online:2019-03-15 Published:2019-03-15
  • Contact: Yue-Hua Wang, PhD, wangyuehua@pku.org.cn; Guan-Hua Du, PhD, dugh@imm.ac.cn.
  • Supported by:

    This study was supported in part by the National Natural Science Foundation of China, No. 81473383 (to YHW); the Significant New-Drugs Creation of Science and Technology Major Projects in China, No. 2018ZX09711001-003-019 (to YHW); the Innovation Fund for Graduate of Beijing Union Medical College of China, No. 2017-1007-02 (to XC).

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摘要：

中药方剂小续命汤由防己、麻黄、防风、杏仁、干姜、人参、甘草、乌头、黄芩、肉桂、川芎和白芍构成，在中国已广泛用于治疗卒中及其后遗症。作者既往研究发现其活性成分可减轻脑缺血损伤，但其调节的总蛋白谱和信号传导通路还不明确。为此，实验以双侧颈总动脉闭塞建立慢性脑低灌注大鼠模型，造模后26d起，连续4周灌胃小续命汤提取物50和150 mg/kg。(1)以Morris水迷宫测试学习记忆能力，以抓握测试检测运动能力，以斜板测试检测协调能力，免疫荧光染色检测神经元可塑性，以无标记蛋白质组学分析海马组织蛋白成分变化及蛋白相互关系，以实时PCR及Western blot验证变化蛋白的变化；(2)结果显示，小续命汤提取物能明显改善慢性脑低灌注大鼠的学习记忆能力、运动能力以及协调能力，海马组织中微管相关蛋白2表达时明显增加。海马组织中13种上调蛋白的表达被抑制， 39种下调蛋白被增加，这些蛋白参与氧化还原过程、细胞内信号级联过程和蛋白质分解代谢过程等多个生理过程，信号通路主要涉及泛素介导的蛋白水解和磷脂酰肌醇信号系统，且其中Rab2a，Ptpn1，Ppm1e，Cdk18，Gorasp2，Eps15，Capza2，Syngap1和Mt-nd1存在相互作用。实时PCR和Western blot验证验证了NADH-泛素氧化还原酶链1的表达情况与蛋白组学检测的结果一致；(3)上述结果揭示了小续命汤提取物治疗慢性脑低灌注的新分子机制。

orcid: 0000-0002-7650-6417(Yue-Hua Wang)

关键词:
慢性脑低灌注, 血管性痴呆, 双侧颈总动脉闭塞, 小续命汤, 无标记定量蛋白组学, Morris水迷宫, 纳升液联用色谱技术, NADH-泛素氧化还原酶链1

Abstract:

Xiao-Xu-Ming decoction has been widely used to treat stroke and sequelae of stroke. We have previously shown that the active fractions of Xiao-Xu-Ming decoction attenuate cerebral ischemic injury. However, the global protein profile and signaling conduction pathways reg­ulated by Xiao-Xu-Ming decoction are still unclear. This study established a two-vessel occlusion rat model by bilateral common carotid artery occlusion. Rats were intragastrically administered 50 or 150 mg/kg Xiao-Xu-Ming decoction for 4 consecutive weeks. Learning and memory abilities were measured with Morris water maze. Motor ability was detected with prehensile test. Coordination ability was exam­ined using the inclined screen test. Neuronal plasticity was observed by immunofluorescent staining. Differentially expressed proteins of rat hippocampus were analyzed by label-free quantitative proteomics. Real time-polymerase chain reaction and western blot assay were used to identify the changes in proteins. Results showed that Xiao-Xu-Ming decoction dramatically alleviated learning and memory deficits, and motor and coordination dysfunction, and increased the expression of microtubule-associated protein 2. Xiao-Xu-Ming decoction extract remarkably decreased 13 upregulated proteins and increased 39 downregulated proteins. The regulated proteins were mainly involved in oxidation reduction process, intracellular signaling cascade process, and protein catabolic process. The signaling pathways were mainly involved in ubiquitin mediated proteolysis and the phosphatidylinositol signaling system. Furthermore, there was an interaction among Rab2a, Ptpn1, Ppm1e, Cdk18, Gorasp2, Eps15, Capza2, Syngap1 and Mt-nd1. Protein analyses confirmed the changes in expression of MT-ND1. The current findings provide new insights into the molecular mechanisms of Xiao-Xu-Ming decoction extract’s effects on chronic cerebral hypoperfusion.

Key words: nerve regeneration, chronic cerebral hypoperfusion, vascular dementia, bilateral common carotid artery occlusion, Xiao-Xu-Ming decoction, label-free quantitative proteomics, Morris water maze test, Nano-LC-ESI LTQ-Orbitrap MS/MS technology, neural regeneration