中国组织工程研究 ›› 2024, Vol. 28 ›› Issue (8): 1186-1192.doi: 10.12307/2023.997

• 组织构建细胞学实验 cytology experiments in tissue construction • 上一篇    下一篇

巴豆霜干预脑缺血再灌注损伤大鼠皮质区JNK/p38 MAPK及神经元凋亡的机制

岳  云1,王佩佩2,袁兆鹤1,何生存1,贾戌生3,刘  倩4,李占涛2,付慧玲2,宋  斐1,贾孟辉2,4   

  1. 1宁夏医科大学中医学院,宁夏回族自治区银川市  750000;2银川市第一人民医院中医科,宁夏回族自治区银川市  750000;3宁夏少数民族医药现代化教育部重点实验室,宁夏回族自治区银川市  750000;4宁夏医科大学临床医学院,宁夏回族自治区银川市  750000
  • 收稿日期:2022-12-26 接受日期:2023-02-04 出版日期:2024-03-18 发布日期:2023-07-18
  • 通讯作者: 贾孟辉,教授,主任医师,银川市第一人民医院中医科,宁夏回族自治区银川市 750000;宁夏医科大学临床医学院,宁夏回族自治区银川市 750000
  • 作者简介:岳云,男,1994年生,宁夏回族自治区中卫市人,汉族,2019年大连医科大学中山学院毕业,主要从事中医药防治脑血管病的研究。
  • 基金资助:
    国家自然科学基金(8176150449),项目负责人:贾孟辉

Effects of croton cream on JNK/p38 MAPK signaling pathway and neuronal apoptosis in cerebral ischemia-reperfusion injury rats

Yue Yun1, Wang Peipei2, Yuan Zhaohe1, He Shengcun1, Jia Xusheng3, Liu Qian4, Li Zhantao2, Fu Huiling2, Song Fei1, Jia Menghui2, 4   

  1. 1College of Traditional Chinese Medicine, Ningxia Medical University, Yinchuan 750000, Ningxia Hui Autonomous Region, China; 2Department of Traditional Chinese Medicine, the First People’s Hospital of Yinchuan, Yinchuan 750000, Ningxia Hui Autonomous Region, China; 3Ningxia Key Laboratory of Modernization of Minority Medicine, Ministry of Education, Yinchuan 750000, Ningxia Hui Autonomous Region, China; 
  • Received:2022-12-26 Accepted:2023-02-04 Online:2024-03-18 Published:2023-07-18
  • Contact: Jia Menghui, Professor, Chief physician, Department of Traditional Chinese Medicine, the First People’s Hospital of Yinchuan, Yinchuan 750000, Ningxia Hui Autonomous Region, China; School of Clinical Medicine, Ningxia Medical University, Yinchuan 750000, Ningxia Hui Autonomous Region, China
  • About author:Yue Yun, College of Traditional Chinese Medicine, Ningxia Medical University, Yinchuan 750000, Ningxia Hui Autonomous Region, China
  • Supported by:
    the National Natural Science Foundation of China, No. 8176150449 (to JMH)

摘要:


文题释义:

巴豆霜:巴豆的炮制加工品,主要治疗峻下积滞,逐水消肿,豁痰利咽。用于寒积便秘、乳食停滞、下腹水肿、二便不通、喉风及喉痹。研究显示巴豆霜成分作用的核心靶点主要作用于神经系统。
脑缺血再灌注损伤:是指脑组织持续一定时间的缺血后恢复血流再灌注,其功能不但得不到恢复,还会出现更为严重的脑组织结构及功能损害。目前认为,脑缺血再灌注损伤的发生机制主要和自由基生成、脑内Ca2+超载、兴奋性氨基酸毒性作用、白细胞大量聚集和ATP合成减少等密切相关。


背景:巴豆霜能够激活ERK通路、对神经元细胞具有抗凋亡作用,是否具有抑制JNK、p38通路的激活而发挥抗凋亡的协同效应尚不清楚。

目的:探讨巴豆霜对脑缺血再灌注损伤大鼠缺血侧皮质区神经元损伤和凋亡的影响及机制。 
方法:①将90只SD大鼠随机分为假手术组,模型组,巴豆霜低、中、高剂量组,尼莫地平组,每组15只,除假手术组外,剩余各组均采取线栓法制备大脑中动脉栓塞大鼠模型,巴豆霜各组大鼠分别按剂量20 ,40,60 mg/kg灌胃;假手术组和模型组大鼠给予等量生理盐水,1次/d,连续给药7 d。运用神经功能缺损评分、TTC染色、脑组织含水量、苏木精-伊红染色及尼氏染色筛选出最佳浓度即巴豆霜高剂量。②将120只SD大鼠随机分为假手术组、模型组、巴豆霜组、JNK抑制剂组、巴豆霜+JNK抑制剂组、p38 MAPK抑制剂组、巴豆霜+p38 MAPK抑制剂组和尼莫地平组,每组15只,除假手术组外,剩余各组大鼠均制备大脑中动脉栓塞大鼠模型,在造模之前30 min,将10 μL JNK抑制剂SP600125或10 μL p38 MAPK抑制剂SB203580分别注射到大鼠侧脑室,巴豆霜各组大鼠灌胃60 mg/kg巴豆霜,7 d 后,采用Western Blot、TUNEL染色及流式细胞术检测各组大鼠脑组织JNK/p38 MAPK信号通路及凋亡相关蛋白水平及细胞凋亡情况。

结果与结论:①与假手术组相比,模型组神经功能缺损评分、脑含水量、脑梗死体积、细胞凋亡率显著升高(P < 0.05),神经细胞呈散乱分布;与模型组相比,巴豆霜中、高剂量组和尼莫地平组大鼠神经功能缺损评分、脑组织含水量、脑梗死体积显著降低(P < 0.05),神经细胞病理形态明显改善;②与JNK抑制剂组相比,巴豆霜+抑制剂组大鼠脑组织p-JNK/JNK、p-p38/p38、Bax表达显著降低(P < 0.05),细胞凋亡率显著降低(P < 0.05),而Bcl-2表达显著升高(P < 0.05);③结果提示,巴豆霜可能通过抑制JNK/p38 MAPK信号通路的激活、减少神经元凋亡等途径,达到对脑缺血再灌注损伤大鼠的神经保护作用。

https://orcid.org/0000-0001-6982-5308(岳云)

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: 巴豆霜, 脑缺血再灌注损伤, 神经元凋亡, 信号通路, 大鼠

Abstract: BACKGROUND: Croton cream can activate ERK pathways and have anti-apoptotic effects on neuronal cells. It is not clear whether it synergistically exerts anti-apoptotic effects by inhibiting the activation of JNK and p38 pathways.
OBJECTIVE: To explore the effects and mechanisms of croton cream on neuronal damage and apoptosis in the ischemic cortex of rats with cerebral ischemia-reperfusion injury.
METHODS: (1) Ninety Sprague-Dawley rats were randomly divided into sham operation group, model group, croton cream low-dose group, croton cream medium-dose group, croton cream high-dose group and nimodipine group, with 15 rats in each group. Except for the sham operation group, animal models of middle cerebral artery occlusion were prepared in rats by the thread method. Rats in the three croton cream groups were given 20, 40, and 60 mg/kg croton cream, respectively. Rats in the sham operation and model groups were given the same amount of normal saline, once a day, for 7 consecutive days. The optimal concentration of croton cream, namely the high dose of croton cream, was selected based on neurological deficit score, TTC staining, brain tissue water content, hematoxylin-eosin staining and Nissl staining. (2) Another 120 Sprague-Dawley rats were randomly divided into sham operation group, model group, croton cream group, JNK inhibitor group, croton cream+JNK inhibitor group, p38 MAPK inhibitor group, croton cream+p38 MAPK inhibitor group, and nimodipine group, with 15 rats in each group. Animal models of middle cerebral artery occlusion were prepared using the thread method in all the groups except in the sham operation group. Thirty minutes before modeling, 10 μL of SP600125 (JNK inhibitor) and 10 μL of SB203580 (p38 MAPK inhibitor) were injected into the lateral ventricle of the rats, respectively. Rats in croton cream groups were intragastrically given 60 mg/kg croton cream. Seven days later, the JNK/p38 MAPK signaling pathway, apoptosis-related proteins and cell apoptosis were detected by western blot, TUNEL staining and flow cytometry, respectively.
RESULTS AND CONCLUSION: (1) Compared with the sham operation group, neurological deficit score, cerebral water content, cerebral infarction volume and apoptosis rate were significantly increased in the model group (P < 0.05), where nerve cells showed scattered distribution. Compared with the model group, neurological deficit score, water content of brain tissue and cerebral infarction volume were significantly decreased in the croton cream medium-dose group, high-dose group and nimodipine group (P < 0.05), and the pathological morphology of nerve cells was significantly improved. (2) Compared with the JNK inhibitor group, p-JNK/JNK, p-p38/p38 and Bax expressions in rat brain tissue and the apoptotic rate were significantly decreased in the croton cream+inhibitor groups (P < 0.05), while the expression of and Bcl-2 was significantly increased (P < 0.05). To conclude, croton cream may inhibit the activation of JNK/p38 MAPK signaling pathway and reduce neuronal apoptosis to achieve neuroprotective effects in rats with cerebral ischemia-reperfusion injury.

Key words: croton cream, cerebral ischemia-reperfusion injury, neuronal apoptosis, signaling pathway, rat

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