督脉电针调控胱氨酸/谷氨酸反向转运体改善脑卒中后肢体痉挛的作用机制

doi:10.11958/20231330

天津医药 ›› 2024, Vol. 52 ›› Issue (5): 463-468.doi: 10.11958/20231330

督脉电针调控胱氨酸/谷氨酸反向转运体改善脑卒中后肢体痉挛的作用机制

张丽红¹(), 李瑞青²^,^△(), 王艺莹¹, 梅紧紧¹, 苏凯奇², 顾昌宇¹, 黄梦玲¹

1 河南中医药大学（邮编450046）
2 河南中医药大学第一附属医院康复中心

收稿日期:2023-09-18 修回日期:2023-11-24 出版日期:2024-05-15 发布日期:2024-05-09
通讯作者: ^△ E-mail：lrq0424@163.com
作者简介:张丽红（1998），女，硕士在读，主要从事中风等常见功能障碍的中医康复治疗及相关研究。E-mail：1176678542@qq.com
基金资助:
国家中医药传承创新专项(2022CCCX010);河南省科技研发计划联合基金项目(222301420066);河南省重点研发与推广专项（科技攻关）(232102310470);河南省中医学“双一流”创建科学研究专项课题(HSRP-DFCTCM-2023-8-34)

The mechanism of Du meridian electroacupuncture regulating cystine/glutamate reverse transporter to improve limb spasm after stroke

ZHANG Lihong¹(), LI Ruiqing²^,^△(), WANG Yiying¹, MEI Jinjin¹, SU Kaiqi², GU Changyu¹, HUANG Mengling¹

1 Henan University of Chinese Medicine, Zhengzhou 450046, China
2 Rehabilitation Center, the First Affiliated Hospital of Henan University of Chinese Medicine

Received:2023-09-18 Revised:2023-11-24 Published:2024-05-15 Online:2024-05-09
Contact: ^△ E-mail：lrq0424@163.com

张丽红, 李瑞青, 王艺莹, 梅紧紧, 苏凯奇, 顾昌宇, 黄梦玲. 督脉电针调控胱氨酸/谷氨酸反向转运体改善脑卒中后肢体痉挛的作用机制[J]. 天津医药, 2024, 52(5): 463-468.

ZHANG Lihong, LI Ruiqing, WANG Yiying, MEI Jinjin, SU Kaiqi, GU Changyu, HUANG Mengling. The mechanism of Du meridian electroacupuncture regulating cystine/glutamate reverse transporter to improve limb spasm after stroke[J]. Tianjin Medical Journal, 2024, 52(5): 463-468.

摘要/Abstract

摘要：

目的探讨督脉电针对大脑中动脉阻塞再灌注（MCAO/R）后肢体痉挛大鼠大脑皮质中胱氨酸/谷氨酸反向转运体[System Xc（-）]的影响，并探讨其可能的作用机制。方法选择SPF级雄性SD大鼠60只，随机分为空白组、假手术组及造模组。空白组不进行任何处理，假手术组仅分离血管，造模组采用大脑中动脉栓塞法制备MCAO/R大鼠模型，并于术后第3天进行Zea-Longa神经功能缺损评分、肌张力评定和电生理描记检测筛选肢体痉挛大鼠，造模组造模成功后随机分为模型组和督脉电针组。其中，督脉电针组于术后第3天开始治疗，每日1次，每次30 min，连续7 d。治疗结束后再次评定其神经功能及肌张力恢复情况；TTC染色测算脑梗死体积；干湿质量法检测各组大鼠脑含水量；比色法检测大脑皮质中谷氨酸（Glu）、半胱氨酸（Cys）和谷胱甘肽（GSH）浓度；蛋白免疫印迹（Western blot）法和实时荧光定量PCR（RT-qPCR）分别从蛋白和基因水平检测大脑皮质中血清溶质载体家族7成员11（SLC7A11）、溶质转运蛋白家族3成员2（SLC3A2）、γ-谷氨酰半胱氨酸合酶（γ-GCS）和谷胱甘肽合成酶（GSS）的表达。结果与模型组相比，治疗后督脉电针组大鼠神经功能缺损评分和改良Ashworth评分降低，肌电信号值增强，脑水肿程度减轻，脑梗死程度有所恢复，Cys和GSH含量显著增加，SLC7A11、SLC3A2、γ-GCS和GSS蛋白和mRNA的表达量显著提高，而Glu浓度显著降低（P<0.05）。结论督脉电针可以改善脑卒中后大鼠神经功能损伤和肢体痉挛程度，其机制可能与调控System Xc（-），改善Glu兴奋性毒性和氧化应激有关。

关键词: 卒中, 痉挛, 督脉电针, 胱氨酸/谷氨酸反向转运体

Abstract:

Objective To investigate the effect and mechanism of Du meridian electroacupuncture on cystine/glutamate reverse transporter [System Xc (-)] in cerebral cortex of middle cerebral artery occlusion/reperfusion (MCAO/R) model rats. Methods A total of 60 SPF male Sprague-Dawley (SD) rats were randomly divided into the blank group, the sham operation group and the model group. The blank group did not do any treatment, the sham operation group only separated the blood vessels and the model group was prepared by middle cerebral artery embolization for MCAO/R rat model. On the third day after operation, Zea-Longa neurological deficit score, muscle tension evaluation and electrophysiological tracing were used to screen rats with limb spasm. After successful modeling, rats was randomly divided into the model group and the Du meridian electroacupuncture group. The Du meridian electroacupuncture group began treatment on the third day after operation, once a day for 7 days. After the end of treatment, the neurological function and muscle tension recovery were evaluated again. TTC staining was used to measure the cerebral infarction volume. The brain water content of rats in each group was detected by wet and dry weight method. The concentrations of glutamic acid (Glu), cysteine (Cys) and glutathione (GSH) in cerebral cortex were detected by colorimetric assay kit. Western blot assay and quantitative real-time PCR (RT-qPCR) were used to detect the expression of SLC7A11, SLC3A2, γ-glutamylcysteine synthetase (γ-GCS) and glutathione synthetase (GSS) in cerebral cortex at the protein level and gene level, respectively. Results Compared with the model group, the neurological deficit score and modified Ashworth score were decreased in the Du meridian electroacupuncture group, the EMG signal value was enhanced, the brain water content was decreased, the degree of cerebral infarction was restored, the contents of Cys and GSH were significantly increased, the expression levels of SLC7A11, SLC3A2, γ-GCS and GSS protein and mRNA were significantly increased, and the concentration of Glu was significantly decreased (P<0.05). Conclusion Du meridian electroacupuncture can improve the degree of neurological impairment and limb spasm in rats after stroke. The mechanism may be related to the regulation of System Xc (-) and improving oxidative stress.

Key words: stroke, spasm, du meridian electroacupuncture, System Xc (-)

中图分类号:

R743.3

图/表 9

参考文献 24

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基因名称	引物序列（5′→3′）	产物大小/ bp
SLC7A11	上游：TCCCGGCGTATTACCTCTTC	122
SLC7A11	下游：TTCTTTAGAGTCTTCTGGTACAACT	122
SLC3A2	上游：CGAGAGGCATAGCTGGTCTG	74
SLC3A2	下游：GCCCAAAACTAGGCCCTTCA	74
γ-GCS	上游：TCAGTCCACAAACTGGCAGA	90
γ-GCS	下游：ACCTCCATTGGTCGGAACTC	90
GSS	上游：ATCTTGCAGGATGAGAAGCAGC	102
GSS	下游：GGAGCTGGGGTTCTTTGCG	102
GAPDH	上游：ACAGCAACAGGGTGGTGGAC	252
GAPDH	下游：TTTGAGGGTGCAGCGAACTT	252

基因名称	引物序列（5′→3′）	产物大小/ bp
SLC7A11	上游：TCCCGGCGTATTACCTCTTC	122
SLC7A11	下游：TTCTTTAGAGTCTTCTGGTACAACT	122
SLC3A2	上游：CGAGAGGCATAGCTGGTCTG	74
SLC3A2	下游：GCCCAAAACTAGGCCCTTCA	74
γ-GCS	上游：TCAGTCCACAAACTGGCAGA	90
γ-GCS	下游：ACCTCCATTGGTCGGAACTC	90
GSS	上游：ATCTTGCAGGATGAGAAGCAGC	102
GSS	下游：GGAGCTGGGGTTCTTTGCG	102
GAPDH	上游：ACAGCAACAGGGTGGTGGAC	252
GAPDH	下游：TTTGAGGGTGCAGCGAACTT	252

组别	干预前	干预后	t
空白组	0±0	0±0	-
假手术组	0±0	0±0	-
模型组	2.08±0.67^ab	1.92±0.67^ab	0.611
督脉电针组	2.00±0.60^ab	0.50±0.67^c	5.745^＊
F	82.300^＊	43.700^＊

组别	干预前	干预后	t
空白组	0±0	0±0	-
假手术组	0±0	0±0	-
模型组	2.08±0.67^ab	1.92±0.67^ab	0.611
督脉电针组	2.00±0.60^ab	0.50±0.67^c	5.745^＊
F	82.300^＊	43.700^＊

组别	干预前	干预后	t
空白组	0±0	0±0	-
假手术组	0±0	0±0	-
模型组	2.42±0.67^ab	2.17±0.72^ab	0.573
督脉电针组	2.25±0.75^ab	0.42±0.67^c	6.303^＊
F	86.000^＊	53.200^＊

督脉电针调控胱氨酸/谷氨酸反向转运体改善脑卒中后肢体痉挛的作用机制

The mechanism of Du meridian electroacupuncture regulating cystine/glutamate reverse transporter to improve limb spasm after stroke

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参考文献 24

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组别	干预前	干预后	t
空白组	2.41±0.63	2.35±0.43	0.277
假手术组	2.37±0.43	2.34±0.53	0.158
模型组	0.24±0.07^ab	0.25±0.06^ab	0.275
督脉电针组	0.20±0.07^ab	1.25±0.30^c	11.760^＊
F	129.000^＊	87.300^＊

组别	Glu/（μmol/L）	Cys/（μmol/g）	GSH/（mg/L）
空白组	172.50±2.17	7.82±0.60	10.36±0.46
假手术组	179.50±1.75	7.68±0.23	9.90±1.24
模型组	241.70±9.64^ab	2.96±0.23^ab	2.58±0.94^ab
督脉电针组	201.20±2.28^c	5.98±0.39^c	7.66±1.69^c
F	109.900^＊	98.890^＊	27.700^＊

组别	SLC7A11	SLC3A2	γ-GCS	GSS
空白组	0.98±0.03	0.62±0.02	0.93±0.05	1.06±0.04
假手术组	0.98±0.01	0.61±0.01	0.93±0.05	1.05±0.07
模型组	0.44±0.14^ab	0.33±0.07^ab	0.44±0.07^ab	0.61±0.08^ab
督脉电针组	0.74±0.07^c	0.52±0.05^c	0.76±0.04^c	0.94±0.03^c
F	32.400^＊	30.800^＊	60.500^＊	36.200^＊

组别	SLC7A11	SLC3A2	γ-GCS	GSS
空白组	1.00±0.04	1.00±0.05	1.00±0.07	1.00±0.03
假手术组	0.92±0.10	0.93±0.03	0.95±0.02	0.92±0.14
模型组	0.50±0.03^ab	0.58±0.04^ab	0.56±0.12^ab	0.50±0.16^ab
督脉电针组	0.81±0.02^c	0.83±0.02^c	0.82±0.02^c	0.85±0.08^c
F	42.010^＊	71.720^＊	24.890^＊	11.060^＊

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