灵芝三萜对癫痫大鼠学习记忆损伤的效果

doi:10.3969/j.issn.1006-9771.2020.04.010

《中国康复理论与实践》 ›› 2020, Vol. 26 ›› Issue (4): 440-446.doi: 10.3969/j.issn.1006-9771.2020.04.010

灵芝三萜对癫痫大鼠学习记忆损伤的效果

农雪娟¹,金佳熹^2a,周冰玉^2a,张丽凤^2b,洪剑威^2a,赵爽^2b()

1.右江民族医学院附属医院检验科,广西百色市 533000
2.右江民族医学院,a临床医学院；b.基础医学院,广西百色市 533000

收稿日期:2019-10-08 修回日期:2019-12-09 出版日期:2020-04-25 发布日期:2020-04-27
通讯作者: 赵爽 E-mail:zhsh8000@163.com
作者简介:农雪娟(1986-),女,壮族,广西南宁市人,硕士研究生,主要研究方向：神经内分泌病理生理学。
基金资助:
1.国家自然科学基金项目(81760249);2.广西自然科学基金项目(2016GXNSFAA380278);2.广西自然科学基金项目(2014GXNSFBA118172);3.广西中医药管理局科研项目(GZZC2019143);4. 广西研究生教育创新计划项目(JGY2018124);5. 国家级大学生创新创业训练计划项目(201810599002);6. 广西自治区级大学生创新创业训练计划项目(201910599048)

Effects of Ganoderma Triterpenoids on Learning and Memory Impairment in Rats with Epilepsy

NONG Xue-juan¹,JIN Jia-xi^2a,ZHOU Bing-yu^2a,ZHANG Li-feng^2b,HONG Jian-wei^2a,ZHAO Shuang^2b()

1. Department of Laboratory Medicine, Affiliated Hospital of Youjiang Medical University for Nationalities, Baise, Guangxi 533000, China
2. a. Clinical Medicine College; b. Preclinical Medicine College, Youjiang Medical University for Nationalities, Baise, Guangxi 533000, China

Received:2019-10-08 Revised:2019-12-09 Published:2020-04-25 Online:2020-04-27
Contact: ZHAO Shuang E-mail:zhsh8000@163.com
Supported by:
National Natural Science Foundation of China(81760249);Guangxi Natural Science Foundation(2016GXNSFAA380278);Guangxi Natural Science Foundation(2014GXNSFBA118172);Scientific Research Project of the Guangxi Chinese Medicine Administration(GZZC2019143);Innovation Project of Guangxi Graduate Education(JGY2018124);National Innovation and Entrepreneurship Training Program for College Students(201810599002);Innovation and Entrepreneurship Training Program for College Students in Guangxi Autonomous Region(201910599048)

摘要/Abstract

摘要：

目的研究灵芝三萜对戊四氮致痫大鼠认知功能的影响及其作用机制。方法 75只Sprague-Dawley大鼠随机分为空白对照组、癫痫模型组、灵芝三萜组、外源性单唾液酸四己糖神经节苷脂(GM1)组和灵芝三萜联合GM1组,每组15只。除空白对照组外,其余各组用戊四氮35 mg/kg腹腔注射,每天1次,持续28 d。各用药组在每天腹腔注射戊四氮的同时进行相应给药。造模后,行Morris水迷宫测试;HE染色及透射电镜观察海马神经元;实时定量聚合酶链反应检测大鼠海马肌动蛋白结合蛋白(Cofilin)、突触素(SYN)、神经生长相关蛋白43 (GAP-43)的mRNA表达水平。结果与空白对照组比较,各时间点癫痫模型组逃避潜伏期延长(P < 0.05);与癫痫模型组比较,各用药组逃避潜伏期均缩短,部分时间点有显著性差异( P < 0.05)。与癫痫模型组比较,各用药组穿越平台次数明显增多( P < 0.01),在目标象限停留时间明显延长( P < 0.01)。各用药组海马神经元数增加,核溶解碎裂减少,核膜结构较清楚,突触小泡增多,突触数量增加,细胞器结构有所改善。与空白对照组相比,癫痫模型组Cofilin mRNA水平上调( P < 0.05),SYN mRNA和GAP-43 mRNA水平降低( P < 0.05);与癫痫模型组比较,各用药组Cofilin mRNA表达水平降低( P < 0.05),SYN mRNA表达水平升高( P < 0.05),仅灵芝三萜联合GM1组GAP-43 mRNA升高( P < 0.05)。结论灵芝三萜和GM1及两者联合用药均可改善癫痫大鼠的学习记忆能力及神经元形态结构,其机制可能与影响海马突触生长重塑相关基因表达有关,以达到保护大脑神经元的作用。

关键词: 癫痫, 突触重塑, 突触素, 肌动蛋白结合蛋白, 神经生长相关蛋白-43, 大鼠

Abstract:

Objective To study the effect of Ganoderma triterpenoids combined with exogenous monosialoteterahexosyl ganglioside (GM1) on cognitive dysfunction in rats with epilepsy.Methods A total of 75 Sprague-Dawley rats were divided randomly into blank control group, epileptic model group, Ganoderma triterpenoids group, GM1 group and GM1 combined with Ganoderma triterpenoids group (combination group), with 15 rats in each group. All the groups, except the blank control group, were intraperitoneally injected with pentylenetetrazol (PTZ) 35 mg/kg once a day for 28 days. Medication groups were given corresponding administration based on daily intraperitoneal injection of PTZ. They were tested with Morris Water Maze; and were observed with transmission electron microscopy and HE staining for hippocampal neurons. Real-time quantitative polymerase chain reaction was used to detect the expression of actin-binding protein (Cofilin), synaptophysin (SYN) and growth-associated protein 43 (GAP-43) mRNA in hippocampus of rats.Results Compared with the blank control group, the escape lantency prolonged in the epileptic model group in all the time points (P < 0.05). Compared with the epileptic model group, the escape lantency shortened in the treatment groups somewhen ( P < 0.05). Compared with the epileptic model group, the number of crossing the platform increased in the treatment groups ( P < 0.01), and the time of staying in the target quadrant prolonged ( P < 0.01); while the number of pyramidal cells increased, the nuclear lysis and fragmentation reduced, the structure of neurons and the number of synapses improved, as well as the organelle structure. Compared with the blank control group, the expression of Cofilin mRNA increased ( P < 0.05), and the expression of SYN mRNA and GAP-43 mRNA decreased ( P < 0.05) in the epileptic model group; compared with the epileptic model group, the expression of Cofilin mRNA decreased ( P < 0.05), and the expression of SYN mRNA increased ( P < 0.05) in all the treatment groups, while the expression of GAP-43 mRNA increased ( P < 0.05) only in the combination group. Conclusion Ganoderma triterpenoids, GM1 and their combination can improve the learning and memory abilities of epileptic rats, which may be associated with increasing the expression of SYN and GAP-43, decreasing the expression of Cofilin, to promote the synaptic remodeling of hippocampal tissue and protect brain neurons from PTZ-induced epilepsy.

Key words: epilepsy, synaptic remodeling, synaptophysin, actin-binding protein, neuronal growth-associated protein 43, rats

中图分类号:

R742.1

农雪娟,金佳熹,周冰玉,张丽凤,洪剑威,赵爽. 灵芝三萜对癫痫大鼠学习记忆损伤的效果[J]. 《中国康复理论与实践》, 2020, 26(4): 440-446.

NONG Xue-juan,JIN Jia-xi,ZHOU Bing-yu,ZHANG Li-feng,HONG Jian-wei,ZHAO Shuang. Effects of Ganoderma Triterpenoids on Learning and Memory Impairment in Rats with Epilepsy[J]. 《Chinese Journal of Rehabilitation Theory and Practice》, 2020, 26(4): 440-446.

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引物名称	引物序列
SYN	F: 5'-GAG TGT GCC AAC AAG ACG GAG AG-3'
SYN	R: 5'-ATT CAG CCG ACG AGG AGT AGT CC-3'
GAP-43	F: 5'-TAC CAC TGA TAA CTC GCC GTC CTC-3'
GAP-43	R: 5'-GTG ACA GCA GCA GGC ACA TCG-3'
Cofilin	F: 5'-AAG GAG AGC AAG AAG GAG GAC CTG-3'
Cofilin	R: 5'-GCA TCC TTG GAG CTG GCA TAG ATC-3'
GAPDH	F: 5'-GAC ATG CCG CCT GGA GAA AC-3'
GAPDH	R: 5'-AGC CCA GGA TGC CCT TTA GT-3'

组别	n	第1天	第2天	第3天	第4天	第5天
空白对照组	10	39.53±7.38	31.38±7.97	23.90±6.52	21.88±7.65	16.70±4.20
癫痫模型组	10	51.45±6.52^b	43.35±8.56^a	37.15±6.05^b	32.88±4.40^a	29.35±5.43^b
灵芝三萜组	10	50.83±5.01^b	37.53±15.65	30.15±10.79	25.85±12.48	21.88±10.68^c
GM1组	10	44.38±5.96^c	36.96±12.73	29.30±8.27^c	26.28±10.34	20.35±5.37^c
灵芝三萜联合GM1组	10	43.93±6.58^c,d	34.27±9.23	28.68±7.62^c	23.93±9.99^c	19.98±4.66^c
F值		6.337	1.574	3.511	1.946	5.216
P值		0.001	0.198	0.014	0.119	0.002

组别	n	穿越平台次数	目标象限停留时间(s)
空白对照组	10	6.4±0.84	40.75±2.55
癫痫模型组	10	3.1±1.37^a	26.78±3.32^a
灵芝三萜组	10	5.0±1.33^b	32.23±4.47^b
GM1组	10	4.9±0.99^b	33.42±5.37^b
灵芝三萜联合GM1组	10	5.7±0.95^b	35.25±4.82^b
F值		12.125	14.225
P值		< 0.001	< 0.001

组别	n	Cofilin	SYN	GAP-43
空白对照组	7	1.01±0.04	0.99±0.05	1.00±0.04
癫痫模型组	7	1.28±0.39^a	0.80±0.08^a	0.76±0.11^a
灵芝三萜组	7	1.02±0.10^b,c	0.95±0.21^b,c	0.77±0.11
GM1组	7	1.06±0.12^b,c	0.97±0.11^b,c	0.82±0.20
灵芝三萜联合GM1组	7	0.99±0.12^b,c	1.04±0.17^b,c	0.96±0.22^b,c
F值		3.780	7.223	3.058
P值		0.019	< 0.001	0.041

灵芝三萜对癫痫大鼠学习记忆损伤的效果

Effects of Ganoderma Triterpenoids on Learning and Memory Impairment in Rats with Epilepsy

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