镇心省睡益智方抗A<italic style="font-style: italic">β</italic><sub>25-35</sub>诱导人脑微血管内皮细胞损伤的作用

吴玲; 郑琴; 郭园园; 张科楠; 罗俊; 肖帅; 李文静; 杨明

doi:10.13422/j.cnki.syfjx.20200204

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镇心省睡益智方抗Aβ_25-35诱导人脑微血管内皮细胞损伤的作用

经典名方 | 更新时间：2021-02-09

- 镇心省睡益智方抗Aβ_25-35诱导人脑微血管内皮细胞损伤的作用
- Effect of Zhenxin Shengshui Yizhi Fang on Damage of Human Brain Microvascular Endothelial Cells Induced by Aβ_25-35
- 中国实验方剂学杂志 2020年26卷第5期页码：26-33
- 作者机构：
  
  江西中医药大学　现代中药制剂教育部重点实验室，南昌　 330004
- 作者简介：
  
  [第一作者]　吴玲，硕士，从事中药新制剂与新技术研究，E-mail: 1061743400@qq.com
  *郑琴，博士生导师，从事中药新制剂与新技术研究，E-mail: zhengqin912006@163.com
- 基金信息：
  
  江西省科技创新人才重点项目（“5511”工程专项）(20171BCB18001);中药学一流学科项目(JXSYLXK-ZHYA0092)
- DOI：10.13422/j.cnki.syfjx.20200204
  中图分类号： R2-0; R22; R285.5; R289
- 收稿日期：2019-07-01，
  
  网络出版日期：2019-10-11，
  
  纸质出版日期：2020-03-05
- 稿件说明：
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吴玲, 郑琴, 郭园园, 等. 镇心省睡益智方抗Aβ_25-35诱导人脑微血管内皮细胞损伤的作用[J]. 中国实验方剂学杂志, 2020,26(5):26-33.

Ling WU, Qin ZHENG, Yuan-yuan GUO, et al. Effect of Zhenxin Shengshui Yizhi Fang on Damage of Human Brain Microvascular Endothelial Cells Induced by Aβ_25-35[J]. Chinese journal of experimental traditional medical formulae, 2020, 26(5): 26-33.
吴玲, 郑琴, 郭园园, 等. 镇心省睡益智方抗Aβ_25-35诱导人脑微血管内皮细胞损伤的作用[J]. 中国实验方剂学杂志, 2020,26(5):26-33. DOI： 10.13422/j.cnki.syfjx.20200204.

Ling WU, Qin ZHENG, Yuan-yuan GUO, et al. Effect of Zhenxin Shengshui Yizhi Fang on Damage of Human Brain Microvascular Endothelial Cells Induced by Aβ_25-35[J]. Chinese journal of experimental traditional medical formulae, 2020, 26(5): 26-33. DOI： 10.13422/j.cnki.syfjx.20200204.

摘要

目的：

观察镇心省睡益智方(省睡方)水提液对抗

淀粉样蛋白25-35(A

25-35

)诱导的人脑微血管内皮细胞(HBMEC)损伤的神经保护作用和潜在机制。

方法：

采用A

25-35

诱导的HBMEC细胞损伤作为阿尔茨海默病(AD)细胞模型。本研究分为空白组，A

25-35

组，省睡方水提液低、中、高剂量组(125，250，500 mg·L

-1

)。对其进行相关治疗后，采用噻唑蓝(MTT)比色法确定不同浓度药物及A

25-35

的细胞毒性，采用Hoechst-33258染色观察细胞凋亡情况，采用比色法检测半胱氨酸天冬氨酸蛋白酶-3(Caspase-3)活性，采用蛋白免疫印迹法(Western blot)检测晚期糖基化终产物受体(RAGE)，低密度脂蛋白受体相关蛋白(LRP1)，葡萄糖转运蛋白1(GLUT1)及葡萄糖转运蛋白3(GLUT3)的表达。

结果：

与空白组比较，A

25-35

组细胞活力显著下降(

0.01)，Hoechst-33258染色观察到明亮的蓝色荧光、染色质固缩、呈致密浓染或碎块状致密浓染，颜色有些发白，凋亡细胞百分比显著增加(

0.01)，Caspase-3活性显著增加(

0.01)，RAGE蛋白表达显著增加(

0.01)，LRP1，GLUT1和GLUT3蛋白表达显著下降(

0.01)；与A

25-35

组比较，省睡方水提液组细胞存活率以剂量依赖性方式显著增加，500 mg·L

-1

省睡方水提液组保护作用比其他组更明显(

0.05)，500 mg·L

-1

省睡方水提液组及显著抑制凋亡细胞的数量(

0.01)，且显著降低Caspase-3活性(

0.01)，125 mg·L

-1

省睡方水提液组RAGE蛋白表达未显著降低，250，500 mg·L

-1

省睡方水提液组RAGE蛋白表达显著下降(

0.01)，而各剂量省睡方水提液组LRP1，GLUT1和GLUT3蛋白表达明显增加(

0.05

0.01)。

结论：

省睡方水提液能够减弱A

25-35

寡聚体诱导的HBMEC细胞毒性，抑制细胞凋亡，降低Caspase-3活性，降低RAGE蛋白的表达，升高LRP1，GLUT1和GLUT3蛋白的表达，从而降低A

在脑内异常聚集和沉积可能是其防治AD的机制。

Abstract

Objective:

To observe the neuroprotective effect and potential mechanism of Zhenxin Shengshui Yizhi Fang(XSF) aqueous extract on human brain microvascular endothelial cells (HBMEC) injury induced by amyloid-

protein(A

)

25-35

Method:

HBMEC cells damage induced by A

25-35

was used as Alzheimer' s disease(AD) cell model. The study included control group

25-35

group

and low

medium and high-dose XSF aqueous extract groups (125

250

500 mg·L

-1

). After treatment

the cytotoxicity of different concentrations of drugs and A

25-35

was determined by methyl thiazolyl tetrazolium(MTT) colorimetry. Apoptosis was observed by Hoechst-33258 staining. The activity of Caspase-3 was detected by colorimetry. Western blot was used to detect the expression levels of the receptor of advanced glycation end products (RAGE) and low-density lipoprotein receptor-related protein (LRP1).

Result:

Compared with the control group

the cell viability of A

25-35

group was significantly decreased (

0.01). Hoechst-33258 staining showed bright blue fluorescence

chromatin condensation

dense staining or fragmentation dense staining

whitening in color

and significant increase of the percentage of apoptotic cells (

0.01). Caspase-3 activity increased significantly (

0.01). Western blot showed that RAGE protein expression increased significantly (

0.01)

while low-density lipoprotein receptor-related protein(LRP1)

glucose transporter 1(GLUT1) and GLUT3 protein expressions decreased significantly (

0.01). Compared with the A

25-35

group

the cell viability of XSF aqueous extract groups was significantly increased in a dose-dependent manner. The XSF aqueous extract had a more significant protective effect of than the other groups (

0.05). The XSF aqueous extract group (500 mg·L

-1

) significantly inhibited the number of apoptotic cells (

0.01)

but significantly reduced the Caspase-3 activity (

0.01). RAGE protein expression was not significantly decreased in XSF aqueous extract group (125 mg·L

-1

)

but significantly decreased in XSF aqueous extract group (250

500 mg·L

-1

0.01)

while LRP1

GLUT1 and GLUT3 protein expression significantly increased (

0.05

0.01) in a dose-dependent manner.

Conclusion:

XSF aqueous extract can attenuate the cytotoxicity of HBMEC induced by A

25-35

oligomer

inhibit apoptosis

decrease caspase-3 activity and RAGE protein expression

increase LRP1

GLUT1 and GLUT3 protein expressions

and reduce the abnormal accumulation and deposition of A

in the brain

which may be its mechanisms for prevention and treatment of AD.

关键词

Keywords

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镇心省睡益智方抗Aβ25-35诱导人脑微血管内皮细胞损伤的作用

Effect of Zhenxin Shengshui Yizhi Fang on Damage of Human Brain Microvascular Endothelial Cells Induced by Aβ25-35

DOI：10.13422/j.cnki.syfjx.20200204

摘要

Abstract

关键词

Keywords

references

镇心省睡益智方抗Aβ_25-35诱导人脑微血管内皮细胞损伤的作用

Effect of Zhenxin Shengshui Yizhi Fang on Damage of Human Brain Microvascular Endothelial Cells Induced by Aβ_25-35