丁苯酞对于糖氧剥夺条件下人脑微血管内皮细胞能量代谢的影响及保护作用

doi:10.12092/j.issn.1009-2501.2018.12.002

中国临床药理学与治疗学 ›› 2018, Vol. 23 ›› Issue (12): 1329-1334.doi: 10.12092/j.issn.1009-2501.2018.12.002

丁苯酞对于糖氧剥夺条件下人脑微血管内皮细胞能量代谢的影响及保护作用

韩晨阳，张晓玲，官俏兵，王琰萍

嘉兴市第二医院，嘉兴 314001，浙江

收稿日期:2018-06-13 修回日期:2018-08-08 出版日期:2018-12-26 发布日期:2018-12-27
通讯作者: 王琰萍，女，博士，主任医师，研究方向：神经病学。 Tel：0573-82056408 E-mail:ypwang73@126.com
作者简介:韩晨阳，男，硕士，药师，研究方向：神经药理学。 Tel：0573-82056408 E-mail:691513770@qq.com
基金资助:
浙江省卫生厅面上项目（2018KY804）

Effects of NBP on energy metabolism of human brain microvascular endothelial cells under the condition of glucose deprivation

HAN Chenyang, ZHANG Xiaoling, GUAN Qiaobin, WANG Yanping

Second Hospital of JiaXing, Jiaxing 314001, Zhejiang, China

Received:2018-06-13 Revised:2018-08-08 Online:2018-12-26 Published:2018-12-27

摘要/Abstract

摘要：

目的: 研究糖氧剥夺（OGD）条件下丁苯酞（NBP）对于人脑微血管内皮细胞（HBMEC）能量代谢的影响及保护作用。方法: 采用糖氧剥夺（OGD）方法处理HBMEC细胞，将细胞分为对照组、模型组、低剂量组、中剂量组和高剂量组。其中对照组为正常培养的HBMEC细胞、模型组为OGD处理的细胞、低剂量组为OGD+5 μmol/L的NBP、中剂量组为OGD+10 μmol/L的NBP、高剂量组为OGD+20 μmol/L的NBP。采用CCK-8试剂盒检测细胞活力；试剂盒检测乳酸脱氢酶（LDH）漏出率、钠-钾-三磷酸腺苷酶（Na ⁺ -K⁺ -ATPase）和钙-三磷酸腺苷酶（Ca ²⁺ -ATPase）活性、细胞内钙离子的浓度；流式细胞术检测细胞凋亡率；Hoechst33342法进行活细胞染色；高效液相色谱法检测细胞中磷酸腺苷（AMP）、二磷酸腺苷（ADP）、三磷酸腺苷（ATP）的含量，能荷EC值。结果: 模型组中细胞活力、Na⁺ -K ⁺ -ATPase、Ca ²⁺ -ATPase的活性降低，而细胞内钙离子浓度、细胞凋亡率、乳酸脱氢酶（LDH）漏出率明显提高，能量代谢中模型组ATP、ADP的含量降低，AMP含量增高，EC值明显降低。NBP干预后可以提高细胞活力，提高Na+-K+-ATPase、Ca2+-ATPase酶活性，降低细胞中钙离子浓度和细胞凋亡率，且还可以提高细胞中ATP、ADP的含量，降低AMP的含量，提高EC值，具有剂量依赖性。结论: NBP可以保护HBMEC在OGD条件下的损伤，其作用机制和改善细胞能量代谢、抑制细胞内超钙有关。

关键词: 丁苯酞, 糖氧剥夺, 人脑微血管内皮细胞, 能量代谢

Abstract:

AIM: To study the effect and protective effect of butylphthalide (NBP) on the energy metabolism of human cerebral microvascular endothelial cells (HBMEC) under the condition of glucose deprivation (OGD). METHODS: HBMEC cells were treated by OGD. The cells were divided into control group, model group, low dose group, middle dose group and high dose group. The control group was normal cultured, the model group were OGD treated, the low dose group received OGD+5 mol/L NBP, the middle dose group received OGD+10 mol/L NBP, and the high dose group received OGD+20 mol/L NBP. The CCK-8 kit was used to detect the cell viability.The leaking rate of lactate dehydrogenase (LDH), sodium potassium ATPase (Na+-K+-ATPase), calcium triphosphatase (Ca2+-ATPase) activity and intracellular calcium concentration were detected acording to the description of kit,respectively. The flow cytometry was used to detect the apoptosis rate and Hoechst33342 staining. The high performance liquid chromatography was used to detect the contents of ATP, ADP and AMP in the cells, and the EC value was measured.RESULTS:The cell activity, Na+-K+-ATPase and Ca2+-ATPase in the model group was decreased, but the intracellular calcium concentration, the rate of apoptosis and the leakage rate of LDH were significantly increased. After NBP intervention, the activity of cell, the activity of Na+-K+-ATPase and Ca2+-ATPase, the concentration of calcium ion in the cell and the rate of cell apoptosis were reduced. The content of ATP, ADP and EC value in the cells was increased while the content of AMP was decreased in a dose-dependent manner. CONCLUSION: NBP can protect HBMEC injury under OGD, and its mechanism is related to improving energy metabolism and inhibiting intracellular calcium.

Key words: butylphthalide, glucose and oxygen deprivation, human brain microvascular endothelial cells, energy metabolism

中图分类号:

R965.2

韩晨阳，张晓玲，官俏兵，王琰萍. 丁苯酞对于糖氧剥夺条件下人脑微血管内皮细胞能量代谢的影响及保护作用[J]. 中国临床药理学与治疗学, 2018, 23(12): 1329-1334.

HAN Chenyang, ZHANG Xiaoling, GUAN Qiaobin, WANG Yanping. Effects of NBP on energy metabolism of human brain microvascular endothelial cells under the condition of glucose deprivation[J]. Chinese Journal of Clinical Pharmacology and Therapeutics, 2018, 23(12): 1329-1334.

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丁苯酞对于糖氧剥夺条件下人脑微血管内皮细胞能量代谢的影响及保护作用

Effects of NBP on energy metabolism of human brain microvascular endothelial cells under the condition of glucose deprivation

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