小檗碱对氧糖剥夺/复氧诱导的大鼠体外血脑屏障损伤的保护作用

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摘要目的探究小檗碱对氧糖剥夺/复氧（OGD/R）诱导大鼠体外血脑屏障损伤的保护作用及其机制。方法将原代培养的大鼠脑微血管内皮细胞（rBMECs）分为对照组、模型组、小檗碱给药组。对照组加入新鲜培养基；模型组和小檗碱给药组给予不同浓度（0、1、3、10、30、100 μmol/L）的小檗碱预孵24 h后更换低糖培养基并将其放在不含氧气的三气培养箱中（95%N2/5%CO2）培养2 h（氧糖剥夺，OGD），之后换成新鲜培养基，置于37℃、5%CO2培养箱内继续培养2 h（复氧，R）。利用MTT法检测细胞存活率，跨膜电阻法测定细胞紧密连接程度，Western blot法检测紧密连接蛋白ZO-1、occludin以及核因子-κB p65（NF-κB p65）的蛋白表达量，ELISA法检测炎性因子肿瘤坏死因子-α（TNF-α）、白细胞介素-1β（IL-1β）含量。结果与对照组比较，模型组rBMECs细胞存活率、ZO-1表达量、occludin表达量显著降低，跨膜电阻值呈下降趋势，NF-κB p65表达量及IL-1β、TNF-α含量显著升高，差异均有高度统计学意义（均P ＜ 0.01）；与模型组比较，小檗碱给药组rBMECs细胞存活率不同程度提高，小檗碱10、30 μmol/L浓度组ZO-1表达量、occludin表达量显著升高，跨膜电阻值显著提高，差异有统计学意义（P ＜ 0.05或P ＜ 0.01），同时NF-κB p65表达量及IL-1β、TNF-α含量显著降低，差异均有高度统计学意义（均P ＜ 0.01）。结论小檗碱通过抑制NF-κB入核、降低炎性因子释放量、改善OGD/R诱导的rBMECs的损伤，发挥对OGD/R损伤的血脑屏障保护作用。

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	王美华王甲韩丹王羽

关键词 ：小檗碱, 血脑屏障, 核因子-&kappa, B p65, 炎症

Abstract：Objective To investigate the protective effect of berberine on oxygen glucose deprivation/reoxygenation (OGD/R) induced blood-brain barrier injury in vitro in rats and its mechanism. Methods Primary cultured rat brain microvascular endothelial cells (rBMECs) were divided into the control group, the model group and the berberine administration group. Fresh medium was added to the control group. The model group and the berberine administration group were preincubated with berberine at different concentrations (0, 1, 3, 10, 30, 100 μmol/L) for 24 h, then the low glucose medium was replaced and the culture medium was placed in a three-gas incubator without oxygen (95%N2/5%CO2) for 2 h (oxygen glucose deprivation, OGD). then replaced with fresh medium, at 37℃ and 5%CO2 incubator continue to develop 2 h (Reoxygenation, R). Cell survival was determined by MTT assay, cell tightness was determined by transendothelial electrical resistance, protein expression levels of tight junction protein ZO-1, occludin and nuclear factor-κB p65 (NF-κB p65) were detected by Western blot assay, and levels of inflammatory factors tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β) were detected by ELISA assay. Results Compared with the the control group, rBMECs cell survival rate, ZO-1 expression level and occludin expression level in the model group were significantly reduced, transendothelial electrical resistance showed a decreasing trend, and NF-κB p65 expression level, IL-1β, and TNF-α levels were significantly increased in the model group, the differences were highly statistically significant (all P ＜ 0.01). Compared with the model group, the survival rate of rBMECs cells in the berberine administration group increased to different degrees, the ZO-1 expression, occludin expression, the transmembrane electrical resistance increased significantly in the berberine 10 and 30 μmol/L concentration groups, with statistically significant differences (P ＜ 0.05 or P ＜ 0.01). Meanwhile, the NF-κB p65 expression and the levels of IL-1β and TNF-α were significantly decreased in the berberine 10 and 30 μmol/L concentration groups, the differences were highly statistically significant (all P ＜ 0.01). Conclusion Berberine plays a protective role of blood brain barrier on OGD/R injury by inhibiting NF-κB entry into the nucleus, reducing the release of inflammatory factors, and improving the damage of OGD/R-induced rBMECs.

Key words： Berberine Blood-brain barrier Nuclear factor-κB p65 Inflammation

基金资助:中央高校基本科研业务费专项基金项目（02141 4380405）。

通讯作者: 王羽（1976.4-），女，硕士，副主任药师；研究方向：药物代谢动力学。

引用本文:

王美华王甲韩丹王羽. 小檗碱对氧糖剥夺/复氧诱导的大鼠体外血脑屏障损伤的保护作用[J]. 中国医药导报, 2020, 17(9): 16-20.
WANG Meihua WANG Jia HAN Dan WANG Yu. Protective effect of berberine on in vitro blood-brain barrier injury induced by oxygen glucose deprivation/reoxygenation in rats. 中国医药导报, 2020, 17(9): 16-20.

链接本文:

http://www.yiyaodaobao.com.cn/CN/ 或 http://www.yiyaodaobao.com.cn/CN/Y2020/V17/I9/16

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