Chinese Journal of Contemporary Neurology and Neurosurgery ›› 2019, Vol. 19 ›› Issue (8): 552-558. doi: 10.3969/j.issn.1672-6731.2019.08.004

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Protective effect of melatonin on blood-brain barrier dysfunction in rats with chronic cerebral hypoperfusion

WANG Shao-peng, WANG Ru, LI Dan-dan, ZHAO Hong-yi, CHI Li-yi, HUANG Yong-hua   

  1. Department of Neurology, the Seventh Medical Center of PLA General Hospital, Beijing 100700, China Department of Neurology, the 986 Hospital of the Air Force PLA, Xi'an 710054, Shaanxi, China
  • Online:2019-08-25 Published:2019-08-27
  • Contact: HUANG Yong-hua (Email: huangyh@163.com)
  • Supported by:

    This study was supported by Wu Jieping Medical Foundation(No.320.6750.18456).

褪黑素对慢性脑低灌注大鼠血-脑屏障的保护作用

王少朋,王茹,李丹丹,赵弘轶,迟丽屹,黄勇华   

  1. 100700 北京,解放军总医院第七医学中心神经内科[王少朋,王茹,李丹丹(现在山东省泰安市市立医院神经内科,邮政编码:271000),赵弘轶,黄勇华];710054 西安,解放军空军第九八六医院神经内科(迟丽屹)
  • 通讯作者: 黄勇华,Email:huangyh@163.com
  • 基金资助:

    吴阶平医学基金会基金资助项目(项目编号:320.6750.18456)

Abstract:

Objective To investigate the mechanism of the effect of melatonin (MT) on permeability of blood-brainbarrier (BBB) in chronic cerebral hypoperfusion (CCH) rats. Methods Therat model of CCH was established by bilateral common carotid artery occlusion (BCCAO). A total of 72 male Sprague-Dawley rats were randomly divided into 4 groups:sham group (N=18), BCCAO group (N=18), the melatonin [5 mg/(kg·d)] treatment model group (MT1 group, N=18), and the melatonin [10 mg/(kg·d)] treatment model group (MT2 group, N=18). Evans blue staining and fluorescein-labeled glucoside (FITC-Dextran) staining marked by fluorescein isothiocyanate (FITC) were used to assess the permeability o fBBB in rats. Expression of matrix metalloproteinase-2 (MMP-2) and MMP-9 mRNA levels in basal ganglia of the brain were measured by quantitative real-time polymerase chain reaction (qRT-PCR). Expression of Occludin and Claudin-5 protein in rat basal ganglia were measured by Western blotting method. Results Compared with the sham group, the fluorescence density in basal ganglia was increased (P = 0.000), expression of MMP-2 and MMP-9 were increased (all P=0.000),and expression of Occludin and Claudin-5 were decreased (all P =0.000) of BCCAO group. Compared with the BCCAO group, the fluorescence density in basal ganglia was decreased (P=0.021), the expression of MMP-2 was decreased (P=0.000) and the expression of Claudin-5 was increased (P=0.000) in MT1 group, and the differences with sham group were statistically significant (P=0.000, 0.006, 0.000). Compared with the BCCAO group, in MT2 group,the fluorescence degree in basal ganglia was further decreased (P=0.000), the expression of MMP-2 and MMP-9 were decreased (all P=0.000),and were similar to the sham group (all P>0.05), while expression of Occludin and Claudin-5 were increased (all P=0.000) but lower than those of sham group (P=0.003, 0.000). Compared with the two melatonin treatment groups, the efficacy of the 10mg/(kg·d) group (MT2 group) was better thanthat of the 5mg/(kg·d ) group (MT1 group, all P<0.05). Conclusions Melatonin may protect the integrity of BBB by inhibiting the degradation of Occludin and Claudin-5 protein through inhibiting the expression of matrix metalloproteinases.

Key words: Cerebral small vessel diseases, Melatonin, Blood-brain barrier, Disease models, animal

摘要:

目的 探讨褪黑素对慢性脑低灌注模型大鼠血-脑屏障通透性的影响及其作用机制。方法 健康雄性Sprague-Dawley(SD)大鼠通过改良双侧颈总动脉结扎法制备慢性脑低灌注模型,随机分为假手术组、模型组、褪黑素5mg/(kg·d)组和10mg/(kg·d)组,以伊文思蓝和异硫氰酸荧光素标记的右旋糖酐双重染色检测血-脑屏障通透性,聚合酶链反应测定基质金属蛋白酶-2和9(MMP-2和MMP-9)、 Westernblotting法测定Occludin蛋白和Claudin-5蛋白表达变化。结果 与假手术组相比,模型组大鼠基底节荧光强度增强(P=0.000),MMP-2(P=0.000)和MMP-9(P=0.000)水平升高,Occludin蛋白(P= 0.000)和Claudin-5蛋白(P=0.000)水平降低;经褪黑素5mg/(kg·d)治疗后,大鼠基底节荧光强度减弱 (P=0.021)、MMP-2水平降低(P=0.000)、Claudin-5蛋白水平升高(P=0.000),与假手术组差异仍有统计学意义(P=0.000,0.006,0.000);而褪黑素10mg/(kg·d)治疗后,大鼠基底节荧光强度进一步减弱(P= 0.000)、MMP-2(P=0.000)和MMP-9(P=0.000)水平持续降低且接近假手术组水平(均 P>0.05),而 Occludin蛋白(P=0.000)和Claudin-5蛋白(P=0.000)水平升高但仍低于假手术组(P=0.003,0.000);褪黑素两剂量组比较,10mg/(kg·d)组疗效优于5mg/(kg·d)组(均P<0.05)。结论 褪黑素通过抑制基质金属蛋白酶的表达而使Occludin和Claudin-5蛋白降解减少,进而保护血-脑屏障的完整性。

关键词: 大脑小血管疾病, 褪黑激素, 血脑屏障, 疾病模型, 动物