目的 探讨川芎Chuanxiong Rhizoma调节血脑屏障通透性的作用特点，并从紧密连接蛋白、外排蛋白表达、埃兹蛋白/根蛋白/膜突蛋白（ezrin/radixin/mesin，ERM）磷酸化水平变化的角度阐述影响血脑屏障通透性的机制。方法 小鼠单次给予川芎提取物（5.8 g/kg）后iv不同相对分子质量（4×10³、4×10⁴、7×10⁴、1.5×10⁵）的异硫氰酸荧光素-葡聚糖（fluorescein isothiocyanate-dextran，FITC-dextran），测定给药后0、0.5、1、2、4、6、8 h脑组织中FITC-dextran的荧光强度，定量描述川芎对体内血脑屏障通透性的影响。建立bEnd.3细胞单培养血脑屏障模型，给予川芎单体成分洋川芎内酯A、洋川芎内酯H、阿魏酸、洋川芎内酯I、藁本内酯、欧当归内酯A、4-羟基-3-丁基苯酞，比较相同作用时间内血脑屏障模型中荧光素钠（fluorescein sodium，Na-F）的表观渗透系数（apparent permeability coefficient，P_app），确定各成分开放血脑屏障的效应。采用Western blotting检测川芎提取物给药前后小鼠脑组织及洋川芎内酯I给药bEnd.3细胞中闭锁小带蛋白1（zonula occluden-1，ZO-1）、闭锁蛋白（Occludin）、P-糖蛋白（P-glycoprotein，P-gp）、血管内皮细胞钙黏合素（VE-cadherin）、p-ERM/ERM的表达水平。结果 川芎提取物给药0.5 h后可显著增加小鼠脑组织中FITC-dextran（相对分子质量4×10³）的透过量（P＜0.001），作用强度随给药时间延长减弱，但对相对分子质量4×10⁴、7×10⁴、1.5×10⁵的FITC-dextran透过量无影响。在脑组织中，川芎提取物给药0.5 h时下调ZO-1、Occludin蛋白表达（P＜0.05），上调P-gp蛋白表达；给药2 h时上调ZO-1、Occludin蛋白表达（P＜0.05、0.01），下调P-gp蛋白表达（P＜0.01）；给药8 h时，ZO-1、Occludin、P-gp的蛋白表达水平与对照组无明显差异。不同浓度（1、10、100 μmol/L）的洋川芎内酯I、洋川芎内酯A、洋川芎内酯H、阿魏酸8 h给药时间内显著增加Na-F透过血脑屏障模型的P_app，其中洋川芎内酯I的促透效应最强。洋川芎内酯I（10 μmol/L）给药bEnd.3细胞0.5 h时可下调Occludin、ZO-1、p-ERM/ERM的蛋白表达（P＜0.01），给药1 h时上调Occludin、ZO-1、p-ERM/ERM的蛋白表达（P＜0.01），给药8 h时Occludin、ZO-1、p-ERM/ERM的蛋白表达水平与对照组无明显差异。结论 川芎在一定程度上可增加血脑屏障通透性，且该作用具有时效性，其机制与调节Occludin、ZO-1、P-gp的蛋白表达及ERM磷酸化水平有关。

Objective To explore the effect of Chuanxiong (Chuanxiong Rhizoma) on blood-brain barrier permeability, and explain the mechanism of its effect on blood-brain barrier permeability from the point of view of the changes of tight junction protein, efflux protein expression and ezrin/radixin/mesin (ERM) phosphorylation level. Methods After a single administration of Chuanxiong Rhizoma extract (5.8 g/kg), different molecular weights (4×10³, 4×10⁴, 7×10⁴, 1.5×10⁵) fluorescein isothiocyanate-dextran (FITC-dextran) were injected intravenously, the fluorescence intensity of FITC-dextran in brain tissue was measured at 0, 0.5, 1, 2, 4, 6, 8 h after administration, and the effect of Chuanxiong Rhizoma on blood-brain barrier permeability in vivo was quantitatively described. The blood-brain barrier model was established by single culture of bEnd.3 cells, monomeric components (senkyunolide A, senkyunolide H, ferulic acid, senkyunolide I, ligustilide, levistilide A, 4-hydroxy-3-butylphthalide) in Chuanxiong Rhizoma were given, the apparent permeability coefficient (P_app) of fluorescein sodium (Na-F) in a blood-brain barrier model for the same duration of action were compared, and the effect of each component on opening the blood-brain barrier was determined. Western blotting was used to detect the expressions of zonula occluden-1 (ZO-1), Occludin, P-glycoprotein (P-gp), VE-cadherin, p-ERM/ERM protein in brain tissue and bEnd.3 cells before and after administration of Chuanxiong Rhizoma and its components. Results After 0.5 h of administration of Chuanxiong Rhizoma extract, it significantly increased the transmittance of FITC-dextran (molecular weight 4×10³) in brain tissue of mice (P < 0.001), and the intensity of action was weaken with prolonged administration time, but had no effect on the transmittance of FITC-dextran (molecular weight of 4×10⁴, 7×10⁴, 1.5×10⁵). In brain tissue, after 0.5 h of administration of Chuanxiong Rhizoma extract, the expressions of ZO-1 and Occludin protein were downregulated (P < 0.05), and the expression of P-gp protein was upregulated; After 2 h of administration, ZO-1 and Occludin protein expressions were upregulated (P < 0.05, 0.01), and P-gp protein expression was downregulation (P < 0.01); At 8 h of administration, there was no significant difference in the protein expression levels of ZO-1, Occludin, and P-gp compared with control group. Within 8 h administration time of different concentrations (1, 10, 100 μmol/L) of senkyunolide I, senkyunolide A, senkyunolide H and ferulic acid, P_app of Na-F penetration through the blood-brain barrier model was significantly increased, with senkyunolide I had the strongest promoting effect. bEnd.3 cells were treated with senkyunolide I (10 μmol/L) for 0.5 h, the protein expressions of Occludin, ZO-1 and p-ERM/ERM were downregulated (P < 0.01), and the protein expressions of Occludin, ZO-1 and p-ERM/ERM were upregulated at 1 h of treatment (P < 0.01). At 8 h of treatment, there was no significant difference in the protein expression levels of Occludin, ZO-1 and p-ERM/ERM compared with control group. Conclusion Chuanxiong Rhizoma can increase the permeability of blood-brain barrier to a limited extent, and the effect is time-dependent, and its mechanism is related to the change of protein expression level of Occludin, ZO-1, P-gp and ERM phosphorylation level.

国家自然科学基金资助项目（81774155）；国家自然科学基金资助项目（82173957）