天麻脑保护活性成分跨血脑屏障的转运方式研究  

Transport mode of brain-protective active components of Gastrodia elata across blood-brain barrier

作　　者：颜明丽 杨茜 宁珑 许春萍 孙云兰 陈明 何芳雁 YAN Mingli;YANG Qian;NING Long;XU Chunping;SUN Yunlan;CHEN Ming;HE Fangyan(College of Traditional Chinese Medicine,Yunnan University of Traditional Chinese Medicine,Yunnan Kunming 650500,China;Yunnan Key Laboratory of Dai&Yi Medicines,Yunnan University of Chinese Medicine,Yunnan Kunming 650500,China)

机构地区：[1]云南中医药大学中药学院,云南昆明650500 [2]云南中医药大学云南省傣医药与彝医药重点实验室,云南昆明650500

出　　处：《中国医院药学杂志》2024年第10期1132-1138,共7页Chinese Journal of Hospital Pharmacy

基　　金：国家自然科学基金项目(编号:82360784);云南省科技厅科技计划-中医联合专项-重点项目(编号:202101AZ070001-009);云南省科技厅科技计划-基础研究专项-面上项目(编号:202101AT070264);云南省傣医药与彝医药重点实验室开放课题(编号:202210SS2210)。

摘　　要：目的:基于药物转运体研究天麻脑保护活性成分对羟基苯甲醛(4-HBd)、对羟基苯甲醇(4-HBA)、3,4-二羟基苯甲醛(3,4-DD)跨血脑屏障(BBB)的转运方式。方法:建立符合要求的4-HBd、4-HBA及3,4-DD于HBSS缓冲液内的HPLC分析方法;用永生化人脑微血管内皮细胞(hCMEC/D3)及Transwell结构建立体外BBB转运模型,通过观察细胞形态、记录跨膜电阻值及荧光素钠转运实验加以评价,进行3个成分的跨膜转运研究;通过记录给予不同BBB关键外排转运体抑制剂(维拉帕米、ko143、MK571)后转运量的变化,对比各自外排率(ER)及渗透系数(Papp),明确4-HBd、4-HBA及3,4-DD跨BBB的转运方式及特征。结果:4-HBd、4-HBA、3,4-DD在2 h内的外排率分别在0.83~0.99、0.74~0.97、0.63~0.85之间,且不同时间点3个成分的吸收速率(Papp AP-BL)与外排速率(Papp BL-AP)相近,三者的Papp均大于1×10^(–6)cm·s^(–1),故判断4-HBd、4-HBA及3,4-DD跨BBB入脑方式为脂溶扩散;加入P-gp抑制剂维拉帕米后,4-HBd外排率降低(P<0.05),而4-HBA及3,4-DD外排率无明显变化,故推断4-HBd为外排转运体P-gp的底物。结论:4-HBd、4-HBA及3,4-DD跨BBB方式均为脂溶扩散,其中4-HBd为P-gp外排转运体的底物。OBJECTIVE To explore the trans-blood-brain barrier(BBB)transport mode of brain-protective active components of Gastrodia elata 4-hydroxybenzaldehyde(4-HBd),4-hydroxybenzyl alcohol(4-HBA),3,4-dihydroxybenzaldehyde(3,4-DD)based upon drug transporter.METHODS The compliant high performance liquid chromatography(HPLC)analytical method was established for 4-HBd,4-HBA and 3,4-DD in HBSS buffer.A BBB transport model in vitro was established by immortalized human brain microvascular endothelial cells(hCMEC/D3)and Transwell structure.The model was evaluated by observing cellular morphology,recording transmembrane resistance value and conducting fluorescein sodium transport assays.And then transmembrane transport of three components was examined.Through recording the changes in transport volume after dosing of different BBB key efflux transporter inhibitors(verapamil,ko143&MK571)and comparing their exhaust rate(ER)and apparent permeability coefficient(Papp),transport modes and characteristics of 4-HBd,4-HBA and 3,4-DD across BBB were clarified.RESULTS The exhaust rates of 4-HBd,4-HBA and 3,4-DD within 2h were 0.83–0.99,0.74–0.97 and 0.63–0.85 and absorption rate(Papp AP-BL)and efflux rate(Papp BL-AP)of three components were similar at different timepoints.Papp of three components were all>1×10^(–6) cm·s^(–1).Therefore the entry mode of 4-HBd,4-HBA and 3,4-DD across BBB was fat-soluble diffusion.After an addition of P-gp inhibitor verapamil,exhaust rate of 4-HBd declined(P<0.05)while those of 4-HBA and 3,4-DD showed no obvious change.Therefore 4-HBd was the substrate of efflux transporter P-gp.CONCLUSION 4-HBd,4-HBA and 3,4-DD exhibit fat-soluble diffusion across BBB.And 4-HBd is the substrate of P-gp efflux transporter.

关 键 词：对羟基苯甲醛 对羟基苯甲醇 3 4-二羟基苯甲醛 血脑屏障 外排转运体 双向转运 

分 类 号：R285[医药卫生—中药学]