洋葱中黄酮类提取物对血脑屏障透过及神经保护作用研究

作者	单位
杨鸣鸣	四川大学华西基础医学与法医学院生物化学与分子生物教研室
所起凤	四川大学华西基础医学与法医学院生物化学与分子生物教研室
刘戟	四川大学华西基础医学与法医学院生物化学与分子生物教研室
何丹	四川中医药高等专科学校药学与检验系医学检验教研室
范雪娇	四川大学华西基础医学与法医学院生物化学与分子生物教研室
杜文婷	四川大学华西基础医学与法医学院生物化学与分子生物教研室

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中文摘要:

目的研究采用乙醇回流法从洋葱中提取的黄酮类物质对血脑屏障(blood-brain barrier,BBB)透过作用以及对原代培养的鼠神经元细胞增殖和凋亡的影响。方法首先原代培养成功SD鼠脑微血管内皮细胞(brain microvascular endothelial cells,BMVECs);然后原代共培养鼠BMVECs和星形胶质细胞(astro-cytes,ACs)以建立BBB体外模型。采用乙醇回流法从洋葱中提取黄酮类物质,通过透射电子显微镜观察和跨膜电阻值测量验证BBB模型;采用高效液相色谱(HPLC)检测黄酮类提取物对BBB的透过率。采用过氧化氢(H2O2)和黄酮提取物处理神经元细胞,MTT法观察其对细胞增殖和凋亡的影响;彗星电泳和免疫荧光法分析神经元细胞DNA损伤情况。结果透射电镜及跨膜电阻仪鉴定BBB体外模型构建成功;HPLC检测洋葱中黄酮类提取物BBB透过率为60.58%;该提取物在10～20μg/mL浓度对H2O2诱导的鼠神经元细胞凋亡有明显的抑制作用,并可减少神经元DNA损伤。结论采用乙醇回流法从洋葱中提取的黄酮类物质可有效透过BBB,并对H2O2诱导的鼠神经元细胞的凋亡和DNA损伤有明显抑制作用。

英文摘要:

Objective To study the flavanoids extracted from onion on the blood-brain barrier(BBB) permeation,and their effects on primary cultured neuron cell proliferation and apoptosis of SD rats using ethanol reflux method.Methods The brain microvascular endothelial cells(BMVECs) were first successfully primary cultured.Then rats BMVECs and astrocytes(ACs) were co-cultured to establish the in vitro BBB model.The flavanoids were extracted from onion using ethanol reflux method.The model was verified by transmission electron microscopy(TEM) and trans-epithelial electric resistance(TEER).The flavanoids permeability was tested using high performance liquid chromatography(HPLC).Meanwhile,rat neuron cells were cultured and exposed to H2O2 and flavanoids.Their effects on the cell proliferation and apoptosis were observed using MTT assay.The injury of neuron DNA was analyzed using single-cell gel electrophoresis(SCGE) and immunofluorescent assay.Results The in vitro BBB model was successfully established by TEM and TEER.Results of HPLC proved flavanoids extracts could effectively permeate the BBB with the permeability of 60.58%.The extractive at 10-20 μg/mL showed obvious inhibition on the apoptosis of neuron cells induced by H2O2,and attenuated the injury of neuron DNA.Conclusions The flavanoids extracted from onion ethanol reflux method could effectively penetrate the BBB.They also showed obvious inhibition on the H2O2 induced neuron cell apoptosis and DNA injury.

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