摘要
目的探讨α细辛醚对大鼠脑微血管内皮细胞P糖蛋白(P-gp)功能与表达的影响。方法采用100μmol/L L-谷氨酸作用于脑微血管内皮细胞30 min,诱导其细胞膜上P-gp高表达,模拟癫痫反复发作诱导的耐药性癫痫血脑屏障中血管内皮细胞P-gp升高。MTT法检测α细辛醚的安全药物浓度范围,设置3个浓度梯度(12.5、25.0、50.0μg/μl)干预模型细胞24 h。干预完成后,采用Western blot法检测P-gp表达,Real-time PCR检测多药耐药基因1a(Mdr1a)mRNA扩增,罗丹明123(Rh123)转运实验测定Rh123在脑微血管内皮细胞内的累积量变化,判断α细辛醚对P-gp功能的影响。结果正常大鼠脑微血管内皮细胞经谷氨酸刺激后,P-gp(F=1.924,P=0.020)与Mdr1a mRNA(F=1.788,P=0.019)表达皆显著升高。25.0(F=1.924,P=0.025;F=1.788,P=0.017)、50.0μg/μl(F=1.924,P=0.035;F=1.788,P=0.026)α细辛醚作用后,p-gp和Mdr1a mRNA的表达较模型组均显著降低。与模型细胞相比,25.0(t=6.085,P=0.000)、50.0μg/μl(t=8.912,P=0.000)α细辛醚作用后,血管内皮细胞内Rh123皆显著升高,升高率分别接近40%和60%。结论α细辛醚可以扭转谷氨酸诱导的大鼠脑微血管内皮细胞P-gp/Mdr1a的高表达,显著增加Rh-123在脑微血管内皮细胞内的蓄积。α细辛醚可能对P-gp介导的耐药性癫痫的耐药性具有一定扭转作用。
Objective To investigate the effect ofα-asarone on the function and expression of P-glycoprotein(P-gp)in rat brain microvascular endothelial cells(rBMECs).Methods rBMECs were exposed to L-glutamate(100μmol/L)for 30 mins to induce the overexpression of P-gp/multidrug resistance gene 1a(Mdr1a)on the cell membranes,which mimicked the overexpression of P-gp/Mdr1a in blood brain barrier(BBB)when drug-resistant epilepsy attacked.MTT assay was used to detect the safe range ofα-asarone concentration.The model cells were intervened with different concentrations ofα-asarone at 12.5,25.0,and 50.0μg/μl for 24 hours.After the treatment ofα-asarone,the expression and the function of P-gp/Mdr1 were measured by Western blotting,real-time PCR,and intracellular rhodamine 123 accumulation assays.Results The rBMECs,stimulated by glutamine,showed a high expression of P-gp(F=1.924,P=0.020)/Mdr1a(F=1.788,P=0.019)compared to the normal rBMECs.The treatment with 25.0(F=1.924,P=0.025;F=1.788,P=0.017)and 50.0μg/μl(F=1.924,P=0.035;F=1.788,P=0.026)α-asarone significantly depressed the expression of P-gp/Mdr1a.The treatment with 25.0 and 50.0μg/μlα-asarone significantly increased intracellular accumulation of Rhodamine 123 by 40%and 60%respectively.Conclusionsα-asarone down-regulates the high expressions of P-gp and Mdr1a mRNA in rBMECs induced by L-glutamate.Moreover and increases intracellular accumulation of rhodamine-123.Thus,α-asarone may reverse drug resistance in P-gp-mediated drug-resistant epilepsy.
作者
袁斯远
孙金
刘金民
YUAN Siyuan;SUN Jin;LIU Jinmin(Department of Neurology,Third Affiliated Hospital,Beijing University of Chinese Medicine,Beijing 100029,China;Department of Neurology,Dongfang Hospital,Beijing University of Chinese Medicine,Beijing 100078,China)
出处
《中国医学科学院学报》
CAS
CSCD
北大核心
2020年第1期1-6,共6页
Acta Academiae Medicinae Sinicae
基金
国家自然科学基金(81774277)~~
关键词
α细辛醚
P糖蛋白
脑微血管内皮细胞
耐药性癫痫
α-asarone
P-glycoprotein
brain microvascular endothelial cells
drug-resistant epilepsy
