基于整合药理学的柴胡不同部位抗癫痫作用研究

目的研究柴胡Bupleuri Radix的抗癫痫活性及其发挥药效的物质基础和潜在的分子作用机制。方法采用活性追踪的方法,考察柴胡低、中、高极性部位分别对戊四唑诱导的小鼠致癫痫模型的影响。并采用LC-MS技术对柴胡抗惊厥活性部位化学成分进行全面表征。通过整合药理学方法对柴胡抗癫痫活性部位潜在作用机制进行初步预测,并构建柴胡潜在靶标与癫痫疾病靶标相互作用网络。最后,对于其中重要的通路进行验证。结果通过小鼠造模剂量筛选,选择戊四唑55.18mg/kg造模,发现柴胡高极性部位(20 g/kg)可以显著延长癫痫小鼠的阵挛潜伏期与强直潜伏期,并能显著减少惊厥次数。化学成分鉴定结果表明,在正、负离子模式下从柴胡抗惊厥活性部位共鉴定出32种化学成分,主要为柴胡皂苷类成分,还包括脂肪酸类、糖类和香豆素类等成分。通过整合药理学平台筛选出柴胡抗癫痫的关键靶标22个,其中疾病与药物共有主要靶点有10个,包括ATP5A1、ATP5B、ATP1A1、ATP5C1、PPP3CA、PTGS2、AR等;其富集的通路包括帕金森病、能量代谢、亨廷顿氏病、卵母细胞减数分裂、神经退行性疾病等。最终,验证了能量代谢中关键节点,结果发现柴胡高极性部位能明显回调癫痫小鼠海马和肝脏中三磷酸腺苷(adenosine triphosphate,ATP)含量。结论柴胡高极性部位具有显著抗惊厥活性,其主要通过调节能量代谢通路来抑制癫痫的发生,其中柴胡皂苷类成分主要调节能量代谢,并通过改善ATP含量水平起到抗癫痫的作用,这可能是柴胡治疗癫痫的主要机制。

Antiepileptic effect and mechanism of different parts of Bupleuri Radix by integrative pharmacology

Objective To investigate the major active components and potential molecular mechanism of Bupleuri Radix in treatment of epilepsy. Methods The activity-tracking method was used to obtain the anticonvulsant active parts of Bupleuri Radix by investigating the effects of three different polar sites on pentylenetetrazol (PTZ) kindled seizures in the rat. And the LC-MS technology was used to fully characterize the chemical components of the anticonvulsant active parts of Bupleuri Radix. The protein targets related with epilepsy were collected by integrated pharmacology method and the interaction network of potential targets of Bupleuri Radix and epilepsy disease targets was constructed to illustrate the molecular mechanism. Results By screening for model dosage, 55.18 mg/kg PTZ was selected to establish the model. The time of clonic latency and tonic latency significantly was prolonged and number of convulsions significantly reduced in rats after high-polar fraction of Bupleuri Radix (20 g/kg) treatment. A total of 32 chemical components was identified with the positive and negative ion mode, primarily including saikosaponins, fatty acids, sugars and coumarins, etc. A total of 22 key targets were screened. The main targets of disease and drug were ATP5A1, ATP5B, ATP1A1, ATP5C1, PPP3CA, PTGS2, AR, etc, which mainly related to Parkinson''s disease, energy metabolism, Huntington''s disease, oocyte meiosis, neurodegenerative diseases and other pathways. Furthermore, the keystone node in energy metabolism was validated by determining ATP content of hippocampus and liver, and the result showed that the decrease of ATP content in hippocampus and liver induced by epilepsy was significantly reversed by the high-polar fraction of Bupleuri Radix. Conclusion The high-polar fraction of Bupleuri Radix has anticonvulsant activity, it inhibited the occurrence and development of seizures mainly by regulating energy metabolism. And saikosaponins and flavonoids mainly regulated energy metabolism by improving ATP content, which may be the main mechanism for the treatment of epilepsy with Bupleuri Radix.