仙灵骨葆胶囊线粒体毒性物质基础及肝损伤机制研究

Xianling Gubao Capsule is a commonly used drug for the treatment of osteoporosis, and the curative effect is remarkable. The classic safety evaluation studies have shown that the whole preparation and the six Chinese herbal medicine are high security, but the long-term use of patients in the serious incidence of drug-induced liver injury is still high, therefore, elucidating the mechanism of liver injury and clearing the toxic substance base of Xianling Gubao Capsule are of great value to ensure the rational use of the drug. Mitochondrial damage is an important target of drug hepatotoxicity. In our preliminary study, we found that Xianling Gubao Capsule has significant mitochondrial inhibitory activity. We speculate that mitochondrial toxicity may be its predisposing factor and pathway for drug-induced liver injury. Therefore, the aim of this study is to track the activity of mitochondrial toxicity by using HPLC-MS/MS, UPLC/Q-TOF-MS and other methods to identify and isolate mitochondrial toxic components quickly and accurately, elucidate the toxic substance basis, and explore the mechanism of liver injury induced by Xianling Gubao by combining the metabolic and kinetic differences of toxic components in animals. This study not only provides the basis for the dose adjustment of the clinical medicine of Xianling Gubao Capsule, but also can provide scientific support for the improvement of its standard and the quality control of the production process.

仙灵骨葆胶囊是临床治疗骨质疏松症的常用药物，疗效显著，经典的安全性评价显示全方和方中六味中药均具有较高的安全性，但长期服用的患者中严重药物性肝损伤事件发生率仍较高，因此，阐明仙灵骨葆胶囊的肝损伤作用机制，明确仙灵骨葆的毒性物质基础，对于保障该药物的合理使用具有重要价值。线粒体损伤是药物肝毒性作用的重要靶标，本课题组前期研究发现仙灵骨葆具有显著的线粒体抑制活性，我们推测，线粒体毒性可能是仙灵骨葆发生药物性肝损伤的诱发因素和经由途径。因此，本课题拟采用线粒体毒性评价技术活性追踪，综合利用HPLC-MS/MS、UPLC/Q-TOF-MS等分析方法，快速、准确的识别和分离线粒体毒性成分，阐明仙灵骨葆引发肝损伤的毒性物质基础，结合毒性成分在动物体内的代谢和动力学差异表征，探索仙灵骨葆诱发肝损伤的作用机制。本研究不仅可为临床用药的剂量调整提供依据，而且可以为其标准提升、生产过程质量控制提供科学支撑。

仙灵骨葆胶囊是临床治疗骨质疏松症的常用药物，疗效显著。经典的安全性评价显示全方和方中六味中药均具有较高的安全性，但仙灵骨葆引起的药源性肝损伤的发生率仍然很高，而引起肝损伤的毒性物质和作用机制尚不清楚。药物诱导的线粒体毒性被认为是药物性肝毒性的重要机制。本课题通过体外Seahorse assay对全方和方中六味中药进行线粒体功能分析确定补骨脂是导致仙灵骨葆发生线粒体毒性的目标中药。对补骨脂中的化合物进行细胞生物能量学研究表明，补骨脂定、补骨脂乙素、补骨脂二氢黄酮甲醚、补骨脂甲素和补骨脂素五个化合物具有线粒体毒性，尤其是补骨脂定，可导致线粒体储备能力和呼吸控制率显著降低。机制研究表明，仙灵骨葆致肝损伤的分子机制与激活PI3K/mTOR信号通路抑制自噬并诱导线粒体凋亡有关。本研究不仅可为仙灵骨葆胶囊的临床用药提供预警，还可为其他药物性肝损伤中药的毒性研究提供借鉴作用。