Mechanistic evaluation of Curcumae rhizoma-Atractylodis macrocephalae rhizoma herb pair enhancing anti-chronic everyinjury through PI3K/AKT/mTOR signaling pathway.

内容摘要

Chronic hepatic injury is a liver disease that poses a threat to human health with increasing morbidity. Chinese herb pairs are the fundamental and concise form of traditional Chinese medicine (TCM) prescriptions, which can effectively explain their underlying concepts. Curcumae rhizoma (Ezhu) and Atractylodis macrocephalae rhizoma (Baizhu) herb-pair (PW) have been frequently used in TCM prescriptions for hundreds of years, which derived from the renowned traditional Chinese medical classic "Yixue Zhongzhong Canxi Lu", primarily to treat liver-related conditions such as hypochondriac pain, abdominal mass, amassment and accumulation that are closely associated with liver diseases including hepatic injury and hepatic fibrosis in modern medicine. As a classic drug pair of invigorating Qi and promoting blood circulation, they align with the TCM syndrome characteristics of chronic hepatic injury with qi deficiency and blood stasis. However, the specific mechanism of the therapeutic effects of PW against chronic hepatic injury remains unclear. The study aimed to assess the enhanced efficacy, underlying targets, and the potential mechanism of PW in treating chronic hepatic injury. UPLC-MS/MS analysis was performed to identify the active components of PW. Subsequently, the therapeutic efficacy as well as latent action mechanisms of PW upon chronic hepatic injury were examined in vivo study in rats with chronic hepatic injury and in vitro in LX-2 cells. A network pharmacology method was used to prognosticate the mechanisms by which PW treats chronic hepatic injury. Integrated application of pharmacokinetic analysis and metabolomics approaches was employed to identify and characterize critical active ingredients and regulatory signaling pathways. Western blot assays and molecular docking were conducted to reveal the primary mechanisms of action. Totally 1,059 metabolites were identified, including 662 detected in positive ion mode and additional 397 detected in negative ion mode. Network pharmacology suggested that the protective effect against chronic hepatic injury of PW may be closely relevant to PI3K/AKT/mTOR signaling pathway. Pharmacodynamics assessment indicated that PW effectively inhibited chronic HI progression in vivo compared with the two herbs alone. PW significantly improved lipid deposition, hepatic function, inflammation and histopathological injury. Additionally, PW significantly decreased the serum AST, ALT and ALP activity levels and effectively reduced the expression level of α-SMA in the liver, significantly inhibited the phosphorylation level of the PI3K/AKT/mTOR pathway. The results of pharmacokinetics and tissue distribution studies showed that PW may increase the concentration of potential bioactive compounds like Curcumol, Curdione, Germacrone, Furanodiene, Atractylenolide III and Atractylon, thereby enhancing the effect of anti-chronic hepatic injury. Combined metabolomics and WB analysis of rat liver tissue revealed that the enhanced anti- chronic hepatic injury effect of PW may be associated with PI3K/AKT/mTOR signaling pathway. Experiments on LX-2 cells indicated that the combination of Curdione and Atractylolide III exerted a hepatoprotective effect by reducing the phosphorylation levels of proteins in the PI3K/AKT/mTOR pathway. PW significantly increases the plasma exposure of potential active compounds and the liver distribution level. It emerges as a potential therapeutic strategy for chronic HI, thereby exerting an enhanced anti-chronic hepatic injury effect accompanying fibrotic lesions through the modulation of PI3K/AKT/mTOR signaling pathway.