基于UPLC-MS/MS、网络药理学与体外实验探讨甲术二草汤抗结直肠癌作用机制OA

Objective Based on ultra-performance liquid chromatography-tandem mass spectrometry(UPLC-MS/MS),network pharmacology,and in vitro experiments,this study aimed to preliminarily characterize the chemical composition of Jia Zhu Er Cao decoction(JZEC),evaluate its effects on the biological behaviors of colorectal cancer(CRC)cells,and explore its potential underlying mechanisms.Methods UHPLC-Q-TOF-MS/MS was performed to identify the major chemical constituents of JZEC.Network pharmacology was applied to screen potential JZEC-CRC targets,construct a compound-target network and protein-protein interaction(PPI)network,and identify core targets using cytoNCA-based topological analysis.GO and KEGG enrichment analyses combined with molecular docking were performed to investigate the potential mechanisms of JZEC.SW480 cells were used as the primary in vitro model,and CCK-8 assay,wound healing assay,Transwell assay,flow cytometry,Mito-Tracker staining,and Western blotting were conducted to evaluate the anti-CRC effects and potential mechanisms of JZEC.Results UHPLC-Q-TOF-MS/MS analysis identified 54 representative candidate compounds in JZEC,including curzerene,oridonin,ponicidin,quercetin,and rutin,which mainly belonged to flavonoids,polyphenols,terpenoids,and organic acids.Network pharmacology analysis identified 123 potential JZEC-CRC targets.The compound-target network suggested that representative chemical constituents,including quercetin,apigenin-related derivatives,and ursolic acid,may contribute to the anti-CRC effects of JZEC.Furthermore,PPI network analysis combined with cytoNCA-based topological analysis identified 19 core targets,including AKT1,EGFR,SRC,and MAPK1.GO and KEGG enrichment analyses revealed that these targets were mainly associated with biological processes such as cell proliferation,migration,apoptosis,and reactive oxygen species metabolism,and were enriched in CRC-related signaling pathways,including the PI3K-Akt and FoxO pathways.Molecular docking results indicated that representative chemical constituents exhibited favorable binding capacities with some key targets.In vitro experiments demonstrated that JZEC reduced SW480 cell viability,inhibited cell migration,and promoted apoptosis,accompanied by alterations in mitochondrial functional status and the expression of migration-and apoptosis-related proteins.Following JZEC treatment,the levels of p-JAK2/JAK2 and p-STAT3/STAT3 were decreased,whereas the expression levels of total JAK2 and STAT3 remained unchanged.Conclusion JZEC can reduce SW480 cell viability,inhibit SW480 cell migration,and promote apoptosis,accompanied by changes in migration-and apoptosis-related protein expression and modulation of JAK2/STAT3 signaling activation status.These findings suggest that the JAK2/STAT3 signaling pathway may be involved in the in vitro anti-CRC effects of JZEC.