Chongqing Three Gorges Medical College, Chongqing 404120, China; Chongqing Key Laboratory of Development and Utilization of Genuine Medicinal Materials in Three Gorges Reservoir Area, Chongqing 404120, China.
Chongqing Three Gorges Medical College, Chongqing 404120, China; Chongqing Key Laboratory of Development and Utilization of Genuine Medicinal Materials in Three Gorges Reservoir Area, Chongqing 404120, China.
Phytomedicine. 2024 Jun;128:155338. doi: 10.1016/j.phymed.2024.155338. Epub 2024 Jan 7.
Liver cancer, one of the most common types of cancer worldwide, accounts for millions of cases annually. With its multi-target and wide-ranging therapeutic effects, traditional Chinese medicine has emerged as a potential approach for treating various tumors. Codonopsis pilosula, a traditional herb, is known for its anti-inflammatory and antioxidant properties. In this study, we investigated the potential molecular mechanisms of Codonopsis pilosula in regulating the inhibition of CDK1 and the modulation of PDK1/β-catenin, which are involved in hepatocellular carcinoma growth and metastasis.
STUDY DESIGN/METHODS: Firstly, we screened the active chemical constituents of Codonopsis pilosula and identified their respective target proteins using the Herb database. Then, we applied the GeneCards database and transcriptome sequencing analysis to screen for critical genes associated with the occurrence and development of liver cancer. The intersection of the target proteins and disease-related genes was used to determine the potential targets of Codonopsis pilosula in hepatocellular carcinoma. Protein-protein interaction analysis and GO/KEGG analysis were subsequently performed to uncover the pathways through which Codonopsis pilosula acts on liver cancer. The Huh-7 cell line, exhibiting the highest sensitivity to Codonopsis pilosula polysaccharide solution (CPP) intervention, was chosen for subsequent studies. Cell viability was evaluated using the CCK-8 assay, colony formation assay was conducted to determine cell proliferation capacity, flow cytometry was used to analyze cell cycle, TUNEL staining was performed to assess cell apoptosis, scratch assay was carried out to evaluate cell migration ability, the expression of EMT-related proteins was detected and analyzed, and cell sphere formation assay was conducted to investigate cell stemness. Finally, a liver cancer animal model was established, and different doses of CPP were administered via gavage the next day. The expression levels of CDK1, PDK1, and β-catenin in mouse liver tissues were detected and analyzed, immunohistochemistry staining was performed to assess the expression of tumor cell proliferation-related proteins Ki67 and PCNA in mouse xenografts, and TUNEL staining was carried out to evaluate cell apoptosis in mouse liver tissues. After intervention with CDK1 expression, the expression levels of CDK1, PDK1, and β-catenin proteins and mRNA in each group of cells were detected using Western blot and RT-qPCR.
Through network pharmacology analysis, transcriptome sequencing, and bioinformatics analysis, 35 target genes through which Codonopsis pilosula acts on liver cancer were identified. Among them, CDK1, with the highest degree in the PPI network, was considered an essential target protein for Codonopsis pilosula in treating liver cancer. In vitro cell experiments revealed that CPP could inhibit the expression of CDK1/PDK1/β-catenin signaling axis factors, suppress cell proliferation, decrease cell migration ability, influence the EMT process, and reduce cell stemness by inhibiting CDK1 and affecting the PDK1/β-catenin signaling axis. Similarly, in vivo experiments demonstrated that CPP could regulate the CDK1/PDK1/β-catenin signaling axis, inhibit tumor growth, and induce cell apoptosis.
Codonopsis pilosula may inhibit hepatocellular carcinoma growth by suppressing CDK1 and affecting the PDK1/β-catenin signaling axis, limiting cell EMT and reducing cell stemness. These findings provide insights into the potential therapeutic role of Codonopsis pilosula in liver cancer.
肝癌是全球最常见的癌症类型之一，每年有数百万人患病。中药具有多靶点、广泛的治疗效果，已成为治疗各种肿瘤的潜在方法。党参是一种传统草药，具有抗炎和抗氧化特性。在这项研究中，我们研究了党参调节 CDK1 抑制和 PDK1/β-连环蛋白调节的潜在分子机制，这些机制参与了肝细胞癌的生长和转移。
研究设计/方法：首先，我们从党参中筛选出活性化学成分，并使用 Herb 数据库鉴定它们各自的靶蛋白。然后，我们应用 GeneCards 数据库和转录组测序分析筛选与肝癌发生和发展相关的关键基因。靶蛋白和疾病相关基因的交集用于确定党参在肝癌中的潜在靶点。随后进行蛋白质-蛋白质相互作用分析和 GO/KEGG 分析，以揭示党参作用于肝癌的途径。选择对党参多糖溶液（CPP）干预最敏感的 Huh-7 细胞系进行后续研究。使用 CCK-8 测定法评估细胞活力，通过集落形成实验测定细胞增殖能力，通过流式细胞术分析细胞周期，通过 TUNEL 染色评估细胞凋亡，通过划痕实验评估细胞迁移能力，检测和分析 EMT 相关蛋白的表达，进行细胞球形成实验以研究细胞干性。最后，建立肝癌动物模型，第二天通过灌胃给予不同剂量的 CPP。检测和分析小鼠肝组织中 CDK1、PDK1 和 β-连环蛋白的表达水平，免疫组织化学染色检测小鼠异种移植中肿瘤细胞增殖相关蛋白 Ki67 和 PCNA 的表达，TUNEL 染色检测小鼠肝组织中的细胞凋亡。用 CDK1 表达干预后，用 Western blot 和 RT-qPCR 检测各组细胞中 CDK1、PDK1 和 β-连环蛋白蛋白和 mRNA 的表达水平。
通过网络药理学分析、转录组测序和生物信息学分析，确定了 35 个党参作用于肝癌的靶基因。其中，PPI 网络中程度最高的 CDK1 被认为是党参治疗肝癌的关键靶蛋白。体外细胞实验表明，CPP 可通过抑制 CDK1 表达并影响 PDK1/β-连环蛋白信号轴，抑制细胞增殖、降低细胞迁移能力、影响 EMT 过程并降低细胞干性，从而抑制 CDK1/PDK1/β-连环蛋白信号轴。同样，体内实验表明，CPP 可以调节 CDK1/PDK1/β-连环蛋白信号轴，抑制肿瘤生长并诱导细胞凋亡。
党参可能通过抑制 CDK1 并影响 PDK1/β-连环蛋白信号轴来抑制肝细胞癌的生长，限制细胞 EMT 并降低细胞干性。这些发现为党参在肝癌治疗中的潜在作用提供了新的思路。
