Britannin通过GSK-3β/β-catenin信号传导途径抑制肝细胞癌发育和转移

肝细胞癌（HCC）是造成癌症相关死亡的重要原因。传统中药（TCM）在治疗HCC方面具有几个优势。 Britannin是Inulae Flos中的一种关键化合物，已经证明了对各种癌症的药理作用，但是对其具体抗HCC效应的研究仍然有限。这项研究旨在探索Britannin及其基本机制的抗HCC效应。MTT分析，克隆形成分析和流感细胞群以较小的蜂窝活动的速度和跨度的活动。通过伤口治疗测定和Transwell迁移和入侵测定法评估了用Britannin处理的HCC细胞系的细胞迁移和侵袭能力。构建了H22异种移植的肿瘤小鼠模型，并进行了Britannin处理以观察Britannin对HCC肿瘤的影响。 ELISA检测到肝癌生物标志物AFP，AFP-L3，APT和TGF-β的表达水平，通过染色观察到组织病理学。网络药理学和分子对接用于预测Britannin抗HCC效应的可能信号传导途径。表面等离子体共振（SPR）实验用于验证Britannin和蛋白质之间的相互作用。使用细胞激酶活性函数实验来检测不列颠宁对酶活性的影响。使用RT-QPCR和西部印迹来验证不列颠宁对与小鼠HCC细胞和肿瘤组织中与GSK-3β/β-catenin途径有关的关键基因和蛋白质水平的mRNA表达的作用。在体外实验中，Britann可以抑制其活性，迁移，迁移和侵袭HCC的活性，同时促进HCC的活性，同时促进HCC的能力。体内实验表明，不列颠宁对移植的肝癌肿瘤的生长施加了抑制作用，从而降低了移植小鼠中肝癌标志物的AFP，AFP-L3，APT和TGF-β的表达水平。网络药理学和分子对接预测细胞粘附因子和GSK-3β/β-catenin途径可能是相关的信号通路，并且具有与关键蛋白质的潜在扩展活性。 SPR实验阐明了Britannin和GSK-3β之间的分子相互作用。酶活性测定表明，不列颠宁可以调节GSK-3β激酶的功能活性。 RT-QPCR表明Britannin可以调节β-catenin，GSK-3β，E-钙粘蛋白和Ncadherin的mRNA表达。 Western-slot进一步证明了不列颠宁可以显着上调GSK-3β的表达，并下调P-GSK-3β和β-catenin的表达。同时，e-钙粘蛋白的表达增加和ncadherin降低，从而减少EMT的发生并抑制HCC的转移。结论，不列颠宁可以抑制HCC的生长，发育和转移，其机制可能与GSK-3β/β-Catnial nir-β-cateenin nime-β-catneenin nir-β-cateenin nir-β-cateenin nirition-lid-emeb-lid-β-catnial ialin-β-catnial ialin-β-catnial ialin sym-β-lid-β-固定蛋白有关有关HCC。

Hepatocellular carcinoma (HCC) stands out as a significant contributor to cancer-related death. Traditional Chinese Medicine (TCM) offers several advantages in the treatment of HCC. Britannin, a pivotal compound in Inulae Flos, has demonstrated pharmacological effects against various cancers, yet research on its specific anti-HCC effects remains limited.This study aims to explore the anti-HCC effects of britannin and its underlying mechanism.MTT assay, clone formation assay and flow cytometry were utilized to detect the cell activity, proliferation ability and apoptosis of britannin against HCC cell lines. Cell migration and invasion abilities of HCC cell lines treated with britannin were evaluated by wound-healing assay and transwell migration and invasion assay. H22 xenografted tumor mouse model was constructed and britannin treatment was performed to observe the effect of britannin on HCC tumors. The expression levels of liver cancer biomarkers AFP, AFP-L3, APT and TGF-β were detected by Elisa, and the histopathology was observed by HE staining. Network pharmacology and molecular docking were used to predict the possible signaling pathway of anti-HCC effect of britannin. The surface plasmon resonance (SPR) experiment was used to verify the interaction between britannin and proteins. The cell kinase activity function experiment was employed to detect the effect of britannin on enzyme activity. RT-qPCR and Western-Blot were used to verify the effect of britannin on the mRNA expressions of key genes and protein levels related to GSK-3β/β-catenin pathway in HCC cells and tumor tissues in mice.In vitro experiments showed that britannin could inhibit the activity, proliferation, migration and invasion abilities of HCC cells, while promoting their apoptosis. In vivo experiments revealed that britannin exerted inhibitory effects on the growth of transplanted liver cancer tumors, reducing the inflammatory infiltration and the expression levels of AFP, AFP-L3, APT and TGF-β of liver cancer markers in transplanted mice. Network pharmacology and molecular docking predicted that cell adhesion factors and GSK-3β/β-catenin pathway might be the related signaling pathway and had potential docking activity with key proteins. The SPR experiments elucidated the molecular interaction between britannin and GSK-3β. Enzyme activity assays indicated that britannin could modulate the functional activity of GSK-3β kinase. RT-qPCR suggested britannin could regulate the mRNA expressions of β-catenin, GSK-3β, E-cadherin and NCadherin. Western-Blot further verified that britannin could significantly up-regulate the expression of GSK-3β and down-regulate the expression of p-GSK-3β and β-catenin. At the same time, the expression of E-cadherin increased and NCadherin decreased, thereby reducing the occurrence of EMT and inhibiting the metastasis of HCC.In conclusion, britannin could inhibit the growth, development and metastasis of HCC, and its mechanism may be related to the regulation of GSK-3β/β-catenin signaling pathway to inhibit epithelial-mesenchymal transition of HCC.