Aim: In this study, we explored the ability of TAMs to affect the malignant phenotype of human hepatoma Huh-7 cells through the Gli2/IGF-II/ERK1/2 pathway.
Methods: The TAMs were characterized by flow cytometry and ELISA assays. Huh-7 cells were treated with conditioned medium of TAMs (TAMs-CM), and the proliferation, migration and invasion abilities were measured by CCK-8, Transwell and scratch assays. The levels of TGF-β1, Gli2, IGF-II and related proteins in the ERK1/2 pathway and the epithelial-mesenchymal transition (EMT) process were examined by RT-qPCR and western blot. Huh-7 cells were injected subcutaneously into nude mice with TAMs to explore the role of TAMs in tumor growth.
Results: The expression levels of TGF-β1, Gli2 and IGF-II and the cell proliferation, migration and invasion abilities were elevated in Huh-7 cells treated with TAMs-CM. TGF-β1 was upregulated in the conditioned medium and was found to be involved in the promotion of migration, invasion and the EMT of Huh-7 cells. The activation of TGF-β1 signaling increased the expression of Gli2. Knockdown of Gli2 decreased the expression of IGF-II and also reversed the promotional effect of the conditioned medium on migration, invasion and the EMT of Huh-7 cells. TGF-β1/Gli2/IGF-II signaling was shown to promote the malignant phenotype of Huh-7 cells by activating the ERK1/2 signaling pathway. Further, TGF-β1 knockdown attenuated the influence of TAMs on tumor growth in mouse model.
Conclusion: The TGF-β1 secreted by TAMs promotes the migration, invasion and EMT of human hepatoma Huh-7 cells through the Gli2/IGF-II/ERK1/2 pathway.
Keywords: ERK1/2 signaling pathway; Gli2; Hepatocellular carcinoma; IGF-II; TGF-β1; tumor growth; tumor-associated macrophages.