KDM6B regulates the epigenetic mechanism of epithelial-mesenchymal transition in differentiated thyroid cancer in response to extracellular matrix stiffness

Objective: This study aims to investigate the epigenetic mechanisms by which KDM6B regulates epithelial-mesenchymal transition (EMT) in differentiated thyroid cancer cells in response to changes in extracellular matrix (ECM) stiffness, and to elucidate its role in tumor invasion and metastasis. Methods: The differentiated thyroid cancer cell line K1 was used to prepare soft matrices (1 kPa), moderately stiff matrices (10 kPa), and stiff matrices (30 kPa). KDM6B was knocked down using siRNA and overexpressed using pcDNA3.1-KDM6B. The expression and activity of EMT markers (E-cadherin, N-cadherin, vimentin) and KDM6B were detected using real-time quantitative polymerase chain reaction (qRT-PCR), Western blot, Chromatin Immunoprecipitation-quantitative Polymerase Chain Reaction (ChIP-qPCR), and RNA sequencing (RNA-seq). Additionally, Transwell migration and invasion assays were performed to assess the migratory and invasive capabilities of the cells. GO and KEGG enrichment analyses were conducted to explore the key genes and signaling pathways regulated by KDM6B. Results: Under different ECM stiffness conditions, the mRNA and protein expression levels and enzymatic activity of KDM6B significantly increased (p < 0.001). Increased matrix stiffness led to a significant decrease in E-cadherin and a significant increase in N-cadherin and vimentin (p < 0.001). Following KDM6B knockdown, E-cadherin significantly decreased, N-cadherin and vimentin significantly increased, and cell migration and invasion capabilities were enhanced (p < 0.01). Conversely, overexpression of KDM6B significantly upregulated E-cadherin, significantly downregulated N-cadherin and vimentin, and inhibited cell migration and invasion capabilities (p < 0.01). ChIP-qPCR analysis indicated that KDM6B binding to the promoters of EMT genes such as Snail and Twist was significantly enhanced in high-stiffness matrices and regulated their transcriptional activity through demethylation of H3K27me3 (p < 0.05). RNA-seq and enrichment analyses revealed that KDM6B regulates key genes and multiple signaling pathways, including PI3K/Akt and MAPK, which are involved in biological processes such as cell proliferation and apoptosis. Conclusion: KDM6B regulates the EMT process in differentiated thyroid cancer cells in response to changes in ECM stiffness, primarily by affecting the expression of EMT-related genes through epigenetic modifications, thereby regulating cell migration and invasion capabilities. KDM6B provides a new theoretical basis as a potential therapeutic target for differentiated thyroid cancer.