Three-Dimensional ECM-Functionalized PAN/C500 Nanofiber Scaffolds Induce Cytoskeletal Remodeling and Stemness-Associated Molecular Changes in Glioblastoma Cells

Glioblastoma (GBM) is the most aggressive primary brain tumor and remains associated with poor clinical outcomes despite advances in surgical and adjuvant therapies. The tumor microenvironment, particularly extracellular matrix (ECM) interactions, plays a crucial role in regulating glioblastoma progression, cellular plasticity, and therapeutic resistance. Therefore, physiologically relevant three-dimensional (3D) models are needed to better recapitulate GBM biology. In this study, we investigated the effects of ECM-functionalized polyacrylonitrile/coumarin-500 (PAN/C500) nanofiber scaffolds on the phenotype of LN-18 and U-87 MG glioblastoma cells cultured under 3D conditions. Cytoskeletal organization was assessed by phalloidin staining and live-cell vimentin imaging, while epithelial–mesenchymal transition (EMT)-associated proteins and stemness-related markers were analyzed by Western blotting. ECM-functionalized 3D PAN/C500 scaffolds promoted significant cytoskeletal remodeling, altered EMT-associated protein expression, and increased the expression of stemness-associated proteins, particularly SOX2, NANOG, and Nestin, compared with conventional 2D cultures. These responses were accompanied by cell line-dependent phenotypic adaptations, indicating that the engineered microenvironment influences glioblastoma cell behavior. This platform may serve as a valuable model for investigating glioblastoma biology and microenvironment-associated molecular adaptations in vitro.