Open micro-valley chip reveals long-term viscosity-induced glioblastoma cellular invasion states

The glioblastoma (GBM) microenvironment exhibits elevated viscosity and spatial confinement that strongly influence tumor invasion, yet these mechanical features are difficult to reproduce in open experimental systems. We developed an open two-layer microfluidic membrane that enables precise control of migration onset and real-time visualization of cellular mechano-adaptation. The detachable cap confines a defined droplet, while the ring-shaped micro-valley topography provides localized confinement that deforms nuclei and activates YAP signaling, recapitulating the mechanical stress experienced by invading tumor cells at the GBM invasive front. Using this platform, we found that long-term culture in a 7.1 cP viscous medium produced smaller, more deformable cells with enhanced migration through confined regions, revealing clear cell-type-dependent differences in motility and adaptive capacity. Transcriptomic analysis further showed that U-251 cells underwent mesenchymal-like reprogramming and gained greater invasive potential, whereas LN-229 cells exhibited limited transcriptional change despite similar structural remodeling. These findings demonstrate that this open microfluidic platform bridges biophysical modeling and cellular mechanobiology, enabling direct investigation of viscosity-driven adaptation in GBM.