Nuclear deformation acts as a mechanical switch to drive breast cancer cell migration in a confined microenvironment

<b>Background:</b> Tumour metastasis is the primary cause of high mortality in cancer patients, and the confined migration of cancer cells is the key step in successful metastasis. However, the biomechanical properties of cancer cells during confined migration and the associated mechanotransduction mechanisms remain elusive. In this study, a hydrogel-based microchannel platform was used to investigate the migratory behaviours of breast cancer cells in wide, medium, and narrow microchannels. <b>Methods:</b> Using fluorescence microscopy, we initially characterized MDA-MB-231 breast cancer cells cultured in three distinct hydrogel-based microchannel systems and assessed both whole-cell and nuclear morphology. In parallel, cell migration dynamics were quantified via time-lapse imaging. Immunofluorescence and laser confocal imaging were subsequently employed to systematically analyse the degree of nuclear envelope unfolding and the differential expression of Piezo1. To elucidate the force-sensing mechanism, live-cell calcium imaging was performed to record responses to mechanical stimuli. Ultimately, by constructing plasmids to regulate Lamin A/C expression (knockdown or overexpression) specifically, we demonstrated its role in controlling restricted migration through targeted interference with nuclear shape changes. <b>Results:</b> The results demonstrated that the breast cancer cells displayed the strongest motility in narrow microchannels. Moreover, upon confinement-induced nuclear deformation, the nuclear membranes unfold and tense, which acts as a mechanical switch to facilitate the rapid migration of breast cancer cells in narrow microchannels. Further investigation revealed that the mechanosensitive ion channel Piezo1 was activated on breast cancer cells in narrow microchannels, thereby accelerating calcium influx. This process not only maintained nuclear membrane tension but also activated the cytosolic calcium-dependent phospholipase A2 (cPLA2)-arachidonic acid (AA) pathway, enhancing cell migration via increased myosin II-driven contractility. <b>Conclusions:</b> This study demonstrates the fundamental importance of nuclear deformation and mechanotransduction in cancer cell migration, providing new perspectives for the development of therapeutic approaches that target nuclear mechanics to inhibit metastatic progression.