Mechanical Stretch Disrupts Calcium Dynamics and Redistributes Piezo1 in Human Astrocytes

PURPOSE: Astrocytes regulate the activity of nearby neurons so disruption of astrocyte calcium dynamics by traumatic brain injury (TBI) could have profound consequences for neural network activity in the brain. This study aimed to define how mechanical stretch injury alters calcium signaling, mitochondrial membrane potential, and mechanosensitive ion channel organization in human induced pluripotent stem cell (hiPSC)-derived astrocytes. METHODS: Human iPSC-derived astrocytes were subjected to controlled two-dimensional stretch injury across multiple severities. Live-cell calcium and mitochondrial membrane potential imaging, Piezo1 immunostaining, and RNA sequencing were used to assess functional and transcriptional responses. RESULTS: Cell viability, mitochondrial membrane potential, and spontaneous calcium transients declined in a severity-dependent manner. At moderate injury levels, reductions in mitochondrial function, calcium dynamics, and Piezo1 spatial distribution were transient. RNA sequencing identified 196 differentially expressed genes, including downregulation of mitochondrial and oxidative metabolic pathways and upregulation of cortical thinning-associated pathways. CONCLUSION: This platform captures functional and molecular hallmarks of astrocyte injury and provides a human in vitro model for studying mechanobiological pathways linking TBI to neurodegenerative processes.