An In Vitro System for Applying Tensile Force to Skeletal Stem Cells Using a Cyclic Cell Stress Tension System

Bone is a major mechanosensitive organ that continuously responds to physical cues in the body. Although mechanical stimulation plays important roles in skeletal development, homeostasis, and injury repair, the specific responses of skeletal stem cells (SSCs) to mechanical stress remain incompletely understood. To address this question, we established an in vitro model to isolate mouse periosteal SSCs and apply mechanical stimulation. Mouse periosteal cells were first obtained by enzymatic digestion, and SSCs were subsequently isolated by fluorescence-activated cell sorting (FACS) based on established surface markers. After attachment to culture, SSCs were subjected to tensile mechanical stimulation using a cyclic cell stress-tension system. This system enables the reproducible application of tensile force to cultured cells and provides a platform for analyzing SSC responses to mechanical input. Using this approach, we found that the expression of the cellular senescence-associated genes p16 and p21 was markedly reduced in SSCs following mechanical stimulation. These findings suggest that tensile stimulation may influence senescence-related changes in SSCs under in vitro conditions. Overall, this protocol provides a useful and suitable platform for studying SSC behavior under mechanical stimulation and for investigating how mechanical cues affect SSC function.