Skeletal Stem Cell Isolation from Cranial Suture Mesenchyme and Maintenance of Stemness in Culture.

Skeletal stem cells residing in the suture mesenchyme are responsible for calvarial development, homeostatic maintenance, and injury-induced repair. These naïve cells exhibit long-term self-renewal, clonal expansion, and multipotency. They possess osteogenic abilities to regenerate bones in a cell-autonomous manner and can directly replace the damaged skeleton. Therefore, the establishment of reliable isolation and culturing methods for skeletal stem cells capable of preserving their stemness promises to further explore their use in cell-based therapy. Our research team is the first to isolate and purify skeletal stem cells from the calvarial suture and demonstrate their potent ability to generate bone at a single-cell level. Here, we describe detailed protocols for suture stem cell (SuSC) isolation and stemness maintenance in culture. These methods are extremely valuable for advancing our knowledge base of skeletal stem cells in craniofacial development, congenital deformity, and tissue repair and regeneration.