IADR Abstract Archives

Establishment of an in vitro culture system for tenogenic/ligamentogenic differentiation using ScxGFP iPS cells

Abstract: Tendons connect muscles to the skeletal components and function as the mechanical force transmitters, while ligaments bind adjacent bones together to stabilize joints. Cells in tendons and ligaments are specialized fibroblasts known as tenocytes and ligamentocytes. Tenocytes and ligamentocytes are derived from progenitor cells that express the basic helix-loop-helix transcription factor Scleraxis (Scx). A certain population of Scx+ cells also express Sry-box 9 (Sox9). These Scx+/Sox9+ cells differentiate into tenocytes and ligamentocytes as well as chondrocytes that contribute to the formation of attachment sites of tendons/ligaments to bone, entheses. Loss of function analysis of Scx revealed that Scx is required for maturation of tendons/ligaments and the entheses. So far, little is known about the molecular mechanism underlying tenogenic and ligamentogenic lineage commitment and differentiation due to a lack of suitable cell culture system. In this study, we established an in vitro culture system that can monitor tenocyte and ligamentocyte differentiation by live imaging using green fluorescence. ScxGFP iPS cells were established from fibroblasts isolated from ScxGFP transgenic mouse embryos that express EGFP in the Scx-expressing region during development. We generated chimeric mice to confirm that ScxGFP iPS derived cells express green fluorescence upon differentiation into the tenogenic and ligamentogenic cells. We then explored the culture conditions for differentiation of ScxGFP iPS cells into tendon/ligament cells monitoring GFP expression. Under optimal conditions, ScxGFP iPS cells are differentiated into mesenchymal progenitor cells and then into Scx+ cells expressing GFP. Scx+ cells mature to express tenomodulin and Mohawk at higher levels than tenocytes isolated from limb tendons. Most of Scx+ cells express Sox9, but Sox9 expression in Scx+/Sox9+ cells decreased in association with tenocyte and ligamentocyte differentiation. Thus, taking advantage of ScxGFP iPS cells, we have successfully established the culture system that recapitulates the process of tendon and ligament formation in vivo.
Japanese Division Meeting
2020 Japanese Division Meeting (Virtual)
Virtual, Japan
Future is now! Stem cell revolution in hard and soft tissue engineering
  • Shukunami, Chisa  ( Department of Molecular Biology and Biochemistry, Graduate School of Biomedical and Health Sciences, Hiroshima University )
  • Symposium II