Scleraxis-expressing progenitor cells are critical for the maturation of the annulus fibrosus and demonstrate therapeutic potential

Background: Annulus fibrosus (AF) is an important part of the intervertebral disc (IVD) and its injury leads to back pain and impaired mobility. The stem/progenitor cells are essential for the maturation and repair of the AF, however, the identity of AF stem/progenitor cells remain elusive. Methods: In this study, we sorted cells from the murine IVDs and performed the single-cell RNA sequencing. Using single-cell transcriptomics, genetic lineage tracing, in vitro stem cell experiment, ablation models and cell transplantation, we elucidate the role of AF progenitor cells in maturation and injury. Results: On the basis of single-cell RNA-sequencing (scRNA-seq) analysis of the intervertebral disc, we found that the transcription factor Scleraxis (Scx) can specifically label a progenitor cell population of the outer AF. By lineage tracing assay, Scx-lineage AF cells proliferate mainly prior to sexual maturity, but barely proliferate after age of 8 weeks. The Scx-expressing AF cells are enriched for stem/progenitor cell markers and show a higher proliferative capacity and differentiation potential than the Scx− cells. The ablation of Scx-expressing AF cells impairs the maturation of AF. The Scx+ AF cells are enriched for TGFβ signaling. Transplantation of Scx-lineage cells to injured AF with Connective tissue growth factor (CTGF) improved the AF healing. Conclusions: Scleraxis-expressing progenitor cells are critical for the maturation of AF and demonstrate therapeutic potential for AF regeneration. The translational potential of this article: These findings expand the important role of stem cells in maturation and repair and provide new strategy for cellular therapy of AF repair.