Sfrp4 is Required to Maintain Ctsk-Lineage Periosteal Stem Cell Niche Function

The periosteum contains cells which function as a reservoir of stem cells and progenitors and contribute to cortical expansion during growth, cortical bone homeostasis and repair. However, the local or paracrine factors that govern stem cell renewal and differentiation within the periosteal niche remains elusive. Cathepsin K (Ctsk) together with additional cell surface markers marks a subset of stem cells in the periosteum (PSC) which possess self-renewal ability and inducible multipotency. These PSCs produce osteoblasts mediating periosteal bone formation and fracture repair. Sfrp4 is expressed in periosteal Ctsk-lineage cells and using CtskCre mice that are either wild type or Sfrp4-/-, we report here that Sfrp4 deletion decreases the pool of PSCs, impairs their self-renewal, their ability to give rise to their derivatives and their clonal multipotency for differentiation into osteoblasts and chondrocytes in vitro and formation of bone organoids in vivo. Bulk RNA sequencing analysis in Ctsk-lineage PSCs demonstrated that Sfrp4 deletion leads to downregulation of signaling pathways associated with skeletal development, positive regulation of bone mineralization and wound healing. Sfrp4 deletion hampers the Ctsk-lineage PSC response and recruitment after bone injury and leads to an impaired periosteal response. Periosteal Ctsk-lineage cells respond to PTH(1-34) treatment with an increase in the % of PSCs, a response not seen in the absence of Sfrp4. Importantly, bone histomorphometry analysis showed that in the absence of Sfrp4, PTH(1-34)-dependent increase in cortical thickness, periosteal bone formation is markedly impaired. Overall design: To gain insight into the function of Sfrp4 in the regulation of the Ctsk-lineage periosteal stem cell (PSC) stemness, differentiation and function, we generated CtskCre;mTmG;wt (GFP+) and CtskCre;mTmG;Sfrp4-/- (GFP+;Sfrp4-/-) mice and investigated Ctsk+ (GFP+) labeled and GFP+;Sfrp4-/- PSC. To better understand the changes occurring in the Ctsk-lineage PSC population in the absence of Sfrp4 we performed bulk RNA-seq analysis. Two biological samples were collected per GFP+ PSC and GFP+Sfrp4-/- PSCs, each composed of the pooled cells yield from n=3 mice.