R-spondin-2 is a Wnt agonist that regulates osteoblast activity and bone mass

Abstract

The R-spondin family of proteins are Wnt agonists, and the complete embryonic disruption of Rspo2 results in skeletal developmental defects that recapitulate the phenotype observed with Lrp5/6 deficiency. Previous work has shown that R-spondin-2 (Rspo2, RSPO2) is both highly expressed in Wnt-stimulated pre-osteoblasts and its overexpression induces osteoblast differentiation in the same cells, supporting its putative role as a positive autocrine regulator of osteoblastogenesis. However, the role of Rspo2 in regulating osteoblastogenesis and bone formation in postnatal bone has not been explored. Here we show that limb-bud progenitor cells from Rspo2 knockout mice undergo reduced mineralization during osteoblastogenesis in vitro and have a corresponding alteration in their osteogenic gene expression profile. We also generated the first Rspo2 conditional knockout (Rspo2^floxed) mouse and disrupted Rspo2 expression in osteoblast-lineage cells by crossing to the Osteocalcin-Cre mouse line (Ocn-Cre + Rspo2^f/f). Ocn-Cre + Rspo2^f/f male and female mice at 1, 3, and 6 months were examined. Ocn-Cre + Rspo2^f/f mice are decreased in overall body size compared to their control littermates and have decreased bone mass. Histomorphometric analysis of 1-month-old mice revealed a similar number of osteoblasts and mineralizing surface per bone surface with a simultaneous decrease in mineral apposition and bone formation rates. Consistent with this observation, serum osteocalcin in 3-month-old Ocn-Cre + Rspo2^f/f was reduced, and bone marrow-mesenchymal stem cells from Ocn-Cre + Rspo2^f/f mice undergo less mineralization in vitro. Finally, gene expression analysis and immunohistochemistry of mature bone shows reduced beta-catenin signaling in Ocn-Cre + Rspo2^f/f. Overall, RSPO2 reduces osteoblastogenesis and mineralization, leading to reduced bone mass.

Bone Physiology: R-spondin-2 reduces mineralization, leading to decreased bone mass

A loss of R-spondin-2 reduces osteoblastogenesis (production of osteoblasts, the cells from which bone develops) and mineralization, thereby leading to decreased bone mass in adults. R-spondin-2 is one of a family of four proteins that are expressed in the developing mouse limb as well as other tissues; each R-spondin family member exerts a different functional effect. R-spondins clearly influence several aspects of skeletal biology, but their specific roles—especially in postnatal bone—remained to be elucidated. A team headed by Kurt Hankenson at the University of Michigan Medical School investigated the role of R-spondin-2 in osteoblastogenesis, both in vitro and in vivo, using a mouse model. For the first time, the team was able to demonstrate that R-spondin-2 promotes osteoblastogenesis, bone development, and consequent bone mass growth in adult mice.

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Funding

U.S. Department of Health & Human Services | NIH | National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIAMS)(P30AR069619)