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 wor...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(OcnCre + Rspo2 ^(f/f)). Ocn-Cre + Rspo2 f/fmale and female mice at 1, 3, and 6 months were examined. Ocn-Cre + Rspo2 f/fmice are decreased in overall body size compared to their control littermates and have decreased bone mass. Histomorphometric analysis of1-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/fwas reduced, and bone marrow-mesenchymal stem cells from Ocn-Cre + Rspo2 f/fmice 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.展开更多
基金supported by the National Institutes of Health (NIH) National Institute of Arthritis and Musculoskeletal Diseases (NIAMS)funding of the University of Pennsylvania Center for Musculoskeletal Disorders (PCMD) (P30AR069619)the University of Michigan Integrative Musculoskeletal Health Core Center (P30AR069620)+1 种基金supported by NIH NIAMS R01AR066028supported by NIH NIAMS F31AR065858
文摘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(OcnCre + Rspo2 ^(f/f)). Ocn-Cre + Rspo2 f/fmale and female mice at 1, 3, and 6 months were examined. Ocn-Cre + Rspo2 f/fmice are decreased in overall body size compared to their control littermates and have decreased bone mass. Histomorphometric analysis of1-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/fwas reduced, and bone marrow-mesenchymal stem cells from Ocn-Cre + Rspo2 f/fmice 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.
