The Orphan Receptor Tyrosine Kinase Ror2 Promotes Osteoblast Differentiation and Enhances ex Vivo Bone Formation

Abstract

Ror2 is a receptor tyrosine kinase, the expression of which increases during differentiation of pluripotent stem cells to osteoblasts and then declines as cells progress to osteocytes. To test whether Ror2 plays a role in osteoblastogenesis, we investigated the effects of Ror2 overexpression and down-regulation on osteoblastic lineage commitment and differentiation. Expression of Ror2 in pluripotent human mesenchymal stem cells (hMSCs) by adenoviral infection caused formation of mineralized extracellular matrix, which is the ultimate phenotype of an osteogenic tissue. Concomitantly, Ror2 over-expression inhibited adipogenic differentiation of hMSCs as monitored by lipid formation. Ror2 shifted hMSC fate toward osteoblastogenesis by inducing osteogenic transcription factor osterix and suppressing adipogenic transcription factors CCAAT/enhancer-binding protein alpha and peroxisome proliferator activated receptor gamma. Infection with Ror2 virus also strongly promoted matrix mineralization in committed osteoblast-like MC3T3-E1 cells. Expression of Ror2 in a human preosteocytic cell line by stable transfection also promoted further differentiation, as judged by inhibited alkaline phosphatase activity, potentiated osteocalcin secretion, and increased cellular apoptosis. In contrast, down-regulation of Ror2 expression by short hairpin RNA essentially abrogated dexamethasone-induced mineralization of hMSCs. Furthermore, down-regulation of Ror2 expression in fully differentiated SaOS-2 osteosarcoma cells inhibited alkaline phosphatase activity. We conclude that Ror2 initiates commitment of MSCs to osteoblastic lineage and promotes differentiation at early and late stages of osteoblastogenesis. Finally, using a mouse calvariae ex vivo organ culture model, we demonstrate that these effects of Ror2 result in increased bone formation, suggesting that it may also activate mature osteoblasts.

The mutationROR2W749X, linked to human BDB, is a recessive mutation in the mouse, causing brachydactyly, mediating patterning of joints and modeling recessive Robinow syndrome

“…proposed that ROR2 promotes both osteoblast differentiation and the commitment of mesenchymal stem cells to the osteoblastic lineage (Liu et al, 2006). Although at 3 weeks of age Ror2 W749FLAG/W749FLAG mice exhibit a reduction in trabecular bone volume, this phenotype is consistent with the delay in embryonic bone development and resolves later in life.…”

“…W749FLAG mutation does not affect bone remodeling in adult mice ROR2 has been reported to have signaling activity in osteoblastic cells and to promote osteoblastic activity in vitro (Billiard et al, 2005;Liu et al, 2006), whereas Ror2 knockout embryos exhibit delayed ossification of endochondral bones . We were therefore interested in determining whether ROR2-W749FLAG has an effect in the regulation of bone formation.…”

Estrogen-related receptor α regulates osteoblast differentiation via Wnt/β-catenin signaling

“…Neonatal mouse calvaria were prepared from 4-day-old pups as described previously (Traianedes et al 1998, Mundy et al 1999) with slight modifications (Liu et al 2007a). Briefly, calvaria were excised and cut into half along the sagittal suture.…”

Knockdown of receptor tyrosine kinase-like orphan receptor 2 inhibits cell proliferation and colony formation in osteosarcoma cells by inducing arrest in cell cycle progression

“…ROR2 is expressed in heart, brain and lung tissue, and is also involved in the development of the nervous system and bones (9)(10)(11). It has been hypothesized that ROR2 is involved in the early formation of chondrocytes as well as the development of cartilage and growth plates (12).…”