RANKL Expression Is Related to the Differentiation State of Human Osteoblasts

Authors

  • Gerald J Atkins PhD,

    Corresponding author
    1. Department of Orthopaedics and Trauma, University of Adelaide, Adelaide, South Australia, Australia
    • Department of Orthopaedics and Trauma, Bone Cell Biology Laboratory, University of Adelaide, North Terrace, Adelaide, South Australia 5000, Australia
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  • Panagiota Kostakis,

    1. Department of Orthopaedics and Trauma, University of Adelaide, Adelaide, South Australia, Australia
    2. Division of Haematology, Institute of Medical and Veterinary Science, Adelaide, South Australia, Australia
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  • Beiqing Pan,

    1. Division of Haematology, Institute of Medical and Veterinary Science, Adelaide, South Australia, Australia
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  • Amanda Farrugia,

    1. Division of Haematology, Institute of Medical and Veterinary Science, Adelaide, South Australia, Australia
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  • Stan Gronthos,

    1. Division of Haematology, Institute of Medical and Veterinary Science, Adelaide, South Australia, Australia
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  • Andreas Evdokiou,

    1. Department of Orthopaedics and Trauma, University of Adelaide, Adelaide, South Australia, Australia
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  • Kate Harrison,

    1. Division of Haematology, Institute of Medical and Veterinary Science, Adelaide, South Australia, Australia
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  • David M Findlay,

    1. Department of Orthopaedics and Trauma, University of Adelaide, Adelaide, South Australia, Australia
    2. These authors share senior author status on this manuscript
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    • Dr Findlay serves as a consultant for the Australian Orthopaedic Innovations Medical Advisory Board. All other authors have no conflict of interest

  • Andrew Cw Zannettino

    1. Division of Haematology, Institute of Medical and Veterinary Science, Adelaide, South Australia, Australia
    2. These authors share senior author status on this manuscript
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Abstract

Human osteoblast phenotypes that support osteoclast differentiation and bone formation are not well characterized. Osteoblast differentiation markers were examined in relation to RANKL expression. RANKL expression was induced preferentially in immature cells. These results support an important link between diverse osteoblast functions.

Cells of the osteoblast lineage support two apparently distinct functions: bone formation and promotion of osteoclast formation. The aim of this study was to examine the relationship between these phenotypes in human osteoblasts (NHBC), in terms of the pre-osteoblast marker, STRO-1, and the mature osteoblast marker, alkaline phosphatase (AP), and the expression of genes involved in osteoclast formation, RANKL and OPG. The osteotropic stimuli, 1α,25(OH)2vitamin D3 (vitD3) and dexamethasone, were found to have profound proliferative and phenotypic effects on NHBCs. VitD3 inhibited NHBC proliferation and increased the percentage of cells expressing STRO-1 over an extended culture period, implying that vitD3 promotes and maintains an immature osteogenic phenotype. Concomitantly, RANKL mRNA expression was upregulated and maintained in NHBC in response to vitD3. Dexamethasone progressively promoted the proliferation of AP-expressing cells, resulting in the overall maturation of the cultures. Dexamethasone had little effect on RANKL mRNA expression and downregulated OPG mRNA expression in a donor-dependent manner. Regression analysis showed that RANKL mRNA expression was associated negatively with the percentage of cells expressing AP (p < 0.01) in vitD3- and dexamethasone-treated NHBCs. In contrast, RANKL mRNA expression was associated positively with the percentage of STRO-1+ cells (p < 0.01). In NHBCs sorted by FACS based on STRO-1 expression (STRO-1bright and STRO-1dim populations), it was found that vitD3 upregulated the expression of RANKL mRNA preferentially in STRO-1bright cells. The results suggest that immature osteoblasts respond to osteotropic factors in a potentially pro-osteoclastogenic manner. Additionally, the dual roles of osteoblasts, in supporting osteoclastogenesis or forming bone, may be performed by the same lineage of cells at different stages of their maturation.

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