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Endostatin Affects Osteoblast Behavior In Vitro, but Collagen XVIII/Endostatin Is Not Essential for Skeletal Development In Vivo

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Abstract

Endostatin, a fragment of collagen XVIII, can inhibit vascular endothelial growth factor (VEGF) signaling. VEGF is known to be crucial for bone development. The aims of this study were to investigate the influences of endostatin on osteoblast behavior in vitro and the roles of collagen XVIII/endostatin on bone development in vivo. For the in vitro experiments, MC3T3-E1 osteoblasts were treated with VEGF-A, 2 μg/ml endostatin, 20 μg/ml endostatin, VEGF-A + 2 μg/ml endostatin, or VEGF-A + 20 μg/ml endostatin. Osteoblast proliferation and matrix mineralization were analyzed. Faxitron, pQCT, and histological analyses were performed on hindleg bones of transgenic mice overexpressing endostatin (ES-tg) and mice lacking collagen XVIII (Col18a1 −/−) to study bone development in vivo. Treatment of cells with endostatin decreased osteoblast proliferation. Moreover, VEGF-A together with endostatin (2 μg/ml) decreased osteoblast proliferation and matrix mineralization. In vivo, Col18a1 −/− and ES-tg mice displayed no differences in bone density or mineral content during bone development, but ES-tg bones grew in length more slowly compared to the controls. The formation of secondary ossification centers was delayed in Col18a1 −/− mice. Immunohistochemistry revealed collagen XVIII in basement membranes of periosteal and bone marrow vessels and at muscle attachment sites. In conclusion, endostatin affects osteoblast behavior in vitro, the effects being boosted by simultaneous treatment with VEGF. In vivo, Col18a1 −/− and ES-tg mice show mild delays in bone development. These changes are transitory and suggest that collagen XVIII/endostatin does not play an indispensable role in skeletal development.

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Acknowledgments

We thank Anna-Maija Ruonala and Jaana Peters for their excellent technical assistance. This work was supported by grants from the Research Council for Health of the Academy of Finland (nos. 115237 and 130795), Sigrid Jusélius Foundation, Emil Aaltonen Foundation, and Graduate School for Musculoskeletal Disorders and Biomaterials, TBGS.

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Authors and Affiliations

  1. Department of Anatomy and Cell Biology, Institute of Biomedicine, University of Oulu, Oulu, Finland

    Annina Sipola & Juha Tuukkanen

  2. Oulu Center for Cell–Matrix Research, Biocenter Oulu, University of Oulu, Oulu, Finland

    Lotta Seppinen & Taina Pihlajaniemi

  3. Department of Medical Biochemistry and Molecular Biology, Institute of Biomedicine, University of Oulu, Oulu, Finland

    Lotta Seppinen & Taina Pihlajaniemi

  4. Department of Anatomy and Cell Biology, Institute of Biomedicine, University of Oulu, P.O. Box 5000, 90014, Oulu, Finland

    Juha Tuukkanen

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  1. Annina Sipola
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Correspondence to Juha Tuukkanen.

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Sipola, A., Seppinen, L., Pihlajaniemi, T. et al. Endostatin Affects Osteoblast Behavior In Vitro, but Collagen XVIII/Endostatin Is Not Essential for Skeletal Development In Vivo. Calcif Tissue Int 85, 412–420 (2009). https://doi.org/10.1007/s00223-009-9287-x

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  • DOI: https://doi.org/10.1007/s00223-009-9287-x

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