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Osteocyte-induced angiogenesis via VEGF–MAPK-dependent pathways in endothelial cells

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Abstract

Recently, it has been suggested osteocytes control the activities of bone formation (osteoblasts) and resorption (osteoclast), indicating their important regulatory role in bone remodelling. However, to date, the role of osteocytes in controlling bone vascularisation remains unknown. Our aim was to investigate the interaction between endothelial cells and osteocytes and to explore the possible molecular mechanisms during angiogenesis. To model osteocyte/endothelial cell interactions, we co-cultured osteocyte cell line (MLOY4) with endothelial cell line (HUVECs). Co-cultures were performed in 1:1 mixture of osteocytes and endothelial cells or by using the conditioned media (CM) transfer method. Real-time cell migration of HUVECs was measured with the transwell migration assay and xCELLigence system. Expression levels of angiogenesis-related genes were measured by quantitative real-time polymerase chain reaction (qRT-PCR). The effect of vascular endothelial growth factor (VEGF) and mitogen-activated phosphorylated kinase (MAPK) signaling were monitored by western blotting using relevant antibodies and inhibitors. During the bone formation, it was noted that osteocyte dendritic processes were closely connected to the blood vessels. The CM generated from MLOY4 cells-activated proliferation, migration, tube-like structure formation, and upregulation of angiogenic genes in endothelial cells suggesting that secretory factor(s) from osteocytes could be responsible for angiogenesis. Furthermore, we identified that VEGF secreted from MLOY4-activated VEGFR2–MAPK–ERK-signaling pathways in HUVECs. Inhibiting VEGF and/or MAPK–ERK pathways abrogated osteocyte-mediated angiogenesis in HUVEC cells. Our data suggest an important role of osteocytes in regulating angiogenesis.

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Acknowledgments

The authors would like to thank staff at Medical Engineering Research Facility (MERF) for assistance in the care of in vivo animals.

Conflict of interest

Authors declare there is no conflict of interest associated with this project.

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Authors and Affiliations

  1. Institute of Health and Biomedical Innovation, Queensland University of Technology, Kelvin Grove Campus, Brisbane, QLD, 4059, Australia

    Indira Prasadam, Yinghong Zhou, Zhibin Du, Jiezhong Chen, Ross Crawford & Yin Xiao

  2. Prince Charles Hospital, Brisbane, QLD, Australia

    Ross Crawford

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  1. Indira Prasadam
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  2. Yinghong Zhou
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  3. Zhibin Du
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  4. Jiezhong Chen
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Correspondence to Indira Prasadam.

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Prasadam, I., Zhou, Y., Du, Z. et al. Osteocyte-induced angiogenesis via VEGF–MAPK-dependent pathways in endothelial cells. Mol Cell Biochem 386, 15–25 (2014). https://doi.org/10.1007/s11010-013-1840-2

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