Analysis of Ca2+ response of osteocyte network by three-dimensional time-lapse imaging in living bone


Osteocytes form a three-dimensional (3D) cellular network within the mineralized bone matrix. The cellular network has important roles in mechanosensation and mechanotransduction related to bone homeostasis. We visualized the embedded osteocyte network in chick calvariae and observed the flow-induced Ca2+ signaling in osteocytes using 3D time-lapse imaging. In response to the flow, intracellular Ca2+ ([Ca2+]i) significantly increased in developmentally mature osteocytes in comparison with young osteocytes in the bone matrix. To investigate the differences in response between young and developmentally mature osteocytes in detail, we evaluated the expression of osteocyte-related genes using the osteocyte-like cell line MLO-Y4, which was 3D-cultured within type I collagen gels. We found that the c-Fos, Cx43, Panx3, Col1a1, and OCN mRNA levels significantly increased on day 15 in comparison with day 7. These findings indicate that developmentally mature osteocytes are more responsive to mechanical stress than young osteocytes and have important functions in bone formation and remodeling.

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This work was performed in part under the Cooperative Research Program of Institute for Frontier Medical Sciences, Kyoto University, Japan, and supported by a Grant-in-Aid for Scientific Research (No. JP 16H05549) from the Ministry of Education, Culture, Sports, Science and Technology (MEXT), Japan.

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Correspondence to Hiroshi Kamioka.

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Tanaka, T., Hoshijima, M., Sunaga, J. et al. Analysis of Ca2+ response of osteocyte network by three-dimensional time-lapse imaging in living bone. J Bone Miner Metab 36, 519–528 (2018).

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  • Osteocyte
  • MLO-Y4
  • Fluid shear stress
  • 3D time-lapse imaging