Elevation of Extracellular Ca2+ Induces Store-Operated Calcium Entry via Calcium-Sensing Receptors: A Pathway Contributes to the Proliferation of Osteoblasts |
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Authors: | Fen Hu Leiting Pan Kai Zhang Fulin Xing Xinyu Wang Imshik Lee Xinzheng Zhang Jingjun Xu |
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Institution: | The Key Laboratory of Weak-Light Nonlinear Photonics, Ministry of Education, TEDA Applied Physics Institute and School of Physics, Nankai University, Tianjin, China.; Faculdade de Medicina Dentária, Universidade do Porto, Portugal, |
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Abstract: | AimsThe local concentration of extracellular Ca2+ (Ca2+]o) in bone microenvironment is accumulated during bone remodeling. In the present study we investigated whether elevating Ca2+]o induced store-operated calcium entry (SOCE) in primary rat calvarial osteoblasts and further examined the contribution of elevating Ca2+]o to osteoblastic proliferation.MethodsCytosolic Ca2+ concentration (Ca2+]c) of primary cultured rat osteoblasts was detected by fluorescence imaging using calcium-sensitive probe fura-2/AM. Osteoblastic proliferation was estimated by cell counting, MTS assay and ATP assay. Agonists and antagonists of calcium-sensing receptors (CaSR) as well as inhibitors of phospholipase C (PLC), SOCE and voltage-gated calcium (Cav) channels were applied to study the mechanism in detail.ResultsOur data showed that elevating Ca2+]o evoked a sustained increase of Ca2+]c in a dose-dependent manner. This Ca2+]c increase was blocked by TMB-8 (Ca2+ release inhibitor), 2-APB and BTP-2 (both SOCE blockers), respectively, whereas not affected by Cav channels blockers nifedipine and verapamil. Furthermore, NPS2143 (a CaSR antagonist) or {"type":"entrez-nucleotide","attrs":{"text":"U73122","term_id":"4098075","term_text":"U73122"}}U73122 (a PLC inhibitor) strongly reduced the Ca2+]o-induced Ca2+]c increase. The similar responses were observed when cells were stimulated with CaSR agonist spermine. These data indicated that elevating Ca2+]o resulted in SOCE depending on the activation of CaSR and PLC in osteoblasts. In addition, high Ca2+]o significantly promoted osteoblastic proliferation, which was notably reversed by BAPTA-AM (an intracellular calcium chelator), 2-APB, BTP-2, TMB-8, NPS2143 and {"type":"entrez-nucleotide","attrs":{"text":"U73122","term_id":"4098075","term_text":"U73122"}}U73122, respectively, but not affected by Cav channels antagonists.ConclusionsElevating Ca2+]o induced SOCE by triggering the activation of CaSR and PLC. This process was involved in osteoblastic proliferation induced by high level of extracellular Ca2+ concentration. |
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