CYP24 Promoter Activity is Affected by Mechanical Stress and Mitogen-Activated Protein Kinase in MG63 Osteoblast-like Cells

Abstract

Bone grafts and improvement in bone quality will increase in the future. Thus, molecular biological knowledge of bone metabolism is essential for prosthetic therapy. This study reveals that expression of CYP24, which inactivates vitamin D, is controlled by mechanical stress and mitogen-activated protein kinase. AbstractPurpose: For bone homeostasis, vitamin D plays an important role in the regulation of calcium. The enzyme CYP24 inactivates vitamin D and is involved in its regulation. However, the mechanism of expression of CYP24 in osteoblastic cells under mechanical stress is not clear. In this study we investigated CYP24 promoter activity in stretched osteoblastic cells and the participation of mitogen-activated protein kinase (MAPK) in expression of CYP24.Methods: MG63 osteoblastic cells were cultured on silicon-bottomed plates. Cells were transfected with a reporter gene that contained a CYP24 promoter. After activated vitamin D, 1,25(OH) 2 D 3 , was added or not added, cells were stretched. Stretched and non-stretched cells were investigated by luciferase dual assay. Cells were also investigated similarly using medium with an ERK1/2 inhibitor or p38 inhibitor. Results:The CYP24 promoter was activated by 1,25(OH) 2 D 3 and the promoter activity decreased in stretched cells. Inhibitor of MAPK decreased CYP24 promoter activity. However, CYP24 promoter activity decreased with mechanical stress after addition of p38 inhibitor, while it did not decrease with mechanical stress after addition of ERK1/2 inhibitor. The CYP24 promoter was not activated without 1,25(OH) 2 D 3 in any case. Conclusion:Mechanical stress and MAPK control CYP24 promoter activity in the presence of Vitamin D in MG63 osteoblast-like cells.

Impact of mechanical stretch on the cell behaviors of bone and surrounding tissues

“…In osteoblasts, this universally leads to stretch-induced activation of mitogen-activated protein kinase (MAPK) pathways and in particular ERK, a key MAPK effector molecule that activates mechanoresponsive transcription factors. 7 , 8 , 10 , 13 , 15 , 18 , 24 , 26 , 164 At the focal adhesion site formed upon mechanical activation of integrins, FAK also undergoes enhanced and sustained association with another tyrosine kinase, proline-rich tyrosine kinase 2 (PYK2), 10 , 16 which seems to stablize FAK and remove the inhibitory Hic5 adaptor protein from the focal adhesion complex upon activation. 16 Collectively, these data confirm that integrins and their associated intracellular partners are necessary for signal transduction downstream of the strain stimulus to be executed.…”