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Biochemistry (Moscow)

, Volume 84, Issue 6, pp 686–692 | Cite as

Resveratrol Promotes in vitro Differentiation of Osteoblastic MC3T3-E1 Cells via Potentiation of the Calcineurin/NFATc1 Signaling Pathway

  • Y. Huang
  • J. Huo
  • F. Q. Liu
  • J. Liu
  • X. J. Zhang
  • C. H. Guo
  • L. H. SongEmail author
Article
  • 1 Downloads

Abstract

Resveratrol has been shown to stimulate differentiation of osteoblastic MC3T3-E1 cells in vitro; however, the mechanisms underlying the anabolic effect of resveratrol on osteoblasts remain largely unknown. Our study was aimed to investigate the molecular mechanism of resveratrol-induced differentiation of MC3T3-E1 cells. MC3T3-E1 cells were treated for 8 days with different concentrations of resveratrol (10−8-10−6 M) and 10−6 M cyclosporine A (CsA), a specific inhibitor of the calcineurin/NFAT pathway. According to the results of pilot studies of cell proliferation and alkaline phos-phatase activity, 10−7 M concentration of resveratrol was used in subsequent experiments. The levels of mRNA expression of the osteosis-related genes CaN, NFATc1, and Runx2 were analyzed by real-time RT-PCR; the levels of the corresponding proteins were estimated by Western blot analysis. Resveratrol upregulated expression of the CaN, NFATc1, and Runx2 genes at both mRNA and protein levels compared to the control group (p < 0.05), while CsA reduced the effects of resveratrol (p < 0.05). Using immunohistochemical staining, we showed that resveratrol induced NFATc1 accumulation in the cell nuclei, and treatment with CsA inhibited resveratrol-mediated induction of NFATc1, suggesting that the calcineurin/NFATc1 signaling pathway plays an important role in the regulatory effect of resveratrol on osteoblasts.

Keywords

resveratrol cyclosporine A calcineurin/NFATc1 osteoblast differentiation 

Abbreviations

ALP

alkaline phosphatase

a-MEM

alpha minimum essential medium

CaN

calcineurin

CsA

cyclosporine A

IHC

immunohistochemistry

MTT

3-(4,5-dimethyl-2-thi-azolyl)-2,5-diphenyl-2-H-tetrazolium bromide

NFATc1

nuclear factor of activated T cells, cytoplasmic 1

NF-κB

nuclear factor kappa B

pNPP

p-nitrophenyl phosphate

PVDF

polyvinylidene fluoride

RIPA

radioimmunoprecipitation assay

Runx2

Runt-related transcription factor 2

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Notes

Acknowledgements

The authors thank Professor Song for her concentrated guidance during this study.

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Copyright information

© Pleiades Publishing, Ltd. 2019

Authors and Affiliations

  • Y. Huang
    • 1
  • J. Huo
    • 2
  • F. Q. Liu
    • 3
  • J. Liu
    • 3
  • X. J. Zhang
    • 4
  • C. H. Guo
    • 4
  • L. H. Song
    • 4
    Email author
  1. 1.Department of BiochemistryChangzhi Medical CollegeChangzhi, ShanxiChina
  2. 2.Department of BiologyChangzhi Medical CollegeChangzhi, ShanxiChina
  3. 3.Changzhi Medical CollegeChangzhi, ShanxiChina
  4. 4.Department of PharmacologyChangzhi Medical CollegeChangzhi, ShanxiChina

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