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Iron excess upregulates SPNS2 mRNA levels but reduces sphingosine-1-phosphate export in human osteoblastic MG-63 cells

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Abstract

Summary

We aimed to study the mechanisms involved in bone-related iron impairment by using the osteoblast-like MG-63 cell line. Our results indicate that iron impact the S1P/S1PR signalizing axis and suggest that iron can affect the S1P process and favor the occurrence of osteoporosis during chronic iron overload.

Introduction

Systemic iron excess favors the development of osteoporosis, especially during genetic hemochromatosis. The cellular mechanisms involved are still unclear despite numerous data supporting a direct effect of iron on bone biology. Therefore, the aim of this study was to characterize mechanisms involved in the iron-related osteoblast impairment.

Methods

We studied, by using the MG-63 cell lines, the effect of iron excess on SPNS2 gene expression which was previously identified by us as potentially iron-regulated. Cell-type specificity was investigated with hepatoma HepG2 and enterocyte-like Caco-2 cell lines as well as in iron-overloaded mouse liver. The SPNS2-associated function was also investigated in MG-63 cells by fluxomic strategy which led us to determinate the S1P efflux in iron excess condition.

Results

We showed in MG-63 cells that iron exposure strongly increased the mRNA level of the SPNS2 gene. This was not observed in HepG2, in Caco-2 cells, and in mouse livers. Fluxomic study performed concomitantly on MG-63 cells revealed an unexpected decrease in the cellular capacity to export S1P. Iron excess did not modulate SPHK1, SPHK2, SGPL1, or SGPP1 gene expression, but decreased COL1A1 and S1PR1 mRNA levels, suggesting a functional implication of low extracellular S1P concentration on the S1P/S1PR signalizing axis.

Conclusions

Our results indicate that iron impacts the S1P/S1PR signalizing axis in the MG-63 cell line and suggest that iron can affect the bone-associated S1P pathway and favor the occurrence of osteoporosis during chronic iron overload.

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Abbreviations

C16d3Carn:

Deuterated C16d3 palmitoyl–carnitine

DFO:

Desferrioxamine

FAC:

Ferric ammonium citrate

FCS:

Fetal calf serum

GH:

Genetic hemochromatosis

IREs:

Iron responsive elements

MEM:

Minimum essential medium

S1P:

Sphingosine-1-phosphate

S1Pd7:

Deuterated sphingosine-1-phosphate d7

S1PR:

Sphingosine-1-phosphate receptor

Sphd7:

Deuterated sphingosine d7

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Acknowledgements

The authors would like to thank Dr. François Gaboriau and Dr. Lenaick Detivaud for helpful advice and assistance, Claudine Rauch and Brigitte Frenot for technical assistance with cell culture and mass spectrometry, Pr. Véronique David and Dr. Nathalie Le Floc’h for helpful discussions, and Dr. Caroline Moreau for global assistance and support in this project.

Funding

This work was supported by the French Society of Rheumatology (SFR) [grant numbers 2320, 2013].

Author information

Author notes

  1. O. Loréal and P. Guggenbuhl contributed equally to this work.

Authors and Affiliations

  1. Service de Biochimie – Toxicologie, CHU Rennes, F-35033, Rennes, France

    L. Peltier, C. Bendavid, T. Cavey & M. Ropert

  2. INSERM, INRA, Univ Rennes1, Univ Bretagne Loire, Nutrition, Metabolism, and Cancer, Rennes, France

    L. Peltier, C. Bendavid, T. Cavey, M.-L. Island, M. Doyard, P. Leroyer, C. Allain, M. Ropert, O. Loréal & P. Guggenbuhl

  3. Faculté de Médecine, Université Rennes 1, F-35043, Rennes, France

    L. Peltier, C. Bendavid, T. Cavey, M. De Tayrac & P. Guggenbuhl

  4. CNRS, UMR 6290, Institut de Génétique et Développement de Rennes (IGdR), F-35043, Rennes, France

    M. De Tayrac

  5. Service de Génétique Moléculaire et Génomique, CHU Rennes, F-35033, Rennes, France

    M. De Tayrac

  6. Service de Rhumatologie, CHU Rennes, F-35203, Rennes, France

    P. Guggenbuhl

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  1. L. Peltier
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Correspondence to P. Guggenbuhl.

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Lucas Peltier, Claude Bendavid, Thibault Cavey, Marie-Laure Island, Mathilde Doyard, Patricia Leroyer, Coralie Allain, Marie De Tayrac, Martine Ropert, Olivier Loréal and Pascal Guggenbuhl declare that they have no conflict of interest.

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Peltier, L., Bendavid, C., Cavey, T. et al. Iron excess upregulates SPNS2 mRNA levels but reduces sphingosine-1-phosphate export in human osteoblastic MG-63 cells. Osteoporos Int 29, 1905–1915 (2018). https://doi.org/10.1007/s00198-018-4531-8

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