Osteoporosis International

, Volume 23, Issue 10, pp 2435–2445 | Cite as

Iron excess limits HHIPL-2 gene expression and decreases osteoblastic activity in human MG-63 cells

  • M. Doyard
  • N. Fatih
  • A. Monnier
  • M. L. Island
  • M. Aubry
  • P. Leroyer
  • R. Bouvet
  • G. Chalès
  • J. Mosser
  • O. Loréal
  • P. Guggenbuhl
Original Article



In order to understand mechanisms involved in osteoporosis observed during iron overload diseases, we analyzed the impact of iron on a human osteoblast-like cell line. Iron exposure decreases osteoblast phenotype. HHIPL-2 is an iron-modulated gene which could contribute to these alterations. Our results suggest osteoblast impairment in iron-related osteoporosis.


Iron overload may cause osteoporosis. An iron-related decrease in osteoblast activity has been suggested.


We investigated the effect of iron exposure on human osteoblast cells (MG-63) by analyzing the impact of ferric ammonium citrate (FAC) and iron citrate (FeCi) on the expression of genes involved in iron metabolism or associated with osteoblast phenotype. A transcriptomic analysis was performed to identify iron-modulated genes.


FAC and FeCi exposure modulated cellular iron status with a decrease in TFRC mRNA level and an increase in intracellular ferritin level. FAC increased ROS level and caspase 3 activity. Ferroportin, HFE and TFR2 mRNAs were expressed in MG-63 cells under basal conditions. The level of ferroportin mRNA was increased by iron, whereas HFE mRNA level was decreased. The level of mRNA alpha 1 collagen type I chain, osteocalcin and the transcriptional factor RUNX2 were decreased by iron. Transcriptomic analysis revealed that the mRNA level of HedgeHog Interacting Protein Like-2 (HHIPL-2) gene, encoding an inhibitor of the hedgehog signaling pathway, was decreased in the presence of FAC. Specific inhibition of HHIPL-2 expression decreased osteoblast marker mRNA levels. Purmorphamine, hedgehog pathway activator, increased the mRNA level of GLI1, a target gene for the hedgehog pathway, and decreased osteoblast marker levels. GLI1 mRNA level was increased under iron exposure.


We showed that in human MG-63 cells, iron exposure impacts iron metabolism and osteoblast gene expression. HHIPL-2 gene expression modulation may contribute to these alterations. Our results support a role of osteoblast impairment in iron-related osteoporosis.


Gene expression Hemochromatosis HHIPL-2 Iron overload Osteoblast Osteoporosis 

Supplementary material

198_2011_1871_MOESM1_ESM.pdf (18 kb)
Fig. S1Iron metabolism gene expression under basal situation in MG-63 cells. SLC40A1 (a), HFE (b) and TFR2 (c) mRNA levels under basal culture conditions. The results are expressed as a percentage of expression in caco-2 cells (PDF 220 kb)
198_2011_1871_MOESM2_ESM.pdf (404 kb)
Fig. S2Impact of FeCi exposure on the expression of osteoblast genes in MG-63 cells. mRNA levels of COL1A1 (a), BGLAP (b) and RUNX2 (c) in cells treated with FeCi and/or DFO for 72 h. The results are expressed as a percentage of their respective control (100%). Asterisk indicates p < 0.05 compared with the corresponding concentration of citrate; plus sign indicates p < 0.05 compared with FeCi 20 μM (PDF 454 kb)
198_2011_1871_MOESM3_ESM.pdf (225 kb)
Fig. S3Inhibition of mRNA HHIPL-2 expression by specific siRNAs. HHIPL-2 mRNA level after transfection of MG-63 cells with two HHIPL-2-specific siRNAs (si1 and si2) or with a control siRNA (Control) for 72 h. The results are expressed as a percentage of the control (100%). Asterisk indicates p < 0.05 compared with the control (PDF 224 kb)
198_2011_1871_MOESM4_ESM.pdf (66 kb)
Fig. S4The impact of iron exposure on GLI1 mRNA expression level in MG-63 cells. Expression of GLI1 mRNA after treatment with FAC and/or DFO for 72 h. The results are expressed as a percentage of the control (100%). Asterisk indicates p < 0.05 compared with the control (PDF 66.0 kb)


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Copyright information

© International Osteoporosis Foundation and National Osteoporosis Foundation 2012

Authors and Affiliations

  • M. Doyard
    • 1
    • 2
  • N. Fatih
    • 1
    • 2
  • A. Monnier
    • 3
  • M. L. Island
    • 1
    • 2
  • M. Aubry
    • 4
  • P. Leroyer
    • 1
    • 2
  • R. Bouvet
    • 4
    • 5
  • G. Chalès
    • 1
    • 2
    • 6
  • J. Mosser
    • 3
    • 4
    • 5
  • O. Loréal
    • 1
    • 2
    • 7
  • P. Guggenbuhl
    • 1
    • 2
    • 6
    • 8
  1. 1.INSERM, UMRU991Rennes CedexFrance
  2. 2.Université de Rennes 1RennesFrance
  3. 3.CNRS UMR6061, Institut de Génétique et Développement, Université de RennesRennesFrance
  4. 4.Plateforme Génomique Santé Biogenouest®RennesFrance
  5. 5.Service de Génétique Moléculaire et GénomiqueRennesFrance
  6. 6.Service de Rhumatologie, Hôpital SudRennesFrance
  7. 7.Service des Maladies du Foie, Hôpital PontchaillouRennesFrance
  8. 8.Service de Rhumatologie, Hôpital SudRennesFrance

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