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Horm Metab Res 2014; 46(08): 537-545
DOI: 10.1055/s-0033-1363265
Endocrine Research
© Georg Thieme Verlag KG Stuttgart · New York

Estrogen Withdrawal from Osteoblasts and Osteocytes Causes Increased Mineralization and Apoptosis

M. Á. Brennan
1   Biomechanics Research Centre (BMEC), Department of Mechanical and Biomedical Engineering, National University of Ireland, Galway, Ireland
2   National Centre for Biomedical Engineering Sciences, NCBES, National University of Ireland, Galway, Ireland
3   Bioengineering Sciences Research Group, Faculty of Engineering and the Environment, University of Southampton, UK
,
M. G. Haugh
1   Biomechanics Research Centre (BMEC), Department of Mechanical and Biomedical Engineering, National University of Ireland, Galway, Ireland
2   National Centre for Biomedical Engineering Sciences, NCBES, National University of Ireland, Galway, Ireland
,
F. J. O’Brien
4   Department of Anatomy, Royal College of Surgeons in Ireland, 123 St. Stephens Green, Dublin 2, Ireland
5   Trinity Centre for Bioengineering, Trinity College, Dublin 2, Ireland
,
L. M. McNamara
1   Biomechanics Research Centre (BMEC), Department of Mechanical and Biomedical Engineering, National University of Ireland, Galway, Ireland
2   National Centre for Biomedical Engineering Sciences, NCBES, National University of Ireland, Galway, Ireland
› Author Affiliations
Further Information

Publication History

received 28 August 2013

accepted 28 November 2013

Publication Date:
20 January 2014 (online)

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Abstract

Recent studies have demonstrated increased bone mineral heterogeneity following estrogen withdrawal in vivo. Such changes likely contribute to fracture risk during post-menopausal osteoporosis since tissue mineralization is correlated with bone strength and stiffness. However, the cellular mechanisms responsible for increased mineral variability have not yet been distinguished. The objective of this study is to elucidate how alterations in mineral distribution are initiated during estrogen depletion. Specifically, we tested two separate hypotheses; (1) estrogen deficiency directly alters osteoblast mineralization and (2) estrogen deficiency increases bone cell apoptosis. Osteoblast-like cells (MC3T3-E1) and osteocyte-like cells (MLO-Y4) were pretreated with or without estrogen (17β-estradiol) for 14 days. Estrogen deficiency was subsequently induced by either withdrawing estrogen from cells or blocking estrogen receptors using an estrogen antagonist, fulvestrant (ICI 182,780). Cell number (Hoechst DNA), alkaline phosphatase activity (p-NPP), mineralization (alizarin red) and apoptosis (Caspase 3/7) were evaluated. Whether estrogen withdrawal altered apoptosis rates in the presence of an apoptosis promoting agent (etoposide) was also determined. Interestingly, estrogen withdrawal from cells accustomed to estrogen exposure caused significantly increased osteoblast mineralization and osteocyte apoptosis compared with continued estrogen treatment. In contrast, blocking estrogen receptors with fulvestrant abrogated the mineralization induced by estrogen treatment. When apoptosis was induced using etoposide, cells undergoing estrogen withdrawal increased apoptosis compared to cells with continued estrogen treatment. Recognizing the underlying mechanisms regulating bone cell mineralization and apoptosis during estrogen deficiency and their consequences is necessary to further our knowledge of osteoporosis.

Key words

estrogen deficiency - osteoblasts - osteocytes - mineralization - apoptosis
 

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