Estrogen deficiency impairs integrin \u03b1v\u03b23-mediated mechanosensation by osteocytes and alters osteoclastogenic paracrine signalling

Ivor P Geoghegan,Laoise M Mcnamara,David A Hoey

doi:10.1038/s41598-019-41095-3

Ivor P Geoghegan, Laoise M Mcnamara + Show 1 more

Open Access

https://doi.org/10.1038/s41598-019-41095-3

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Abstract

The integrin αvβ3 has been shown to play an important role in osteocyte mechanotransduction. It has been reported that there are fewer β3 integrin-containing cells in osteoporotic bone cells. Osteocytes cultured in vitro under estrogen deficient conditions demonstrate altered mechanotransduction. However, it is unknown whether the altered mechanotransduction in estrogen deficient osteocytes is directly associated with defective αvβ3 expression or signalling. The objective of this study is to investigate the role of estrogen deficiency for regulating MLO-Y4 cell morphology, αvβ3 expression, focal adhesion formation and mechanotransduction by osteocytes. Here, we report that estrogen withdrawal leads to a smaller focal adhesion area and reduced αvβ3 localisation at focal adhesion sites, resulting in an increased Rankl/Opg ratio and defective Cox-2 responses to oscillatory fluid flow. Interestingly, αvβ3 antagonism had a similar effect on focal adhesion assembly, Rankl/Opg ratio, and Cox-2 responses to oscillatory fluid flow. Taken together, our results provide the first evidence for a relationship between estrogen withdrawal and defective αvβ3-mediated signalling. Specifically, this study implicates estrogen withdrawal as a putative mechanism responsible for altered αvβ3 expression and resultant changes in downstream signalling in osteocytes during post-menopausal osteoporosis, which might provide an important, but previously unidentified, contribution to the bone loss cascade.

Highlights

It is unknown whether the altered mechanotransduction in estrogen deficient osteocytes is directly associated with defective αvβ[3] expression or signalling
In vivo estrogen is important for normal bone cell function and so, to more closely mimic the physiological environment of osteocytes in vivo, in this study we first investigated the effect of supplementing culture media with pre-menopausal levels of estrogen (10 nM) on MLO-Y4 osteocytes under static and oscillatory flow conditions in vitro
Estrogen supplementation leads to larger osteocytes with more mature focal adhesions (FA) assembly and a more robust actin cytoskeleton, and these changes are accompanied by a reduction in gene expression associated with osteoclast differentiation (Rankl/Opg)

Summary

Introduction

It is unknown whether the altered mechanotransduction in estrogen deficient osteocytes is directly associated with defective αvβ[3] expression or signalling. We report that estrogen withdrawal leads to a smaller focal adhesion area and reduced αvβ[3] localisation at focal adhesion sites, resulting in an increased Rankl/Opg ratio and defective Cox-2 responses to oscillatory fluid flow. Focal adhesions are involved in Focal Adhesion Kinase (FAK) and shc signalling[13,14] and are widely understood to play a role in mechanosensation for many distinct cell types[15,16,17,18,19,20,21] Osteocytes express both β1 and β3 integrins[4,11] and it has been shown that β1 integrins localise around osteocyte cell bodies, whereas osteocyte cell processes have αvβ[3] integrins and both interact with the surrounding pericellular matrix[4,22]. Most in vitro studies of osteocyte biology use culture media without exogenous estrogen, and there is a limited understanding of pre-menopausal levels of estrogen on osteocyte biology

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