Primary cilia respond to fluid shear stress and mediate flow‐induced calcium deposition in osteoblasts

Robin M Delaine‐Smith,Anuphan Sittichokechaiwut,Gwendolen C Reilly

doi:10.1096/fj.13-231894

Robin M Delaine‐Smith, Anuphan Sittichokechaiwut + Show 1 more

Open Access

https://doi.org/10.1096/fj.13-231894

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Journal: The FASEB Journal	Publication Date: Oct 4, 2013
Citations: 71	License type: CC BY 3.0

Affiliation: Naresuan University, University of Sheffield

Abstract

Bone turnover in vivo is regulated by mechanical forces such as shear stress originating from interstitial oscillatory fluid flow (OFF), and bone cells in vitro respond to mechanical loading. However, the mechanisms by which bone cells sense mechanical forces, resulting in increased mineral deposition, are not well understood. The aim of this study was to investigate the role of the primary cilium in mechanosensing by osteoblasts. MLO-A5 murine osteoblasts were cultured in monolayer and subjected to two different OFF regimens: 5 short (2 h daily) bouts of OFF followed by morphological analysis of primary cilia; or exposure to chloral hydrate to damage or remove primary cilia and 2 short bouts (2 h on consecutive days) of OFF. Primary cilia were shorter and there were fewer cilia per cell after exposure to periods of OFF compared with static controls. Damage or removal of primary cilia inhibited OFF-induced PGE2 release into the medium and mineral deposition, assayed by Alizarin red staining. We conclude that primary cilia are important mediators of OFF-induced mineral deposition, which has relevance for the design of bone tissue engineering strategies and may inform clinical treatments of bone disorders causes by load-deficiency.—Delaine-Smith, R. M., Sittichokechaiwut, A., Reilly, G. C. Primary cilia respond to fluid shear stress and mediate flow-induced calcium deposition in osteoblasts.

Highlights

Bone turnover in vivo is regulated by mechanical forces such as shear stress originating from interstitial oscillatory fluid flow (OFF), and bone cells in vitro respond to mechanical loading
Bone turnover and homeostasis are regulated by mechanical forces, and it is predicted that bone cells in vivo are exposed to shear stresses originating from interstitial oscillatory fluid flow (OFF; refs. 1, 2)
It has been clearly demonstrated that bone cells respond to Abbreviations: Alizarin red S (AR), Alizarin red; COX-2, cyclooxygenase 2; CH, chloral hydrate; DAPI, 4=-6-diamidino-2-phenylindole; ECM, extracellular matrix; FITC, fluorescein isothiocyanate; FSS, fluid shear stress; GAG, glycosaminoglycan; HA, hyaluronan; intraflagellar transport (IFT), intraflagella transport; Microtubule-associated protein 1 (MAP1), microtubule associated protein 1; MSC, mesenchymal stem cell; OFF, oscillatory fluid flow; OPN, osteopontin; PGE2, prostaglandin E2; PG, proteoglycan

Summary

Introduction

Bone turnover in vivo is regulated by mechanical forces such as shear stress originating from interstitial oscillatory fluid flow (OFF), and bone cells in vitro respond to mechanical loading. Primary cilia have been seen to exist in most mammalian tissue types and usually form several hours after growth arrest when cultured in vitro, taking several days to grow to full size [12], with cell spatial confinement being a major regulator of ciliogenesis [13] They are populated with receptors that participate in numerous signaling events [14] and play important roles in chemosensation [15, 16], thermosensation [17], and mechanosensation [18], and are believed to act as sensors to fluid flow. Application of a continuous 0.036 Pa flow caused primary cilia to deflect in osteoblasts [10], while the absence of a healthy primary cilium in osteoblasts and osteocytes resulted in a loss of the

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