Calcium-containing scaffolds induce bone regeneration by regulating mesenchymal stem cell differentiation and migration

Rubén Aquino-Martínez,Francesc Ventura Pujol,Alcira P Angelo

doi:10.1186/s13287-017-0713-0

Abstract

BackgroundOsteoinduction and subsequent bone formation rely on efficient mesenchymal stem cell (MSC) recruitment. It is also known that migration is induced by gradients of growth factors and cytokines. Degradation of Ca2+-containing biomaterials mimics the bone remodeling compartment producing a localized calcium-rich osteoinductive microenvironment. The aim of our study was to determine the effect of calcium sulfate (CaSO4) on MSC migration. In addition, to evaluate the influence of CaSO4 on MSC differentiation and the potential molecular mechanisms involved.MethodsA circular calvarial bone defect (5 mm diameter) was created in the parietal bone of 35 Balb-C mice. We prepared and implanted a cell-free agarose/gelatin scaffold alone or in combination with different CaSO4 concentrations into the bone defects. After 7 weeks, we determined the new bone regenerated by micro-CT and histological analysis. In vitro, we evaluated the CaSO4 effects on MSC migration by both wound healing and agarose spot assays. Osteoblastic gene expression after BMP-2 and CaSO4 treatment was also evaluated by qPCR.ResultsCaSO4 increased MSC migration and bone formation in a concentration-dependent manner. Micro-CT analysis showed that the addition of CaSO4 significantly enhanced bone regeneration compared to the scaffold alone. The histological evaluation confirmed an increased number of endogenous cells recruited into the cell-free CaSO4-containing scaffolds. Furthermore, MSC migration in vitro and active AKT levels were attenuated when CaSO4 and BMP-2 were in combination. Addition of LY294002 and Wortmannin abrogated the CaSO4 effects on MSC migration.ConclusionsSpecific CaSO4 concentrations induce bone regeneration of calvarial defects in part by acting on the host’s undifferentiated MSCs and promoting their migration. Progenitor cell recruitment is followed by a gradual increment in osteoblast gene expression. Moreover, CaSO4 regulates BMP-2-induced MSC migration by differentially activating the PI3K/AKT pathway. Altogether, these results suggest that CaSO4 scaffolds could have potential applications for bone regeneration.

Highlights

Osteoinduction and subsequent bone formation rely on efficient mesenchymal stem cell (MSC) recruitment
Several studies have demonstrated that extracellular calcium, TGF-β, Bone morphogenetic protein 2 (BMP-2), BMP-4, PDGF, and other growth factors have a promigratory effect over the MSCs [5,6,7,8,9]
CaSO4 induces MSC migration in vitro In order to evaluate the MSC migration response, the cells were exposed to different CaSO4 concentrations and allowed to migrate for 24 hours

Summary

Introduction

Osteoinduction and subsequent bone formation rely on efficient mesenchymal stem cell (MSC) recruitment. Osteoinduction is initiated by guided attraction of the host’s mesenchymal stem cells (MSCs) from adjacent tissues in response to chemotactic cues released from the bone graft or the implanted biomaterial. During remodeling osteoclasts release a myriad of signaling molecules from the bone matrix. These soluble signals diffuse and create an osteoinductive microenvironment that promotes the osteoprogenitor cell recruitment into the resorbed lacunae. Osteoprogenitor cell motility relies on a single growth factor but on a chemoattractant gradient formed by multiple biochemical signals. Several studies have demonstrated that extracellular calcium (the main component of the mineralized bone), TGF-β, BMP-2, BMP-4, PDGF, and other growth factors have a promigratory effect over the MSCs [5,6,7,8,9]

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