Fabrication of Biomimetic Bone Tissue Using Mesenchymal Stem Cell-Derived Three-Dimensional Constructs Incorporating Endothelial Cells.

Jun-Ichi Sasaki,Itsumi Yoshimoto,Yoshihiro Itoh,Satoshi Imazato,Masanori Hashimoto,Satoshi Yamaguchi,Takuya Matsumoto,Xing-Ming Shi

doi:10.1371/journal.pone.0129266

Jun-Ichi Sasaki, Itsumi Yoshimoto + Show 6 more

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https://doi.org/10.1371/journal.pone.0129266

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Journal: PloS one	Publication Date: Jun 5, 2015
Citations: 31	License type: CC BY 4.0

Affiliation: Osaka University, Okayama University

Abstract

The development of technologies to promote vascularization of engineered tissue would drive major developments in tissue engineering and regenerative medicine. Recently, we succeeded in fabricating three-dimensional (3D) cell constructs composed of mesenchymal stem cells (MSCs). However, the majority of cells within the constructs underwent necrosis due to a lack of nutrients and oxygen. We hypothesized that incorporation of vascular endothelial cells would improve the cell survival rate and aid in the fabrication of biomimetic bone tissues in vitro. The purpose of this study was to assess the impact of endothelial cells combined with the MSC constructs (MSC/HUVEC constructs) during short- and long-term culture. When human umbilical vein endothelial cells (HUVECs) were incorporated into the cell constructs, cell viability and growth factor production were increased after 7 days. Furthermore, HUVECs were observed to proliferate and self-organize into reticulate porous structures by interacting with the MSCs. After long-term culture, MSC/HUVEC constructs formed abundant mineralized matrices compared with those composed of MSCs alone. Transmission electron microscopy and qualitative analysis revealed that the mineralized matrices comprised porous cancellous bone-like tissues. These results demonstrate that highly biomimetic bone tissue can be fabricated in vitro by 3D MSC constructs incorporated with HUVECs.

Highlights

Approaches that attempt to fabricate living tissues ex vivo are increasing with the development of tissue engineering technologies and biomaterials [1, 2]
Human umbilical vein endothelial cells (HUVECs; Riken) were cultured in Endothelial Cell Basal Medium-2 supplemented with SingleQuots (EBM-2; Lonza, Walkersville, MD). human-derived mesenchymal stem cells (hMSC) and human umbilical vein endothelial cells (HUVECs) were maintained in a humidified incubator at 37°C with 5% CO2
Cavities between the cells were observed by scanning electron microscopy (SEM) and Transmission electron microscopy (TEM). These results indicated that HUVECs self-organized under the influence of hMSCs and formed a luminal-like structure. Because these results demonstrated that HUVECs were able to survive and self-organize under the influence of hMSCs, we investigated the distribution of VEGF and HGF, two well-known angiogenic factors [37,38,39]

Summary

Introduction

Approaches that attempt to fabricate living tissues ex vivo are increasing with the development of tissue engineering technologies and biomaterials [1, 2]. In vitro tissue engineering techniques are considered to be applicable to regenerative medicine and to drug-discovery technology and histogenetics research [3,4,5,6]. PLOS ONE | DOI:10.1371/journal.pone.0129266 June 5, 2015

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