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Role of embryonic origin on osteogenic potential and bone repair capacity of rat calvarial osteoblasts

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Abstract

Introduction

The aim of this study was to evaluate the in vitro osteogenic potential of osteoblasts from neural crest-derived frontal bone (OB-NC) and mesoderm-derived parietal bone (OB-MS) and the bone formation induced by them when injected into calvarial defects.

Materials and methods

Calvarial bones were collected from newborn Wistar rats (3-day old) and characterized as frontal and parietal prior to OB-NC and OB-MS harvesting. The cells were cultured, and several parameters of osteoblast differentiation were evaluated. These cells, or PBS without cells (control), were locally injected into 5-mm rat calvarial defects (5 × 106 cells/defect) and after 4 weeks bone formation was evaluated by morphometric and histological analyses.

Results

The characterization of frontal and parietal bones assured the different embryonic origin of both cell populations, OB-NC and OB-MS. The OB-NC presented higher proliferation while the OB-MS presented higher alkaline phosphatase (ALP) activity, extracellular matrix mineralization and gene expression of runt-related transcription factor 2, Alp, bone sialoprotein and osteocalcin revealing their high osteogenic potential. µCT analysis indicated that there was higher amount of bone formation in defects injected with both OB-NC and OB-MS compared to the control. Moreover, the bone tissue formed by both cells displayed the same histological characteristics.

Conclusions

Despite the distinct in vitro osteogenic potential, OB-NC and OB-MS induced similar bone repair in a rat calvarial defect model. Thus, osteoblasts, irrespective of their in vitro osteogenic potential linked to embryonic origins, seem to be suitable for cell-based therapies aiming to repair bone defects.

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Acknowledgements

Fabiana Rossetto de Moraes and Roger Rodrigo Fernandes are acknowledged for technical assistance during the experiments. This research was supported by the Coordination for the Improvement of Higher Education Personnel (CAPES, Grant # 00.889.834/0001-08), National Council for Scientific and Technological Development (CNPq, Grant # 404318/2016-9) and São Paulo Research Foundation (FAPESP, Grants # 2009/54142-5 and 2015/21439-6).

Author information

Authors and Affiliations

  1. Bone Research Lab, School of Dentistry of Ribeirão Preto, University of São Paulo, Av do Café s/n, Ribeirão Preto, SP, 14040-904, Brazil

    Alann Thaffarell Portilho Souza, Helena Bacha Lopes, Gileade Pereira Freitas, Fabiola Singaretti Oliveira, Adriana Luisa Gonçalves Almeida, Denise Weffort, Marcio Mateus Beloti & Adalberto Luiz Rosa

  2. Department of Maxillofacial Surgery, Prosthesis and Traumatology, School of Dentistry, University of São Paulo, São Paulo, SP, Brazil

    Emanuela Prado Ferraz

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  1. Alann Thaffarell Portilho Souza
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Contributions

ATPS undertook the experiments, analysed the data and drafted the manuscript. HBL and GPF worked with animal experiments, analysed the data and revised the manuscript. EPF, FSO and DW worked with in vitro experiments and revised the manuscript. ALGA performed the histological and microtomographic analysis and revised the manuscript. MMB and ALR designed the study, analysed the data and drafted the manuscript.

Corresponding author

Correspondence to Adalberto Luiz Rosa.

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Conflict of interest

All authors have no conflicts of interest.

Animal rights

All applicable international, national, and/or institutional guidelines for the care and use of animals were followed. The Committee of Ethics in Animal Research of the School of Dentistry of Ribeirão Preto, University of São Paulo approved all animal procedures performed during the experiments (Protocol # 2016.668.58.0, approved on September 21th, 2016).

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774_2020_1090_MOESM3_ESM.tif

Fig. S1 Gene expression of the bone morphogenetic proteins 2 (Bmp2, a) and 4 (Bmp4, b), and distal-less homeobox 5 (Dlx5, c) in osteoblasts derived from neural crest (frontal bone, OB-NC) and mesoderm (parietal bone, OB-MS) of newborn rats, grown under osteogenic conditions, on 3, 7 and 10 days. Asterisks (*) indicate statistically significant differences (ANOVA, p≤0.05). Data are presented as mean ± standard deviation (n=3) (TIF 2026 kb)

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Souza, A.T.P., Lopes, H.B., Freitas, G.P. et al. Role of embryonic origin on osteogenic potential and bone repair capacity of rat calvarial osteoblasts. J Bone Miner Metab 38, 481–490 (2020). https://doi.org/10.1007/s00774-020-01090-5

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