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Tissue Eng Part A. 2018 Aug 20. doi: 10.1089/ten.TEA.2017.0465. [Epub ahead of print]

Frontal Bone Healing Is Sensitive to Wnt Signaling Inhibition via Lentiviral-Encoded Beta-Catenin Short Hairpin RNA.

Author information

1
1 Department of Pathology, Johns Hopkins University , Baltimore, Maryland.
2
2 Department of Oral and Maxillofacial Surgery, School of Stomatology, China Medical University , Shenyang, China .
3
3 University of California San Diego School of Medicine , La Jolla, California.
4
4 UCLA and Orthopaedic Hospital Department of Orthopaedic Surgery and the Orthopaedic Hospital Research Center , Los Angeles, California.
5
5 Division of Growth and Development and Section of Orthodontics, School of Dentistry , UCLA, Los Angeles, California.
6
6 Division of Plastic and Reconstructive Surgery, Department of Surgery, David Geffen School of Medicine, University of California , Los Angeles, California.

Abstract

The Wnt/β-catenin signaling pathway plays an integral role in skeletal biology, spanning from embryonic skeletal patterning through bone maintenance and bone repair. Most experimental methods to antagonize Wnt signaling in vivo are either systemic or transient, including genetic approaches, use of small-molecule inhibitors, or neutralizing antibodies. We sought to develop a novel, localized model of prolonged Wnt/β-catenin signaling blockade by the application and validation of a lentivirus encoding β-catenin short hairpin RNA (shRNA). Efficacy of lentiviral-encoded β-catenin shRNA was first confirmed in vitro using bone marrow mesenchymal stromal cells, and in vivo using an intramedullary long bone injection model in NOD SCID mice. Next, the effects of β-catenin knockdown were assessed in a calvarial bone defect model, in which the frontal bone demonstrates enhanced bone healing associated with heightened Wnt/β-catenin signaling. Lentivirus encoding either β-catenin shRNA or random sequence shRNA with enhanced green fluorescent protein (control) was injected overlying the calvaria of NOD SCID mice and bone defects were created in either the frontal or parietal bones. Among mice treated with lentivirus encoding β-catenin shRNA, frontal bone defect healing was significantly reduced by all radiographic and histologic metrics. In contrast, parietal bone healing was minimally impacted by β-catenin shRNA. In aggregate, our data document the application and validation of a lentivirus encoding β-catenin shRNA model that represents an easily replicable tool for examining the importance of locoregional Wnt/β-catenin signaling in bone biology and regeneration.

KEYWORDS:

Wnt signaling; beta-catenin shRNA; bone healing; bone repair; calvaria; calvarial bone defect; intramembranous bone; membranous bone; osteogenesis; skull

PMID:
29929440
PMCID:
PMC6302677
[Available on 2019-12-01]
DOI:
10.1089/ten.TEA.2017.0465

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