Netrin-4 Promotes Differentiation and Migration of Osteoblasts

Yuichiro Enoki; Tsuyoshi Sato; Shoichiro Kokabu; Naoki Hayashi; Takanori Iwata; Masayuki Yamato; Michihiko Usui; Masahito Matsumoto; Taketo Tomoda; Wataru Ariyoshi; Tatsuji Nishihara; Tetsuya Yoda

doi:10.21873/invivo.11132

Netrin-4 Promotes Differentiation and Migration of Osteoblasts

In Vivo. 2017 Sep-Oct;31(5):793-799. doi: 10.21873/invivo.11132.

Authors

Affiliations

¹ Department of Oral and Maxillofacial Surgery, Saitama Medical University, Saitama, Japan.
² Department of Oral and Maxillofacial Surgery, Saitama Medical University, Saitama, Japan tsato@saitama-med.ac.jp.
³ Division of Molecular Signaling and Biochemistry, Department of Health Promotion and Kyushu Dental University, Fukuoka, Japan.
⁴ Institute of Advanced Biomedical Engineering and Science, Tokyo Women's Medical University, Tokyo, Japan.
⁵ Division of Periodontology, Department of Cardiology and Periodontology, Kyushu Dental University, Fukuoka, Japan.
⁶ Division of Functional Genomics and Systems Medicine, Research Center for Genomic Medicine, Saitama Medical University, Saitama, Japan.
⁷ Division of Infections and Molecular Biology, Department of Health Promotion, Kyushu Dental University, Fukuoka, Japan.

Abstract

Background/aim: While netrin-4 plays a vital role in the vascular system, the role of netrin-1 in osteoblast differentiation is not well understood. In this study we explored whether netrin-4 has functional roles in osteoblasts.

Materials and methods: Quantitative reverse-transcriptase polymerase chain reaction (PCR), RNA interference, the generation of plasmids, transfections, measurement of alkaline phosphatase activity, a mineralization assay, a migration assay and a cell proliferation assay were performed.

Results: Netrin-4 expression was up-regulated during osteoblast differentiation and an RNA interference experiment showed that small interfering RNA used to silence netrin-4 inhibited osteoblast differentiation. Recombinant mouse netrin-4 promoted alkaline phosphatase (ALP) activity of osteoblasts and enhancement of calcium deposits. Moreover, we constructed a vector containing the netrin-4 gene on the basis of the plasmid pcDNA3.1/V5-His. Overexpression of netrin-4 enhanced differentiation of osteoblasts. Finally, recombinant mouse netrin-4 promoted cell migration of osteoblasts.

Conclusion: Netrin-4 promotes differentiation and migration of osteoblasts.

Keywords: Netrin-4; differentiation; migration; osteoblast.

MeSH terms

Animals
Cell Differentiation / genetics*
Cell Line
Cell Movement / genetics
Cell Proliferation / genetics
Gene Expression
Gene Knockdown Techniques
Mice
Netrins / genetics*
Netrins / metabolism
Osteoblasts / cytology*
Osteoblasts / metabolism*
RNA Interference
RNA, Small Interfering / genetics

Substances

Netrins
Ntn4 protein, mouse
RNA, Small Interfering