Dual usage of a stage-specific fluorescent reporter system based on a helper-dependent adenoviral vector to visualize osteogenic differentiation

Takefumi Sone, Masashi Shin, Takehito Ouchi, Hiroki Sasanuma, Arei Miyamoto, Satoshi Ohte, Sho Tsukamoto, Mahito Nakanishi, Hideyuki Okano, Takenobu Katagiri, Kohnosuke Mitani

Research output: Contribution to journalArticle

Abstract

We developed a reporter system that can be used in a dual manner in visualizing mature osteoblast formation. The system is based on a helper-dependent adenoviral vector (HDAdV), in which a fluorescent protein, Venus, is expressed under the control of the 19-kb human osteocalcin (OC) genomic locus. By infecting human and murine primary osteoblast (POB) cultures with this reporter vector, the cells forming bone-like nodules were specifically visualized by the reporter. In addition, the same vector was utilized to efficiently knock-in the reporter into the endogenous OC gene of human induced pluripotent stem cells (iPSCs), by homologous recombination. Neural crest-like cells (NCLCs) derived from the knock-in reporter iPSCs were differentiated into osteoblasts forming bone-like nodules and could be visualized by the expression of the fluorescent reporter. Living mature osteoblasts were then isolated from the murine mixed POB culture by fluorescence-activated cell sorting (FACS), and their mRNA expression profile was analyzed. Our study presents unique utility of reporter HDAdVs in stem cell biology and related applications.

Original languageEnglish
Article number9705
JournalScientific reports
Volume9
Issue number1
DOIs
Publication statusPublished - 2019 Dec 1

    Fingerprint

ASJC Scopus subject areas

  • General

Cite this

Sone, T., Shin, M., Ouchi, T., Sasanuma, H., Miyamoto, A., Ohte, S., Tsukamoto, S., Nakanishi, M., Okano, H., Katagiri, T., & Mitani, K. (2019). Dual usage of a stage-specific fluorescent reporter system based on a helper-dependent adenoviral vector to visualize osteogenic differentiation. Scientific reports, 9(1), [9705]. https://doi.org/10.1038/s41598-019-46105-y