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Establishment of Immortalized BMP2/4 Double Knock‐Out Osteoblastic Cells Is Essential for Study of Osteoblast Growth, Differentiation, and Osteogenesis
Author(s) -
Wu LiAn,
Wang Feng,
Donly Kevin J.,
Baker Andrew,
Wan Chunyan,
Luo Daoshu,
MacDougall Mary,
Chen Shuo
Publication year - 2016
Publication title -
journal of cellular physiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.529
H-Index - 174
eISSN - 1097-4652
pISSN - 0021-9541
DOI - 10.1002/jcp.25266
Subject(s) - bone morphogenetic protein 2 , osteoblast , microbiology and biotechnology , immortalised cell line , cellular differentiation , cell culture , extracellular matrix , cell growth , chemistry , biology , gene , genetics , biochemistry , in vitro
Bone morphogenetic proteins 2 and 4 (BMP2/4) are essential for osteoblast differentiation and osteogenesis. Generation of a BMP2/4 dual knock‐out ( ko/ko ) osteoblastic cell line is a valuable asset for studying effects of BMP2/4 on skeletal development. In this study, our goal was to create immortalized mouse deleted BMP2/4 osteoblasts by infecting adenoviruses with Cre recombinase and green fluorescent protein genes into immortalized murine floxed BMP2/4 osteoblasts. Transduced BMP2/4 ko/ko cells were verified by green immunofluorescence and PCR. BMP2/4 ko/ko osteoblasts exhibited small size, slow cell proliferation rate and cell growth was arrested in G1 and G2 phases. Expression of bone‐relate genes was reduced in the BMP2/4 ko/ko cells, resulting in delay of cell differentiation and mineralization. Importantly, extracellular matrix remodeling was impaired in the BMP2/4 ko/ko osteoblasts as reflected by decreased Mmp‐2 and Mmp‐9 expressions. Cell differentiation and mineralization were rescued by exogenous BMP2 and/or BMP4. Therefore, we for the first time described establishment of an immortalized deleted BMP2/4 osteoblast line useful for study of mechanisms in regulating osteoblast lineages. J. Cell. Physiol. 231: 1189–1198, 2016. © 2015 The Authors. Journal of Cellular Physiology Published by Wiley Periodicals, Inc.