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Development and characterization of a mouse floxed Bmp2 osteoblast cell line that retains osteoblast genotype and phenotype

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Abstract

Bone morphogenetic protein 2 (Bmp2) is essential for osteoblast differentiation and osteogenesis. Generation of floxed Bmp2 osteoblast cell lines is a valuable tool for studying the effects of Bmp2 on osteoblast differentiation and its signaling pathways during skeletal metabolism. Due to relatively limited sources of primary osteoblasts, we have developed cell lines that serve as good surrogate models for the study of osteoblast cell differentiation and bone mineralization. In this study, we established and characterized immortalized mouse floxed Bmp2 osteoblast cell lines. Primary mouse floxed Bmp2 osteoblasts were transfected with pSV3-neo and clonally selected. These transfected cells were verified by PCR and immunohistochemistry. To determine the genotype and phenotype of the immortalized cells, cell morphology, proliferation, differentiation and mineralization were analyzed. Also, expression of osteoblast-related gene markers including Runx2, Osx, ATF4, Dlx3, bone sialoprotein, dentin matrix protein 1, osteonectin, osteocalcin and osteopontin were examined by quantitative RT-PCR and immunohistochemistry. These results showed that immortalized floxed Bmp2 osteoblasts had a higher proliferation rate but preserved their genotypic and phenotypic characteristics similar to the primary cells. Thus, we, for the first time, describe the development of immortalized mouse floxed Bmp2 osteoblast cell lines and present a useful model to study osteoblast biology mediated by BMP2 and its downstream signaling transduction pathways.

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Abbreviations

Bmp2 :

Bone morphogenetic protein 2

BrdU:

5-bromo-2'-deoxyuridine

ALP :

Alkaline phosphatase

ATF4 :

Activating transcription factor 4

Bsp :

Bone sialoprotein

Col1α1 :

Alpha 1 collagen type

Dlx3 :

Distal-less 3

Dmp1 :

Dentin matrix protein 1

Gadph :

Glyceraldehyde-3-phosphate dehydrogenase

Oc :

Osteocalcin

Opn :

Osteopontin

Osn :

Osteonectin

Osx :

Osterix

SV40:

SV40 large T-antigen.

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Acknowledgments

This work was supported in part by National Institute of Health Grant DE019892 (S.C.), San Antonio Area Foundation (S.C.) and the grant from the Natural Science Foundation of China 30801293 (L.A. Wu) and 2009-10 3 M ESPE Preventative Pediatric Dentistry Postdoctoral Research Fellowship of AAPD (L.A. Wu).

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Authors and Affiliations

  1. Department of Pediatric Dentistry, Dental School, The University of Texas Health Science Center at San Antonio, 7703 Floyd Curl Drive, San Antonio, TX, 78229-3900, USA

    Li-an Wu, Junsheng Feng, Lynn Wang, Yan-dong Mu, Andrew Baker, Kevin J. Donly & Shuo Chen

  2. Department of Pediatric Dentistry, School of Stomatology, the Fourth Military Medical University, Xi-an, China

    Li-an Wu

  3. Department of Anatomy & Embryology, Fujian Medical University, Fuzhou, China

    Junsheng Feng

  4. Department of Periodontics, Dental School, The University of Texas Health Science Center at San Antonio, San Antonio, TX, USA

    Andrew Baker & Stephen E. Harris

  5. Department of Oral/Maxillofacial Surgery, University of Alabama, Birmingham, AL, USA

    Mary MacDougall

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  1. Li-an Wu
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  2. Junsheng Feng
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Correspondence to Shuo Chen.

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Wu, La., Feng, J., Wang, L. et al. Development and characterization of a mouse floxed Bmp2 osteoblast cell line that retains osteoblast genotype and phenotype. Cell Tissue Res 343, 545–558 (2011). https://doi.org/10.1007/s00441-010-1120-3

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  • DOI: https://doi.org/10.1007/s00441-010-1120-3

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