title: The Prodomain-Containing BMP9 Produced from a Stable Line Effectively Regulates the Differentiation of Mesenchymal Stem Cells.
abstract: BACKGROUND
  BMPs play important roles in regulating stem cell proliferation and differentiation. Using adenovirus-mediated expression of the 14 types of BMPs we demonstrated that BMP9 is one of the most potent BMPs in inducing osteogenic differentiation of mesenchymal stem cells (MSCs), which was undetected in the early studies using recombinant BMP9 proteins. Endogenous BMPs are expressed as a precursor protein that contains an N-terminal signal peptide, a prodomain and a C-terminal mature peptide. Most commercially available recombinant BMP9 proteins are purified from the cells expressing the mature peptide. It is unclear how effectively these recombinant BMP9 proteins functionally recapitulate endogenous BMP9.
  
  
  METHODS
  A stable cell line expressing the full coding region of mouse BMP9 was established in HEK-293 cells by using the piggyBac transposon system. The biological activities and stability of the conditioned medium generated from the stable line were analyzed.
  
  
  RESULTS
  The stable HEK-293 line expresses a high level of mouse BMP9. BMP9 conditioned medium (BMP9-cm) was shown to effectively induce osteogenic differentiation of MSCs, to activate BMP-R specific Smad signaling, and to up-regulate downstream target genes in MSCs. The biological activity of BMP9-cm is at least comparable with that induced by AdBMP9 in vitro. Furthermore, BMP9-cm exhibits an excellent stability profile as its biological activity is not affected by long-term storage at -80ºC, repeated thawing cycles, and extended storage at 4ºC.
  
  
  CONCLUSIONS
  We have established a producer line that stably expresses a high level of active BMP9 protein. Such producer line should be a valuable resource for generating biologically active BMP9 protein for studying BMP9 signaling mechanism and functions.
authors.affiliation: 1. Department of Neurology, Hubei Zhongshan Hospital, Wuhan, China; 2. Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, IL, USA.
authors.forename: Ruifang
authors.initials: R
authors.lastname: Li
authors.affiliation: 2. Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, IL, USA; 3. Ministry of Education Key Laboratory of Diagnostic Medicine, and the Affiliated Hospitals of Chongqing Medical University, Chongqing, China.
authors.forename: Zhengjian
authors.initials: Z
authors.lastname: Yan
authors.affiliation: 2. Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, IL, USA; 4. Department of Biomedical Engineering, School of Bioengineering, Chongqing University, Chongqing, China.
authors.forename: Jixing
authors.initials: J
authors.lastname: Ye
authors.affiliation: 5. Ben May Department for Cancer Research, The University of Chicago Medical Center, Chicago, IL 60637, USA.
authors.forename: He
authors.initials: H
authors.lastname: Huang
authors.affiliation: 2. Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, IL, USA; 3. Ministry of Education Key Laboratory of Diagnostic Medicine, and the Affiliated Hospitals of Chongqing Medical University, Chongqing, China.
authors.forename: Zhongliang
authors.initials: Z
authors.lastname: Wang
authors.affiliation: 2. Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, IL, USA; 3. Ministry of Education Key Laboratory of Diagnostic Medicine, and the Affiliated Hospitals of Chongqing Medical University, Chongqing, China.
authors.forename: Qiang
authors.initials: Q
authors.lastname: Wei
authors.affiliation: 2. Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, IL, USA; 3. Ministry of Education Key Laboratory of Diagnostic Medicine, and the Affiliated Hospitals of Chongqing Medical University, Chongqing, China.
authors.forename: Jing
authors.initials: J
authors.lastname: Wang
authors.affiliation: 2. Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, IL, USA; 6. Department of Orthopaedic Surgery, the Second Affiliated Hospital of Lanzhou University, Lanzhou, China.
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authors.affiliation: 2. Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, IL, USA; 7. Department of Orthopaedic Surgery, Shandong Provincial Hospital and Shandong University School of Medicine, Jinan, China.
authors.forename: Shun
authors.initials: S
authors.lastname: Lu
authors.affiliation: 2. Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, IL, USA; 8. Department of Surgery, West China Hospital of Sichuan University, Chengdu, China;
authors.forename: Xin
authors.initials: X
authors.lastname: Wang
authors.affiliation: 1. Department of Neurology, Hubei Zhongshan Hospital, Wuhan, China; 9. Department of General Surgery, the Research Center of Digestive Diseases, Zhongnan Hospital of Wuhan University, Wuhan, China.
authors.forename: Shengli
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authors.lastname: Tang
authors.affiliation: 2. Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, IL, USA; 3. Ministry of Education Key Laboratory of Diagnostic Medicine, and the Affiliated Hospitals of Chongqing Medical University, Chongqing, China.
authors.forename: Jiaming
authors.initials: J
authors.lastname: Fan
authors.affiliation: 2. Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, IL, USA; 3. Ministry of Education Key Laboratory of Diagnostic Medicine, and the Affiliated Hospitals of Chongqing Medical University, Chongqing, China.
authors.forename: Fugui
authors.initials: F
authors.lastname: Zhang
authors.affiliation: 2. Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, IL, USA; 3. Ministry of Education Key Laboratory of Diagnostic Medicine, and the Affiliated Hospitals of Chongqing Medical University, Chongqing, China.
authors.forename: Yulong
authors.initials: Y
authors.lastname: Zou
authors.affiliation: 2. Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, IL, USA; 8. Department of Surgery, West China Hospital of Sichuan University, Chengdu, China;
authors.forename: Dongzhe
authors.initials: D
authors.lastname: Song
authors.affiliation: 2. Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, IL, USA; 3. Ministry of Education Key Laboratory of Diagnostic Medicine, and the Affiliated Hospitals of Chongqing Medical University, Chongqing, China.
authors.forename: Junyi
authors.initials: J
authors.lastname: Liao
authors.affiliation: 2. Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, IL, USA; 3. Ministry of Education Key Laboratory of Diagnostic Medicine, and the Affiliated Hospitals of Chongqing Medical University, Chongqing, China.
authors.forename: Minpeng
authors.initials: M
authors.lastname: Lu
authors.affiliation: 2. Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, IL, USA; 3. Ministry of Education Key Laboratory of Diagnostic Medicine, and the Affiliated Hospitals of Chongqing Medical University, Chongqing, China.
authors.forename: Feng
authors.initials: F
authors.lastname: Liu
authors.affiliation: 2. Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, IL, USA.
authors.forename: Lewis L
authors.initials: LL
authors.lastname: Shi
authors.affiliation: 2. Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, IL, USA.
authors.forename: Aravind
authors.initials: A
authors.lastname: Athiviraham
authors.affiliation: 2. Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, IL, USA.
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Lee
authors.affiliation: 2. Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, IL, USA.
authors.forename: Tong-Chuan
authors.initials: TC
authors.lastname: He
authors.affiliation: 2. Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, IL, USA; 9. Department of General Surgery, the Research Center of Digestive Diseases, Zhongnan Hospital of Wuhan University, Wuhan, China.
authors.forename: Zhonglin
authors.initials: Z
authors.lastname: Zhang
mesh_terms: D000818:Animals; D002454:Cell Differentiation / Q000235:genetics*; D017077:Culture Media, Conditioned / Q000378:metabolism; D055427:Growth Differentiation Factor 2 / Q000096:biosynthesis* / Q000235:genetics; D057809:HEK293 Cells; D006801:Humans; D059630:Mesenchymal Stem Cells / Q000166:cytology*; D051379:Mice; D010012:Osteogenesis / Q000235:genetics*; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics
chemical_list: D017077:Culture Media, Conditioned; C526108:Gdf2 protein, mouse; D055427:Growth Differentiation Factor 2; D011994:Recombinant Proteins
keywords: BMP9; Mesenchymal stem cells; conditioned medium; osteogenic differentiation; recombinant proteins; stem cells
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