title: Bone morphogenetic protein 9 (BMP9) induces effective bone formation from reversibly immortalized multipotent adipose-derived (iMAD) mesenchymal stem cells.
abstract: Regenerative medicine and bone tissue engineering using mesenchymal stem cells (MSCs) hold great promise as an effective approach to bone and skeletal reconstruction. While adipose tissue harbors MSC-like progenitors, or multipotent adipose-derived cells (MADs), it is important to identify and characterize potential biological factors that can effectively induce osteogenic differentiation of MADs. To overcome the time-consuming and technically challenging process of isolating and culturing primary MADs, here we establish and characterize the reversibly immortalized mouse multipotent adipose-derived cells (iMADs). The isolated mouse primary inguinal MAD cells are reversibly immortalized via the retrovirus-mediated expression of SV40 T antigen flanked with FRT sites. The iMADs are shown to express most common MSC markers. FLP-mediated removal of SV40 T antigen effectively reduces the proliferative activity and cell survival of iMADs, indicating the immortalization is reversible. Using the highly osteogenic BMP9, we find that the iMADs are highly responsive to BMP9 stimulation, express multiple lineage regulators, and undergo osteogenic differentiationin vitroupon BMP9 stimulation. Furthermore, we demonstrate that BMP9-stimulated iMADs form robust ectopic bone with a thermoresponsive biodegradable scaffold material. Collectively, our results demonstrate that the reversibly immortalized iMADs exhibit the characteristics of multipotent MSCs and are highly responsive to BMP9-induced osteogenic differentiation. Thus, the iMADs should provide a valuable resource for the study of MAD biology, which would ultimately enable us to develop novel and efficacious strategies for MAD-based bone tissue engineering.
authors.affiliation: Shandong Provincial Orthopaedics Hospital, The Provincial Hospital Affiliated to Shandong UniversityJinan 250021, China; Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical CenterChicago, IL 60637, USA.
authors.forename: Shun
authors.initials: S
authors.lastname: Lu
authors.affiliation: Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical CenterChicago, IL 60637, USA; Ministry of Education Key Laboratory of Diagnostic Medicine, The Affiliated Hospitals of Chongqing Medical UniversityChongqing 400016, China.
authors.forename: Jing
authors.initials: J
authors.lastname: Wang
authors.affiliation: Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical CenterChicago, IL 60637, USA; Department of Biomedical Engineering, School of Bioengineering, Chongqing UniversityChongqing 400044, China.
authors.forename: Jixing
authors.initials: J
authors.lastname: Ye
authors.affiliation: Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical CenterChicago, IL 60637, USA; Ministry of Education Key Laboratory of Diagnostic Medicine, The Affiliated Hospitals of Chongqing Medical UniversityChongqing 400016, China.
authors.forename: Yulong
authors.initials: Y
authors.lastname: Zou
authors.affiliation: Department of Biomedical Engineering and Simpson Querrey Institute for BioNanotechnology in Medicine, Northwestern University Evanston, IL 60208, USA.
authors.forename: Yunxiao
authors.initials: Y
authors.lastname: Zhu
authors.affiliation: Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical CenterChicago, IL 60637, USA; Ministry of Education Key Laboratory of Diagnostic Medicine, The Affiliated Hospitals of Chongqing Medical UniversityChongqing 400016, China.
authors.forename: Qiang
authors.initials: Q
authors.lastname: Wei
authors.affiliation: Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical CenterChicago, IL 60637, USA; Departments of Surgery, Conservative Dentistry and Endodontics, West China Hospital and West China School of Stomatology, Sichuan UniversityChengdu 610041, China.
authors.forename: Xin
authors.initials: X
authors.lastname: Wang
authors.affiliation: Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical CenterChicago, IL 60637, USA; Department of General Surgery and Neurosurgery, The Affiliated Zhongnan Hospital of Wuhan UniversityWuhan 430071, China.
authors.forename: Shengli
authors.initials: S
authors.lastname: Tang
authors.affiliation: Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical CenterChicago, IL 60637, USA; Ministry of Education Key Laboratory of Diagnostic Medicine, The Affiliated Hospitals of Chongqing Medical UniversityChongqing 400016, China.
authors.forename: Hao
authors.initials: H
authors.lastname: Liu
authors.affiliation: Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical CenterChicago, IL 60637, USA; Ministry of Education Key Laboratory of Diagnostic Medicine, The Affiliated Hospitals of Chongqing Medical UniversityChongqing 400016, China.
authors.forename: Jiaming
authors.initials: J
authors.lastname: Fan
authors.affiliation: Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical CenterChicago, IL 60637, USA; Ministry of Education Key Laboratory of Diagnostic Medicine, The Affiliated Hospitals of Chongqing Medical UniversityChongqing 400016, China.
authors.forename: Fugui
authors.initials: F
authors.lastname: Zhang
authors.affiliation: Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center Chicago, IL 60637, USA.
authors.forename: Evan M
authors.initials: EM
authors.lastname: Farina
authors.affiliation: Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center Chicago, IL 60637, USA.
authors.forename: Maryam M
authors.initials: MM
authors.lastname: Mohammed
authors.affiliation: Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical CenterChicago, IL 60637, USA; Departments of Surgery, Conservative Dentistry and Endodontics, West China Hospital and West China School of Stomatology, Sichuan UniversityChengdu 610041, China.
authors.forename: Dongzhe
authors.initials: D
authors.lastname: Song
authors.affiliation: Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical CenterChicago, IL 60637, USA; Ministry of Education Key Laboratory of Diagnostic Medicine, The Affiliated Hospitals of Chongqing Medical UniversityChongqing 400016, China.
authors.forename: Junyi
authors.initials: J
authors.lastname: Liao
authors.affiliation: Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical CenterChicago, IL 60637, USA; Ministry of Education Key Laboratory of Diagnostic Medicine, The Affiliated Hospitals of Chongqing Medical UniversityChongqing 400016, China.
authors.forename: Jiayi
authors.initials: J
authors.lastname: Huang
authors.affiliation: Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical CenterChicago, IL 60637, USA; Ministry of Education Key Laboratory of Diagnostic Medicine, The Affiliated Hospitals of Chongqing Medical UniversityChongqing 400016, China.
authors.forename: Dan
authors.initials: D
authors.lastname: Guo
authors.affiliation: Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical CenterChicago, IL 60637, USA; Ministry of Education Key Laboratory of Diagnostic Medicine, The Affiliated Hospitals of Chongqing Medical UniversityChongqing 400016, China.
authors.forename: Minpeng
authors.initials: M
authors.lastname: Lu
authors.affiliation: Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical CenterChicago, IL 60637, USA; Ministry of Education Key Laboratory of Diagnostic Medicine, The Affiliated Hospitals of Chongqing Medical UniversityChongqing 400016, China.
authors.forename: Feng
authors.initials: F
authors.lastname: Liu
authors.affiliation: Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical CenterChicago, IL 60637, USA; Department of Orthopaedic Surgery, Union Hospital of Tongji Medical College, Huazhong University of Science & TechnologyWuhan 430022, China.
authors.forename: Jianxiang
authors.initials: J
authors.lastname: Liu
authors.affiliation: Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical CenterChicago, IL 60637, USA; Department of Biomedical Engineering, School of Bioengineering, Chongqing UniversityChongqing 400044, China.
authors.forename: Li
authors.initials: L
authors.lastname: Li
authors.affiliation: Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical CenterChicago, IL 60637, USA; Department of General Surgery and Neurosurgery, The Affiliated Zhongnan Hospital of Wuhan UniversityWuhan 430071, China.
authors.forename: Chao
authors.initials: C
authors.lastname: Ma
authors.affiliation: Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical CenterChicago, IL 60637, USA; Ministry of Education Key Laboratory of Diagnostic Medicine, The Affiliated Hospitals of Chongqing Medical UniversityChongqing 400016, China.
authors.forename: Xue
authors.initials: X
authors.lastname: Hu
authors.affiliation: Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center Chicago, IL 60637, USA.
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Lee
authors.affiliation: Section of Plastic Surgery, Department of Surgery, The University of Chicago Medical Center Chicago, IL 60637, USA.
authors.forename: Russell R
authors.initials: RR
authors.lastname: Reid
authors.affiliation: Department of Biomedical Engineering and Simpson Querrey Institute for BioNanotechnology in Medicine, Northwestern UniversityEvanston, IL 60208, USA; Department of Surgery, Feinberg School of MedicineChicago, IL 60616, USA.
authors.forename: Guillermo A
authors.initials: GA
authors.lastname: Ameer
authors.affiliation: Shandong Provincial Orthopaedics Hospital, The Provincial Hospital Affiliated to Shandong University Jinan 250021, China.
authors.forename: Dongsheng
authors.initials: D
authors.lastname: Zhou
authors.affiliation: Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical CenterChicago, IL 60637, USA; Ministry of Education Key Laboratory of Diagnostic Medicine, The Affiliated Hospitals of Chongqing Medical UniversityChongqing 400016, China.
authors.forename: Tongchuan
authors.initials: T
authors.lastname: He
keywords: BMP9; adipose-derived stem cells; bone formation; immortalized progenitor cells; mesenchymal stem cells; tissue engineering