title: TrAmplification of Human Dental Follicle Cells by piggyBac Transposon - Mediated Reversible Immortalization System.
abstract: Dental follicle cells (DFCs) are the precursor cells of periodontium. Under certain differentiation conditions, DFCs can be induced to differentiate into chondrogenic, osteogenic and adipogenic cells. However, DFCs has limited lifespan in vitro, so it's difficult to harvest enough cells for basic research and translational application. pMPH86 is a piggyBac transposon-mediated vector which contains SV40 T-Ag cassette that can be removed by flippase recognition target (FRT) recombinase. Here we demonstrated the pMPH86 can effectively amplify human DFCs through reversible immortalization. The immortalized DFCs (iDFCs) exhibit higher proliferate activity, which can be reversed to its original level before immortalization when deimmortalized by FLP recombinase. The iDFCs and deimmortalized DFCs (dDFCs) express most DFC markers and maintain multiple differentiation potential in vitro as they can be induced by BMP9 to differentiate into chondrogenic, osteogenic and adipogenic cells evidenced by gene expression and protein marker. We also proved telomerase activity of iDFCs are significantly increased and maintained at a high level, while the telomerase activity of primary DFCs was relatively low and decreased with every passage. After SV40 T-Ag was removed to deimmortalize the cells, telomerase activity was reduced to its original level before immortalization and decreased with passages just the same as primary DFCs. These results suggest that piggyBac immortalization system could be a potential strategy to amplify primary cells, which is critical for regenerative research and further clinical application.
authors.affiliation: Department of Orthodontics, Stomatological Hospital of Chongqing Medical University, Chongqing, P. R. China; Chongqing Key Laboratory of Oral Disease and Biomedical Sciences, Chongqing, P. R. China; Chongqing Municipal Key Laboratory of Oral Biomedical Engineering of Higher Education, Chongqing, P. R. China.
authors.forename: Yan
authors.initials: Y
authors.lastname: Wu
authors.affiliation: Department of Orthodontics, Stomatological Hospital of Chongqing Medical University, Chongqing, P. R. China; Chongqing Key Laboratory of Oral Disease and Biomedical Sciences, Chongqing, P. R. China; Chongqing Municipal Key Laboratory of Oral Biomedical Engineering of Higher Education, Chongqing, P. R. China.
authors.forename: Ge
authors.initials: G
authors.lastname: Feng
authors.affiliation: Department of Orthodontics, Stomatological Hospital of Chongqing Medical University, Chongqing, P. R. China; Chongqing Key Laboratory of Oral Disease and Biomedical Sciences, Chongqing, P. R. China; Chongqing Municipal Key Laboratory of Oral Biomedical Engineering of Higher Education, Chongqing, P. R. China.
authors.forename: Jinlin
authors.initials: J
authors.lastname: Song
authors.affiliation: Wake Forest Institute for Regenerative Medicine (WFIRM), Wake Forest University Health Sciences, Winston-Salem, North Carolina, United States of America.
authors.forename: Yuanyuan
authors.initials: Y
authors.lastname: Zhang
authors.affiliation: Department of Orthodontics, Stomatological Hospital of Chongqing Medical University, Chongqing, P. R. China; Chongqing Key Laboratory of Oral Disease and Biomedical Sciences, Chongqing, P. R. China; Chongqing Municipal Key Laboratory of Oral Biomedical Engineering of Higher Education, Chongqing, P. R. China.
authors.forename: Yong
authors.initials: Y
authors.lastname: Yu
authors.affiliation: Department of Orthodontics, Stomatological Hospital of Chongqing Medical University, Chongqing, P. R. China; Chongqing Key Laboratory of Oral Disease and Biomedical Sciences, Chongqing, P. R. China; Chongqing Municipal Key Laboratory of Oral Biomedical Engineering of Higher Education, Chongqing, P. R. China.
authors.forename: Lan
authors.initials: L
authors.lastname: Huang
authors.affiliation: Department of Orthodontics, Stomatological Hospital of Chongqing Medical University, Chongqing, P. R. China; Chongqing Key Laboratory of Oral Disease and Biomedical Sciences, Chongqing, P. R. China; Chongqing Municipal Key Laboratory of Oral Biomedical Engineering of Higher Education, Chongqing, P. R. China.
authors.forename: Leilei
authors.initials: L
authors.lastname: Zheng
authors.affiliation: Department of Orthodontics, Stomatological Hospital of Chongqing Medical University, Chongqing, P. R. China; Chongqing Key Laboratory of Oral Disease and Biomedical Sciences, Chongqing, P. R. China; Chongqing Municipal Key Laboratory of Oral Biomedical Engineering of Higher Education, Chongqing, P. R. China.
authors.forename: Feng
authors.initials: F
authors.lastname: Deng
mesh_terms: D050156:Adipogenesis; D015415:Biomarkers / Q000378:metabolism; D002454:Cell Differentiation; D049109:Cell Proliferation; D020219:Chondrogenesis; D004251:DNA Transposable Elements / Q000235:genetics*; D003795:Dental Sac / Q000166:cytology* / Q000378:metabolism; D005822:Genetic Vectors / Q000235:genetics; D006801:Humans; D010012:Osteogenesis; D019098:Telomerase / Q000378:metabolism
chemical_list: D015415:Biomarkers; D004251:DNA Transposable Elements; D019098:Telomerase