title: Efficient preparation and labeling of human induced pluripotent stem cells by nanotechnology.
abstract: Efficient preparation and labeling of human induced pluripotent stem (iPS) cells is a great challenge in stem cell research and development. With the aim of investigating the feasibility of using nanotechnology to enhance the preparation efficiency of iPS cells and to label iPS cells for long-term tracing and imaging, in this paper, four transcription factor genes, ie, Oct4, Sox2, LIN28, and Nanog, and packaging plasmids such as PSPAX2 and PMD2.G were cotransfected into 293T cells using Generation 5.0 polyamidoamine dendrimer-modified magnetic nanoparticles (dMNPs) as a delivery system. The resultant supernatant liquids were incubated with human fibroblast cells at 37°C for 21 days, then the embryonic stem (ES) cell-like clones were screened, cultured, and identified. Finally, the prepared iPS cells were labeled with fluorescent magnetic nanoparticles (FMNPs). The results showed that dMNPs can efficiently deliver all vectors into 293T cells. The resultant lentiviruses' titers were 10-fold more than those based on Lipofectamine™ 2000. Reverse transcription polymerase chain reaction analysis showed that four genes (Oct4, Sox2, LIN28, and Nanog) exhibited different expressions in iPS cells. Immunostaining analysis showed that specific surface markers of ES cells such as SSEA-3, SSEA-4, Tra-1-60, and Tra-1-81 were positive in iPS cells, and the terotomas were formed in NOD-SCID mice that were implanted with iPS cells. Red fluorescent signals could be observed in iPS cells labeled with FMNPs by fluorescent microscopy, and the magnetic signals were detected in labeled iPS cells by magnetic resonance imaging. In conclusion, human iPS cells can be efficiently generated using polyamidoamine dMNPs and lentivirus and labeled with FMNPs for long-term observation and tracking, which has great potential application in the research and development of stem cells in the near future.
authors.affiliation: Department of Bio-Nano Science and Engineering, Key Laboratory for Thin Film and Microfabrication Technology of the Ministry of Education, National Key Laboratory of Micro/Nano Fabrication Technology, Research Institute of Micro/Nano Science and Technology, Shanghai Jiao Tong University, Shanghai, People's Republic of China.
authors.forename: Jing
authors.initials: J
authors.lastname: Ruan
authors.forename: Jie
authors.initials: J
authors.lastname: Shen
authors.forename: Zheng
authors.initials: Z
authors.lastname: Wang
authors.forename: Jiajia
authors.initials: J
authors.lastname: Ji
authors.forename: Hua
authors.initials: H
authors.lastname: Song
authors.forename: Kan
authors.initials: K
authors.lastname: Wang
authors.forename: Bin
authors.initials: B
authors.lastname: Liu
authors.forename: Jinhui
authors.initials: J
authors.lastname: Li
authors.forename: Daxiang
authors.initials: D
authors.lastname: Cui
mesh_terms: D000818:Animals; D001483:Base Sequence; D002469:Cell Separation; D002478:Cells, Cultured; D017931:DNA Primers / Q000235:genetics; D050091:Dendrimers / Q000737:chemistry; D005434:Flow Cytometry; D005456:Fluorescent Dyes / Q000008:administration & dosage; D015870:Gene Expression; D005822:Genetic Vectors; D006801:Humans; D057026:Induced Pluripotent Stem Cells / Q000166:cytology* / Q000378:metabolism / Q000637:transplantation; D007621:Karyotyping; D016086:Lentivirus / Q000235:genetics; D008279:Magnetic Resonance Imaging; D058185:Magnetite Nanoparticles / Q000008:administration & dosage / Q000737:chemistry; D051379:Mice; D016688:Mice, Inbred NOD; D016513:Mice, SCID; D008856:Microscopy, Fluorescence; D050997:Nanomedicine; D036103:Nanotechnology; D014157:Transcription Factors / Q000235:genetics; D014162:Transfection; D014183:Transplantation, Heterologous
chemical_list: D017931:DNA Primers; D050091:Dendrimers; D005456:Fluorescent Dyes; D058185:Magnetite Nanoparticles; C104700:PAMAM Starburst; D014157:Transcription Factors