title: Lentiviral transduction and subsequent loading with nanoparticles do not affect cell viability and proliferation in hair-follicle-bulge-derived stem cells in vitro.
abstract: The application of stem cells in the treatment of various degenerative diseases is highly promising. However, cell-based therapy could be limited by the problem of low viability of grafted cells and uncertainty about their fate. The combination of molecular imaging and contrast-enhanced MRI may give more insight into the survival and behavior of grafted stem cells. We explore hair-follicle-bulge-derived stem cells (HFBSCs) as a potential candidate for autologous cell-based therapy. HFBSCs are transduced with a lentiviral construct with genes coding for bioluminescent (Luc2) and fluorescent (copGFP) reporter proteins, and subsequently loaded with magnetic nanoparticles to enable MRI visualization. Thus, we investigate for the first time if lentiviral transduction and cellular loading with nanoparticles have a cytotoxic effect upon these stem cells. Transduction efficiency, proliferation rate, cell viability and reporter protein co-expression during long-term culture of transduced HFBSCs were studied using fluorescence and bioluminescence microscopy. In addition, the effect of TMSR50 nanoparticles on proliferation and viability was investigated using the MTS assay and bioluminescence microscopy. The amount of TMSR50-loaded HFBSCs needed to reach signal threshold for MRI was assessed using an agarose phantom. Transduction with the Luc2-copGFP construct did not influence senescence, proliferation, doubling time, and differentiation of the HFBSCs. CopGFP expression was visible immediately after transduction and persisted for at least 15 passages, concomitantly with Luc2 expression. Cellular loading with TMSR50 nanoparticles did not affect cell viability and proliferation. The results imply that combined MRI and bioluminescence imaging may enable in vivo localization and long-term monitoring of grafted viable HFBSCs. Copyright © 2016 John Wiley & Sons, Ltd.
authors.affiliation: Auditory Neurobiology Laboratory, Department of Otorhinolaryngology and Head & Neck Surgery, Leiden University Medical Center, 2333 ZA, Leiden, The Netherlands.
authors.forename: Timo
authors.identifier: 0000-0002-0948-6209
authors.initials: T
authors.lastname: Schomann
authors.affiliation: Optical Molecular Imaging, Department of Radiology, Erasmus Medical Center, 3015 CE, Rotterdam, The Netherlands.
authors.forename: Laura
authors.initials: L
authors.lastname: Mezzanotte
authors.affiliation: Auditory Neurobiology Laboratory, Department of Otorhinolaryngology and Head & Neck Surgery, Leiden University Medical Center, 2333 ZA, Leiden, The Netherlands.
authors.forename: Ierry-Ann-Lym M
authors.initials: IM
authors.lastname: Lourens
authors.affiliation: Auditory Neurobiology Laboratory, Department of Otorhinolaryngology and Head & Neck Surgery, Leiden University Medical Center, 2333 ZA, Leiden, The Netherlands.
authors.forename: John C M J
authors.initials: JC
authors.lastname: de Groot
authors.affiliation: Auditory Neurobiology Laboratory, Department of Otorhinolaryngology and Head & Neck Surgery, Leiden University Medical Center, 2333 ZA, Leiden, The Netherlands.
authors.forename: Johan H M
authors.initials: JH
authors.lastname: Frijns
authors.affiliation: Auditory Neurobiology Laboratory, Department of Otorhinolaryngology and Head & Neck Surgery, Leiden University Medical Center, 2333 ZA, Leiden, The Netherlands.
authors.forename: Margriet A
authors.initials: MA
authors.lastname: Huisman
mesh_terms: D000818:Animals; D049109:Cell Proliferation; D002470:Cell Survival; D017930:Genes, Reporter; D006085:Graft Survival; D049452:Green Fluorescent Proteins / Q000235:genetics; D018859:Hair Follicle / Q000166:cytology; D008279:Magnetic Resonance Imaging / Q000379:methods*; D008280:Magnetics; D051379:Mice; D057054:Molecular Imaging / Q000379:methods*; D053758:Nanoparticles / Q000737:chemistry; D033581:Stem Cell Transplantation; D013234:Stem Cells / Q000737:chemistry / Q000166:cytology* / Q000378:metabolism; D014161:Transduction, Genetic / Q000379:methods
chemical_list: D049452:Green Fluorescent Proteins
keywords: bioluminescence; fluorescence; hair follicle; magnetic nanoparticles; magnetic resonance imaging; stem cells