title: Label-free nonlinear optical microscopy detects early markers for osteogenic differentiation of human stem cells.
abstract: Tissue engineering by stem cell differentiation is a novel treatment option for bone regeneration. Most approaches for the detection of osteogenic differentiation are invasive or destructive and not compatible with live cell analysis. Here, non-destructive and label-free approaches of Raman spectroscopy, coherent anti-Stokes Raman scattering (CARS) and second harmonic generation (SHG) microscopy were used to detect and image osteogenic differentiation of human neural crest-derived inferior turbinate stem cells (ITSCs). Combined CARS and SHG microscopy was able to detect markers of osteogenesis within 14 days after osteogenic induction. This process increased during continued differentiation. Furthermore, Raman spectroscopy showed significant increases of the PO4(3-) symmetric stretch vibrations at 959 cm(-1) assigned to calcium hydroxyapatite between days 14 and 21. Additionally, CARS microscopy was able to image calcium hydroxyapatite deposits within 14 days following osteogenic induction, which was confirmed by Alizarin Red-Staining and RT- PCR. Taken together, the multimodal label-free analysis methods Raman spectroscopy, CARS and SHG microscopy can monitor osteogenic differentiation of adult human stem cells into osteoblasts with high sensitivity and spatial resolution in three dimensions. Our findings suggest a great potential of these optical detection methods for clinical applications including in vivo observation of bone tissue-implant-interfaces or disease diagnosis.
authors.affiliation: Cell Biology, University of Bielefeld, D-33501 Bielefeld, Germany.|Biomolecular Photonics, University of Bielefeld, D-33501 Bielefeld, Germany.
authors.forename: Arne D
authors.initials: AD
authors.lastname: Hofemeier
authors.affiliation: Biomolecular Photonics, University of Bielefeld, D-33501 Bielefeld, Germany.
authors.forename: Henning
authors.initials: H
authors.lastname: Hachmeister
authors.affiliation: Biomolecular Photonics, University of Bielefeld, D-33501 Bielefeld, Germany.
authors.forename: Christian
authors.initials: C
authors.lastname: Pilger
authors.affiliation: Cell Biology, University of Bielefeld, D-33501 Bielefeld, Germany.
authors.forename: Matthias
authors.initials: M
authors.lastname: Schürmann
authors.affiliation: Cell Biology, University of Bielefeld, D-33501 Bielefeld, Germany.|Department of Otolaryngology, Head and Neck Surgery, Klinikum Bielefeld, D-33604 Bielefeld, Germany.
authors.forename: Johannes F W
authors.initials: JF
authors.lastname: Greiner
authors.affiliation: Biomolecular Photonics, University of Bielefeld, D-33501 Bielefeld, Germany.
authors.forename: Lena
authors.initials: L
authors.lastname: Nolte
authors.affiliation: Department of Otolaryngology, Head and Neck Surgery, Klinikum Bielefeld, D-33604 Bielefeld, Germany.
authors.forename: Holger
authors.initials: H
authors.lastname: Sudhoff
authors.affiliation: Cell Biology, University of Bielefeld, D-33501 Bielefeld, Germany.
authors.forename: Christian
authors.initials: C
authors.lastname: Kaltschmidt
authors.affiliation: Biomolecular Photonics, University of Bielefeld, D-33501 Bielefeld, Germany.
authors.forename: Thomas
authors.initials: T
authors.lastname: Huser
authors.affiliation: Cell Biology, University of Bielefeld, D-33501 Bielefeld, Germany.|Molecular Neurobiology, University of Bielefeld, D-33501 Bielefeld, Germany.
authors.forename: Barbara
authors.initials: B
authors.lastname: Kaltschmidt
mesh_terms: D000943:Antigens, Differentiation / Q000378:metabolism*; D002454:Cell Differentiation; D017886:Durapatite / Q000378:metabolism*; D006801:Humans; D008853:Microscopy; D010012:Osteogenesis; D013059:Spectrum Analysis, Raman; D013234:Stem Cells / Q000166:cytology / Q000378:metabolism
chemical_list: D000943:Antigens, Differentiation; D017886:Durapatite