Detection of the osteogenic differentiation of mesenchymal stem cells in 2D and 3D cultures by electrochemical impedance spectroscopy.

Human mesenchymal stem cells are promising candidates for cell-based therapies since they have the capacity to differentiate into a variety of cell types. However, the acceptance of hMSCs for clinical applications as well as in vitro tissue models will depend on strategies for standard characterisations. Impedance spectroscopy is a proven and powerful tool for non-invasive monitoring of cellular processes. The aim of this study was to prove the hypothesis, that the process of osteogenic differentiation can be monitored non-invasively and time-continuously by using impedance spectroscopy. This hypothesis was examined for 2D cell layers of hMSCs by continuous impedance spectroscopy employing a planar electrode-based chip and for 3D aggregates of hMSCs after 21 and 25 days of osteogenic treatment by using a capillary measurement system. The impedance spectra of osteogenic treated hMSCs reported a significant increase of the magnitude of impedance compared to controls cultivated in normal growth medium. The osteogenic status of the cells was determined by alkaline phosphatase expression and von Kossa staining. In respect to that finding it is concluded that impedance spectroscopy is an appropriate method for non-invasive characterisation of osteogenic differentiation of hMSCs, which is relevant for quality control of cell-based implants and cell-based test systems for drug development.

[1]  H. Schwan.  Mechanisms responsible for electrical properties of tissues and cell suspensions. , 1993, Medical progress through technology.

[2]  Sungbo Cho,et al.  Impedance monitoring of herpes simplex virus-induced cytopathic effect in Vero cells , 2007 .

[3]  D. Prockop.  Marrow Stromal Cells as Stem Cells for Nonhematopoietic Tissues , 1997, Science.

[4]  A M Mackay,et al.  Chondrogenic differentiation of cultured human mesenchymal stem cells from marrow. , 1998, Tissue engineering.

[5]  J H Luong,et al.  Monitoring motility, spreading, and mortality of adherent insect cells using an impedance sensor. , 2001, Analytical chemistry.

[6]  S. Takahashi,et al.  A synthetic porous ceramic as a bone graft substitute in the surgical management of scoliosis: a prospective, randomized study. , 2000, Spine.

[7]  Sungbo Cho,et al.  Chip-based time-continuous monitoring of toxic effects on stem cell differentiation. , 2009, Annals of anatomy = Anatomischer Anzeiger : official organ of the Anatomische Gesellschaft.

[8]  Bernard Lachance,et al.  Assessment of cytotoxicity using electric cell-substrate impedance sensing: concentration and time response function approach. , 2002, Analytical chemistry.

[9]  D. Butler,et al.  Use of mesenchymal stem cells in a collagen matrix for achilles tendon repair , 1998, Journal of orthopaedic research : official publication of the Orthopaedic Research Society.

[10]  Maurilio Marcacci,et al.  Stem cells associated with macroporous bioceramics for long bone repair: 6- to 7-year outcome of a pilot clinical study. , 2007, Tissue engineering.

[11]  Ivar Giaever,et al.  A morphological biosensor for mammalian cells , 1993, Nature.

[12]  Su‐Li Cheng,et al.  αvβ Integrins Play an Essential Role in BMP‐2 Induction of Osteoblast Differentiation , 2004 .

[13]  Hagen Thielecke,et al.  3D-biohybrid systems: applications in drug screening. , 2002, Trends in biotechnology.

[14]  K. Satomura,et al.  Repair of craniotomy defects using bone marrow stromal cells. , 1998, Transplantation.

[15]  J. Jansen,et al.  Ectopic bone formation in rat marrow stromal cell/titanium fiber mesh scaffold constructs: effect of initial cell phenotype. , 2005, Biomaterials.

[16]  H. Galla,et al.  Impedance analysis of epithelial and endothelial cell monolayers cultured on gold surfaces. , 1996, Journal of biochemical and biophysical methods.

[17]  P. Ducheyne,et al.  In vivo evaluation of a bioactive scaffold for bone tissue engineering. , 2002, Journal of biomedical materials research.

[18]  Hermann Eichler,et al.  Comparative Analysis of Mesenchymal Stem Cells from Bone Marrow, Umbilical Cord Blood, or Adipose Tissue , 2006, Stem cells.

[19]  K. Kraus,et al.  Mesenchymal stem cells in osteobiology and applied bone regeneration. , 1998, Clinical orthopaedics and related research.

[20]  Sungbo Cho,et al.  Electrical characterization of human mesenchymal stem cell growth on microelectrode , 2008 .

[21]  Robert Plonsey,et al.  Bioelectromagnetism: Principles and Applications of Bioelectric and Biomagnetic Fields , 1995 .

[22]  W. Davros,et al.  Spine Fusion Using Cell Matrix Composites Enriched in Bone Marrow-Derived Cells , 2003, Clinical orthopaedics and related research.

[23]  Joachim Wegener,et al.  Bioelectrical impedance assay to monitor changes in cell shape during apoptosis. , 2004, Biosensors & bioelectronics.

[24]  A. Caplan,et al.  Dexamethasone inhibition of confluence‐induced apoptosis in human mesenchymal stem cells , 2009, Journal of orthopaedic research : official publication of the Orthopaedic Research Society.

[25]  A. Robitzki,et al.  A multicellular spheroid-based sensor for anti-cancer therapeutics. , 2001, Biosensors & bioelectronics.

[26]  A. Robitzki,et al.  A More Aggressive Breast Cancer Spheroid Model Coupled to an Electronic Capillary Sensor System for a High-Content Screening of Cytotoxic Agents in Cancer Therapy: 3-Dimensional In Vitro Tumor Spheroids as a Screening Model , 2005, Journal of biomolecular screening.

[27]  M. Pittenger,et al.  Multilineage potential of adult human mesenchymal stem cells. , 1999, Science.

[28]  N. Jørgensen,et al.  Dexamethasone, BMP-2, and 1,25-dihydroxyvitamin D enhance a more differentiated osteoblast phenotype: validation of an in vitro model for human bone marrow-derived primary osteoblasts , 2004, Steroids.

[29]  L. Muul,et al.  Isolated allogeneic bone marrow-derived mesenchymal cells engraft and stimulate growth in children with osteogenesis imperfecta: Implications for cell therapy of bone , 2002, Proceedings of the National Academy of Sciences of the United States of America.

[30]  I. Martin,et al.  Prefabricated Engineered Bone Flaps: An Experimental Model of Tissue Reconstruction in Plastic Surgery , 1998, Plastic and reconstructive surgery.

[31]  W. Koo,et al.  Clinical responses to bone marrow transplantation in children with severe osteogenesis imperfecta. , 2001, Blood.

[32]  A. Friedenstein,et al.  Stromal stem cells: marrow-derived osteogenic precursors. , 1988, Ciba Foundation symposium.

[33]  R Cancedda,et al.  Repair of large bone defects with the use of autologous bone marrow stromal cells. , 2001, The New England journal of medicine.

[34]  Q. Shang,et al.  Tissue-Engineered Bone Repair of Sheep Cranial Defects with Autologous Bone Marrow Stromal Cells , 2001, The Journal of craniofacial surgery.

[35]  A. Oosterom,et al.  Electrical properties of platinum electrodes: Impedance measurements and time-domain analysis , 2006, Medical and Biological Engineering and Computing.

[36]  A. Osyczka,et al.  Bone morphogenetic protein regulation of early osteoblast genes in human marrow stromal cells is mediated by extracellular signal-regulated kinase and phosphatidylinositol 3-kinase signaling. , 2005, Endocrinology.

[37]  A I Caplan,et al.  In vitro chondrogenesis of bone marrow-derived mesenchymal progenitor cells. , 1998, Experimental cell research.

[38]  P. Byers,et al.  Gene Targeting in Stem Cells from Individuals with Osteogenesis Imperfecta , 2004, Science.

[39]  S. Mann,et al.  An ex vivo model for chondrogenesis and osteogenesis. , 2007, Biomaterials.

[40]  S. Bruder,et al.  Osteogenic differentiation of purified, culture‐expanded human mesenchymal stem cells in vitro , 1997, Journal of cellular biochemistry.

[41]  A Boyde,et al.  Autologous bone marrow stromal cells loaded onto porous hydroxyapatite ceramic accelerate bone repair in critical-size defects of sheep long bones. , 2000, Journal of biomedical materials research.

[42]  Darwin J. Prockop,et al.  Transplantability and therapeutic effects of bone marrow-derived mesenchymal cells in children with osteogenesis imperfecta , 1999, Nature Medicine.

[43]  I. Giaever,et al.  Micromotion of mammalian cells measured electrically. , 1991, Proceedings of the National Academy of Sciences of the United States of America.

[44]  Thomas Braschler,et al.  Two-dimensional impedance imaging of cell migration and epithelial stratification. , 2006, Lab on a chip.