Differentiation of Osteoprogenitor Cells Is Induced by High-Frequency Pulsed Electromagnetic Fields

Craniofacial defect repair is often limited by a finite supply of available autologous tissue (ie, bone) and less than ideal alternatives. Therefore, other methods to produce bony healing must be explored. Several studies have demonstrated that low-frequency pulsed electromagnetic field (PEMF) stimulation (ie, 5–30 Hz) of osteoblasts enhances bone formation. The current study was designed to investigate whether a Food and Drug Administration–approved, high-frequency PEMF–emitting device is capable of inducing osteogenic differentiation of osteoprogenitor cells. Osteoprogenitor cells (commercially available C3H10T1/2 and mouse calvarial) in complete Dulbecco modified Eagle medium were continuously exposed to PEMF stimulation delivered by the ActiPatch at a frequency of 27.1 MHz. Markers of cellular proliferation and early, intermediate, and terminal osteogenic differentiation were measured and compared with unstimulated controls. All experiments were performed in triplicate. High-frequency PEMF stimulation increases alkaline phosphatase activity in both cell lines. In addition, high-frequency PEMF stimulation augments osteopontin and osteocalcin expression as well as mineral nodule formation in C3H10T1/2 cells, indicating late and terminal osteogenic differentiation, respectively. Cellular proliferation, however, was unaffected by high-frequency PEMF stimulation. Mechanistically, high-frequency PEMF-stimulated osteogenic differentiation is associated with elevated mRNA expression levels of osteogenic bone morphogenetic proteins in C3H10T1/2 cells. Our findings suggest that high-frequency PEMF stimulation of osteoprogenitor cells may be explored as an effective tissue engineering strategy to treat critical-size osseous defects of the craniofacial and axial skeleton. AbbreviationsALP, alkaline phosphatase; BMP, bone morphogenetic protein; ERK-1, extracellular signal–regulated kinase 1; iCALs, immortalized calvarial cells; IHC, immunohistochemical; MAP, mitogen-activated protein; MSC, mesenchymal stem cell; OCN, osteocalcin; OPN, osteopontin; p38&agr;, p38-reactivating kinase; PBS, phosphate-buffered saline; PEMF, pulsed electromagnetic field

[1]  T. He,et al.  Epigenetic Regulation of Mesenchymal Stem Cells: A Focus on Osteogenic and Adipogenic Differentiation , 2011, Stem cells international.

[2]  T. He,et al.  Tetrandrine Inhibits Wnt/β-Catenin Signaling and Suppresses Tumor Growth of Human Colorectal Cancer , 2011, Molecular Pharmacology.

[3]  Erik Neovius,et al.  Craniofacial reconstruction with bone and biomaterials: review over the last 11 years. , 2010, Journal of plastic, reconstructive & aesthetic surgery : JPRAS.

[4]  H. Weinans,et al.  Stimulation of osteogenic differentiation in human osteoprogenitor cells by pulsed electromagnetic fields: an in vitro study , 2010, BMC musculoskeletal disorders.

[5]  T. He,et al.  Insulin-like Growth Factor 2 (IGF-2) Potentiates BMP-9-Induced Osteogenic Differentiation and Bone Formation , 2010, Journal of bone and mineral research : the official journal of the American Society for Bone and Mineral Research.

[6]  T. He,et al.  Synergistic Antitumor Effect of the Activated PPARγ and Retinoid Receptors on Human Osteosarcoma , 2010, Clinical Cancer Research.

[7]  T. He,et al.  Activation of RXR and RAR signaling promotes myogenic differentiation of myoblastic C2C12 cells. , 2009, Differentiation; research in biological diversity.

[8]  T. He,et al.  Wnt antagonist SFRP3 inhibits the differentiation of mouse hepatic progenitor cells , 2009, Journal of cellular biochemistry.

[9]  T. He,et al.  BMP‐9‐induced osteogenic differentiation of mesenchymal progenitors requires functional canonical Wnt/β‐catenin signalling , 2009, Journal of cellular and molecular medicine.

[10]  T. He,et al.  A comprehensive analysis of the dual roles of BMPs in regulating adipogenic and osteogenic differentiation of mesenchymal progenitor cells. , 2009, Stem cells and development.

[11]  T. He,et al.  Establishment and characterization of a new highly metastatic human osteosarcoma cell line , 2009, Clinical & Experimental Metastasis.

[12]  T. He,et al.  Hey1 Basic Helix-Loop-Helix Protein Plays an Important Role in Mediating BMP9-induced Osteogenic Differentiation of Mesenchymal Progenitor Cells* , 2009, Journal of Biological Chemistry.

[13]  A. Gosain,et al.  Application-Specific Selection of Biomaterials for Pediatric Craniofacial Reconstruction: Developing a Rational Approach to Guide Clinical Use , 2009, Plastic and reconstructive surgery.

[14]  A. Gosain,et al.  Biomaterials in Craniofacial Surgery: Experimental Studies and Clinical Application , 2009, The Journal of craniofacial surgery.

[15]  G. Barabino,et al.  Pulsed electromagnetic fields enhance BMP‐2 dependent osteoblastic differentiation of human mesenchymal stem cells , 2008, Journal of orthopaedic research : official publication of the Orthopaedic Research Society.

[16]  T. He,et al.  S100A6 Expression and Function in Human Osteosarcoma , 2008, Clinical orthopaedics and related research.

[17]  M. Costa,et al.  The role of electromagnetic stimulation in the management of established non-union of long bone fractures: what is the evidence? , 2008, Injury.

[18]  Walter H. Chang,et al.  Pulsed electromagnetic fields affect osteoblast proliferation and differentiation in bone tissue engineering , 2007, Bioelectromagnetics.

[19]  N. Selvamurugan,et al.  Effects of BMP‐2 and pulsed electromagnetic field (PEMF) on rat primary osteoblastic cell proliferation and gene expression , 2007, Journal of Orthopaedic Research.

[20]  A. Montag,et al.  Distinct roles of bone morphogenetic proteins in osteogenic differentiation of mesenchymal stem cells , 2007, Journal of orthopaedic research : official publication of the Orthopaedic Research Society.

[21]  K. Kinzler,et al.  A protocol for rapid generation of recombinant adenoviruses using the AdEasy system , 2007, Nature Protocols.

[22]  G. Evans,et al.  Current Options in Head and Neck Reconstruction , 2006, Plastic and reconstructive surgery.

[23]  J. Hollinger,et al.  Commercially Available Demineralized Bone Matrix Compositions to Regenerate Calvarial Critical-Sized Bone Defects , 2006, Plastic and reconstructive surgery.

[24]  A. Montag,et al.  CCN1/Cyr61 Is Regulated by the Canonical Wnt Signal and Plays an Important Role in Wnt3A-Induced Osteoblast Differentiation of Mesenchymal Stem Cells , 2006, Molecular and Cellular Biology.

[25]  Y Raulo,et al.  Autogenous Bone Grafts and Bone Substitutes—Tools and Techniques: I. A 20,000-Case Experience in Maxillofacial and Craniofacial Surgery , 2005, Plastic and reconstructive surgery.

[26]  T. He.  Distinct osteogenic activity of BMPs and their orthopaedic applications. , 2005, Journal of musculoskeletal & neuronal interactions.

[27]  James M. Smartt,et al.  Repair of the Immature and Mature Craniofacial Skeleton with a Carbonated Calcium Phosphate Cement: Assessment of Biocompatibility, Osteoconductivity, and Remodeling Capacity , 2005, Plastic and reconstructive surgery.

[28]  A. Montag,et al.  Connective Tissue Growth Factor (CTGF) Is Regulated by Wnt and Bone Morphogenetic Proteins Signaling in Osteoblast Differentiation of Mesenchymal Stem Cells* , 2004, Journal of Biological Chemistry.

[29]  J. Szatkowski,et al.  Characterization of the distinct orthotopic bone-forming activity of 14 BMPs using recombinant adenovirus-mediated gene delivery , 2004, Gene Therapy.

[30]  Wei Jiang,et al.  Inhibitor of DNA Binding/Differentiation Helix-Loop-Helix Proteins Mediate Bone Morphogenetic Protein-induced Osteoblast Differentiation of Mesenchymal Stem Cells* , 2004, Journal of Biological Chemistry.

[31]  Deborah McK Ciombor,et al.  Treatment of nonunions with electric and electromagnetic fields. , 2004, Clinical orthopaedics and related research.

[32]  Hongwei Cheng,et al.  Transcriptional characterization of bone morphogenetic proteins (BMPs)‐mediated osteogenic signaling , 2003, Journal of cellular biochemistry.

[33]  R. Tuan,et al.  Activation of p38 and Smads mediates BMP-2 effects on human trabecular bone-derived osteoblasts. , 2003, Experimental cell research.

[34]  J. Boehm,et al.  p38 MAP kinases: key signalling molecules as therapeutic targets for inflammatory diseases , 2003, Nature Reviews Drug Discovery.

[35]  Baojie Li,et al.  Activation of p38 mitogen-activated protein kinase is required for osteoblast differentiation. , 2003, Endocrinology.

[36]  A. Montag,et al.  Nuclear receptor agonists as potential differentiation therapy agents for human osteosarcoma. , 2002, Clinical cancer research : an official journal of the American Association for Cancer Research.

[37]  G. Xiao,et al.  Bone Morphogenetic Proteins, Extracellular Matrix, and Mitogen‐Activated Protein Kinase Signaling Pathways Are Required for Osteoblast‐Specific Gene Expression and Differentiation in MC3T3‐E1 Cells , 2002, Journal of bone and mineral research : the official journal of the American Society for Bone and Mineral Research.

[38]  Su‐Li Cheng,et al.  Erk Is Essential for Growth, Differentiation, Integrin Expression, and Cell Function in Human Osteoblastic Cells* , 2001, The Journal of Biological Chemistry.

[39]  B. Boyan,et al.  Pulsed electromagnetic field stimulation of MG63 osteoblast‐like cells affects differentiation and local factor production , 2000, Journal of orthopaedic research : official publication of the Orthopaedic Research Society.

[40]  M. Pittenger,et al.  Adult Human Mesenchymal Stem Cell Differentiation to the Osteogenic or Adipogenic Lineage Is Regulated by Mitogen-activated Protein Kinase* , 2000, The Journal of Biological Chemistry.

[41]  A. Gosain,et al.  Biomaterials in the face: benefits and risks. , 1999, The Journal of craniofacial surgery.

[42]  D. Kucharzyk.  A controlled prospective outcome study of implantable electrical stimulation with spinal instrumentation in a high-risk spinal fusion population. , 1999, Spine.

[43]  J. Kanczler,et al.  Pulsed Electromagnetic Fields Simultaneously Induce Osteogenesis and Upregulate Transcription of Bone Morphogenetic Proteins 2 and 4 in Rat Osteoblastsin Vitro , 1998 .

[44]  M. Ishiyama,et al.  A combined assay of cell viability and in vitro cytotoxicity with a highly water-soluble tetrazolium salt, neutral red and crystal violet. , 1996, Biological & pharmaceutical bulletin.

[45]  P. Leboulch,et al.  Reversible immortalization of mammalian cells mediated by retroviral transfer and site-specific recombination. , 1996, Proceedings of the National Academy of Sciences of the United States of America.

[46]  J. Swez,et al.  A mechanism for action of extremely low frequency electromagnetic fields on biological systems. , 1996, Biochemical and biophysical research communications.

[47]  Stephen D. Smith,et al.  Ion resonance electromagnetic field stimulation of fracture healing in rabbits with a fibular ostectomy , 1994, Journal of orthopaedic research : official publication of the Orthopaedic Research Society.

[48]  C. Rubin,et al.  Optimization of electric field parameters for the control of bone remodeling: Exploitation of an indigenous mechanism for the prevention of osteopenia , 1993, Journal of bone and mineral research : the official journal of the American Society for Bone and Mineral Research.

[49]  L. Sargent,et al.  Reconstruction of Internal Orbital Fractures with Vitallium Mesh , 1991, Plastic and reconstructive surgery.

[50]  Sargent La,et al.  Reconstruction of internal orbital fractures with Vitallium mesh. , 1991 .

[51]  C. Bassett,et al.  Bone density changes in osteoporosis‐prone women exposed to pulsed electromagnetic fields (PEMFs) , 1990, Journal of bone and mineral research : the official journal of the American Society for Bone and Mineral Research.

[52]  R. J. Pawluk,et al.  ACCELERATION OF FRACTURE REPAIR BY ELECTROMAGNETIC FIELDS. A SURGICALLY NONINVASIVE METHOD , 1974, Annals of the New York Academy of Sciences.

[53]  D. Savitz,et al.  INTERNATIONAL COMMISSION ON NON-IONIZING RADIATION PROTECTION , 2011 .

[54]  Antonios G Mikos,et al.  Tissue engineering strategies for bone regeneration. , 2005, Advances in biochemical engineering/biotechnology.

[55]  B. Kinney,et al.  Pulsed Electromagnetic Field Therapy in Plastic Surgery , 2007 .

[56]  Jiahuai Han,et al.  Activation and signaling of the p38 MAP kinase pathway , 2005, Cell Research.

[57]  J. Szatkowski,et al.  Osteogenic activity of the fourteen types of human bone morphogenetic proteins (BMPs). , 2003, The Journal of bone and joint surgery. American volume.

[58]  J. Kanczler,et al.  Pulsed electromagnetic fields simultaneously induce osteogenesis and upregulate transcription of bone morphogenetic proteins 2 and 4 in rat osteoblasts in vitro. , 1998, Biochemical and biophysical research communications.