Reversibly immortalized human umbilical cord–derived mesenchymal stem cells (UC‐MSCs) are responsive to BMP9‐induced osteogenic and adipogenic differentiation

Human mesenchymal stem cells (MSCs) are a heterogeneous subset of nonhematopoietic multipotent stromal stem cells and can differentiate into mesodermal lineage, such as adipocytes, osteocytes, and chondrocytes, as well as ectodermal and endodermal lineages. Human umbilical cord (UC) is one of the most promising sources of MSCs. However, the molecular and cellular characteristics of UC‐derived MSCs (UC‐MSCs) require extensive investigations, which are hampered by the limited lifespan and the diminished potency over passages. Here, we used the piggyBac transposon–based simian virus 40 T antigen (SV40T) immortalization system and effectively immortalized UC‐MSCs, yielding the iUC‐MSCs. A vast majority of the immortalized lines are positive for MSC markers but not for hematopoietic markers. The immortalization phenotype of the iUC‐MSCs can be effectively reversed by flippase recombinase–induced the removal of SV40T antigen. While possessing long‐term proliferation capability, the iUC‐MSCs are not tumorigenic in vivo. Upon bone morphogenetic protein 9 (BMP9) stimulation, the iUC‐MSC cells effectively differentiate into osteogenic, chondrogenic, and adipogenic lineages both in vitro and in vivo, which is indistinguishable from that of primary UC‐MSCs, indicating that the immortalized UC‐MSCs possess the characteristics similar to that of their primary counterparts and retain trilineage differentiation potential upon BMP9 stimulation. Therefore, the engineered iUC‐MSCs should be a valuable alternative cell source for studying UC‐MSC biology and their potential utilities in immunotherapies and regenerative medicine.

[1]  T. He,et al.  BMP9 induces osteogenesis and adipogenesis in the immortalized human cranial suture progenitors from the patent sutures of craniosynostosis patients , 2017, Journal of cellular and molecular medicine.

[2]  T. He,et al.  Characterization of retroviral infectivity and superinfection resistance during retrovirus-mediated transduction of mammalian cells , 2017, Gene Therapy.

[3]  T. He,et al.  Engineering the Rapid Adenovirus Production and Amplification (RAPA) Cell Line to Expedite the Generation of Recombinant Adenoviruses , 2017, Cellular Physiology and Biochemistry.

[4]  T. He,et al.  Notch Signaling Augments BMP9-Induced Bone Formation by Promoting the Osteogenesis-Angiogenesis Coupling Process in Mesenchymal Stem Cells (MSCs) , 2017, Cellular Physiology and Biochemistry.

[5]  T. He,et al.  Noncanonical Wnt signaling plays an important role in modulating canonical Wnt-regulated stemness, proliferation and terminal differentiation of hepatic progenitors , 2017, Oncotarget.

[6]  F. Liu,et al.  NEL-Like Molecule-1 (Nell1) Is Regulated by Bone Morphogenetic Protein 9 (BMP9) and Potentiates BMP9-Induced Osteogenic Differentiation at the Expense of Adipogenesis in Mesenchymal Stem Cells , 2017, Cellular Physiology and Biochemistry.

[7]  Rong Zhang,et al.  Therapeutic Effects of Human Umbilical Cord-Derived Mesenchymal Stem Cells in Acute Lung Injury Mice , 2017, Scientific Reports.

[8]  T. He,et al.  A Blockade of IGF Signaling Sensitizes Human Ovarian Cancer Cells to the Anthelmintic Niclosamide-Induced Anti-Proliferative and Anticancer Activities , 2016, Cellular Physiology and Biochemistry.

[9]  Maryam K. Mohammed,et al.  A thermoresponsive polydiolcitrate-gelatin scaffold and delivery system mediates effective bone formation from BMP9-transduced mesenchymal stem cells , 2016, Biomedical materials.

[10]  T. He,et al.  Immortalized Mouse Achilles Tenocytes Demonstrate Long-Term Proliferative Capacity While Retaining Tenogenic Properties. , 2016, Tissue engineering. Part C, Methods.

[11]  Evan M. Farina,et al.  Bone morphogenetic protein 9 (BMP9) induces effective bone formation from reversibly immortalized multipotent adipose-derived (iMAD) mesenchymal stem cells. , 2016, American journal of translational research.

[12]  T. He,et al.  The Anthelmintic Drug Niclosamide Inhibits the Proliferative Activity of Human Osteosarcoma Cells by Targeting Multiple Signal Pathways. , 2015, Current cancer drug targets.

[13]  Yinglin Xia,et al.  TqPCR: A Touchdown qPCR Assay with Significantly Improved Detection Sensitivity and Amplification Efficiency of SYBR Green qPCR , 2015, PloS one.

[14]  T. He,et al.  Reversibly Immortalized Mouse Articular Chondrocytes Acquire Long-Term Proliferative Capability While Retaining Chondrogenic Phenotype , 2015, Cell transplantation.

[15]  T. He,et al.  Sustained high level transgene expression in mammalian cells mediated by the optimized piggyBac transposon system , 2015, Genes & diseases.

[16]  T. He,et al.  Canonical Wnt signaling acts synergistically on BMP9-induced osteo/odontoblastic differentiation of stem cells of dental apical papilla (SCAPs). , 2015, Biomaterials.

[17]  P. Bianco.  "Mesenchymal" stem cells. , 2014, Annual review of cell and developmental biology.

[18]  T. He,et al.  Functional Characteristics of Reversibly Immortalized Hepatic Progenitor Cells Derived from Mouse Embryonic Liver , 2014, Cellular Physiology and Biochemistry.

[19]  X. Chen,et al.  Overexpression of Ad5 precursor terminal protein accelerates recombinant adenovirus packaging and amplification in HEK-293 packaging cells , 2014, Gene Therapy.

[20]  T. He,et al.  Bone morphogenetic protein-9 effectively induces osteo/odontoblastic differentiation of the reversibly immortalized stem cells of dental apical papilla. , 2014, Stem cells and development.

[21]  T. He,et al.  The piggyBac Transposon-Mediated Expression of SV40 T Antigen Efficiently Immortalizes Mouse Embryonic Fibroblasts (MEFs) , 2014, PloS one.

[22]  T. Nagamura-Inoue,et al.  Umbilical cord-derived mesenchymal stem cells: Their advantages and potential clinical utility. , 2014, World journal of stem cells.

[23]  T. He,et al.  Adenovirus-Mediated Gene Transfer in Mesenchymal Stem Cells Can Be Significantly Enhanced by the Cationic Polymer Polybrene , 2014, PloS one.

[24]  王金华,et al.  Bone morphogenetic protein-9 effectively induces osteo/odontoblastic differentiation of the reversibly immortalized stem cells of dental apical papilla , 2014 .

[25]  T. He,et al.  BMP9 signaling in stem cell differentiation and osteogenesis. , 2018, American journal of stem cells.

[26]  T. He,et al.  IGFBP5 domains exert distinct inhibitory effects on the tumorigenicity and metastasis of human osteosarcoma. , 2013, Cancer letters.

[27]  T. He,et al.  Cross-talk between EGF and BMP9 signalling pathways regulates the osteogenic differentiation of mesenchymal stem cells , 2013, Journal of cellular and molecular medicine.

[28]  T. He,et al.  Establishment and Characterization of the Reversibly Immortalized Mouse Fetal Heart Progenitors , 2013, International journal of medical sciences.

[29]  H. Papadaki,et al.  Mesenchymal stem cells derived from Wharton's Jelly of the umbilical cord: biological properties and emerging clinical applications. , 2013, Current stem cell research & therapy.

[30]  C. Fong,et al.  The Therapeutic Potential, Challenges and Future Clinical Directions of Stem Cells from the Wharton’s Jelly of the Human Umbilical Cord , 2012, Stem Cell Reviews and Reports.

[31]  T. He,et al.  Conditional immortalization establishes a repertoire of mouse melanocyte progenitors with distinct melanogenic differentiation potential. , 2012, The Journal of investigative dermatology.

[32]  T. He,et al.  Growth hormone synergizes with BMP9 in osteogenic differentiation by activating the JAK/STAT/IGF1 pathway in murine multilineage cells , 2012, Journal of bone and mineral research : the official journal of the American Society for Bone and Mineral Research.

[33]  T. He,et al.  Conditionally Immortalized Mouse Embryonic Fibroblasts Retain Proliferative Activity without Compromising Multipotent Differentiation Potential , 2012, PloS one.

[34]  J. Qin,et al.  Insulin-like growth factor binding protein 5 suppresses tumor growth and metastasis of human osteosarcoma , 2011, Oncogene.

[35]  Jerome Ritz,et al.  The elusive nature and function of mesenchymal stem cells , 2011, Nature Reviews Molecular Cell Biology.

[36]  Qing-Jun Zhang,et al.  Therapeutic Potentials of Mesenchymal Stem Cells Derived from Human Umbilical Cord , 2011, Stem Cell Reviews and Reports.

[37]  T. He,et al.  Mesenchymal Progenitor Cells and Their Orthopedic Applications: Forging a Path towards Clinical Trials , 2010, Stem cells international.

[38]  Yang Bi,et al.  Mesenchymal stem cells: Molecular characteristics and clinical applications. , 2010, World journal of stem cells.

[39]  T. He,et al.  Pre‐activation of retinoid signaling facilitates neuronal differentiation of mesenchymal stem cells , 2010, Development, growth & differentiation.

[40]  T. He,et al.  Retinoic acid signalling induces the differentiation of mouse fetal liver‐derived hepatic progenitor cells , 2009, Liver international : official journal of the International Association for the Study of the Liver.

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

[42]  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.

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

[44]  M. Weiss,et al.  A comparison of human bone marrow-derived mesenchymal stem cells and human umbilical cord-derived mesenchymal stromal cells for cartilage tissue engineering. , 2009, Tissue engineering. Part A.

[45]  A. Montag,et al.  Osteogenic BMPs promote tumor growth of human osteosarcomas that harbor differentiation defects , 2008, Laboratory Investigation.

[46]  A. Uccelli,et al.  Mesenchymal stem cells in health and disease , 2008, Nature Reviews Immunology.

[47]  M. Weiss,et al.  Concise Review: Wharton's Jelly‐Derived Cells Are a Primitive Stromal Cell Population , 2008, Stem cells.

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

[49]  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.

[50]  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.

[51]  G. Sukhikh,et al.  Mesenchymal Stem Cells , 2002, Bulletin of Experimental Biology and Medicine.