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Genetic Modification of Mesenchymal Stem Cells

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Somatic Stem Cells

Part of the book series: Methods in Molecular Biology ((MIMB,volume 879))

Abstract

Mesenchymal stem cells (MSC) are currently considered the most promising type of adult stem cells for therapeutic applications, because they can be easily isolated from the bone marrow and other tissues, and manipulated for different applications. The genetic transformation of MSC using genes that enhance their homing ability, as well as their proliferation and survival capacities when transplanted to sites of injury, is an important alternative to improve MSC function, especially for tissue regeneration. This chapter describes protocols for the transformation of MSC using plasmid vectors by lipofection and electroporation, as well as retroviral vectors representing viral transformations.

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References

  1. da Silva Meirelles L, Nardi NB (2009) Methodology, biology and clinical applications of mesenchymal stem cells. Front Biosci 14:4281–4298

    Article  Google Scholar 

  2. da Silva Meirelles L, Chagastelles PC, Nardi NB (2006) Mesenchymal stem cells reside in virtually all post-natal organs and tissues. J Cell Sci 119:2204–2213

    Article  PubMed  Google Scholar 

  3. Huang NF, Li S (2008) Mesenchymal stem cells for vascular regeneration. Regen Med 3:877–892

    Article  PubMed  Google Scholar 

  4. Montzka K, Lassonczyk N, Tschöke B, Neuss S, Führmann T, Franzen R et al (2009) Neural differentiation potential of human bone marrow-derived mesenchymal stromal cells: misleading marker gene expression. BMC Neurosci 10:16

    Article  PubMed  Google Scholar 

  5. Orciani M, Morabito C, Emanuelli M, Guarnieri S, Sartini D, Giannubilo SR et al (2011) Neurogenic potential of mesenchymal-like stem cells from human amniotic fluid: the influence of extracellular growth factors. J Biol Regul Homeost Agents 25:115–130

    PubMed  CAS  Google Scholar 

  6. Snykers S, De Kock J, Tamara V, Rogiers V (2011) Hepatic differentiation of mesenchymal stem cells: in vitro strategies. Methods Mol Biol 698:305–314

    Article  PubMed  CAS  Google Scholar 

  7. Iyer SS, Rojas M (2008) Anti-inflammatory effects of mesenchymal stem cells: novel concept for future therapies. Expert Opin Biol Ther 8:569–581

    Article  PubMed  CAS  Google Scholar 

  8. Schenk S, Mal N, Finan A, Zhang M, Kiedrowski M, Popovic Z et al (2007) Monocyte chemotactic protein-3 is a myocardial mesenchymal stem cell homing factor. Stem Cells 25:245–251

    Article  PubMed  CAS  Google Scholar 

  9. Giordano A, Galderisi U, Marino IR (2007) From the laboratory bench to the patient’s bedside: an update on clinical trials with mesenchymal stem cells. J Cell Physiol 211:27–35

    Article  PubMed  CAS  Google Scholar 

  10. Dimmeler S, Leri A (2008) Aging and disease as modifiers of efficacy of cell therapy. Circ Res 102:1319–1330

    Article  PubMed  CAS  Google Scholar 

  11. Madeira C, Mendes RD, Ribeiro SC, Boura JS, Aires-Barros MR, da Silva CL et al (2010) Nonviral gene delivery to mesenchymal stem cells using cationic liposomes for gene and cell therapy. J Biomed Biotechnol 2010:735349

    Article  PubMed  CAS  Google Scholar 

  12. Ricks DM, Kutner R, Zhang X-Y, Welsh DA, Reiser J (2008) Optimized lentiviral transduction of mouse bone marrow-derived mesenchymal stem cells. Stem Cells Dev 17:441–450

    Article  PubMed  CAS  Google Scholar 

  13. Young RL, Shinojima N, Fueyo J, Gumin J, Vecil GG, Marini FC et al (2009) Human bone marrow-derived mesenchymal stem cells for intravascular delivery of oncolytic adenovirus delta-24-RGD to human gliomas. Cancer Res 69:8932–8940

    Article  Google Scholar 

  14. Kim JH, Park SN, Suh H (2007) Generation of insulin-producing human mesenchymal stem cells using recombinant adeno-associated virus. Yonsei Med J 48:109–119

    Article  PubMed  CAS  Google Scholar 

  15. Yang J, Tang T, Li F, Zhou W, Liu J, Tan Z et al (2009) Experimental study of the effects of marrow mesenchymal stem cells transfected with hypoxia-inducible factor-1a gene. J Biomed Biotechnol 2009:128627

    PubMed  Google Scholar 

  16. Gehl J (2003) Electroporation: theory and methods, perspectives for drug delivery, gene therapy and research. Acta Physiol Scand 177:437–447

    Article  PubMed  CAS  Google Scholar 

  17. Andreason GL, Evans GA (1989) Optimization of electroporation for transfection of mammalian cell lines. Anal Biochem 180:269–275

    Article  PubMed  CAS  Google Scholar 

  18. Trompeter HI, Weinhold S, Thiel C, Wernet P, Uhrberg M (2003) Rapid and highly efficient gene transfer into natural killer cells by nucleofection. J Immunol Methods 274:245–256

    Article  PubMed  CAS  Google Scholar 

  19. Ohta S, Suzuki K, Ogino Y, Miyagawa S, Murashima A, Matsumaru D et al (2008) Gene transduction by sonoporation. Dev Growth Differ 50:517–520

    Article  PubMed  CAS  Google Scholar 

  20. Ziv R, Steinhardt Y, Pelled G, Gazit D, Rubinsky B (2009) Micro-electroporation of mesenchymal stem cells with alternating electrical current pulses. Biomed Microdevices 11:95–101

    Article  PubMed  Google Scholar 

  21. Aiuti A, Bachoud-Lévi AC, Blesch A, Brenner MK, Cattaneo F, Chiocca EA et al (2007) Progress and prospects: gene therapy clinical trials (part 2). Gene Ther 14:1555–1563

    Article  PubMed  Google Scholar 

  22. Alexander BL, Ali RR, Alton EW, Bainbridge JW, Braun S, Cheng SH et al (2007) Progress and prospects: gene therapy clinical trials (part 1). Gene Ther 14:1439–1447

    Article  PubMed  CAS  Google Scholar 

  23. Bushman FD (2007) Retroviral integration and human gene therapy. J Clin Invest 117: 2083–2086

    Article  PubMed  CAS  Google Scholar 

  24. Zentilin L, Qin G, Tafuro S, Dinauer MC, Baum C, Giacca M (2000) Variegation of retroviral vector gene expression in myeloid cells. Gene Ther 7:153–166

    Article  PubMed  CAS  Google Scholar 

  25. Li CL, Emery DW (2008) The cHS4 chromatin insulator reduces gammaretroviral vector silencing by epigenetic modifications of integrated provirus. Gene Ther 15:49–53

    Article  PubMed  Google Scholar 

  26. Nienhuis AW (2006) Assays to evaluate the genotoxicity of retroviral vectors. Mol Ther 14:459–460

    Article  PubMed  CAS  Google Scholar 

  27. Roelants V, Labar D, de Meester C, Havaux X, Tabilio A, Gambhir SS et al (2008) Comparison between adenoviral and retroviral vectors for the transduction of the thymidine kinase PET reporter gene in rat mesenchymal stem cells. J Nucl Med 49:1836–1844

    Article  PubMed  CAS  Google Scholar 

  28. Gnecchi M, Melo LG (2009) Bone marrow-derived mesenchymal stem cells: isolation, expansion, characterization, viral transduction, and production of conditioned medium. Methods Mol Biol 482:281–294

    Article  PubMed  CAS  Google Scholar 

  29. Nardi NB, Camassola M (2011) Isolation and culture of rodent bone marrow-derived multipotent mesenchymal stromal cells. Methods Mol Biol 698:151–160

    Article  PubMed  CAS  Google Scholar 

  30. Beyer Nardi N, da Silva Meirelles L (2006) Mesenchymal stem cells: isolation, in vitro expansion and characterization. Handb Exp Pharmacol 174:249–282

    Google Scholar 

  31. Helledie T, Nurcombe V, Cool SM (2008) A simple and reliable electroporation method for human bone marrow mesenchymal stem cells. Stem Cells Dev 17:837–848

    Article  PubMed  CAS  Google Scholar 

  32. Yalvac ME, Ramazanoglu M, Gumru OZ, Sahin F, Palotás A, Rizvanov AA (2009) Comparison and optimisation of transfection of human dental follicle cells, a novel source of stem cells, with different chemical methods and electroporation. Neurochem Res 34:1272–1277

    Article  PubMed  CAS  Google Scholar 

  33. Colleoni S, Donofrio G, Lagutina I, Duchi R, Galli C, Lazzari G (2005) Establishment, differentiation, electroporation, viral transduction, and nuclear transfer of bovine and porcine mesenchymal stem cells. Cloning Stem Cells 7:154–166

    Article  PubMed  CAS  Google Scholar 

  34. Friedman R, Betancur M, Boissel L, Tuncer H, Cetrulo C, Klingemann H (2007) Umbilical cord mesenchymal stem cells: adjuvants for human cell transplantation. Biol Blood Marrow Transplant 13:1477–1486

    Article  PubMed  Google Scholar 

  35. Jin EJ, Lee SY, Jung JC, Bang OS, Kang SS (2008) TGF-beta3 inhibits chondrogenesis of cultured chick leg bud mesenchymal cells via downregulation of connexin 43 and integrin beta4. J Cell Physiol 214:345–353

    Article  PubMed  CAS  Google Scholar 

  36. Jin EJ, Choi YA, Sonn JK, Kang SS (2007) Suppression of ADAM 10-induced delta-1 shedding inhibits cell proliferation during the chondro-inhibitory action of TGF-beta3. Mol Cells 24:139–147

    PubMed  CAS  Google Scholar 

  37. Choi YA, Kim DK, Kang SS, Sonn JK, Jin EJ (2009) Integrin signaling and cell spreading alterations by rottlerin treatment of chick limb bud mesenchymal cells. Biochimie 91:624–631

    Article  PubMed  CAS  Google Scholar 

  38. Yao S, Rana S, Liu D, Wise GE (2009) Electroporation optimization to deliver plasmid DNA into dental follicle cells. Biotechnol J 4:1488–1496

    Article  PubMed  CAS  Google Scholar 

  39. Potapova I, Plotnikov A, Lu Z, Danilo P Jr, Valiunas V, Qu J et al (2004) Human mesenchymal stem cells as a gene delivery system to create cardiac pacemakers. Circ Res 94:952–959

    Article  PubMed  CAS  Google Scholar 

  40. Aluigi M, Fogli M, Curti A, Isidori A, Gruppioni E, Chiodoni C et al (2006) Nucleofection is an efficient nonviral transfection technique for human bone marrow-derived mesenchymal stem cells. Stem Cells 24:454–461

    Article  PubMed  Google Scholar 

  41. Mok PL, Cheong SK, Leong CF, Othman A (2008) In vitro expression of erythropoietin by transfected human mesenchymal stromal cells. Cytotherapy 10:116–124

    Article  PubMed  CAS  Google Scholar 

  42. Haleem-Smith H, Derfoul A, Okafor C, Tuli R, Olsen D, Hall DJ, Tuan RS (2005) Optimization of high-efficiency transfection of adult human mesenchymal stem cells in vitro. Mol Biotechnol 30:9–20

    Article  PubMed  CAS  Google Scholar 

  43. Wang QW, Chen ZL, Piao YJ (2005) Mesenchymal stem cells differentiate into tenocytes by bone morphogenetic protein (BMP) 12 gene transfer. J Biosci Bioeng 100:418–422

    Article  PubMed  CAS  Google Scholar 

  44. Schambach A, Galla M, Modlich U, Will E, Chandra S, Reeves L et al (2006) Lentiviral vectors pseudotyped with murine ecotropic envelope: increased biosafety and convenience in preclinical research. Exp Hematol 34:588–592

    Article  PubMed  CAS  Google Scholar 

  45. Pear WS, Nolan GP, Scott ML, Baltimore D (1993) Production of high-titer helper-free retroviruses by transient transfection. Proc Natl Acad Sci U S A 90:8392–8396

    Article  PubMed  CAS  Google Scholar 

  46. Tiscornia G, Singer O, Verma IM (2006) Production and purification of lentiviral vectors. Nat Protoc 1:241–245

    Article  PubMed  CAS  Google Scholar 

  47. Flasshove M, Banerjee D, Mineishi S, Li MX, Bertino JR, Moore MA (1995) Ex vivo expansion and selection of human CD34+ peripheral blood progenitor cells after introduction of a mutated dihydrofolate reductase cDNA via retroviral gene transfer. Blood 85:566–574

    PubMed  CAS  Google Scholar 

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Acknowledgments

The authors would like to thank Drs. Giorgio Palù, Claudia Del Vecchio and Sang Won Han, whose work was important for the construction of this knowledge. The artwork of Pedro Cesar Chagastelles is gratefully acknowledged. This work was supported by Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq), Fundação de Amparo à Pesquisa do Estado do Rio Grande do Sul (FAPERGS), Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP), and Instituto Nacional de Ciencia e Tecnologia-INCT-NanoBiofar.

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Authors and Affiliations

  1. Laboratório de Cardiologia Molecular e Celular, Instituto de Cardiologia do Rio Grande do Sul, Porto Alegre, RS, Brazil

    Andréia Escosteguy Vargas & Melissa Medeiros Markoski

  2. Universidade Federal do Pampa, São Gabriel, RS, Brazil

    Andrés Delgado Cañedo

  3. Grupo Fleury, São Paulo, SP, Brazil

    Flávia Helena da Silva

  4. Universidade Luterana do Brasil, Canoas, RS, Brazil

    Nance Beyer Nardi

  5. Instituto de Cardiologia do Rio Grande do Sul, Porto Alegre, RS, Brazil

    Nance Beyer Nardi

Authors
  1. Andréia Escosteguy Vargas
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  2. Melissa Medeiros Markoski
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  3. Andrés Delgado Cañedo
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  4. Flávia Helena da Silva
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  5. Nance Beyer Nardi
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Corresponding author

Correspondence to Nance Beyer Nardi .

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Editors and Affiliations

  1. Mouse Cancer Genetics Program, National Cancer Institute, Center Drive 31, Frederick, 21702, Maryland, USA

    Shree Ram Singh

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Vargas, A.E., Markoski, M.M., Cañedo, A.D., da Silva, F.H., Nardi, N.B. (2012). Genetic Modification of Mesenchymal Stem Cells. In: Singh, S. (eds) Somatic Stem Cells. Methods in Molecular Biology, vol 879. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-61779-815-3_29

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  • DOI: https://doi.org/10.1007/978-1-61779-815-3_29

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  • Publisher Name: Humana Press, Totowa, NJ

  • Print ISBN: 978-1-61779-814-6

  • Online ISBN: 978-1-61779-815-3

  • eBook Packages: Springer Protocols

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