Adult Stem Cells in Small Animal Wound Healing Models

  • Allison C. Nauta
  • Geoffrey C. Gurtner
  • Michael T. Longaker
Part of the Methods in Molecular Biology book series (MIMB, volume 1037)


This chapter broadly reviews the use of stem cells as a means to accelerate wound healing, focusing first on the properties of stem cells that make them attractive agents to influence repair, both alone and as vehicles for growth factor delivery. Major stem cell reservoirs are described, including adult, embryonic, and induced pluripotent cell sources, outlining the advantages and limitations of each source as wound healing agents, as well as the possible mechanisms responsible for wound healing acceleration. Finally, the chapter includes a materials and methods section that provides an in-depth description of adult tissue harvest techniques.

Key words

Adult stem cells Adipose derived stromal cells Bone marrow derived mesenchymal cells 


  1. 1.
    Hernandez A, Evers BM (1999) Functional genomics: clinical effect and the evolving role of the surgeon. Arch Surg 134:1209–1215PubMedCrossRefGoogle Scholar
  2. 2.
    Yoder MC, Mead LE, Prater D, Krier TR, Mroueh KN, Li F, Krasich R, Temm CJ, Prchal JT, Ingram DA (2007) Redefining endothelial progenitor cells via clonal analysis and hematopoietic stem/progenitor cell principals. Blood 109:1801–1809PubMedCentralPubMedCrossRefGoogle Scholar
  3. 3.
    Thomson JA, Itskovitz-Eldor J, Shapiro SS, Waknitz MA, Swiergiel JJ, Marshall VS, Jones JM (1998) Embryonic stem cell lines derived from human blastocysts. Science 282:1145–1147PubMedCrossRefGoogle Scholar
  4. 4.
    Evans MJ, Kaufman MH (1981) Establishment in culture of pluripotential cells from mouse embryos. Nature 292:154–156PubMedCrossRefGoogle Scholar
  5. 5.
    Reubinoff BE, Pera MF, Fong CY, Trounson A, Bongso A (2000) Embryonic stem cell lines from human blastocysts: somatic differentiation in vitro. Nat Biotechnol 18:399–404PubMedCrossRefGoogle Scholar
  6. 6.
    Fuegemann CJ, Samraj AK, Walsh S, Fleischmann BK, Jovinge S, Breitbach M (2010) Differentiation of mouse embryonic stem cells into cardiomyocytes via the hanging-drop and mass culture methods. Curr Protoc Stem Cell Biol. Chapter 1, Unit 1F.11Google Scholar
  7. 7.
    Troy TC, Turksen K (2002) ES cell differentiation into the hair follicle lineage in vitro. Methods Mol Biol 185:255–260PubMedGoogle Scholar
  8. 8.
    Sulzbacher S, Schroeder IS, Truong TT, Wobus AM (2009) Activin A-induced differentiation of embryonic stem cells into endoderm and pancreatic progenitors-the influence of differentiation factors and culture conditions. Stem Cell Rev 5:159–173PubMedCrossRefGoogle Scholar
  9. 9.
    Coraux C, Hilmi C, Rouleau M, Spadafora A, Hinnrasky J, Ortonne JP, Dani C, Aberdam D (2003) Reconstituted skin from murine embryonic stem cells. Curr Biol 13:849–853PubMedCrossRefGoogle Scholar
  10. 10.
    Turksen K, Troy TC (1998) Epidermal cell lineage. Biochem Cell Biol 76:889–898PubMedCrossRefGoogle Scholar
  11. 11.
    Segre J (2003) Complex redundancy to build a simple epidermal permeability barrier. Curr Opin Cell Biol 15:776–782PubMedCrossRefGoogle Scholar
  12. 12.
    Troy TC, Turksen K (2005) Commitment of embryonic stem cells to an epidermal cell fate and differentiation in vitro. Dev Dyn 232:293–300PubMedCrossRefGoogle Scholar
  13. 13.
    McGowan KM, Coulombe PA (1998) Onset of keratin 17 expression coincides with the definition of major epithelial lineages during skin development. J Cell Biol 143:469–486PubMedCentralPubMedCrossRefGoogle Scholar
  14. 14.
    Yanai J, Doetchman T, Laufer N, Maslaton J, Mor-Yosef S, Safran A, Shani M, Sofer D (1995) Embryonic cultures but not embryos transplanted to the mouse's brain grow rapidly without immunosuppression. Int J Neurosci 81:21–26PubMedCrossRefGoogle Scholar
  15. 15.
    Glaser T, Perez-Bouza A, Klein K, Brüstle O (2005) Generation of purified oligodendrocyte progenitors from embryonic stem cells. FASEB J 19:112–114PubMedGoogle Scholar
  16. 16.
    Klug MG, Soonpaa MH, Koh GY, Field LJ (1996) Genetically selected cardiomyocytes from differentiating embronic stem cells form stable intracardiac grafts. J Clin Invest 98:216–224PubMedCentralPubMedCrossRefGoogle Scholar
  17. 17.
    Xu C, Inokuma MS, Denham J, Golds K, Kundu P, Gold JD, Carpenter MK (2001) Feeder-free growth of undifferentiated human embryonic stem cells. Nat Biotechnol 19:971–974PubMedCrossRefGoogle Scholar
  18. 18.
    Richards M, Fong CY, Chan WK, Wong PC, Bongso A (2002) Human feeders support prolonged undifferentiated growth of human inner cell masses and embryonic stem cells. Nat Biotechnol 20:933–936PubMedCrossRefGoogle Scholar
  19. 19.
    Amit M, Margulets V, Segev H, Shariki K, Laevsky I, Coleman R, Itskovitz-Eldor J (2003) Human feeder layers for human embryonic stem cells. Biol Reprod 68:2150–2156PubMedCrossRefGoogle Scholar
  20. 20.
    Takahashi K, Yamanaka S (2006) Induction of pluripotent stem cells from mouse embryonic and adult fibroblast cultures by defined factors. Cell 126:663–676PubMedCrossRefGoogle Scholar
  21. 21.
    Takahashi K, Tanabe K, Ohnuki M, Narita M, Ichisaka T, Tomoda K, Yamanaka S (2007) Induction of pluripotent stem cells from adult human fibroblasts by defined factors. Cell 131:861–872PubMedCrossRefGoogle Scholar
  22. 22.
    Kiskinis E, Eggan K (2010) Progress toward the clinical application of patient-specific pluripotent stem cells. J Clin Invest 120:51–59PubMedCentralPubMedCrossRefGoogle Scholar
  23. 23.
    Jia F, Wilson KD, Sun N, Gupta DM, Huang M, Li Z, Panetta NJ, Chen ZY, Robbins RC, Kay MA, Longaker MT, Wu JC (2010) A nonviral minicircle vector for deriving human iPS cells. Nat Methods 7:197–199PubMedCentralPubMedCrossRefGoogle Scholar
  24. 24.
    Lee CH, Kim JH, Lee HJ, Jeon K, Lim H, Choi H, Lee ER, Park SH, Park JY, Hong S, Kim S, Cho SG (2011) The generation of iPS cells using non-viral magnetic nanoparticle based transfection. Biomaterials 32:6683–6691PubMedCrossRefGoogle Scholar
  25. 25.
    Zhou H, Wu S, Joo JY, Zhu S, Han DW, Lin T, Trauger S, Bien G, Yao S, Zhu Y, Siuzdak G, Schöler HR, Duan L, Ding S (2009) Generation of induced pluripotent stem cells using recombinant proteins. Cell Stem Cell 4:381–384PubMedCrossRefGoogle Scholar
  26. 26.
    Mikkelsen TS, Hanna J, Zhang X, Ku M, Wernig M, Schorderet P, Bernstein BE, Jaenisch R, Lander ES, Meissner A (2008) Dissecting direct reprogramming through integrative genomic analysis. Nature 454:49–55PubMedCentralPubMedCrossRefGoogle Scholar
  27. 27.
    Huangfu D, Maehr R, Guo W, Eijkelenboom A, Snitow M, Chen AE, Melton DA (2008) Induction of pluripotent stem cells by defined factors is greatly improved by small-molecule compounds. Nat Biotechnol 26:795–797PubMedCrossRefGoogle Scholar
  28. 28.
    Huangfu D, Osafune K, Maehr R, Guo W, Eijkelenboom A, Chen S, Muhlestein W, Melton DA (2008) Induction of pluripotent stem cells from primary human fibroblasts with only Oct4 and Sox2. Nat Biotechnol 26:1269–1275PubMedCrossRefGoogle Scholar
  29. 29.
    Shi Y, Do JT, Desponts C, Hahm HS, Schöler HR, Ding S (2008) A combined chemical and genetic approach for the generation of induced pluripotent stem cells. Cell Stem Cell 2:525–528PubMedCrossRefGoogle Scholar
  30. 30.
    Lyssiotis CA, Foreman RK, Staerk J, Garcia M, Mathur D, Markoulaki S, Hanna J, Lairson LL, Charette BD, Bouchez LC, Bollong M, Kunick C, Brinker A, Cho CY, Schultz PG, Jaenisch R (2009) Reprogramming of murine fibroblasts to induced pluripotent stem cells with chemical complementation of Klf4. Proc Natl Acad Sci U S A 106:8912–8917PubMedCentralPubMedCrossRefGoogle Scholar
  31. 31.
    Okita K, Ichisaka T, Yamanaka S (2007) Generation of germline-competent induced pluripotent stem cells. Nature 448:313–317PubMedCrossRefGoogle Scholar
  32. 32.
    Vierbuchen T, Ostermeier A, Pang ZP, Kokubu Y, Südhof TC, Wernig M (2010) Direct conversion of fibroblasts to functional neurons by defined factors. Nature 463:1035–1041PubMedCentralPubMedCrossRefGoogle Scholar
  33. 33.
    Yamanaka S (2009) Elite and stochastic models for induced pluripotent stem cell generation. Nature 460:49–52PubMedCrossRefGoogle Scholar
  34. 34.
    Friedenstein AJ, Petrakova KV, Kurolesova AI, Frolova GP (1968) Heterotopic of bone marrow. Analysis of precursor cells for osteogenic and hematopoietic tissues. Transplantation 6:230–247PubMedCrossRefGoogle Scholar
  35. 35.
    Dominici M, Le Blanc K, Mueller I, Slaper-Cortenbach I, Marini F, Krause D, Deans R, Keating A, Prockop D, Horwitz E (2006) Minimal criteria for defining multipotent mesenchymal stromal cells. The International Society for Cellular Therapy position statement. Cytotherapy 8:315–317PubMedCrossRefGoogle Scholar
  36. 36.
    Zuk PA, Zhu M, Mizuno H, Huang J, Futrell JW, Katz AJ, Benhaim P, Lorenz HP, Hedrick MH (2001) Multilineage cells from human adipose tissue: implications for cell-based therapies. Tissue Eng 7:211–228PubMedCrossRefGoogle Scholar
  37. 37.
    Dimmeler S, Zeiher AM (2009) Cell therapy of acute myocardial infarction: open questions. Cardiology 113:155–160PubMedCrossRefGoogle Scholar
  38. 38.
    Wang Y, Hu X, Xie X, He A, Liu X, Wang JA (2011) Effects of mesenchymal stem cells on matrix metalloproteinase synthesis in cardiac fibroblasts. Exp Biol Med (Maywood) 236:1197–1204CrossRefGoogle Scholar
  39. 39.
    Jung H, Kim HH, Lee DH, Hwang YS, Yang HC, Park JC (2011) Transforming growth factor-beta 1 in adipose derived stem cells conditioned medium is a dominant paracrine mediator determines hyaluronic acid and collagen expression profile. Cytotechnology 63:57–66PubMedCentralPubMedCrossRefGoogle Scholar
  40. 40.
    Maherali N, Hochedlinger K (2008) Guidelines and techniques for the generation of induced pluripotent stem cells. Cell Stem Cell 3:595–605PubMedCrossRefGoogle Scholar
  41. 41.
    Xu S, De Becker A, Van Camp B, Vanderkerken K, Van Riet I (2010) An improved harvest and in vitro expansion protocol for murine bone marrow-derived mesenchymal stem cells. J Biomed Biotechnol 2010:105940Google Scholar
  42. 42.
    Lennon DP, Caplan AI (2006) Isolation of rat marrow-derived mesenchymal stem cells. Exp Hematol 34:1606–1607PubMedCrossRefGoogle Scholar
  43. 43.
    Yu G, Wu X, Kilroy G, Halvorsen YD, Gimble JM, Floyd ZE (2011) Isolation of murine adipose-derived stem cells. Methods Mol Biol 702:29–36PubMedCrossRefGoogle Scholar
  44. 44.
    Mitchell JB, McIntosh K, Zvonic S, Garrett S, Floyd ZE, Kloster A, Di Halvorsen Y, Storms RW, Goh B, Kilroy G, Wu X, Gimble JM (2006) Immunophenotype of human adipose-derived cells: temporal changes in stromal-associated and stem cell-associated markers. Stem Cells 24:376–385PubMedCrossRefGoogle Scholar
  45. 45.
    Chioni AM, Grose R (2008) Organotypic modelling as a means of investigating epithelial-stromal interactions during tumourigenesis. Fibrogenesis Tissue Repair 1:8PubMedCentralPubMedCrossRefGoogle Scholar
  46. 46.
    Garlick JA (2007) Engineering skin to study human disease–tissue models for cancer biology and wound repair. Adv Biochem Eng Biotechnol 103:207–239PubMedGoogle Scholar
  47. 47.
    Paus R, Müller-Röver S, Van Der Veen C, Maurer M, Eichmüller S, Ling G, Hofmann U, Foitzik K, Mecklenburg L, Handjiski B (1999) A comprehensive guide for the recognition and classification of distinct stages of hair follicle morphogenesis. J Invest Dermatol 113:523–532PubMedCrossRefGoogle Scholar
  48. 48.
    Porter RM (2003) Mouse models for human hair loss disorders. J Anat 202:125–131PubMedCentralPubMedCrossRefGoogle Scholar
  49. 49.
    Azzi L, El-Alfy M, Martel C, Labrie F (2005) Gender differences in mouse skin morphology and specific effects of sex steroids and dehydroepiandrosterone. J Invest Dermatol 124:22–27PubMedCrossRefGoogle Scholar
  50. 50.
    Galiano RD, Michaels JT, Dobryansky M, Levine JP, Gurtner GC (2004) Quantitative and reproducible murine model of excisional wound healing. Wound Repair Regen 12:485–492PubMedCrossRefGoogle Scholar
  51. 51.
    Wong VW, Sorkin M, Glotzbach JP, Longaker MT, Gurtner GC (2011) Surgical approaches to create murine models of human wound healing. J Biomed Biotechnol 2011:969618.Google Scholar
  52. 52.
    Li F, Huang Q, Chen J, Peng Y, Roop DR, Bedford JS, Li CY (2010) Apoptotic cells activate the “phoenix rising” pathway to promote wound healing and tissue regeneration. Sci Signal 3:ra13PubMedCentralPubMedGoogle Scholar
  53. 53.
    Kim WS, Park BS, Kim HK, Park JS, Kim KJ, Choi JS, Chung SJ, Kim DD, Sung JH (2008) Evidence supporting antioxidant action of adipose-derived stem cells: protection of human dermal fibroblasts from oxidative stress. J Dermatol Sci 49:133–142PubMedCrossRefGoogle Scholar
  54. 54.
    Valle-Prieto A, Conget PA (2010) Human mesenchymal stem cells efficiently manage oxidative stress. Stem Cells Dev 19:1885–1893PubMedCrossRefGoogle Scholar
  55. 55.
    Schäfer M, Werner S (2008) Oxidative stress in normal and impaired wound repair. Pharmacol Res 58:165–171PubMedCrossRefGoogle Scholar
  56. 56.
    Lund AW, Yener B, Stegemann JP, Plopper GE (2009) The natural and engineered 3D microenvironment as a regulatory cue during stem cell fate determination. Tissue Eng Part B Rev 15:371–380PubMedCentralPubMedCrossRefGoogle Scholar
  57. 57.
    Rustad KC, Wong VW, Sorkin M, Glotzbach JP, Major MR, Rajadas J, Longaker MT, Gurtner GC (2011) Enhancement of mesenchymal stem cell angiogenic capacity and stemness by a biomimetic hydrogel scaffold. BiomaterialsGoogle Scholar
  58. 58.
    Dunnwald M, Tomanek-Chalkley A, Alexandrunas D, Fishbaugh J, Bickenbach JR (2001) Isolating a pure population of epidermal stem cells for use in tissue engineering. Exp Dermatol 10:45–54PubMedCrossRefGoogle Scholar
  59. 59.
    Lichti U, Anders J, Yuspa SH (2008) Isolation and short-term culture of primary keratinocytes, hair follicle populations and dermal cells from newborn mice and keratinocytes from adult mice for in vitro analysis and for grafting to immunodeficient mice. Nat Protoc 3:799–810PubMedCrossRefGoogle Scholar
  60. 60.
    Gharzi A, Reynolds AJ, Jahoda CA (2003) Plasticity of hair follicle dermal cells in wound healing and induction. Exp Dermatol 12:126–136PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2013

Authors and Affiliations

  • Allison C. Nauta
    • 1
    • 2
  • Geoffrey C. Gurtner
    • 1
  • Michael T. Longaker
    • 1
    • 2
  1. 1.Hagey Laboratory for Pediatric and Regenerative Medicine, Division of Plastic and Reconstructive Surgery, Department of Surgery, Institute of Stem Cell Biology and Regenerative MedicineStanford University School of MedicineStanfordUSA
  2. 2.Department of SurgeryStanford UniversityStanfordUSA

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