Journal of Pharmaceutical Investigation

, Volume 48, Issue 2, pp 215–223 | Cite as

Artificial skin models for animal-free testing

  • Ye Eun Yun
  • Youn Jae Jung
  • Yeo Jin Choi
  • Ji Suk Choi
  • Yong Woo ChoEmail author


Since 2013, as ethics for animal experiments has been strengthened in the European Union, artificial skin models have attracted attention as an alternative to animal testing for assessing the safety and toxicity of products. Driven by regulatory authorities and industry demands, various artificial skin models have been developed by combining various biomaterials and human cells as well as using a variety of techniques, including freeze-drying, 3-D printing, electrospinning, and microfluidic system. Elaborately designed artificial skin models which closely mimic the human skin can be highly valuable and effective tools to replace in vivo animal tests for the evaluation of the safety and efficacy in the field of cosmetic and pharmaceutical industries as well as for the basic studies on cell to cell interactions, cell to extracellular matrix interactions, tissue formation and development. This review recapitulates diverse fabrication techniques for artificial skin models and their main applications.


Artificial skin models Animal alternatives Safety evaluation Toxicity Animal-free testing 



This research was supported by the Ministry of Trade, Industry and Energy (MOTIE, Korea) under Industrial Technology Innovation Program (Grant No.10062127). This research was also supported by the Basic Science Research Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Science and ICT & Future Planning (NRF 20100027955 and NRF-2016R1C1B1006882).

Compliance with ethical standards

Conflict of interest

All authors (Y. E. Yun, Y. J. Jung, Y. J. Choi, J. S. Choi, and Y. W. Cho) declare that they have no conflict of interest.


  1. Abd E, Yousef SA, Pastore MN, Telaprolu K, Mohammed YH, Namjoshi S, Grice JE, Roberts MS (2016) Skin models for the testing of transdermal drugs. Clin Pharmacol 8:163–176PubMedPubMedCentralGoogle Scholar
  2. Ackermann K, Borgia SL, Korting HC, Mewes KR, Schafer-Korting M (2010) The phenion full-thickness skin model for percutaneous absorption testing. Skin Pharmacol Physiol 23(2):105–112PubMedCrossRefGoogle Scholar
  3. Ahadian S, Civitarese R, Bannerman D, Mohammadi MH, Lu R, Wang E, Davenport-Huyer L, Lai B, Zhang B, Zhao Y, Mandla S, Korolj A, Radisic M (2018) Organ-On-A-Chip platforms: a convergence of advanced materials, cells, and microscale technologies. Adv Healthc Mater 7(2)Google Scholar
  4. Ahn SH, Yoon H, Kim GH, Kin YY, Lee SH, Chun W (2010) Designed three-deimensional collagen scaffolds for skin tissue regeneration. Tissue Eng C 16(5):813–821CrossRefGoogle Scholar
  5. Atac B, Wagner I, Horland R, Lauster R, Marx U, Tonevitsky AG, Azar RP, Lindner G (2013) Skin and hair on-a-chip: in vitro skin models versus ex vivo tissue maintenance with dynamic perfusion. Lab Chip 13(18):3555–3561PubMedCrossRefGoogle Scholar
  6. Bernard G, Auger M, Soucy J, Pouliot R (2007) Physical characterization of the stratum corneum of an in vitro psoriatic skin model by ATR-FTIR and Raman spectroscopies. Biochim Biophys Acta 1770(9):1317–1323PubMedCrossRefGoogle Scholar
  7. Brohem CA, Cardeal LB, Tiago M, Soengas MS, Barros SB, Maria-Engler SS (2011) Artificial skin in perspective: concepts and applications. Pigment Cell Melanoma Res 24(1):35–50PubMedCrossRefGoogle Scholar
  8. Cheluvappa R, Scowen P, Eri R (2017) Ethics of animal research in human disease remediation, its institutional teaching; and alternatives to animal experimentation. Pharmacol Res Perspect 5(4):e00332PubMedCentralCrossRefGoogle Scholar
  9. Chen H, Peng Y, Wu S, Tan LP (2016) Electrospun 3D fibrous scaffolds for chronic wound repair. Materials (Basel) 9(4):272CrossRefGoogle Scholar
  10. Cubo N, Garcia M, Del Canizo JF, Velasco D, Jorcano JL (2016) 3D bioprinting of functional human skin: production and in vivo analysis. Biofabrication 9(1):015006PubMedCrossRefGoogle Scholar
  11. Do AV, Khorsand B, Geary SM, Salem AK (2015) 3D printing of scaffolds for tissue regeneration applications. Adv Healthc Mater 4(12):1742–1762PubMedPubMedCentralCrossRefGoogle Scholar
  12. Driskell RR, Lichtenberger BM, Hoste E, Kretzschmar K, Simons BD, Charalambous M, Ferron SR, Herault Y, Pavlovic G, Ferguson-Smith AC, Watt FM (2013) Distinct fibroblast lineages determine dermal architecture in skin development and repair. Nature 504(7479):277–281PubMedPubMedCentralCrossRefGoogle Scholar
  13. Flaten GE, Palac Z, Engesland A, Filipovic-Grcic J, Vanic Z, Skalko-Basnet N (2015) In vitro skin models as a tool in optimization of drug formulation. Eur J Pharm Sci 75:10–24PubMedCrossRefGoogle Scholar
  14. Fu L, Xie JW, Carlson MA, Reilly DA (2017) Three-dimensional nanofiber scaffolds with arrayed holes for engineering skin tissue constructs. MRS Commun 7(03):361–366CrossRefGoogle Scholar
  15. Gangatirkar P, Paquet-Fifield S, Li A, Rossi R, Kaur P (2007) Establishment of 3D organotypic cultures using human neonatal epidermal cells. Nat Protoc 2(1):178–186PubMedCrossRefGoogle Scholar
  16. Garland MJ, Migalska K, Tuan-Mahmood TM, Singh TRR, Majithija R, Caffarel-Salvador E, McCrudden CM, McCarthy HO, Woolfson AD, Donnelly RF (2012) Influence of skin model on in vitro performance of drug-loaded soluble microneedle arrays. Int J Pharm 434(1–2):80–89PubMedCrossRefGoogle Scholar
  17. Groeber F, Holeiter M, Hampel M, Hinderer S, Schenke-Layland K (2011) Skin tissue engineering—in vivo and in vitro applications. Adv Drug Deliv Rev 63(4–5):352–366PubMedCrossRefGoogle Scholar
  18. Haslik W, Kamolz LP, Nathschlager G, Andel H, Meissl G, Frey M (2007) First experiences with the collagen-elastin matrix Matriderm as a dermal substitute in severe burn injuries of the hand. Burns 33(3):364–368PubMedCrossRefGoogle Scholar
  19. Haslik W, Kamolz LP, Manna F, Hladik M, Rath T, Frey M (2010) Management of full-thickness skin defects in the hand and wrist region: first long-term experiences with the dermal matrix Matriderm. J Plast Reconstr Aesthet Surg 63(2):360–364PubMedCrossRefGoogle Scholar
  20. Hilmi AB, Halim AS, Hassan A, Lim CK, Noorsal K, Zainol I (2013) In vitro characterization of a chitosan skin regenerating template as a scaffold for cells cultivation. SpringerPlus 2(79):1–9Google Scholar
  21. Hussain SH, Limthongkul B, Humphreys TR (2013) The biomechanical properties of the skin. Dermatol Surg 39(2):193–203PubMedCrossRefGoogle Scholar
  22. Kempf M, Miyamura Y, Liu PY, Chen AC, Nakamura H, Shimizu H, Tabata Y, Kimble RM, McMillan JR (2011) A denatured collagen microfiber scaffold seeded with human fibroblasts and keratinocytes for skin grafting. Biomaterials 32(21):4782–4792PubMedCrossRefGoogle Scholar
  23. Khorshidi S, Solouk A, Mirzadeh H, Mazinani S, Lagaron JM, Sharifi S, Ramakrishna S (2016) A review of key challenges of electrospun scaffolds for tissue-engineering applications. J Tissue Eng Regen Med 10(9):715–738PubMedCrossRefGoogle Scholar
  24. Killat J, Reimers K, Choi CY, Jahn S, Vogt PM, Radtke C (2013) Cultivation of keratinocytes and fibroblasts in a three-dimensional bovine collagen-elastin matrix (Matriderm(R)) and application for full thickness wound coverage in vivo. Int J Mol Sci 14(7):14460–14474PubMedPubMedCentralCrossRefGoogle Scholar
  25. Kim SH, Nam YS, Lee TS, Park WH (2003) Silk fibroin nanofiber electrospinning, properties, and structure. Polym J 35:185–190CrossRefGoogle Scholar
  26. Kljenak A (2016) Fibrin gel as a scaffold for skin substitute—production and clinical experience. Acta Clin Croat 55:279–289PubMedCrossRefGoogle Scholar
  27. Koch L, Kuhn S, Sorg H, Gruene N, Schlie S, Gaebel R, Polchow B, Reimers K, Stoelting S, Ma N, Vogt PM, Steinhoff G, Chichkov B (2010) Laser printing of skin cells and human stem cells. Tissue Eng C 16(5):847–854CrossRefGoogle Scholar
  28. Koch L, Deiwick A, Schlie S, Michael S, Gruene M, Coger V, Zychlinski D, Schambach A, Reimers K, Vogt PM, Chichkov B (2012) Skin tissue generation by laser cell printing. Biotechnol Bioeng 109(7):1855–1863PubMedCrossRefGoogle Scholar
  29. Kuchler S, Henkes D, Eckl KM, Ackermann K, Plendl J, Korting HC, Hennies HC, Korting HC (2011) Hallmarks of atopic skin mimicked in vitro by means of a skin disease model based on FLG knock-down. Altern Lab Anim 39:471–480PubMedGoogle Scholar
  30. Lee S, Sung J (2018) Organ-on-a-Chip technology for reproducing multiorgan physiology. Adv Healthc Mater. PubMedCrossRefGoogle Scholar
  31. Lee SB, Kim YH, Chong MS, Hong SH, Lee YM (2005) Study of gelatin-containing artificial skin V: fabrication of gelatin scaffolds using a salt-leaching method. Biomaterials 26(14):1961–1968PubMedCrossRefGoogle Scholar
  32. Lee W, Debasitis JC, Lee VK, Lee JH, Fischer K, Edminster K, Park JK, Yoo SS (2009) Multi-layered culture of human skin fibroblasts and keratinocytes through three-dimensional freeform fabrication. Biomaterials 30(8):1587–1595PubMedCrossRefGoogle Scholar
  33. Lee OJ, Ju HW, Kim JH, Lee JM, Ki CS, Kim JH, Moon BM, Park HJ, Sheikh FA, Park CH (2014a) Development of artificial dermis using 3D electrospun silk fibroin nanofiber matrix. J Biomed Nanotechnol 10(7):1294–1303PubMedCrossRefGoogle Scholar
  34. Lee V, Singh G, Trasatti JP, Bjornsson C, Xu X, Tran TN, Yoo SS, Dai G, Karande P (2014b) Design and fabrication of human skin by three-dimensional bioprinting. Tissue Eng C 20(6):473–484CrossRefGoogle Scholar
  35. Lee S, Jin SP, Kim YK, Sung GY, Chung JH, Sung JH (2017) Construction of 3D multicellular microfluidic chip for an in vitro skin model. Biomed Microdevices 19(2):22PubMedCrossRefGoogle Scholar
  36. Leiros GJ, Kusinsky AG, Drago H, Bossi S, Sturla F, Castellanos ML, Stella IY, Balana ME (2014) Dermal papilla cells improve the wound healing process and generate hair bud-like structures in grafted skin substitutes using hair follicle stem cells. Stem Cells Transl Med 3(10):1209–1219PubMedPubMedCentralCrossRefGoogle Scholar
  37. Li WJ, Shanti RM, Tuan RS (2006a) Nanotechnologies for the life sciences. Tissue Cell Organ Eng 9:135–187Google Scholar
  38. Li J, He A, Zheng J, Han CC (2006b) Gelatin and gelatin-hyaluronic acid nanofibrous membranes produced by electrospinning of their aqueous solutions. Biomacromolecules 7:2243–2247PubMedCrossRefGoogle Scholar
  39. Liu H, Mao JS, Yao KD, Yang GH, Cui L, Cao YL (2004) A study on a chitosan-gelatin-hyaluronic acid scaffold as artificial skin in vitro and its tissue engineering applications. J Biomater Sci Polym Ed 15(1):25–40PubMedCrossRefGoogle Scholar
  40. Liu H, Yin Y, Yao K (2007) Construction of chitosan-gelatin-hyaluronic acid artificial skin in vitro. J Biomater Appl 21(4):413–430PubMedCrossRefGoogle Scholar
  41. Lopez-Camarillo C, Ocampo EA, Casamichana ML, Perez-Plasencia C, Alvarez-Sanchez E, Marchat LA (2012) Protein kinases and transcription factors activation in response to UV-radiation of skin: implications for carcinogenesis. Int J Mol Sci 13(1):142–172PubMedCrossRefGoogle Scholar
  42. Lu T, Li Y, Chen T (2013) Techniques for fabrication and construction of three-dimensional scaffolds for tissue engineering. Int J Nanomed 8:337–350CrossRefGoogle Scholar
  43. Ma J, Wang H, He B, Chen J (2001) A preliminary in vitro study on the fabrication and tissue engineering applications of a novel chitosan bilayer material as a scaffold of human neofetal dermal fibroblasts. Biomaterials 22:331–336PubMedCrossRefGoogle Scholar
  44. Mao J, Zhao L, de Yao K, Shang Q, Yang G, Cao Y (2003) Study of novel chitosan-gelatin artificial skin in vitro. J Biomed Mater Res A 64A(2):301–308CrossRefGoogle Scholar
  45. Marionnet C, Vioux-Chagoleau C, Pierrard C, Sok J, Asselineau D, Bernerd F (2006) Morphogenesis of dermal-epidermal junction in a model of reconstructed skin: beneficial effects of vitamin C. Exp Dermatol 15:625–633PubMedCrossRefGoogle Scholar
  46. Mazlyzam AL, Aminuddin BS, Fuzina NH, Norhayati MM, Fauziah O, Isa MR, Saim L, Ruszymah BH (2007) Reconstruction of living bilayer human skin equivalent utilizing human fibrin as a scaffold. Burns 33(3):355–363PubMedCrossRefGoogle Scholar
  47. Min BM, Jeong L, Lee KY, Park WH (2006) Regenerated silk fibroin nanofibers: water vapor-induced structural changes and their effects on the behavior of normal human cells. Macromol Biosci 6(4):285–292PubMedCrossRefGoogle Scholar
  48. Netzlaff F, Lehr CM, Wertz PW, Schaefer UF (2005) The human epidermis models EpiSkin, SkinEthic and EpiDerm: an evaluation of morphology and their suitability for testing phototoxicity, irritancy, corrosivity, and substance transport. Eur J Pharm Biopharm 60(2):167–178PubMedCrossRefGoogle Scholar
  49. Ng WL, Yeong WY, Naing MW (2016) Polyelectrolyte gelatin-chitosan hydrogel optimized for 3D bioprinting in skin tissue engineering. Int J Bioprint 2(1):53–62Google Scholar
  50. Noh HK, Lee SW, Kim JM, Oh JE, Kim KH, Chung CP, Choi SC, Park WH, Min BM (2006) Electrospinning of chitin nanofibers: degradation behavior and cellular response to normal human keratinocytes and fibroblasts. Biomaterials 27(21):3934–3944PubMedCrossRefGoogle Scholar
  51. O’Brien FJ, Harley BA, Yannas IV, Gibson LJ (2005) The effect of pore size on cell adhesion in collagen-GAG scaffolds. Biomaterials 26:433PubMedCrossRefGoogle Scholar
  52. Pan H, Jiang H, Chen W (2006) Interaction of dermal fibroblasts with electrospun composite polymer scaffolds prepared from dextran and poly lactide-co-glycolide. Biomaterials 27(17):3209–3220PubMedCrossRefGoogle Scholar
  53. Patra S, Young V (2016) A review of 3D printing techniques and the future in biofabrication of bioprinted tissue. Cell Biochem Biophys 74(2):93–98PubMedCrossRefGoogle Scholar
  54. Planz V, Lehr CM, Windbergs M (2016) In vitro models for evaluating safety and efficacy of novel technologies for skin drug delivery. J Control Release 242:89–104PubMedCrossRefGoogle Scholar
  55. Rho KS, Jeong L, Lee G, Seo BM, Park YJ, Hong SD, Roh S, Cho JJ, Park WH, Min BM (2006) Electrospinning of collagen nanofibers: effects on the behavior of normal human keratinocytes and early-stage wound healing. Biomaterials 27(8):1452–1461PubMedCrossRefGoogle Scholar
  56. Satoshi K, Hiroaki T, Kenichi S, Hidenori F, Keiichi N, Yoshihiro T, Fumie H, Kenji S (2010) Utilization of reconstructed cultured human skin models as an alternative skin for permeation studies of chemical compounds. Altern Anim Test Exp 15(2):61–70Google Scholar
  57. Semlin L, Schafer-Korting M, Borelli C, Korting HC (2011) In vitro models for human skin disease. Drug Discov Today 16(3–4):132–139PubMedCrossRefGoogle Scholar
  58. Silver FH, Freeman JW, DeVore D (2001) Viscoelastic properties of human skin and processed dermis. Skin Res Technol 7(1):18–23PubMedCrossRefGoogle Scholar
  59. Song HJ, Lim HY, Chun W, Choi KC, Sung JH, Sung GY (2017) Fabrication of a pumpless, microfluidic skin chip from different collagen sources. J Ind Eng Chem 56:375–381CrossRefGoogle Scholar
  60. Sorrell JM, Baber MA, Caplan AI (2004) Site-matched papillary and reticular human dermal fibroblasts differ in their release of specific growth factors/cytokines and in their interaction with keratinocytes. J Cell Physiol 200(1):134–145PubMedCrossRefGoogle Scholar
  61. Sun T, Jackson S, Haycock JW, MacNeil S (2006) Culture of skin cells in 3D rather than 2D improves their ability to survive exposure to cytotoxic agents. J Biotechnol 122(3):372–381PubMedCrossRefGoogle Scholar
  62. Uchida T, Kadhum WR, Kanai S, Todo H, Oshizaka T, Sugibayashi K (2015) Prediction of skin permeation by chemical compounds using the artificial membrane, Strat-M. Eur J Pharm Sci 67:113–118PubMedCrossRefGoogle Scholar
  63. Van Gele M, Geusens B, Brochez L, Speeckaert R, Lambert J (2011) Three-dimensional skin models as tools for transdermal drug delivery: challenges and limitations. Expert Opin Drug Deliv 8(6):705–720PubMedCrossRefGoogle Scholar
  64. van den Bogaard EH, Tjabringa GS, Joosten I, Vonk-Bergers M, van Rijssen E, Tijssen HJ, Erkens M, Schalkwijk J, Koenen H (2014) Crosstalk between keratinocytes and T cells in a 3D microenvironment: a model to study inflammatory skin diseases. J Invest Dermatol 134(3):719–727PubMedCrossRefGoogle Scholar
  65. Venugopal J, Ramakrishna S (2005) Biocompatible nanofiber matrices for the engineering of a dermal substitute for skin regeneration. Tissue Eng 11(5–6):847–854PubMedCrossRefGoogle Scholar
  66. Vorsmann H, Groeber F, Walles H, Busch S, Beissert S, Walczak H, Kulms D (2013) Development of a human three-dimensional organotypic skin-melanoma spheroid model for in vitro drug testing. Cell Death Dis 11(4):e719CrossRefGoogle Scholar
  67. Whang KK, Kim MJ, Song WK, Cho SY (2005) Comparative treatment of giant congenital melanocytic nevi with curettage or Er:YAG laser ablation alone versus with cultured epithelial autografts. Dermatol Surg 31(12):1660–1667PubMedGoogle Scholar
  68. Woo CH, Choi YC, Choi JS, Lee HY, Cho YW (2015) A bilayer composite composed of TiO2-incorporated electrospun chitosan membrane and human extracellular matrix sheet as a wound dressing. J Biomater Sci Polym Ed 26(13):841–854PubMedCrossRefGoogle Scholar
  69. Wufuer M, Lee G, Hur W, Jeon B, Kim BJ, Choi TH, Lee S (2016) Skin-on-a-chip model simulating inflammation, edema and drug-based treatment. Sci Rep 6:37471PubMedPubMedCentralCrossRefGoogle Scholar
  70. You HJ, Han SK, Lee JW, Chang H (2012) Treatment of diabetic foot ulcers using cultured allogeneic keratinocytes—a pilot study. Wound Repair Regen 20(4):491–499PubMedGoogle Scholar
  71. Zhang YZ, Venugopal J, Huang Z-M, Lim CT, Ramakrishna S (2005) Characterization of the surface biocompatiblity of the electrospun PCL-collagen nanofibers using fibroblasts. Biomacromolecules 6:2583–2589PubMedCrossRefGoogle Scholar
  72. Zhang YZ, Venugopal J, Huang ZM, Lim CT, Ramakrishna S (2006) Crosslinking of the electrospun gelatin nanofibers. Polymer 47(8):2911–2917CrossRefGoogle Scholar
  73. Zhong S, Teo WE, Zhu X, Beuerman RW, Ramakrishna S, Yung LY (2006) An aligned nanofibrous collagen scaffold by electrospinning and its effects on in vitro fibroblast culture. J Biomed Mater Res A 79(3):456–463PubMedCrossRefGoogle Scholar
  74. Zhu W, Ma X, Gou M, Mei D, Zhang K, Chen S (2016) 3D printing of functional biomaterials for tissue engineering. Curr Opin Biotechnol 40:103–112PubMedCrossRefGoogle Scholar

Copyright information

© The Korean Society of Pharmaceutical Sciences and Technology 2018

Authors and Affiliations

  • Ye Eun Yun
    • 1
  • Youn Jae Jung
    • 1
  • Yeo Jin Choi
    • 1
  • Ji Suk Choi
    • 1
  • Yong Woo Cho
    • 1
    Email author
  1. 1.Department of Materials Science and Chemical EngineeringHanyang UniversityAnsanRepublic of Korea

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