Current Transplantation Reports

, Volume 3, Issue 1, pp 100–108 | Cite as

3D Printing for Liver Tissue Engineering: Current Approaches and Future Challenges

  • Phillip L. Lewis
  • Ramille N. Shah
Tissue Engineering and Regeneration (JA Wertheim, Section Editor)
Part of the following topical collections:
  1. Topical Collection on Tissue Engineering and Regeneration


Recent developments in 3D printing have greatly accelerated progress in the field of liver tissue engineering by enabling the fabrication of more tissue-mimetic structures capable of restoring function. A variety of 3D printing and additive manufacturing techniques ranging from stereolithography to direct ink writing have shown great promise in liver tissue engineering and the study of cellular interactions. Despite these advances, however, there is significant room for improvement. Furthermore, because of the enormous capabilities of 3D printing, methods to analyze complex heterogeneous tissues in vitro have yet to be perfected. Investigations into the ability of 3D printing to recreate the macro- and microstructural components of the liver are still in their infancy. These specific issues need to be addressed in combination with massive scale up if 3D-printed tissue-engineered livers are to reach clinical relevance.


Bioprinting 3D printing Additive manufacturing Liver tissue engineering 


Compliance with Ethical Standards

Conflict of Interest

Phillip L. Lewis and Ramille N. Shah declare that they have no conflict of interest.

Human and Animal Rights and Informed Consent

This article does not contain any studies with human or animal subjects performed by any of the authors.


Papers of particular interest, published recently, have been highlighted as: •• Of major importance

  1. 1.
    Vernon G, Baranova A, Younossi ZM. Systematic review: the epidemiology and natural history of non-alcoholic fatty liver disease and non-alcoholic steatohepatitis in adults. Aliment Pharmacol Ther. 2011;34(3):274–85.CrossRefPubMedGoogle Scholar
  2. 2.
    Atala A, Kasper FK, Mikos AG. Engineering complex tissues. Sci Transl Med. 2012;4(160):160rv12.CrossRefPubMedGoogle Scholar
  3. 3.
    Griffith LG, Wells A, Stolz DB. Engineering liver. Hepatology. 2014;60(4):1426–34.CrossRefPubMedPubMedCentralGoogle Scholar
  4. 4.
    Murphy SV, Atala A. 3D bioprinting of tissues and organs. Nat Biotechnol. 2014;32(8):773–85.CrossRefPubMedGoogle Scholar
  5. 5.••
    Jakus AE, Rutz AL, Shah RN. Advancing the field of 3D biomaterial printing. Biomed Mater. 2015. A review of trends and limitations in 3D printing for tissue engineering applications, drawing on concerns associated with materials science and engineering as well as biological components.Google Scholar
  6. 6.
    Wang X, Yan Y, Zhang R. Rapid prototyping as a tool for manufacturing bioartificial livers. Trends Biotechnol. 2007;25(11):505–13.CrossRefPubMedGoogle Scholar
  7. 7.
    Yoshida H, Katayose Y, Unno M, et al. Segmentectomy of the liver. J Hepatobiliary Pancreat Sci. 2012;19(1):67–71.CrossRefPubMedGoogle Scholar
  8. 8.
    Gibson I, Rosen D, Stucker B. Additive manufacturing technologies: 3D printing, rapid prototyping, and direct digital manufacturing. vol. 2. 2010.Google Scholar
  9. 9.
    Huang H, Oizumi S, Sakai Y, et al. Avidin-biotin binding-based cell seeding and perfusion culture of liver-derived cells in a porous scaffold with a three-dimensional interconnected flow-channel network. Biomaterials. 2007;28(26):3815–23.CrossRefPubMedGoogle Scholar
  10. 10.
    Ho CT, Lin RZ, Liu CH, et al. Liver-cell patterning Lab Chip: mimicking the morphology of liver lobule tissue. Lab Chip. 2013;13(18):3578–87.CrossRefPubMedGoogle Scholar
  11. 11.
    Albrecht DR, Sah RL, Bhatia SN. Dielectrophoretic cell patterning within tissue engineering scaffolds. Proc Second Jt 24th Annu Conf Annu Fall Meet Biomed Eng Soc Eng Med Biol vol. 2, pp. 1708–1709. 2002.Google Scholar
  12. 12.
    Starly B, Chang R, Sun W. UV-photolithography fabrication of poly-ethylene glycol hydrogels encapsulated with hepatocytes. Proc 17th Annu Solid Free Fabr Symp Univ Texas Austin pp. 102–110. 2006.Google Scholar
  13. 13.
    Jiankang H, Dichen L, Yi L, et al. Preparation of chitosan-gelatin hybrid scaffolds with well-organized microstructures for hepatic tissue engineering. Acta Biomater. 2009;5(1):453–61.CrossRefPubMedGoogle Scholar
  14. 14.
    Kaihara S, Borenstein J, Vacanti JP, et al. Silicon micromachining to tissue engineer branched vascular channels for liver fabrication. Tissue Eng. 2000;6(2):105–17.CrossRefPubMedGoogle Scholar
  15. 15.
    Liu Tsang V, Chen AA, Bhatia SN, et al. Fabrication of 3D hepatic tissues by additive photopatterning of cellular hydrogels. FASEB J. 2007;21(3):790–801.CrossRefPubMedGoogle Scholar
  16. 16.
    Li CY, Stevens KR, Bhatia SN, et al. Micropatterned cell-cell interactions enable functional encapsulation of primary hepatocytes in hydrogel microtissues. Tissue Eng A. 2014;20:2200–12.CrossRefGoogle Scholar
  17. 17.
    Boland T, Xu T, Damon B, Cui X. Application of inkjet printing to tissue engineering. Biotechnol J. 2006;1(9):910–7.CrossRefPubMedGoogle Scholar
  18. 18.
    Parsa S, Gupta M, Loizeau F, Cheung KC. Effects of surfactant and gentle agitation on inkjet dispensing of living cells. Biofabrication. 2010;2(2):025003.CrossRefPubMedGoogle Scholar
  19. 19.
    Matsusaki M, Sakaue K, Akashi M, et al. Three-dimensional human tissue chips fabricated by rapid and automatic inkjet cell printing. Adv Healthc Mater. 2013;2(4):534–9.CrossRefPubMedGoogle Scholar
  20. 20.
    Woodrow KA, Wood MJ, Saltzman WM, et al. Biodegradable meshes printed with extracellular matrix proteins support micropatterned hepatocyte cultures. Tissue Eng A. 2009;15(5):1169–79.CrossRefGoogle Scholar
  21. 21.
    Vozzi G, Previti A, Ahluwalia A, et al. Microsyringe-based deposition of two-dimensional and three-dimensional polymer scaffolds with a well-defined geometry for application to tissue engineering. Tissue Eng. 2002;8(6):1089–98.CrossRefPubMedGoogle Scholar
  22. 22.
    Zein I, Hutmacher DW, Teoh SH, et al. Fused deposition modeling of novel scaffold architectures for tissue engineering applications. Biomaterials. 2002;23(4):1169–85.CrossRefPubMedGoogle Scholar
  23. 23.
    Kim SS, Utsunomiya H, Vacanti JP, et al. Survival and function of hepatocytes on a novel three-dimensional synthetic biodegradable polymer scaffold with an intrinsic network of channels. Ann Surg. 1998;228(1):8–13.CrossRefPubMedPubMedCentralGoogle Scholar
  24. 24.
    Shim JH, Kim JY, Cho DW, et al. Development of a hybrid scaffold with synthetic biomaterials and hydrogel using solid freeform fabrication technology. Biofabrication. 2011;3:034102–11.CrossRefPubMedGoogle Scholar
  25. 25.
    Tamayol A, Najafabadi AH, Khademhosseini A, et al. Hydrogel templates for rapid manufacturing of bioactive fibers and 3D constructs. Adv Health Mater. 2015.Google Scholar
  26. 26.
    Chang R, Nam J, Sun W. Effects of dispensing pressure and nozzle diameter on cell survival from solid freeform fabrication-based direct cell writing. Tissue Eng A. 2008;14(1):41–8.CrossRefGoogle Scholar
  27. 27.
    Yan Y, Wang X, Lu Q, et al. Direct construction of a three-dimensional structure with cells and hydrogel. J Bioact Compat Polym. 2005;20(3):259–69.CrossRefGoogle Scholar
  28. 28.
    Wang X, Yan Y, Lu Q, et al. Generation of three-dimensional hepatocyte/gelatin structures with rapid prototyping system. Tissue Eng. 2006;12(1):83–90.CrossRefPubMedGoogle Scholar
  29. 29.
    Yan Y, Wang X, Lu Q, et al. Fabrication of viable tissue-engineered constructs with 3D cell-assembly technique. Biomaterials. 2005;26(29):5864–71.CrossRefPubMedGoogle Scholar
  30. 30.
    Xu W, Wang X, Zhang R, et al. Rapid prototyping three-dimensional cell/gelatin/fibrinogen constructs for medical regeneration. J Bioact Compat Polym. 2007;22(4):363–77.CrossRefGoogle Scholar
  31. 31.
    Rutz AL, Hyland KE, Shah RN, et al. A multimaterial bioink method for 3D printing tunable, cell-compatible hydrogels. Adv Mater pp 1–8. 2015.Google Scholar
  32. 32.
    Skardal A, Devarasetty M, Atala A, et al. A hydrogel bioink toolkit for mimicking native tissue biochemical and mechanical properties in bioprinted tissue constructs. Acta Biomater. 2015;25:24–34.CrossRefPubMedGoogle Scholar
  33. 33.
    Billiet T, Gevaert E, Dubruel P, et al. The 3D printing of gelatin methacrylamide cell-laden tissue-engineered constructs with high cell viability. Biomaterials. 2014;35(1):49–62.CrossRefPubMedGoogle Scholar
  34. 34.
    Bertassoni LE, Cardoso JC, Khademhosseini A, et al. Direct-write bioprinting of cell-laden methacrylated gelatin hydrogels. Biofabrication. 2014;6(2):024105.CrossRefPubMedPubMedCentralGoogle Scholar
  35. 35.
    Norotte C, Niklason LE, Forgacs G, et al. Biomaterials Scaffold-free vascular tissue engineering using bioprinting. Biomaterials. 2009;30(30):5910–7.CrossRefPubMedPubMedCentralGoogle Scholar
  36. 36.
    Jakab K, Norotte C, Forgacs G, et al. Tissue engineering by self-assembly and bio-printing of living cells. Biofabrication. 2010;2(2):022001.CrossRefPubMedPubMedCentralGoogle Scholar
  37. 37.
    Marga F, Jakab K, Forgacs G, et al. Toward engineering functional organ modules by additive manufacturing. Biofabrication. 2012;4(2):022001.CrossRefPubMedGoogle Scholar
  38. 38.
    Lazar A, Mann H, Remmel R. Extended liver-specific functions of porcine hepatocyte spheroids entrapped in collagen gel. Vitr Cell. 1995;31(5):340–6.CrossRefGoogle Scholar
  39. 39.
    Thorling CA, Crawford D, Roberts MS, et al. Multiphoton microscopy in defining liver function. J Biomed Opt. 2014;19(9):090901.CrossRefGoogle Scholar
  40. 40.••
    Miller JS, Stevens KR, Chen CS, et al. Rapid casting of patterned vascular networks for perfusable engineered three-dimensional tissues. Nat Mater. 2012;11(9):768–74. Demonstrated microvascular patterning, capillary branching, and enahnced hepatocyte survival and function within gels cast around sacrificial 3D printed microvessel networks.CrossRefPubMedPubMedCentralGoogle Scholar
  41. 41.
    Buesa RJ, Peshkov MV. Histology without xylene. Ann Diagn Pathol. 2009;13(4):246–56.CrossRefPubMedGoogle Scholar
  42. 42.
    Underhill GH, Chen AA, Bhatia SN, et al. Assessment of hepatocellular function within PEG hydrogels. Biomaterials. 2007;28:256–70.CrossRefPubMedGoogle Scholar
  43. 43.
    Baranski JD, Chaturvedi RR, Chen CS, et al. Geometric control of vascular networks to enhance engineered tissue integration and function. Proc Natl Acad Sci U S A. 2013;110(19):7586–91.CrossRefPubMedPubMedCentralGoogle Scholar
  44. 44.
    Ohashi K, Waugh JM, Kay MA, et al. Liver tissue engineering at extrahepatic sites in mice as a potential new therapy for genetic liver diseases. Hepatology. 2005;41(1):132–40.CrossRefPubMedGoogle Scholar
  45. 45.
    Chaturvedi RR, Stevens KR, Chen CS, et al. Patterning vascular networks in vivo for tissue engineering applications. Tissue Eng Part C Methods. 2015;21(5):509–17.CrossRefPubMedGoogle Scholar
  46. 46.
    Kolesky DB, Truby RL, Lewis JA, et al. 3D bioprinting of vascularized, heterogeneous cell-laden tissue constructs. Adv Mater. 2014;26(19):3124–30.CrossRefPubMedGoogle Scholar
  47. 47.
    Takebe T, Sekine K, Taniguchi H, et al. Vascularized and functional human liver from an iPSC-derived organ bud transplant. Nature pp 1–5. 2013.Google Scholar
  48. 48.
    Zein NN, Hanouneh IA, Klatte R, et al. Three-dimensional print of a liver for preoperative planning in living donor liver transplantation. Liver Transplant. 2013;19(12):1304–10.CrossRefGoogle Scholar
  49. 49.
    Si-Tayeb K, Lemaigre FP, Duncan SA. Organogenesis and development of the liver. Dev Cell. 2010;18(2):175–89.CrossRefPubMedGoogle Scholar
  50. 50.
    Bae H, Peppas NA, Khademhosseini A, et al. Building vascular networks. Sci Transl Med. 2012;4(160):160ps23.CrossRefPubMedPubMedCentralGoogle Scholar
  51. 51.
    Aird WC. Phenotypic heterogeneity of the endothelium: I. Structure, function, and mechanisms. Circ Res. 2007;100(2):158–73.CrossRefPubMedGoogle Scholar
  52. 52.
    Aird WC. Phenotypic heterogeneity of the endothelium: II. Representative vascular beds. Circ Res. 2007;100(2):174–90.CrossRefPubMedGoogle Scholar
  53. 53.
    Arnaoutova I, George J, Benton G, et al. The endothelial cell tube formation assay on basement membrane turns 20: state of the science and the art. Angiogenesis. 2009;12:267–74.CrossRefPubMedGoogle Scholar
  54. 54.
    Nerem RM, Seliktar D. Vascular tissue engineering. Annu Rev Biomed Eng. 2001;3:225–43.CrossRefPubMedGoogle Scholar
  55. 55.
    Nguyen DHT, Stapleton SC, Chen CS, et al. Biomimetic model to reconstitute angiogenic sprouting morphogenesis in vitro. Proc Natl Acad Sci. 2013;110(17):6712–7.CrossRefPubMedPubMedCentralGoogle Scholar
  56. 56.
    Lee VK, Kim DY, Dai G, et al. Creating perfused functional vascular channels using 3D bio-printing technology. Biomaterials. 2014;35(28):8092–102.CrossRefPubMedPubMedCentralGoogle Scholar
  57. 57.
    Sellaro TL, Ravindra AK, Badylak SF, et al. Maintenance of hepatic sinusoidal endothelial cell phenotype in vitro using organ-specific extracellular matrix scaffolds. Tissue Eng. 2007;13(9):2301–10.CrossRefPubMedGoogle Scholar
  58. 58.
    Kim Y, Rajagopalan P. 3D hepatic cultures simultaneously maintain primary hepatocyte and liver sinusoidal endothelial cell phenotypes. PLoS One. 2010;5(11):e15456.CrossRefPubMedPubMedCentralGoogle Scholar
  59. 59.
    Detzel CJ, Kim Y, Rajagopalan P. Engineered three-dimensional liver mimics recapitulate critical rat-specific bile acid pathways. Tissue Eng A. 2011;17(5–6):677–89.CrossRefGoogle Scholar
  60. 60.••
    Godoy P, Hewitt NJ, Hengstler JG, et al. Recent advances in 2D and 3D in vitro systems using primary hepatocytes, alternative hepatocyte sources and non-parenchymal liver cells and their use in investigating mechanisms of hepatotoxicity, cell signaling and ADME. Arch Toxicol. 2013;87:1315–530. Comprehensive review of liver biology and engineering strategies to study liver cell and molecular biology as well as approaches for liver assist devices and tissue engineering.CrossRefPubMedPubMedCentralGoogle Scholar
  61. 61.
    Bhatia S, Toner M, Rotem A, et al. Zonal liver cell heterogeneity: effects of oxygen on metabolic functions of hepatocytes. Cell Eng. 1996;2:125–33.Google Scholar
  62. 62.
    Gebhardt R, Hovhannisyan A. Organ patterning in the adult stage: the role of Wnt/β-catenin signaling in liver zonation and beyond. Dev Dyn. 2010;239(1):45–55.PubMedGoogle Scholar
  63. 63.
    McClelland R, Wauthier E, Reid L, et al. Gradients in the liver’s extracellular matrix chemistry from periportal to pericentral zones: influence on human hepatic progenitors. Tissue Eng A. 2008;14(1):59–70.CrossRefGoogle Scholar
  64. 64.
    Wang Y, Cui CB, Reid LM, et al. Lineage restriction of human hepatic stem cells to mature fates is made efficient by tissue-specific biomatrix scaffolds. Hepatology. 2011;53(1):293–305.CrossRefPubMedGoogle Scholar
  65. 65.
    Allen JW. In vitro zonation and toxicity in a hepatocyte bioreactor. Toxicol Sci. 2005;84(1):110–9.CrossRefPubMedGoogle Scholar
  66. 66.
    Gumucio JJ, May M, Massey V, et al. The isolation of functionally heterogeneous hepatocytes of the proximal and distal half of the liver acinus in the rat. Hepatology. 1986;6(5):932–44.CrossRefPubMedGoogle Scholar
  67. 67.
    Bhatia SN, Underhill GH, Fox IJ, et al. Cell and tissue engineering for liver disease. Sci Transl Med. 2014;6(245):245sr2.CrossRefPubMedPubMedCentralGoogle Scholar
  68. 68.
    Miyazawa M, Torii T, Ikada Y, et al. A tissue-engineered artificial bile duct grown to resemble the native bile duct. Am J Transplant. 2005;5(6):1541–7.CrossRefPubMedGoogle Scholar
  69. 69.
    Miyazawa M, Aikawa M, Ikada Y, et al. Regeneration of extrahepatic bile ducts by tissue engineering with a bioabsorbable polymer. J Artif Organs. 2012;15(1):26–31.CrossRefPubMedGoogle Scholar
  70. 70.
    Barralet JE, Wallace LL, Strain AJ. Tissue engineering of human biliary epithelial cells on polyglycolic acid/polycaprolactone scaffolds maintains long-term phenotypic stability. Tissue Eng. 2003;9(5):1037–45.CrossRefPubMedGoogle Scholar
  71. 71.
    Li Q, Tao L, Ding Y, et al. Extrahepatic bile duct regeneration in pigs using collagen scaffolds loaded with human collagen-binding bFGF. Biomaterials. 2012;33(17):4298–308.CrossRefPubMedGoogle Scholar
  72. 72.
    Auth MK, Joplin RE, Strain AJ, et al. Morphogenesis of primary human biliary epithelial cells: induction in high-density culture or by coculture with autologous human hepatocytes. Hepatology. 2001;33(3):519–29.CrossRefPubMedGoogle Scholar
  73. 73.
    Ueno Y, Alpini G, Shimosegawa T, et al. Evaluation of differential gene expression by microarray analysis in small and large cholangiocytes isolated from normal mice. Liver Int. 2003;23(6):449–59.CrossRefPubMedGoogle Scholar
  74. 74.
    Venter J, Francis H, Alpini G, et al. Development and functional characterization of extrahepatic cholangiocyte lines from normal rats. Dig Liver Dis. 2015;47(11):964–72.CrossRefPubMedGoogle Scholar

Copyright information

© Springer International Publishing AG 2016

Authors and Affiliations

  1. 1.Department of Biomedical EngineeringNorthwestern UniversityEvanstonUSA
  2. 2.Simpson Querrey InstituteNorthwestern UniversityChicagoUSA
  3. 3.Department of Materials Science and EngineeringNorthwestern UniversityEvanstonUSA
  4. 4.Department of Surgery (Transplant Division), Feinberg School of MedicineNorthwestern UniversityChicagoUSA
  5. 5.Comprehensive Transplant Center, Feinberg School of MedicineNorthwestern UniversityChicagoUSA

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