Advertisement

Investigating the effect of sterilisation methods on the physical properties and cytocompatibility of methyl cellulose used in combination with alginate for 3D-bioplotting of chondrocytes

  • Ella HodderEmail author
  • Sarah Duin
  • David Kilian
  • Tilman Ahlfeld
  • Julia Seidel
  • Carsten Nachtigall
  • Peter Bush
  • Derek Covill
  • Michael Gelinsky
  • Anja LodeEmail author
S.I.: Biofabrication and Bioinks for Tissue Engineering Original Research
Part of the following topical collections:
  1. S.I.: Biofabrication and Bioinks for Tissue Engineering

Abstract

For both the incorporation of cells and future therapeutic applications the sterility of a biomaterial must be ensured. However, common sterilisation techniques are intense and often negatively impact on material physicochemical attributes, which can affect its suitability for tissue engineering and 3D printing. In the present study four sterilisation methods, autoclave, supercritical CO2 (scCO2) treatment, UV- and gamma (γ) irradiation were evaluated regarding their impact on material properties and cellular responses. The investigations were performed on methyl cellulose (MC) as a component of an alginate/methyl cellulose (alg/MC) bioink, used for bioprinting embedded bovine primary chondrocytes (BPCs). In contrast to the autoclave, scCO2 and UV-treatments, the γ-irradiated MC resulted in a strong reduction in alg/MC viscosity and stability after extrusion which made this method unsuitable for precise bioprinting. Gel permeation chromatography analysis revealed a significant reduction in MC molecular mass only after γ-irradiation, which influenced MC chain mobility in the Ca2+-crosslinked alginate network as well as gel composition and microstructure. With regard to cell survival and proteoglycan matrix production, the results determined UV-irradiation and autoclaving as the best candidates for sterilisation. The scCO2-treatment of MC resulted in an unfavourable cell response indicating that this method needs careful optimisation prior to application for cell encapsulation. As proven by consistent FT-IR spectra, chemical alterations could be excluded as a cause for the differences seen between MC treatments on alg/MC behaviour. This investigation provides knowledge for the development of a clinically appropriate 3D-printing-based fabrication process to produce bioengineered tissue for cartilage regeneration.

Notes

Acknowledgements

The authors thank Ms Ortrud Zieschang for preparation of SEM samples, the microscopy facility CFCI of the TU Dresden for providing equipment and support in cell imaging as well as the Institute of Natural Materials Technology, Chair of Food Engineering at TU Dresden for the opportunity to perform gel permeation chromatography (GPC). This work was supported by the European Social Fund (ESF) and Free State of Saxony (Young Researchers Group IndivImp at Technische Universität Dresden), the German Research Society (DFG) as part of the priority program SPP 1934 as well as the German Center for Diabetes Research (DZD). Partial funding was received from a Santander Research Scholarship, awarded to Ella Hodder.

Compliance with ethical standards

Conflict of interest

The authors declare that there is no conflict of interest.

Supplementary material

10856_2018_6211_MOESM1_ESM.tif (10.1 mb)
Supplementary figure1
10856_2018_6211_MOESM2_ESM.tif (78 kb)
Supplementary figure2
10856_2018_6211_MOESM3_ESM.tif (68 kb)
Supplementary figure3
10856_2018_6211_MOESM4_ESM.tif (59 kb)
Supplementary figure4
10856_2018_6211_MOESM5_ESM.tif (416 kb)
Supplementary figure5
10856_2018_6211_MOESM6_ESM.docx (14 kb)
Supplementary figures

References

  1. 1.
    Liaw C-Y, Guvendiren M. Current and emerging applications of 3D printing in medicine. Biofabrication. 2017;9(2):24102.CrossRefGoogle Scholar
  2. 2.
    Ventola CL. Medical applications for 3D printing: current and projected uses. P T. 2014;39(10):704–11.Google Scholar
  3. 3.
    Billiet T, Vandenhaute M, Schelfhout J, Van Vlierberghe S, Dubruel P. A review of trends and limitations in hydrogel-rapid prototyping for tissue engineering. Biomaterials. 2012;33(26):6020–41.CrossRefGoogle Scholar
  4. 4.
    Malda J, Visser J, Melchels FP, Jüngst T, Hennink WE, Dhert WJA et al. 25th anniversary article: engineering hydrogels for biofabrication. Adv Mater. 2013;25(36):5011–28.CrossRefGoogle Scholar
  5. 5.
    Lee KY, Mooney DJ. Alginate: properties and biomedical applications. Prog Polym Sci. 2012;37(1):106–26.CrossRefGoogle Scholar
  6. 6.
    Lee KY, Rowley JA, Eiselt P, Moy EM, Bouhadir KH, Mooney DJ. Controlling mechanical and swelling properties of alginate hydrogels independently by cross-linker type and cross-linking density. Macromolecules. 2000;33(11):4291–4.CrossRefGoogle Scholar
  7. 7.
    Joydip K, Jin-Hyung S, Jinah, Jang S, Sung WK, Dong-Woo C. An additive manufacturing-based PCL–alginate– chondrocyte bioprinted scaffold for cartilage tissue engineering. J Tissue Eng Regen Med. 2013;9(11):1286–97.Google Scholar
  8. 8.
    Fedorovich NE, Schuurman W, Wijnberg HM, Prins H-J, van Weeren PR, Malda J et al. Biofabrication of osteochondral tissue equivalents by printing topologically defined, cell-laden hydrogel scaffolds. Tissue Eng Part C Methods. 2012;18(1):33–44.CrossRefGoogle Scholar
  9. 9.
    Maher PS, Keatch RP, Donnelly K, Mackay RE, Paxton JZ. Construction of 3D biological matrices using rapid prototyping technology. Rapid Prototyp J. 2009;15(3):204–10.CrossRefGoogle Scholar
  10. 10.
    Schütz K, Placht A-M, Paul B, Brüggemeier S, Gelinsky M, Lode A. Three-dimensional plotting of a cell-laden alginate/ methylcellulose blend: towards biofabrication of tissue engineering constructs with clinically relevant dimensions. J Tissue Eng Regen Med. 2017;11(5):1574–87.CrossRefGoogle Scholar
  11. 11.
    Zehnder T, Sarker B, Boccaccini AR, Detsch R. Evaluation of an alginate-gelatine crosslinked hydrogel for bioplotting. Biofabrication. 2015;7(2):25001.CrossRefGoogle Scholar
  12. 12.
    Hwang CM, Ay B, Kaplan DL, Rubin JP, Marra KG, Atala A et al. Assessments of injectable alginate particle-embedded fibrin hydrogels for soft tissue reconstruction. Biomed Mater. 2013;8(1):14105.CrossRefGoogle Scholar
  13. 13.
    Lode A, Krujatz F, Brüggemeier S, Quade M, Schütz K, Knaack S et al. Green bioprinting: fabrication of photosynthetic algae-laden hydrogel scaffolds for biotechnological and medical applications. Eng Life Sci. 2015;15(2):177–83.CrossRefGoogle Scholar
  14. 14.
    Seidel J, Ahlfeld T, Adolph M, Kümmritz S, Steingroewer J, Krujatz F et al. Green bioprinting: extrusion-based fabrication of plant cell-laden biopolymer hydrogel scaffolds. Biofabrication. 2017;9(4):45011.CrossRefGoogle Scholar
  15. 15.
    Li H, Tan YJ, Leong KF, Li L. 3D Bioprinting of highly thixotropic alginate/methyl cellulose hydrogel with strong interface bonding. ACS Appl Mater Interfaces. 2017;9(23):20086–97.CrossRefGoogle Scholar
  16. 16.
    Ahlfeld T, Cidonio G, Kilian D, Duin S, Akkineni AR, Dawson JI et al. Development of a clay based bioink for 3D cell printing for skeletal application. Biofabrication. 2017;9(3):e034103.CrossRefGoogle Scholar
  17. 17.
    Wang Q, Sun J, Yao Q, Ji C, Liu J, Zhu Q. 3D printing with cellulose materials. Cellulose. 2018;25:1–27.CrossRefGoogle Scholar
  18. 18.
    Nasatto PL, Pignon F, Silveira JLM, Duarte MER, Noseda MD, Rinaudo M. Methylcellulose, a cellulose derivative with original physical properties and extended applications. Polymers. 2015;7(5):777–803.CrossRefGoogle Scholar
  19. 19.
    Sarma NJ, Takeda A, Yaseen NR. Colony forming cell (CFC) assay for human hematopoietic cells. J Vis Exp. 2010;46:3–7.Google Scholar
  20. 20.
    Munarin F, Bozzini S, Visai L, Tanzi MC, Petrini P. Sterilization treatments on polysaccharides: effects and side effects on pectin. Food Hydrocoll. 2013;31(1):74–84.CrossRefGoogle Scholar
  21. 21.
    Mendes GCC, Brandão TRS, Silva CLM. Ethylene oxide sterilization of medical devices: a review. Am J Infect Control. 2007;35(9):574–81.CrossRefGoogle Scholar
  22. 22.
    Dai Z, Ronholm J, Tian Y, Sethi B, Cao X. Sterilization techniques for biodegradable scaffolds in tissue engineering applications. J Tissue Eng. 2016;7:1–13.CrossRefGoogle Scholar
  23. 23.
    Ahmed EM. Hydrogel: Preparation, characterization, and applications: a review. J Adv Res. 2015;6(2):105–21.CrossRefGoogle Scholar
  24. 24.
    Karajanagi SS, Yoganathan R, Mammucari R, Park H, Cox J, Zeitels SM et al. Application of a dense gas technique for sterilizing soft biomaterials. Biotechnol Bioeng. 2011;108(7):1716–25.CrossRefGoogle Scholar
  25. 25.
    Damar S, Balaban MO. Review of dense phase CO2 technology: microbial and enzyme inactivation. J Food Sci. 2006;71(1):1–11.CrossRefGoogle Scholar
  26. 26.
    Bernhardt A, Wehrl M, Paul B, Hochmuth T, Schumacher M, Schütz K et al. Improved sterilization of sensitive biomaterials with supercritical carbon dioxide at low temperature. PLoS ONE. 2015;10(6):e0129205. 19 pagesCrossRefGoogle Scholar
  27. 27.
    Spilimbergo S, Dehghani F, Bertucco A, Foster NR. Inactivation of bacteria and spores by pulse electric field and high-pressure CO2 at low temperature. Biotechnol Bioeng. 2003;82(1):118–25.CrossRefGoogle Scholar
  28. 28.
    Collier JP, Sperling DK, Currier JH, Sutula LC, Saum KA, Mayor MB. Impact of gamma sterilization on clinical performance of polyethylene in the knee. J Arthroplast. 1996;11(4):377–89.CrossRefGoogle Scholar
  29. 29.
    El-ashhab F, Sheha L, Abdalkhalek M, Khalaf HA. The influence of gamma irradiation on the intrinsic properties of cellulose acetate polymers. J Assoc Arab Univ Basic Appl Sci. 2013;14(1):46–50.Google Scholar
  30. 30.
    Palmer I, Clarke S, Nelson J, Schatton W, Dunne N, Buchanan F. Identification of a suitable sterilisation method for collagen derived from a marine Demosponge. Int Nano Biomater. 2012;4(2):148–63.CrossRefGoogle Scholar
  31. 31.
    Stoppel WL, White JC, Horava SD, Henry AC, Roberts SC, Bhatia SR. Terminal sterilization of alginate hydrogels: efficacy and impact on mechanical properties. J Biomed Mater Res B Appl Biomater. 2014;102(4):877–84.CrossRefGoogle Scholar
  32. 32.
    Goldman M, Gronsky R, Ranganathan R, Pruitt L. The effects of gamma radiation sterilization and ageing on the structure and morphology of medical grade ultra high molecular weight polyethylene. Polym. 1996;37(14):2909–13.CrossRefGoogle Scholar
  33. 33.
    Maslennikova A, Kochueva M, Ignatieva N, Vitkin A, Zakharkina O, Kamensky V et al. Effects of gamma irradiation on collagen damage and remodeling. Int J Radiat Biol. 2015;91(3):240–7.CrossRefGoogle Scholar
  34. 34.
    Meechan P, Wilson C. Use of ultraviolet lights in biological safety cabinets: a contrarian view. Appl Biosaf. 2006;11(4):222–7.CrossRefGoogle Scholar
  35. 35.
    Huebsch N, Gilbert M, Healy KE. Analysis of sterilization protocols for peptide-modified hydrogels. J Biomed Mater Res B Appl Biomater. 2005;74(1):440–7.CrossRefGoogle Scholar
  36. 36.
    Wilda H, Gough JE. In vitro studies of annulus fibrosus disc cell attachment, differentiation and matrix production on PDLLA/45S5 Bioglass composite films. Biomaterials. 2006;27(30):5220–9.CrossRefGoogle Scholar
  37. 37.
    Reza A T, Nicoll S. B. Hydrostatic pressure differentially regulates outer and inner annulus fibrosus cell matrix production in 3D scaffolds. Ann Biomed Eng. 2008;36(2):204–13.CrossRefGoogle Scholar
  38. 38.
    Bryant SJ, Anseth KS. Hydrogel properties influence ECM production by chondrocytes photoencapsulated in poly (ethylene glycol) hydrogels. J Biomed Mater Res. 2002;59(1):63–72.CrossRefGoogle Scholar
  39. 39.
    Pan T, Song W, Cao X, Wang Y. 3D Bioplotting of gelatin/alginate scaffolds for tissue engineering: influence of crosslinking degree and pore architecture on physicochemical properties. J Mater Sci Technol. 2016;32(9):889–900.CrossRefGoogle Scholar
  40. 40.
    Leo WJ, Mcloughlin AJ, Malone DM. Effects of sterilization treatments on some properties of alginate solutions and gels. Biotechnol. 1990;6(1):51–3.Google Scholar
  41. 41.
    Young J, So J, Park H, Won J. Effects of intermittent hydrostatic pressure magnitude on the chondrogenesis of MSCs without biochemical agents under 3D co-culture. J Mater Sci Mater Med. 2012;23(11):2773–81.CrossRefGoogle Scholar
  42. 42.
    Park JY, Choi YJ, Shim JH, Park JH, Cho DWJ. Development of a 3D cell printed structure as an alternative to autologs cartilage for auricular reconstruction. Biomed Mater Res B Appl Biomater. 2016;105(5):1–13.Google Scholar
  43. 43.
    Kundu J, Shim JH, Jang J, Kim SW, Cho DW. An additive manufacturing-based PCL–alginate– chondrocyte bioprinted scaffold for cartilage tissue engineering. Tissue Eng Regen Med. 2013;9(11):1286–97.CrossRefGoogle Scholar
  44. 44.
    Hall AC, Starks I, Shoults CL, Rashidbigi S. Pathways for K+transport across the bovine articular chondrocyte membrane and their sensitivity to cell volume. Am J Physiol. 1996;270(5):C1300–10.CrossRefGoogle Scholar
  45. 45.
    Fischbach C, Tessmar J, Lucke A, Schnell E, Schmeer G, Blunk T et al. Irradiation affect polymer properties relevant to tissue engineering? Surf Sci. 2001;491(3):333–45.CrossRefGoogle Scholar
  46. 46.
    Janorkar A, Metters A, Hirt D. Degradation of Poly(L-Lactide) films under ultraviolet- induced photografting and sterilization conditions. Surf Sci. 2006;106(2):1042–7.Google Scholar
  47. 47.
    Yixiang D, Yong T, Liao S, Chan CK, Ramakrishna S. Degradation of electrospun nanofiber scaffold by short wave length ultraviolet radiation treatment and its potential applications in tissue engineering. Tissue Eng Part A. 2008;14(8):1321–9.CrossRefGoogle Scholar
  48. 48.
    Koch HH, Pimsler M. Evaluation of Uvitex 2B: a nonspecific fluorescent stain for detecting and identifying fungi and algae in tissue. Lab Med. 1987;18(9):603–6.CrossRefGoogle Scholar
  49. 49.
    Rasconi S, Jobard M, Jouve L, Sime-Ngando T. Appl Environ Microbiol. 2009;75(8):2545–53.CrossRefGoogle Scholar
  50. 50.
    O’Brien FJ. Biomaterials & scaffolds for tissue engineering. Mater Today. 2011;14(3):88–95.CrossRefGoogle Scholar
  51. 51.
    Law N, Doney, Glover H, Qin Y, Aman ZM, Sercombe TB et al. J Mech Behav Biomed Mater. 2018;77:389–99.CrossRefGoogle Scholar
  52. 52.
    Hugo WB. A brief history of heat, chemical and radiation preservation and disinfection. Int Biodeterior Biodegrad. 1995;36(3–4):197–217.CrossRefGoogle Scholar
  53. 53.
    Hermannova M, Bezáková Z, Ebringerova A, Malovı A, Dřímalová E, Malovíková A et al. Effect of microwave irradiation on the molecular and structural properties of hyaluronan. Carbohydr Polym. 2008;73:640–6.CrossRefGoogle Scholar
  54. 54.
    Hoffman AS. Hydrogels for biomedical applications. Adv Drug Deliv Rev. 2002;54:3–12.CrossRefGoogle Scholar
  55. 55.
    Shahriari D, Koffler J, Lynam DA, Tuszynski MH, Sakamoto JS. Characterizing the degradation of alginate hydrogel for use in multilumen scaffolds for spinal cord repair. J Biomed Mater Res A. 2016;104(3):611–9.CrossRefGoogle Scholar
  56. 56.
    Ersch C, Linden E, Martin A, Venema P. Food Hydrocoll. 2016;52:991–1002.CrossRefGoogle Scholar
  57. 57.
    Meyer M, Prade I, Leppchen-Fröhlich K, Felix A, Herdegen V, Haseneder R et al. Sterilisation of collagen materials using hydrogen peroxide doted supercritical carbon dioxide and its effects on the materials properties. J Supercrit Fluids. 2015;102:32–9.CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  1. 1.School of Computing, Engineering and MathematicsUniversity of BrightonBrightonUK
  2. 2.School of Pharmacy and Biomolecular ScienceUniversity of BrightonBrightonUK
  3. 3.Centre for Translational Bone, Joint and Soft Tissue Research, Faculty of Medicine Carl Gustav CarusTechnische Universität DresdenDresdenGermany
  4. 4.Institute of Natural Materials TechnologyTechnische Universität DresdenDresdenGermany

Personalised recommendations