Drug Delivery and Translational Research

, Volume 7, Issue 1, pp 132–146 | Cite as

A methylcellulose and collagen based temperature responsive hydrogel promotes encapsulated stem cell viability and proliferation in vitro

  • Christina Payne
  • Eimear B. Dolan
  • Janice O’Sullivan
  • Sally-Ann Cryan
  • Helena M. KellyEmail author
Original Article


With the number of stem cell-based therapies emerging on the increase, the need for novel and efficient delivery technologies to enable therapies to remain in damaged tissue and exert their therapeutic benefit for extended periods, has become a key requirement for their translation. Hydrogels, and in particular, thermoresponsive hydrogels, have the potential to act as such delivery systems. Thermoresponsive hydrogels, which are polymer solutions that transform into a gel upon a temperature increase, have a number of applications in the biomedical field due to their tendency to maintain a liquid state at room temperature, thereby enabling minimally invasive administration and a subsequent ability to form a robust gel upon heating to physiological temperature. However, various hurdles must be overcome to increase the clinical translation of hydrogels as a stem cell delivery system, with barriers including their low tensile strength and their inadequate support of cell viability and attachment. In order to address these issues, a methylcellulose based hydrogel was formulated in combination with collagen and beta glycerophosphate, and key development issues such as injectability and sterilisation processes were examined. The polymer solution underwent thermogelation at ~36 °C as determined by rheological analysis, and when gelled, was sufficiently robust to resist significant disintegration in the presence of phosphate buffered saline (PBS) while concomitantly allowing for diffusion of methylene blue dye solution into the gel. We demonstrate that human mesenchymal stem cells (hMSCs) encapsulated within the gel remained viable and showed raised levels of dsDNA at increasing time points, an indication of cell proliferation. Mechanical testing showed the “injectability”, i.e. force required for delivery of the polymer solution through devices such as a syringe, needle or catheter. Sterilisation of the freeze-dried polymer wafer via gamma irradiation showed no adverse effects on the formed hydrogel characteristics. Taken together, these results indicate the potential of this gel as a clinically translatable delivery system for stem cells and therapeutic molecules in vivo.


Mesenchymal stem cells Thermoresponsive Hydrogel Methylcellulose Collagen Gamma irradiation Injectability 



human mesenchymal stem cells






internal diameter


storage modulus


loss modulus


standard error of mean



A bursary to allow for this work to be carried out was provided by the School of Pharmacy, Royal College of Surgeons in Ireland.

Compliance with ethical standards

Conflict of interests

The authors declare that they have no conflict of interest.


  1. 1.
    Orlando G, Wood KJ, Stratta RJ, Yoo JJ, Atala A, Soker S. Regenerative medicine and organ transplantation: past, present, and future. Transplantation. 2011;91(12):1310–7.CrossRefPubMedGoogle Scholar
  2. 2.
    Lau AN, Goodwin M, Kim CF, Weiss DJ. Stem cells and regenerative medicine in lung biology and diseases. Mol Ther. 2012;20(6):1116–30.CrossRefPubMedPubMedCentralGoogle Scholar
  3. 3.
    Krause DS, Theise ND, Collector MI, Henegariu O, Hwang S, Gardner R, Neutzel S, Sharkis SJ. Multi-organ, multi-lineage engraftment by a single bone marrow-derived stem cell. Cell. 2001;105(3):369–77.CrossRefPubMedGoogle Scholar
  4. 4.
    D. N. Kotton, B. Y. Ma, W. V Cardoso, E. A. Sanderson, R. S. Summer, M. C. Williams, and A. Fine, “Bone marrow-derived cells as progenitors of lung alveolar epithelium.,” Development, vol. 128, no. 24, pp. 5181–5188, 2001.Google Scholar
  5. 5.
    Orlic D, Kajstura J, Chimenti S, Jakoniuk I, Anderson SM, Li B, Pickel J, McKay R, Nadal-Ginard B, Bodine DM, Leri A, Anversa P. Bone marrow cells regenerate infarcted myocardium. Nature. 2001;410(6829):701–5.CrossRefPubMedGoogle Scholar
  6. 6.
    Antunes MA, Laffey JG, Pelosi P, Rocco PRM. Mesenchymal stem cell trials for pulmonary diseases. J Cell Biochem. 2014;115(6):1023–32.CrossRefPubMedGoogle Scholar
  7. 7.
    O’Neill HS, Gallagher LB, O’Sullivan J, Whyte W, Curley C, Dolan E, Hameed A, O’Dwyer J, Payne C, O’Reilly D, Ruiz-Hernandez E, Roche ET, O’Brien FJ, Cryan SA, Kelly H, Murphy B, Duffy GP. Biomaterial-enhanced cell and drug delivery: lessons learned in the cardiac field and future perspectives. Adv Mater. 2016:5648–61.Google Scholar
  8. 8.
    Ye Z, Zhou Y, Cai H, Tan W. Myocardial regeneration: roles of stem cells and hydrogels. Adv Drug Deliv Rev. 2011;63(8):688–97.CrossRefPubMedGoogle Scholar
  9. 9.
    Laflamme MA, Murry CE. Regenerating the heart. Nat Biotechnol. 2005;23(7):845–56.CrossRefPubMedGoogle Scholar
  10. 10.
    Zhang M, Methot D, Poppa V, Fujio Y, Walsh K, Murry CE. Cardiomyocyte grafting for cardiac repair: graft cell death and anti-death strategies. J Mol Cell Cardiol. 2001;33(5):907–21.CrossRefPubMedGoogle Scholar
  11. 11.
    Malliaras K, Marbán E. Cardiac cell therapy: where we’ve been, where we are, and where we should be headed. Br Med Bull. 2011;98:161–85.CrossRefPubMedPubMedCentralGoogle Scholar
  12. 12.
    Wang Q-D, Sjöquist P-O. Myocardial regeneration with stem cells: pharmacological possibilities for efficacy enhancement. Pharmacol Res. 2006;53(4):331–40.CrossRefPubMedGoogle Scholar
  13. 13.
    Hastings CL, Roche ET, Ruiz-Hernandez E, Schenke-Layland K, Walsh CJ, Duffy GP. Drug and cell delivery for cardiac regeneration. Adv Drug Deliv Rev. 2014;84:85–106.CrossRefPubMedGoogle Scholar
  14. 14.
    Liu Z, Wang H, Wang Y, Lin Q, Yao A, Cao F, Li D, Zhou J, Duan C, Du Z, Wang Y, Wang C. The influence of chitosan hydrogel on stem cell engraftment, survival and homing in the ischemic myocardial microenvironment. Biomaterials. 2012;33(11):3093–106.CrossRefPubMedGoogle Scholar
  15. 15.
    A. C. Gaffey, M. H. Chen, C. M. Venkataraman, A. Trubelja, C. B. Rodell, P. V Dinh, G. Hung, J. W. MacArthur, R. V Soopan, J. A. Burdick, and P. Atluri, “Injectable shear-thinning hydrogels used to deliver endothelial progenitor cells, enhance cell engraftment, and improve ischemic myocardium.,” J Thorac Cardiovasc Surg, vol. 150, no. 5, pp. 1268–1277, 2015.Google Scholar
  16. 16.
    Hoare TR, Kohane DS. Hydrogels in drug delivery: Progress and challenges. Polymer (Guildf). 2008;49(8):1993–2007.CrossRefGoogle Scholar
  17. 17.
    Klouda L, Mikos AG. Thermoresponsive hydrogels in biomedical applications. Eur J Pharm Biopharm. 2008;68(1):34–45.CrossRefPubMedGoogle Scholar
  18. 18.
    Chenite A, Chaput C, Wang D, Combes C, Buschmann MD, Hoemann CD, Leroux JC, Atkinson BL, Binette F, Selmani A. Novel injectable neutral solutions of chitosan form biodegradable gels in situ. Biomaterials. 2000;21(21):2155–61.CrossRefPubMedGoogle Scholar
  19. 19.
    Hastings CL, Kelly HM, Murphy MJ, Barry FP, O’Brien FJ, Duffy GP. Development of a thermoresponsive chitosan gel combined with human mesenchymal stem cells and desferrioxamine as a multimodal pro-angiogenic therapeutic for the treatment of critical limb ischaemia. J Control Release. 2012;161(1):73–80.CrossRefPubMedGoogle Scholar
  20. 20.
    Zhang Z, Ni J, Chen L, Yu L, Xu J, Ding J. Biodegradable and thermoreversible PCLA-PEG-PCLA hydrogel as a barrier for prevention of post-operative adhesion. Biomaterials. 2011;32(21):4725–36.CrossRefPubMedGoogle Scholar
  21. 21.
    Elstad NL, Fowers KD. OncoGel (ReGel/paclitaxel)--clinical applications for a novel paclitaxel delivery system. Adv Drug Deliv Rev. 2009;61(10):785–94.CrossRefPubMedGoogle Scholar
  22. 22.
    Shive MS, Hoemann CD, Restrepo A, Hurtig MB, Duval N, Ranger P, Stanish W, Buschmann MD. BST-CarGel: in situ ChondroInduction for cartilage repair. Oper Tech Orthop. 2006;16(4):271–8.CrossRefGoogle Scholar
  23. 23.
    Shalhoub J, Thapar A, Davies AH. The use of reverse thermosensitive polymer (LeGoo) for temporary vessel occlusion in clampless peripheral vascular surgery. Vasc Endovasc Surg. 2011;45(5):422–5.CrossRefGoogle Scholar
  24. 24.
    Buwalda SJ, Boere KWM, Dijkstra PJ, Feijen J, Vermonden T, Hennink WE. Hydrogels in a historical perspective: from simple networks to smart materials. J Control Release. 2014;190:254–73.Google Scholar
  25. 25.
    Higuchi A, Sugiyama K, Yoon BO, Sakurai M, Hara M, Sumita M, Sugawara S, Shirai T. Serum protein adsorption and platelet adhesion on pluronic™-adsorbed polysulfone membranes. Biomaterials. 2003;24(19):3235–45.CrossRefPubMedGoogle Scholar
  26. 26.
    Ruel-Gariépy E, Leroux J-C. In situ-forming hydrogels--review of temperature-sensitive systems. Eur J Pharm Biopharm. 2004;58(2):409–26.CrossRefPubMedGoogle Scholar
  27. 27.
    Yu L, Ding J. Injectable hydrogels as unique biomedical materials. Chem Soc Rev. 2008;37(8):1473–81.CrossRefPubMedGoogle Scholar
  28. 28.
    Supper S, Anton N, Seidel N, Riemenschnitter M, Curdy C, Vandamme T. Thermosensitive chitosan/glycerophosphate-based hydrogel and its derivatives in pharmaceutical and biomedical applications. Expert Opin Drug Deliv. 2014;11(2):249–67.CrossRefPubMedGoogle Scholar
  29. 29.
    Tang Y, Wang X, Li Y, Lei M, Du Y, Kennedy JF, Knill CJ. Production and characterisation of novel injectable chitosan/methylcellulose/salt blend hydrogels with potential application as tissue engineering scaffolds. Carbohydr Polym. 2010;82(3):833–41.CrossRefGoogle Scholar
  30. 30.
    Chenite A, Buschmann M, Wang D, Chaput C, Kandani N. Rheological characterisation of thermogelling chitosan / glycerol-phosphate solutions. Carbohydr Polym. 2001;46:39–47.CrossRefGoogle Scholar
  31. 31.
    Huang CC, Liao ZX, Chen DY, Hsiao CW, Chang Y, Sung HW. Injectable cell constructs fabricated via culture on a thermoresponsive methylcellulose hydrogel system for the treatment of ischemic diseases. Adv Healthc Mater. 2014;3(8):1133–48.CrossRefPubMedGoogle Scholar
  32. 32.
    Thirumala S, Gimble J, Devireddy R. Methylcellulose based thermally reversible hydrogel system for tissue engineering applications. Cells. 2013;2:460–75.CrossRefPubMedPubMedCentralGoogle Scholar
  33. 33.
    Wallace D. Collagen gel systems for sustained delivery and tissue engineering. Adv Drug Deliv Rev. 2003;55(12):1631–49.CrossRefPubMedGoogle Scholar
  34. 34.
    Wang L, Stegemann JP. Thermogelling chitosan and collagen composite hydrogels initiated with beta-glycerophosphate for bone tissue engineering. Biomaterials. 2010;31(14):3976–85.CrossRefPubMedPubMedCentralGoogle Scholar
  35. 35.
    Lee KY, Mooney DJ. Hydrogels for Tissue Engineering. Chem Rev. 2001;101(7):1869–80.CrossRefPubMedGoogle Scholar
  36. 36.
    Camci-Unal G, Annabi N, Dokmeci MR, Liao R, Khademhosseini A. Hydrogels for cardiac tissue engineering. NPG Asia Mater. 2014;6(5):e99.CrossRefGoogle Scholar
  37. 37.
    Liu X, Ma PX. Polymeric scaffolds for bone tissue engineering. Ann Biomed Eng. 2004;32(3):477–86.CrossRefPubMedGoogle Scholar
  38. 38.
    Burdick JA, Anseth KS. Photoencapsulation of osteoblasts in injectable RGD-modified PEG hydrogels for bone tissue engineering. Biomaterials. 2002;23(22):4315–23.CrossRefPubMedGoogle Scholar
  39. 39.
    Nichols JE, Kojima K, Bonassar LJ, Dargon P, Roy AK, Vacant MP, et al. Tissue-engineered lung: an in vivo and in vitro comparison of polyglycolic acid and pluronic F-127 hydrogel/somatic lung progenitor cell constructs to support tissue growth. Tissue Eng. 2006;12(5):1213–25.Google Scholar
  40. 40.
    O’Brien F. Influence of freezing rate on pore structure in freeze-dried collagen-GAG scaffolds. Biomaterials. Mar. 2004;25(6):1077–86.CrossRefPubMedGoogle Scholar
  41. 41.
    Duffy GP, Ahsan T, O’Brien T, Barry F, Nerem RM. Bone marrow-derived mesenchymal stem cells promote angiogenic processes in a time- and dose-dependent manner in vitro. Tissue Eng. Part A. 2009;15(9):2459–70.CrossRefPubMedGoogle Scholar
  42. 42.
    C. of Europe, European Pharmacopoeia, 5.0., no. 1. Strasbourg, 2005.Google Scholar
  43. 43.
    Bramley K, Pisani MA, Murphy TE, Araujo KL, Homer RJ, Puchalski JT. Endobronchial ultrasound-guided cautery-assisted transbronchial forceps biopsies: safety and sensitivity relative to transbronchial needle aspiration. Ann Thorac Surg. 2016;101(5):1870–6.CrossRefPubMedGoogle Scholar
  44. 44.
    Nilsen T, Hermann M, Eriksen CS, Dagfinrud H, Mowinckel P, Kjeken I. Grip force and pinch grip in an adult population: reference values and factors associated with grip force. Scand J Occup Ther. 2012;19(3):288–96.CrossRefPubMedGoogle Scholar
  45. 45.
    Rungseevijitprapa W, Bodmeier R. Injectability of biodegradable in situ forming microparticle systems (ISM). Eur J Pharm Sci. 2009;36(4–5):524–31.CrossRefPubMedGoogle Scholar
  46. 46.
    Lutolf MP. Biomaterials: spotlight on hydrogels. Nat Mater. 2009;8(6):451–3.CrossRefPubMedGoogle Scholar
  47. 47.
    E. T. Roche, C. L. Hastings, S. A. Lewin, D. E. Shvartsman, Y. Brudno, N. V Vasilyev, F. J. O’Brien, C. J. Walsh, G. P. Duffy, and D. J. Mooney, “Comparison of biomaterial delivery vehicles for improving acute retention of stem cells in the infarcted heart.,” Biomaterials, vol. 35, no. 25, pp. 6850–6858, 2014.Google Scholar
  48. 48.
    Aguado BA, Mulyasasmita W, Su J, Lampe KJ, Heilshorn SC. Improving viability of stem cells during syringe needle flow through the design of hydrogel cell carriers. Tissue Eng Part A. 2012;18(7–8):806–15.CrossRefPubMedGoogle Scholar
  49. 49.
    O’Brien FJ. Biomaterials & scaffolds for tissue engineering. Mater Today. 2011;14(3):88–95.CrossRefGoogle Scholar
  50. 50.
    Burdick JA, Mauck RL, Gerecht S. To serve and protect: hydrogels to improve stem cell-based therapies. Cell Stem Cell. 2016;18(1):13–5.CrossRefPubMedGoogle Scholar
  51. 51.
    Tate MC, Shear DA, Hoffman SW, Stein DG, LaPlaca MC. Biocompatibility of methylcellulose-based constructs designed for intracerebral gelation following experimental traumatic brain injury. Biomaterials. 2001;22(10):1113–23.CrossRefPubMedGoogle Scholar
  52. 52.
    Xu Y, Wang C, Tam KC, Li L. Salt-assisted and salt-suppressed sol-gel transitions of methylcellulose in water. Langmuir. 2004;20(3):646–52.CrossRefPubMedGoogle Scholar
  53. 53.
    Kobayashi K, Huang C, Lodge TP. Thermoreversible gelation of aqueous methylcellulose solutions. Macromolecules. 1999;32(21):7070–7.CrossRefGoogle Scholar
  54. 54.
    Chen K, Baker AN, Vyazovkin S. Concentration effect on temperature dependence of gelation rate in aqueous solutions of methylcellulose. Macromol Chem Phys. 2009;210(3–4):211–6.CrossRefGoogle Scholar
  55. 55.
    Yang Y-L, Leone LM, Kaufman LJ. Elastic moduli of collagen gels can be predicted from two-dimensional confocal microscopy. Biophys J. 2009;97(7):2051–60.CrossRefPubMedPubMedCentralGoogle Scholar
  56. 56.
    Helary C, Bataille I, Abed A, Illoul C, Anglo A, Louedec L, Letourneur D, Meddahi-Pellé A, Giraud-Guille MM. Concentrated collagen hydrogels as dermal substitutes. Biomaterials. 2010;31(3):481–90.CrossRefPubMedGoogle Scholar
  57. 57.
    Friess W. Collagen--biomaterial for drug delivery. Eur J Pharm Biopharm. 1998;45(2):113–36.CrossRefPubMedGoogle Scholar
  58. 58.
    Tang Y-F, Du Y-M, Hu X-W, Shi X-W, Kennedy JF. Rheological characterisation of a novel thermosensitive chitosan/poly(vinyl alcohol) blend hydrogel. Carbohydr Polym. 2007;67(4):491–9.CrossRefGoogle Scholar
  59. 59.
    Edsman K, Carlfors J, Petersson R. Rheological evaluation of poloxamer as an in situ gel for ophthalmic use. Eur J Pharm Sci. 1998;6(2):105–12.CrossRefPubMedGoogle Scholar
  60. 60.
    Nowak AP, Breedveld V, Pakstis L, Ozbas B, Pine DJ, Pochan D, et al. Rapidly recovering hydrogel scaffolds from self-assembling diblock copolypeptide amphiphiles. Nature. 2002;417(6887):424–8.Google Scholar
  61. 61.
    Lin H-R, Sung K. Carbopol/pluronic phase change solutions for ophthalmic drug delivery. J Control Release. 2000;69(3):379–88.CrossRefPubMedGoogle Scholar
  62. 62.
    Jeong B, Kim SW, Bae YH. Thermosensitive sol–gel reversible hydrogels. Adv Drug Deliv Rev. 2002;54(1):37–51.CrossRefPubMedGoogle Scholar
  63. 63.
    Kleinman HK, Klebe RJ, Martin GR. Role of collagenous matrices in the adhesion and growth of cells. J Cell Biol. 1981;88(3):473–85.CrossRefPubMedGoogle Scholar
  64. 64.
    Gupta D, Tator CH, Shoichet MS. Fast-gelling injectable blend of hyaluronan and methylcellulose for intrathecal, localized delivery to the injured spinal cord. Biomaterials. 2006;27(11):2370–9.CrossRefPubMedGoogle Scholar
  65. 65.
    Lee DW, Choi WS, Byun MW, Park HJ, Yu Y-M, Lee CM. Effect of gamma-irradiation on degradation of alginate. J Agric Food Chem. 2003;51:4819–23.CrossRefPubMedGoogle Scholar
  66. 66.
    Wasikiewicz JM, Yoshii F, Nagasawa N, Wach RA, Mitomo H. Degradation of chitosan and sodium alginate by gamma radiation, sonochemical and ultraviolet methods. Radiat Phys Chem. 2005;73(5):287–95.CrossRefGoogle Scholar
  67. 67.
    Sanina C, Hare JM. Mesenchymal stem cells as a biological drug for heart disease: where are We with cardiac cell-based therapy? Circ Res. 2015;117(3):229–33.CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© Controlled Release Society 2016

Authors and Affiliations

  • Christina Payne
    • 1
    • 2
  • Eimear B. Dolan
    • 1
    • 2
  • Janice O’Sullivan
    • 2
  • Sally-Ann Cryan
    • 1
    • 2
    • 3
    • 4
  • Helena M. Kelly
    • 1
    • 2
    Email author
  1. 1.School of PharmacyRoyal College of Surgeons in IrelandDublin 2Ireland
  2. 2.Tissue Engineering Research Group, Department of AnatomyRoyal College of Surgeons in IrelandDublin 2Ireland
  3. 3.Trinity Centre for BioengineeringTrinity College DublinDublin 2Ireland
  4. 4.Centre for Research in Medical Devices (CÚRAM)National University of Ireland GalwayGalwayIreland

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