Abstract
Collagen-elastin (CE) scaffolds are frequently used for dermal replacement in the treatment of full-thickness skin defects such as burn wounds. But little is known about the optimal pore size and level of cross-linking. Different formulations of dermal substitutes with unidirectional pores were tested in porcine full-thickness wounds in combination with autologous split skin mesh grafts (SSG). Effect on wound healing was evaluated both macro- and microscopically. CE scaffolds with a pore size of 80 or 100 μm resulted in good wound healing after one-stage grafting. Application of scaffolds with a larger average pore size (120 μm) resulted in more myofibroblasts and more foreign body giant cells (FBGC). Moderate crosslinking impaired wound healing as it resulted in more wound contraction, more FBGC and increased epidermal thickness compared to no cross-linking. In addition, take rate and redness were negatively affected compared to SSG only. Vascularization and the number of myofibroblasts were not affected by cross-linking. Surprisingly, stability of cross-linked scaffolds was not increased in the wound environment, in contrast to in vitro results. Cross-linking reduced the proliferation of fibroblasts in vitro, which might explain the reduced clinical outcome. The non-cross-linked CE substitute with unidirectional pores allowed one-stage grafting of SSG, resulting in good wound healing. In addition, only a very mild foreign body reaction was observed. Cross-linking of CE scaffolds negatively affected wound healing on several important parameters. The optimal non-cross-linked CE substitute is a promising candidate for future clinical evaluation.
Similar content being viewed by others
References
van Zuijlen PPM, Vloemans JFPM, van Trier AJ, Suijker MH, van Unen E, Groenevelt F, et al. Dermal substitution in acute burns and reconstructive surgery: a subjective and objective long-term follow-up. Plast Reconstr Surg. 2001;108(7):1938–46.
Haslik W, Kamolz LP, Manna F, Hladik M, Rath T, Frey M. 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. 2010;63(2):360–4.
Ryssel H, Germann G, Kloeters O, Gazyakan E, Radu CA. Dermal substitution with Matriderm((R)) in burns on the dorsum of the hand. Burns. 2010;36(8):1248–53.
van Zuijlen PPM, van Trier AJ, Vloemans JFPM, Groenevelt F, Kreis RW, Middelkoop E. Graft survival and effectiveness of dermal substitution in burns and reconstructive surgery in a one-stage grafting model. Plast Reconstr Surg. 2000;106(3):615–23.
Bloemen MC, van Leeuwen MC, van Vucht NE, van Zuijlen PPM, Middelkoop E. Dermal substitution in acute burns and reconstructive surgery: a 12-year follow-up. Plast Reconstr Surg. 2010;125(5):1450–9.
Wang H, Pieper J, Peters F, van Blitterswijk CA, Lamme EN. Synthetic scaffold morphology controls human dermal connective tissue formation. J Biomed Mater Res, Part A. 2005;74(4):523–32.
Salem AK, Stevens R, Pearson RG, Davies MC, Tendler SJ, Roberts CJ, et al. Interactions of 3T3 fibroblasts and endothelial cells with defined pore features. J Biomed Mater Res. 2002;61(2):212–7.
Zeltinger J, Sherwood JK, Graham DA, Mueller R, Griffith LG. Effect of pore size and void fraction on cellular adhesion, proliferation, and matrix deposition. Tissue Eng. 2001;7(5):557–72.
O’Brien FJ, Harley BA, Yannas IV, Gibson LJ. The effect of pore size on cell adhesion in collagen-GAG scaffolds. Biomaterials. 2005;26(4):433–41.
Yannas IV, Lee E, Orgill DP, Skrabut EM, Murphy GF. Synthesis and characterization of a model extracellular matrix that induces partial regeneration of adult mammalian skin. Proc Natl Acad Sci USA. 1989;86(3):933–7.
Jarman-Smith ML, Bodamyali T, Stevens C, Howell JA, Horrocks M, Chaudhuri JB. Porcine collagen crosslinking, degradation and its capability for fibroblast adhesion and proliferation. J Mater Sci. 2004;15(8):925–32.
Grabarek Z, Gergely J. Zero-length crosslinking procedure with the use of active esters. Anal Biochem. 1990;185(1):131–5.
Olde Damink LHH, Dijkstra PJ, van Luyn MJA, van Wachem PB, Nieuwenhuis P, Feijen J. Cross-linking of dermal sheep collagen using a water-soluble carbodiimide. Biomaterials. 1996;17(8):765–73.
Olde Damink LHH, Dijkstra PJ, van Luyn MJA, van Wachem PB, Nieuwenhuis P, Feijen J. In vitro degradation of dermal sheep collagen cross-linked using a water-soluble carbodiimide. Biomaterials. 1996;17(7):679–84.
Zeeman R, Dijkstra PJ, van Wachem PB, van Luyn MJA, Hendriks M, Cahalan PT, et al. Successive epoxy and carbodiimide cross-linking of dermal sheep collagen. Biomaterials. 1999;20(10):921–31.
Marois Y, Chakfe N, Deng X, Marois M, How T, King MW, et al. Carbodiimide cross-linked gelatin: a new coating for porous polyester arterial prostheses. Biomaterials. 1995;16(15):1131–9.
Pieper JS, Oosterhof A, Dijkstra PJ, Veerkamp JH, van Kuppevelt TH. Preparation and characterization of porous crosslinked collagenous matrices containing bioavailable chondroitin sulphate. Biomaterials. 1999;20(9):847–58.
Thompson JI, Czernuszka JT. The effect of two types of cross-linking on some mechanical properties of collagen. Bio-Med Mater Eng. 1995;5(1):37–48.
Hafemann B, Ghofrani K, Gattner HG, Stieve H, Pallua N. Cross-linking by 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide (EDC) of a collagen/elastin membrane meant to be used as a dermal substitute: effects on physical, biochemical and biological features in vitro. J Mater Sci. 2001;12(5):437–46.
Hafemann B, Ensslen S, Erdmann C, Niedballa R, Zuhlke A, Ghofrani K, et al. Use of a collagen/elastin-membrane for the tissue engineering of dermis. Burns. 1999;25(5):373–84.
Hanthamrongwit M, Reid WH, Grant MH. Chondroitin-6-sulphate incorporated into collagen gels for the growth of human keratinocytes: the effect of cross-linking agents and diamines. Biomaterials. 1996;17(8):775–80.
Powell HM, Boyce ST. EDC cross-linking improves skin substitute strength and stability. Biomaterials. 2006;27(34):5821–7.
Buijtenhuijs P, Buttafoco L, Poot AA, Daamen WF, van Kuppevelt TH, Dijkstra PJ, et al. Tissue engineering of blood vessels: characterization of smooth-muscle cells for culturing on collagen-and-elastin-based scaffolds. Biotechnol Appl Biochem. 2004;39(Pt 2):141–9.
Park SN, Park JC, Kim HO, Song MJ, Suh H. Characterization of porous collagen/hyaluronic acid scaffold modified by 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide cross-linking. Biomaterials. 2002;23(4):1205–12.
Pieper JS, van Wachem PB, van Luyn MJA, Brouwer LA, Hafmans T, Veerkamp JH, et al. Attachment of glycosaminoglycans to collagenous matrices modulates the tissue response in rats. Biomaterials. 2000;21(16):1689–99.
Bozkurt A, Brook GA, Moellers S, Lassner F, Sellhaus B, Weis J, et al. In vitro assessment of axonal growth using dorsal root ganglia explants in a novel three-dimensional collagen matrix. Tissue Eng. 2007;13(12):2971–9.
Kroehne V, Heschel I, Schugner F, Lasrich D, Lasrich D, Bartsch JW, Jockusch H. Use of a novel collagen matrix with oriented pore structure for muscle cell differentiation in cell culture and in grafts. J Cel Mol Med. 2008;12(5A):1640–8.
Kuberka M, von Heimburg D, Schoof H, Heschel I, Rau G. Magnification of the pore size in biodegradable collagen sponges. Int J Artif Organs. 2002;25(1):67–73.
Bozkurt A, Deumens R, Beckmann C, Olde Damink L, Schugner F, Heschel I, et al. In vitro cell alignment obtained with a Schwann cell enriched microstructured nerve guide with longitudinal guidance channels. Biomaterials. 2009;30(2):169–79.
Heijmen FH, du Pont JS, Middelkoop E, Kreis RW, Hoekstra MJ. Cross-linking of dermal sheep collagen with tannic acid. Biomaterials. 1997;18(10):749–54.
Li CQ, Huang B, Luo G, Zhang CZ, Zhuang Y, Zhou Y. Construction of collagen II/hyaluronate/chondroitin-6-sulfate tri-copolymer scaffold for nucleus pulposus tissue engineering and preliminary analysis of its physico-chemical properties and biocompatibility. J Mater Sci. 2010;21(2):741–51.
Middelkoop E, van den Bogaerdt AJ, Lamme EN, Hoekstra MJ, Brandsma K, Ulrich MMW. Porcine wound models for skin substitution and burn treatment. Biomaterials. 2004;25(9):1559–67.
van den Bogaerdt AJ, van Zuijlen PP, van Galen M, Lamme EN, Middelkoop E. The suitability of cells from different tissues for use in tissue-engineered skin substitutes. Arch Dermatol Res. 2002;294(3):135–42.
Coolen NA, Vlig M, van den Bogaerdt AJ, Middelkoop E, Ulrich MMW. Development of an in vitro burn wound model. Wound Repair Regen. 2008;16(4):559–67.
Yang P, Chen L, Zwart R, Kijlstra A. Immune cells in the porcine retina: distribution, characterization and morphological features. Invest Ophthalmol Vis Sci. 2002;43(5):1488–92.
Dominguez J, Ezquerra A, Alonso F, McCullough K, Summerfield A, Bianchi A, et al. Porcine myelomonocytic markers: summary of the Second International Swine CD Workshop. Vet Immunol Immunopathol. 1998;60(3–4):329–41.
van Wachem PB, van Luyn MJA, Olde Damink LHH, Dijkstra PJ, van Luyn MJA, van Wachem PB, Nieuwenhuis P. Biocompatibility and tissue regenerating capacity of crosslinked dermal sheep collagen. J Biomed Mater Res. 1994;28(3):353–63.
Osborne CS, Reid WH, Grant MH. Investigation into cell growth on collagen/chondroitin-6-sulphate gels: the effect of crosslinking agents and diamines. J Mater Sci. 1997;8(4):179–84.
Dainiak MB, Allan IU, Savina IN, Cornelio L, James ES, James SL, et al. Gelatin-fibrinogen cryogel dermal matrices for wound repair: preparation, optimisation and in vitro study. Biomaterials. 2010;31(1):67–76.
McKegney M, Taggart I, Grant MH. The influence of crosslinking agents and diamines on the pore size, morphology and the biological stability of collagen sponges and their effect on cell penetration through the sponge matrix. J Mater Sci. 2001;12(9):833–44.
Harrison CA, Gossiel F, Layton CM, Bullock AJ, Johnson T, Blumsohn A, et al. Use of an in vitro model of tissue-engineered skin to investigate the mechanism of skin graft contraction. Tissue Eng. 2006;12(11):3119–33.
Chevallay B, Abdul-Malak N, Herbage D. Mouse fibroblasts in long-term culture within collagen three-dimensional scaffolds: influence of crosslinking with diphenylphosphorylazide on matrix reorganization, growth, and biosynthetic and proteolytic activities. J Biomed Mater Res. 2000;49(4):448–59.
Powell HM, Boyce ST. Wound closure with EDC cross-linked cultured skin substitutes grafted to athymic mice. Biomaterials. 2007;28(6):1084–92.
Badylak SF, Valentin JE, Ravindra AK, McCabe GP, Stewart-Akers AM. Macrophage phenotype as a determinant of biologic scaffold remodeling. Tissue Eng Part A. 2008;14(11):1835–42.
Ye Q, Harmsen MC, van Luyn MJ, Bank RA. The relationship between collagen scaffold cross-linking agents and neutrophils in the foreign body reaction. Biomaterials. 2010;31(35):9192–201.
Anderson JM, Rodriguez A, Chang DT. Foreign body reaction to biomaterials. Semin Immunol. 2008;20(2):86–100.
Bloemen MC, van der Wal MB, Verhaegen PD, Nieuwenhuis MK, van Baar ME, van Zuijlen PP, et al. Clinical effectiveness of dermal substitution in burns by topical negative pressure: a multicenter randomized controlled trial. Wound Repair Regen. 2012;20(6):797–805.
Hinz B. Masters and servants of the force: the role of matrix adhesions in myofibroblast force perception and transmission. Eur J Cell Biol. 2006;85(3–4):175–81.
Dagalakis N, Flink J, Stasikelis P, Burke JF, Yannas IV. Design of an artificial skin. Part III. Control of pore structure. J Biomed Mater Res. 1980;14(4):511–28.
Suzuki S, Matsuda K, Isshiki N, Tamada Y, Ikada Y. Experimental study of a newly developed bilayer artificial skin. Biomaterials. 1990;11(5):356–60.
O’Brien FJ, Harley BA, Waller MA, Yannas IV, Gibson LJ, Prendergast PJ. The effect of pore size on permeability and cell attachment in collagen scaffolds for tissue engineering. Technol Health Care. 2007;15(1):3–17.
Roten SV, Bhat S, Bhawan J. Elastic fibers in scar tissue. J Cutan Pathol. 1996;23(1):37–42.
Sullivan TP, Eaglstein WH, Davis SC, Mertz P. The pig as a model for human wound healing. Wound Repair Regen. 2001;9(2):66–76.
de Vries HJC, Middelkoop E, Mekkes JR, Dutrieux RP, Wildevuur CH, Westerhof H. Dermal regeneration in native non-cross-linked collagen sponges with different extracellular matrix molecules. Wound Repair Regen. 1994;2(1):37–47.
Ryssel H, Gazyakan E, Germann G, Ohlbauer M. The use of MatriDerm in early excision and simultaneous autologous skin grafting in burns—a pilot study. Burns. 2008;34(1):93–7.
Acknowledgments
This work was supported by the Dutch Burns Foundation in the Netherlands (Grant #06-203).
Author information
Authors and Affiliations
Corresponding author
Electronic supplementary material
Below is the link to the electronic supplementary material.
Rights and permissions
About this article
Cite this article
Boekema, B.K.H.L., Vlig, M., Olde Damink, L. et al. Effect of pore size and cross-linking of a novel collagen-elastin dermal substitute on wound healing. J Mater Sci: Mater Med 25, 423–433 (2014). https://doi.org/10.1007/s10856-013-5075-2
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s10856-013-5075-2