Abstract
Tissue-derived decellularized biomaterials are ideal for tissue engineering applications as they mimic the biochemical composition of the native tissue. These materials can be used as hydrogels for cell encapsulation and delivery. The decellularization process can alter the composition of the extracellular matrix (ECM) and thus influence the hydrogels characteristics. The aim of this study was to examine the impact of decellularization protocols in ECM-derived hydrogels obtained from porcine corneas. Porcine corneas were isolated and decellularized with SDS, Triton X-100 or by freeze-thaw cycles. All decellularization methods decreased DNA significantly when measured by PicoGreen and visually assessed by the absence of cell nuclei. Collagen and other ECM components were highly retained, as quantified by hydroxyproline content and sGAG, by histological analysis and by SDS-PAGE. Hydrogels obtained by freeze-thaw decellularization were the most transparent. The method of decellularization impacted gelation kinetics assessed by turbidimetric analysis. All hydrogels showed a fibrillary and porous structure determined by cryoSEM. Human corneal stromal cells were embedded in the hydrogels to assess cytotoxicity. SDS decellularization rendered cytotoxic hydrogels, while the other decellularization methods produced highly cytocompatible hydrogels. Freeze-thaw decellularization produced hydrogels with the overall best properties.
Highlights
Tissue-derived decellularized biomaterials are ideal for tissue engineering applications as they mimic the biochemical composition of the native tissue
PicoGreen was used to quantify DNA remnants, collagen content was measured by hydroxyproline quantification and sulphated glycosaminoglycans were quantified by dimethylmethylene blue assay (DMMB)
Collagen levels remained constant in all treatments (Fig. 1B). sulphated glycosaminoglycans (sGAG) levels were maintained when decellularization was performed with Triton or the freeze-thaw methods, while sodium dodecyl sulphate (SDS) resulted in significant loss of sGAG (Fig. 1C)
Summary
Tissue-derived decellularized biomaterials are ideal for tissue engineering applications as they mimic the biochemical composition of the native tissue. For many tissue-engineering applications, healthy cells need to be embedded into the ECM to generate a functional and viable tissue To overcome this problem decellularized organs and tissues can be transformed into hydrogels that allow cells to be encapsulated throughout their structure. Two detergent-based techniques (SDS and Triton X-100) and a freeze-thaw cycling technique were used to decellularize corneas and hydrogels were fabricated from the resulting ECM. The impact of these decellularization protocols were evaluated in terms of biochemical composition, transparency, gelation kinetics, mechanical properties and cytocompatibility
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