ABSTRACT
For tissue engineering and cell therapy applications, expansion of cells such as chondrocytes on beads in spinner culture can provide advantages compared with monolayer culture. The use of resorbable beads that can be included as an integral part of the construct provides the advantage of minimizing the extent of cell handling and eliminating a final trypsin treatment to detach cells from the bead. In this study, we have made various types of beads based on native collagen and denatured collagen (gelatin). The beads have been stabilized by different extents of glutaraldehyde cross-linking, and characterized by a combination of chemical analysis, thermal stability, and microscopy. In vitro examination in the presence and absence of chondrocytes showed that stability increased with the extent of crosslinking and could also be influenced by the manner of fabrication. On the basis of the in vitro stability studies, gelatin beads of a defined stability were shown to resorb over time in subcutaneous implants in nude mice compared with more stable demineralized bone particle (DMB) carriers. These data indicate that for direct use in tissue engineering or cell therapy applications, where resorbable beads can be used for cell expansion and then direct delivery of cells, it is possible to design suitable carrier beads with a range of stabilities that match the implant requirements.
Subject(s)
Absorbable Implants , Biocompatible Materials/metabolism , Cell- and Tissue-Based Therapy , Collagen/metabolism , Tissue Engineering , Animals , Biocompatible Materials/chemistry , Cell- and Tissue-Based Therapy/instrumentation , Cell- and Tissue-Based Therapy/methods , Cells, Cultured , Chondrocytes/cytology , Chondrocytes/metabolism , Collagen/chemistry , Gelatin/chemistry , Materials Testing , Mice , Mice, Nude , Tissue Engineering/instrumentation , Tissue Engineering/methodsABSTRACT
We recently reported the generation of a highly elastic, crosslinked protein biomaterial via a rapid photochemical process using visible light illumination. In light of these findings, we predicted that other unmodified, tyrosine-rich, self-associating proteins might also be susceptible to this covalent crosslinking method. Here we show that unmodified native fibrinogen can also be photochemically crosslinked into an elastic hydrogel biomaterial through the rapid formation of intermolecular dityrosine. Photochemically crosslinked fibrinogen forms tissue sealant bonds at least 5-fold stronger than commercial fibrin glue and is capable of producing maximum bond strength within 20s. In vitro studies showed that components of the photochemical crosslinking reaction are non-toxic to cells. This material will find useful application in various surgical procedures where rapid curing for high strength tissue sealing is required.
Subject(s)
Fibrinogen/chemistry , Photochemistry/methods , Tissue Adhesives/chemistry , Animals , Cattle , Chromatography, High Pressure Liquid , Electrophoresis, Polyacrylamide Gel , Hydrogel, Polyethylene Glycol Dimethacrylate/chemistry , Mass SpectrometryABSTRACT
A method for the preparation of bioresorbable collagen beads with an open porous structure is presented. These beads were prepared from collagen-alginate composite beads by removal of the alginate component. These collagen beads were suitable for rapid proliferation of chondrocytes in a dynamic, spinner culture system. Histology and immuno-histology showed that biochemical markers of chondrocytes are present during this cell proliferation, indicating that there was control of phenotype and that cell de-differentiation had not occurred. Unlike other 3-D scaffolds that have been used, the cells were amplified from very low cell densities and were able to proliferate freely without loss of phenotype.
