Optimisation of UV irradiation as a binding site conserving method for crosslinking collagen-based scaffolds

Abstract

Short wavelength (λ = 254 nm) UV irradiation was evaluated over a range of intensities (0.06 to 0.96 J/cm2) as a means of cross-linking collagen- and gelatin-based scaffolds, to tailor their material characteristics whilst retaining biological functionality. Zero-link carbodiimide treatments are commonly applied to collagen-based materials, forming cross-links from carboxylate anions (for example the acidic E of GFOGER) that are an essential part of integrin binding sites on collagen. Cross-linking these amino acids therefore disrupts the bioactivity of collagen. In contrast, UV irradiation forms bonds from less important aromatic tyrosine and phenylalanine residues. We therefore hypothesised that UV cross-linking would not compromise collagen cell reactivity. Here, highly porous (~99 %) isotropic, collagen-based scaffolds were produced via ice-templating. A series of scaffolds (pore diameters ranging from 130–260 μm) with ascending stability in water was made from gelatin, two different sources of collagen I, or blends of these materials. Glucose, known to aid UV crosslinking of collagen, was added to some lower-stability formulations. These scaffolds were exposed to different doses of UV irradiation, and the scaffold morphology, dissolution stability in water, resistance to compression and cell reactivity was assessed. Stabilisation in aqueous media varied with both the nature of the collagen-based material employed and the UV intensity. Scaffolds made from the most stable materials showed the greatest stability after irradiation, although the levels of cross-linking in all cases were relatively low. Scaffolds made from pure collagen from the two different sources showed different optimum levels of irradiation, suggesting altered balance between stabilisation from cross-linking and destabilisation from denaturation. The introduction of glucose into the scaffold enhanced the efficacy of UV cross-linking. Finally, as hypothesized, cell attachment, spreading and proliferation on collagen materials were unaffected by UV cross-linking. UV irradiation may therefore be used to provide relatively low level cross-linking of collagen without loss of biological functionality.Electronic supplementary materialThe online version of this article (doi:10.1007/s10856-015-5627-8) contains supplementary material, which is available to authorized users.

Comprehensive Identification of RNA-Binding Domains in Human Cells

“…First, UV light is applied to cell monolayers to covalently stabilize native protein-RNA interactions taking place at “zero” distance (Pashev et al., 1991). While UV exposure using dosages exceeding those used here can potentially promote protein-protein crosslinking (Davidenko et al., 2016, Suchanek et al., 2005), we could not detect such crosslinks under our conditions, evidenced by the lack of UV-dependent, high molecular weight complexes in RNase-treated samples (Figures S1A and S4A; Strein et al., 2014). …”

Rose Bengal Crosslinking to Stabilize Collagen Sheets and Generate Modulated Collagen Laminates

“…Some crosslinkers like glutaraldehyde may also be cytotoxic and lead to calcification [ 18 , 23 ]. For physical crosslinking, dehydrothermal treatment (DHT) [ 24 , 25 ] and UV irradiation [ 10 , 26 , 27 ] are commonly used. However, these methods can lead to partial degradation of collagen and to protein denaturation [ 24 ].…”

Collagen type I and hyaluronic acid based hybrid scaffolds for heart valve tissue engineering

“…Therefore, a hybrid scaffold with heterogeneous microstructure taking advantage of its constituent natural polymers has the potential to meet these challenges. Collagen/HA scaffolds have been developed with hybrid microstructures using physical methods (dehydrothermal treatment and irradiation techniques) or chemical methods . Physical methods involve the risk of denaturation of collagen triple helix structure at higher temperature whereas glutaraldehyde and genipin impart undesirable coloration to the scaffold .…”