Long-term stabilization of polysaccharide electrospun fibres by in situ cross-linking

Cross-linking of polysaccharide electrospun constructs using currently available techniques results in poor scaffold structural stability. In general, cross-linked substrates lose their nanofibrous architecture within a short time under physiological conditions. In this study, we introduce an in situ cross-linking electrospinning technique to fabricate and stabilize pullulan/dextran fibres. Pullulan/dextran (4:1 weight ratio, 16.7 and 20 wt%) solutions were preloaded with the chemical cross-linker, trisodium trimetaphosphate (STMP), to enable cross-linking during electrospinning. By increasing STMP from 4 to 16 wt%, the average diameter of electrospun fibres increased significantly from 268 ± 35 nm to 416 ± 74 nm (P < 0.05). Additionally, the enhanced cross-linking effectively decreased the swelling extent of the scaffolds. In particular, in the presence of 10 wt% gelatin, a significant decrease in scaffold swelling ratio was observed (208.5 ± 31.3% at 4 wt% STMP vs 133.1 ± 9.1% at 16 wt% STMP, P < 0.05). In vitro stability studies demonstrated the retention of scaffold fibrous morphology and negligible weight loss in all samples after 28 days. Environmental SEM analysis revealed that at least 16 wt% STMP was required in order to retain the nanofibrous structure of the scaffolds under hydrated conditions. Compared with hydrogels of similar chemical content, the nanofibrous architecture of electrospun scaffolds significantly enhanced human dermal fibroblast (HDF) viability at days 3 and 7 (P < 0.05). The incorporation of gelatin and the increase in scaffold cross-linking density favoured HDF cell attachment and spreading. In particular, 16 wt% STMP promoted actin stress fibre formation. Taken together, the results support the promise of using STMP in situ cross-linking for long-term stabilization of polysaccharide electrospun fibres and the advantage of polysaccharide nanofibrous constructs for tissue engineering.