Plant-derived human collagen scaffolds for skin tissue engineering.

Tissue engineering scaffolds are commonly formed using proteins extracted from animal tissues, such as bovine hide. Risks associated with the use of these materials include hypersensitivity and pathogenic contamination. Human-derived proteins lower the risk of hypersensitivity, but possess the risk of disease transmission. Methods engineering recombinant human proteins using plant material provide an alternate source of these materials without the risk of disease transmission or concerns regarding variability. To investigate the utility of plant-derived human collagen (PDHC) in the development of engineered skin (ES), PDHC and bovine hide collagen were formed into tissue engineering scaffolds using electrospinning or freeze-drying. Both raw materials were easily formed into two common scaffold types, electrospun nonwoven scaffolds and lyophilized sponges, with similar architectures. The processing time, however, was significantly lower with PDHC. PDHC scaffolds supported primary human cell attachment and proliferation at an equivalent or higher level than the bovine material. Interleukin-1 beta production was significantly lower when activated THP-1 macrophages where exposed to PDHC electrospun scaffolds compared to bovine collagen. Both materials promoted proper maturation and differentiation of ES. These data suggest that PDHC may provide a novel source of raw material for tissue engineering with low risk of allergic response or disease transmission.

Dehydrothermal crosslinking of electrospun collagen.

Electrospun collagen scaffolds must be crosslinked to improve stability. Chemical crosslinking methods are often associated with cytotoxicity and can require lengthy rinsing procedures to remove the crosslinker. Physical crosslinking using dehydrothermal (DHT) treatment is utilized to stabilize fibrous collagen sponges; however, little is known regarding the effect of DHT crosslinking on electrospun collagen. To investigate the efficacy of DHT crosslinking, soluble type I collagen was electrospun and exposed to DHT crosslinking, chemical crosslinking with N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride (EDC; 5 mM), and DHT+EDC. DHT crosslinking produced no change in scaffold fiber diameter or interfiber distance and reduced scaffold degradation. Strength was significantly improved by DHT (139.0 ± 34.9 kPa) compared to control but was weaker than EDC or DHT+EDC (222.7 ± 58.4, 353.3 ± 19.0 kPa, respectively). Fourier transform infrared spectroscopy (FTIR) indicated increased amide bond formation with DHT compared to control but a lower amide bond density than EDC or DHT+EDC. After crosslinking, sterilization, and rinsing (a total of 50 h for DHT, 98 h for EDC, and 122 h for DHT+EDC), fibroblasts adhered and proliferated on all scaffolds; however, cell metabolism was 12% less on DHT scaffolds. These data indicate that DHT crosslinking can be utilized to stabilize electrospun collagen scaffolds; however, a tradeoff exists between scaffold stability/strength and rapid processing.