Functionalised Hybrid Collagen-Elastin for Acellular Cutaneous Substitute Applications

Abstract

Wound contracture, which commonly happens after wound healing, may lead to physical distortion, including skin constriction. Therefore, the combination of collagen and elastin as the most abundant extracellular matrix (ECM) skin matrices may provide the best candidate biomaterials for cutaneous wound injury. This study aimed to develop a hybrid scaffold containing green natural resources (ovine tendon collagen type-I and poultry-based elastin) for skin tissue engineering. Briefly, freeze-drying was used to create the hybrid scaffolds, which were then crosslinked with 0.1% (w/v) genipin (GNP). Next, the physical characteristics (pore size, porosity, swelling ratio, biodegradability and mechanical strength) of the microstructure were assessed. Energy dispersive X-ray spectroscopy (EDX) and Fourier transform infrared (FTIR) spectrophotometry were used for the chemical analysis. The findings showed a uniform and interconnected porous structure with acceptable porosity (>60%) and high-water uptake capacity (>1200%), with pore sizes ranging between 127 ± 22 and 245 ± 35 µm. The biodegradation rate of the fabricated scaffold containing 5% elastin was lower (<0.043 mg/h) compared to the control scaffold (collagen only; 0.085 mg/h). Further analysis with EDX identified the main elements of the scaffold: it contained carbon (C) 59.06 ± 1.36-70.66 ± 2.89%, nitrogen (N) 6.02 ± 0.20-7.09 ± 0.69% and oxygen (O) 23.79 ± 0.65-32.93 ± 0.98%. FTIR analysis revealed that collagen and elastin remained in the scaffold and exhibited similar functional amides (amide A: 3316 cm^-1, amide B: 2932 cm^-1, amide I: 1649 cm^-1, amide II: 1549 cm^-1 and amide III: 1233 cm^-1). The combination of elastin and collagen also produced a positive effect via increased Young's modulus values. No toxic effect was identified, and the hybrid scaffolds significantly supported human skin cell attachment and viability. In conclusion, the fabricated hybrid scaffolds demonstrated optimum physicochemical and mechanical properties and may potentially be used as an acellular skin substitute in wound management.

Keywords: acellular skin substitute; collagen; elastin; hybrid bioscaffold; tissue engineering.

Figure 1

(a) Gross appearance of the genipin-crosslinked scaffolds after crosslinking at 25 °C. (b) The cross-sectional FESEM images of the scaffolds at 100× magnification (Scale bar = 200 µm). (c) Measured porosity of Col/Elas scaffolds. (d) Mean pore sizes of Col/Elas scaffolds. (*) represents a significant difference (p < 0.05; n = 3) between the scaffolds.

Figure 2

(a) The degree of crosslinking of Col/Elas scaffolds crosslinked by 0.1% genipin. (b) The enzymatic degradation of Col/Elas scaffolds in 0.0006% collagenase at 37 °C. (c) The water absorption capacity (%) of Col/Elas scaffolds. (d) Young’s modulus of Col/Elas scaffolds. (*) represents a significant difference (p < 0.05; n = 3) between fabricated scaffolds.

Figure 3

The FTIR spectra of Col/Elas scaffolds crosslinked by 0.1% genipin.

Figure 4

Cell-scaffold interaction in 3D scaffolds. (a) Live/Dead assay of the fabricated scaffolds; Scale bar: 100 µm. (b) The quantification of cell attachment at 24 h. (c) Cell viability of HDF on the scaffolds at day 1, 5 and 7 quantified by MTT assay. (d) Cross-sectional morphological features of HDF at 500× magnification; Scale bar: 20 µm. (*) represents a significant difference (p < 0.05; n = 3) between fabricated scaffolds.

Figure 4

Cell-scaffold interaction in 3D scaffolds. (a) Live/Dead assay of the fabricated scaffolds; Scale bar: 100 µm. (b) The quantification of cell attachment at 24 h. (c) Cell viability of HDF on the scaffolds at day 1, 5 and 7 quantified by MTT assay. (d) Cross-sectional morphological features of HDF at 500× magnification; Scale bar: 20 µm. (*) represents a significant difference (p < 0.05; n = 3) between fabricated scaffolds.