A novel egg-shell membrane based hybrid nanofibrous scaffold for cutaneous tissue engineering

Abstract

BackgroundThe main issue in cutaneous regeneration is to develop engineered scaffolds based on natural extracellular matrix to promote dynamics of skin progenitor cells and accelerate differentiation into mature keratinocytes.MethodsIn this study, nanofibrous scaffolds composed of a blend poly (ɛ-caprolactone) (PCL), silk fibroin (SF), soluble eggshell membrane (SESM), and Aloe vera (AV) gel were developed by electrospinning method and human basal cells were used to examine differentiation capacity toward keratinocyte-like cells. For this propose, cells were allocated to four distinct groups; control, PCL/SF, PCL/SF/SESM, and PCL/SF/SESM/AV. In all groups, cells were incubated with differentiation medium. Morphology, composition, hydrophilicity and mechanical features of PCL/SF, PCL/SF/SESM and PCL/SF/SESM/AV nanofibers were studied by scanning electron microscopy (SEM), Fourier transforms infrared spectroscopy (FT-IR), water contact angle and tensile tests. To examine the orientation of basal cells to mature keratinocytes, we performed immunofluorescence analysis by monitoring cytokeratin-19. The expression of genes such as involucrin, keratin-14 and -5 was monitored by real-time PCR assay.ResultsPCL/SF, PCL/SF/SESM, and PCL/SF/SESM/AV had suitable physic chemical indices and biological activities to be applied as biomimetic scaffolds for the restoration cutaneous tissue. Compared to control, we found an increased basal cell proliferation at 7 and 14 days after plating on scaffolds and reach maximum levels in group PCL/SF/SESM/AV on day 14 (p < 0.05). Electron microscopy showed cell flattening, morphological adaptation. An integrated cell-to-cell connection was generated after cell seeding on scaffolds in all groups. Immunofluorescence imaging showed the ability of basal cells to synthesize cytokeratin-19 in PCL/SF, PCL/SF/SESM, and positive control cells after exposure to differentiation medium. However, these values were less in PCL/SF/SESM/AV compared to other groups. Real-time PCR analysis showed the potency of all scaffolds to induce the transcription of involucrin, keratin-14 and -5, especially involucrin in PCL/SF/SESM/AV group compared to the negative control.ConclusionModulation of scaffolds with natural biopolymers could enable us to synthesize structures appropriate for cutaneous regeneration.

State-of-the-Art of Eggshell Waste in Materials Science: Recent Advances in Catalysis, Pharmaceutical Applications, and Mechanochemistry

“…It was suggested as an important component of various wound-care dressings (Guarderas et al, 2016 ; Kiselioviene et al, 2016 ; Li et al, 2016 , 2019b , c ; Krishnan et al, 2020 ). ESM was also used to produce hybrid nanofibrous scaffolds for cutaneous tissue engineering (Mohammadzadeh et al, 2019 ), materials for periodontal tissues (Motoji et al, 2020 ), and for sciatic nerve regeneration (Farjah et al, 2020 ). The nanofibrous scaffolds based on the natural extracellular matrix promote the dynamics of skin progenitor cells and accelerate differentiation into mature keratinocytes (Mohammadzadeh et al, 2019 ).…”

“…Each of them can have a pronounced influence on wound healing as a biologically active compound. Proteoglycans in ESM have been used successfully in treating non-healing wounds and burns, due to their biocompatibility, biodegradability, and similarity to macromolecules found in the human body (Benson et al, 2016 ; Mohammadzadeh et al, 2019 ).…”

“…Collagen (several types, in particular I, V, X) was named as one of the components that accounts for the biological activity of ESM (Yamauchi et al, 2013 ; Benson et al, 2016 ; Tummalapalli et al, 2016 ; Ahmed et al, 2017 , 2019a , b ; Dubourdieu, 2019 ; Mohammadzadeh et al, 2019 ; Farjah et al, 2020 ; Liu et al, 2020b ; Motoji et al, 2020 ). ESM exerts an anti-aging effect by increasing type III collagen levels (Motoji et al, 2020 ).…”

“…The presence of essential structural proteins such as osteopontin, sialoprotein, keratin, proteoglycans, and glycoproteins has been previously proved (Wong et al, 1984 ; Chowdhury, 1990 ; Arias et al, 1991 ; Nakano et al, 2003 ; Zhao and Chi, 2009 ; Hincke et al, 2012 ; Dombre et al, 2017 ; Silva et al, 2018 ). Also, ESM harbors numerous natural glycoproteins, notably glucosamine, chondroitin, and hyaluronic acid, which are applicable for the cutaneous wound dressings (Mohammadzadeh et al, 2019 ).…”

“…As another example, one can mention the nanofibrous scaffolds composed of a blend of poly(ε-caprolactone) (PCL), silk fibroin (SF), soluble eggshell membrane (SESM), and Aloe vera (AV) gel (Mohammadzadeh et al, 2019 ). Such composites were also developed by electrospinning.…”