title: Fabrication and characterization of nano-fibrous bilayer composite for skin regeneration application.
abstract: Full thickness wound healing with minimal scarring and complete restoration of normal skin properties still remains as a clinical challenge. In this study, a bilayer skin substitute has been fabricated to biomimic the microstructure of natural extracellular matrix of the skin. Human amniotic membrane (HAM) and silk fibroin nano-fibers were combined to produce bilayer construct, which was further treated and characterized. HAM was obtained from healthy mothers and de-epithelized by means of fine enzymatic method to preserve the extracellular structure. Fibroin protein was extracted from fresh Bombyx mori cocoons and transformed to uniform nano-fiberous structure, which was used as a coating layer on the de-epithelized membrane. Surface modification through oxygen plasma treatment was attempted to further induce hydrophilicity. Subsequently, scaffolds were fully characterized in terms of morphology, mechanical properties, hydrophilicity and cell culture response. Histological and immunohistological staining demonstrated localization of fibronectin, cell denudation and structural integrity of HAM after de-epithelization. Scanning electron microscopy images showed bead-free silk fibroin nano-fibers with the average diameter of 250nm. Water contact angle of bilayer scaffolds reduced dramatically to 26.34° after oxygen plasma treatment, which is correlated with more hydrophilic surface. Due to fibroin nano-fiber coating, mechanical properties of HAM improved significantly. Tensile Young's modulus and tensile strength increased from 16.14MPa and 68.46MPa to 25.69MPa and 108.03MPa, respectively. 14days in vitro cultivation of mouse embryonic fibroblasts on the scaffolds revealed that bilayer scaffolds are able to support cell attachment and proliferation. Plasma-etched scaffolds provided the best niche for cell-matrix crosstalk by allowing cells to penetrate beneath the pores and to integrate in fibers direction. The obtained results suggest that the presented nano-fibrous bilayer composite based on HAM is a potential substitute for skin regeneration application.
authors: Reproductive Biotechnology Research Center, Avicenna Research Institute, ACECR, Tehran, Iran.
authors: Shaghayegh
authors: S
authors: Arasteh
authors: Reproductive Biotechnology Research Center, Avicenna Research Institute, ACECR, Tehran, Iran.
authors: Somaieh
authors: S
authors: Kazemnejad
authors: Reproductive Biotechnology Research Center, Avicenna Research Institute, ACECR, Tehran, Iran.
authors: Sayeh
authors: S
authors: Khanjani
authors: Reproductive Biotechnology Research Center, Avicenna Research Institute, ACECR, Tehran, Iran.
authors: Hamed
authors: H
authors: Heidari-Vala
authors: Reproductive Biotechnology Research Center, Avicenna Research Institute, ACECR, Tehran, Iran.
authors: Mohammad Mehdi
authors: MM
authors: Akhondi
authors: Reproductive Biotechnology Research Center, Avicenna Research Institute, ACECR, Tehran, Iran; Experimental Orthopedics & Trauma Surgery, Frankfurt Initiative for Regenerative Medicine, J.W. Goethe-University Frankfurt, Germany. Electronic address: sahba.mobini@manchester.ac.uk.
authors: Sahba
authors: S
authors: Mobini
mesh_terms: D000650:Amnion / Q000166:cytology; D000818:Animals; D001672:Biocompatible Materials / Q000737:chemistry*; D012831:Bombyx; D018929:Cell Culture Techniques; D002478:Cells, Cultured; D055119:Elastic Modulus; D005260:Female; D005349:Fibroins / Q000737:chemistry; D006801:Humans; D057927:Hydrophobic and Hydrophilic Interactions; D051379:Mice; D053761:Nanocomposites / Q000737:chemistry / Q000648:ultrastructure; D057139:Nanofibers / Q000737:chemistry* / Q000648:ultrastructure; D062326:Re-Epithelialization; D023822:Tissue Engineering; D054457:Tissue Scaffolds / Q000737:chemistry*
chemical_list: D001672:Biocompatible Materials; D005349:Fibroins
keywords: Bilayer scaffold; Electrospinning; Human amniotic membrane; Silk; Skin tissue engineering