# Fabricating a Novel Three-Dimensional Skin Model Using Silica Nonwoven Fabrics (SNF)
## 1 Introduction  
In tissue engineering, especially for regenerated tissues, it is important for cell scaffolds to function as a replacement for the extracellular matrix (ECM). The native ECM supports cells by providing structural [1] and biochemical conditions for cell adhesion, maintenance, proliferation, self-replication, and differentiation [2]. Therefore, the main goal of tissue engineering is to reproduce the cellular microenvironment that controls and regulates cellular functions in vivo using an in vitro culture system. The behavior of individual cells is regulated by their interactions with neighboring cells and ECM [3]. The complex biological systems in the in vivo microenvironment cannot be reproduced by cells cultured in 2D monolayers [4]. There are various culture methods to grow cells in a three-dimensional system, such as seeding in scaffolds comprising protein-based natural biomaterials [5], polymer hydrogels [6], and porous and fibrous materials [7, 8]. Protein-based natural biomaterials can be fabricated using collagen, gelatin, and fibrinogen. Collagen, the most abundant protein in the human dermis, is secreted by fibroblasts and is responsible for tensile rigidity of the ECM [9]. In a collagen matrix skin model, epidermal growth is enhanced when the collagen matrix is organized by fibroblasts [10]. Thus, the use of a collagen matrix as a scaffold for skin models leads to epidermal keratinization and mimicry of skin tissue. However, models using collagen matrices have problems such as shrinkage after long-term culture and inability to form elastin fibers important for the elasticity of the dermis [11]. As the cultivation period lengthens, the surviving epidermis shows all the signs of a normal differentiation program [12], so that long-term culture is important for skin models. Contraction of the human collagen matrix is regulated by tension homeostasis--the balance between the ECM and intracellular traction forces [13; 14; 15; 16]. To improve the poor mechanical properties of collagen matrices [17], investigations has been made to add poly (caprolactone) (PCL) to collagen [18], used gelatin instead of collagen [19], and reproduced the tension equilibrium by pulling the collagen matrix [20]. Silica nonwoven fabrics (SNF) prepared by electric field spinning via a sol-gel process developed by [PERSON] et al. [21; 22; 23] show high biocompatibility [24] and high thermal stability [25]. SNF with interconnected highly porous microstructures [26] mimic the fibrillar ECM [27; 28] and are effective as a three-dimensional scaffold for cell culture [29; 30]. Studies with NIH3T3 and Normal human dermal fibroblasts (NHDF) cells also showed that SNF contributed to the high functionality of these cells [30; 31]. Furthermore, their mechanical properties showed sufficient mechanical strength and permeability for implantation [32; 33]. The SNFs' mechanical strength and high elasticity makes them suitable for long-term cell cultivation as they do not shrink even when cells migrate and grow. Therefore, SNF are a useful biomimetic material with high cell functionality that can morphologically mimic the ECM and function as a cell scaffold for skin models [30; 31]. With this SNF, the problem of collagen matrices which has shrinkage after long-term cultivation [11] can be improved. Also, collagen matrices lack the ability to form elastin fibers, would be able to solve by applying SNF, which contains high porous microstructures [26], that has the potential to enhance fibroblast function within SNFs. Using SNF to construct a three-dimensional artificial skin model can improve the problems of known models such as the collagen matrix. In this study, we evaluated the usefulness of SNF by creating a three-dimensional artificial skin model. We created skin models using immortalized and primary cultured cell lines and conducted morphological observations, gene expression analysis, and skin corrosion tests.