Micro-architectural analysis of decellularised unscarred and scarred dermis provides insight into the organization and ultrastructure of the human skin with implications for future dermal substitute scaffold design

The three-dimensional (3-D) spatial arrangement of dermal tissue plays a crucial role in directing cellular behaviour during wound healing. Thus, it is crucial to elucidate a better understanding of the three-dimensional dermal architecture of the human skin. The aim of this project was to understand the configuration in morphological structure of decellularised human dermis between unscarred skin and normal cutaneous scars. Skin samples were obtained from ethically consented volunteer patients undergoing Abdominoplasty surgery. All skin samples underwent decellularisation (DNA removal = 88%.). Histological sections of cellular and decellularised dermis were subsequently analysed using standard haematoxylin and eosin (H&E), and 4’,6-diamidino-2- phenylindole (DAPI) stains. In addition, extent of decellularisation was quantified using an Easy-DNATM isolation kit. Biomechanical and structural evaluations were performed using atomic force microscopy (AFM) and Multiphoton Microscopy (MPM). Interestingly, there was no change in the gross morphology of decellularised unscarred and scarred dermis, under light microscopy. However, MPM and AFM showed that collagen fibers in unscarred decellularised dermis were arranged randomly. Collagen fibers of decellularised unscarred dermis appeared to have a significantly rougher (Rq- 16.5, Ra-12.5, Rmax-198; p<0.0001) surface topography. Based on AFM elastic modulus values, collagen fibers of unscarred decellularised dermis were less stiff (mean 2.155 MPa ± 0.9595; p<0.0001) compared to decellularised scarred dermis. MPM demonstrated that collagen fibers in unscarred dermis are interwoven, akin to a mesh-like structure. Furthermore, scarred dermis had a higher collagen volume density (papillary dermis, p<0.0082; reticular dermis, p<0.0332). Decellularisation of unscarred and scarred dermis was successfully achieved which enabled evaluation of unique micro-architectural differences between intact and injured skin. The parameters addressed in this study may help in developing engineered scaffolds for dermal wound repair. Ideally, the scaffolds should exhibit a mesh-like structure with a rough surface and low stiffness, which represents the microenvironment of unscarred dermal tissue.