Effect of 3D-Fibroblast Dermis Constructed by Layer-by-Layer Cell Coating Technique on Tight Junction Formation and Function in Full-Thickness Skin Equivalent

Human skin equivalents (HSEs) consisting of an epidermis and dermis have been used as promising tools for drug evaluation and for clinical applications in regenerative medicine. Normal human dermal fibroblasts (NHDFs) are essential for the fabrication of HSEs because they play an important role in the maturation of the epidermis. Recently, epidermal tight junctions (TJs), which are complex cell-cell junctions, have attracted much attention as a second barrier and regulator for other barrier functions. In a previous study, we revealed the expression of TJ-related proteins and the time course of formation of TJ structure in the HSE (layer-by-layer (LbL)-three-dimensional (3D) Skin) constructed by layer-by-layer (LbL) cell coating technique that have a unique dermis consisting of NHDFs only (3D-fibroblast dermis). However, the effect of the 3D-fibroblast dermis on the formation of functional epidermal TJs is unknown. In this study, we investigated the effect of the 3D-fibroblast dermis on the expression of TJ-related proteins and TJ function in LbL-3D Skin. We demonstrated that the 3D-fibroblast dermis affects the long-term expression of TJ-related proteins and the formation of TJ with barrier function in the epidermis. These results show that the 3D-fibroblast dermis in LbL-3D Skin contributes to the formation and maintenance of functional TJs as in native human skin by direct contact with KCs.