Strategies and challenges in developing pre-vascularized three-dimensional skin substitutes using layer-by-layer cell coating technology.

Irreversible cutaneous injuries must be treated by skin transplantation. Autologous skin grafts are the gold standard; however, donor sites are often limited in patients with extensive injuries. Conventional artificial skin substitutes are also effective in several clinical cases. However, a major limitation is the lack of vascular structures, which results in delayed perfusion, increased risk of infection, and engraftment failure, especially in intractable wounds. To address this challenge, various advanced tissue engineering technologies have been explored to develop novel three-dimensional (3D) skin substitutes with vascular networks. In this review, we first discuss recent progress in the development of tissue-engineered 3D skin models. In particular, we highlight a 3D culture technique based on the layer-by-layer (LbL) method, which forms ultrathin films of extracellular matrix proteins on cell surfaces and promotes the self-assembly of cells. We also describe the developmental history of the LbL method for fabricating 3D biological tissue models, including skin constructs. Furthermore, we introduce our original application of photobiomodulation (PBM), an optical activation method, to LbL-3D skin during cultivation. Finally, we present the results of transplanting LbL-3D skin onto a mouse wound model and discuss the future prospects for this skin substitute.