Suzuki Daisuke, Pinto Filipa, Senoo Makoto
Department of Physiology and Biochemistry, Faculty of Nutrition, Kobe Gakuin University, Kobe, Japan.
Sapphiros Ai Bio., Boston, MA, USA.
Methods Mol Biol. 2025;2922:3-20. doi: 10.1007/978-1-0716-4510-9_1.
The co-culture of human epidermal keratinocytes with mouse 3T3-J2 feeder cells, a method developed by Green and colleagues, has been widely used since the early 1980s to generate skin autografts. Additionally, co-culture with 3T3-J2 cells has been a crucial tool in skin stem cell biology, as it facilitates the evaluation of self-renewal capacity and differentiation of epidermal stem cells. This chapter presents a recent enhancement of the Green method, aimed at further promoting the expansion of human epidermal keratinocytes by utilizing a small molecule inhibitor of TGF-β signaling. This new protocol also enables the rapid expansion of human epidermal keratinocytes in co-culture with human feeder cells, including human dermal fibroblasts and human preadipocytes-two key alternatives to 3T3-J2 cells-with the long-term goal of developing customized Xeno-free skin autografts.
自20世纪80年代初以来，格林及其同事开发的一种将人表皮角质形成细胞与小鼠3T3-J2饲养细胞共培养的方法，已被广泛用于生成皮肤自体移植物。此外，与3T3-J2细胞共培养一直是皮肤干细胞生物学中的关键工具，因为它有助于评估表皮干细胞的自我更新能力和分化情况。本章介绍了对格林方法的最新改进，旨在通过使用一种TGF-β信号小分子抑制剂来进一步促进人表皮角质形成细胞的扩增。这个新方案还能够在与人饲养细胞（包括人真皮成纤维细胞和人前脂肪细胞——3T3-J2细胞的两种关键替代细胞）共培养时快速扩增人表皮角质形成细胞，其长期目标是开发定制的无动物源皮肤自体移植物。
