Optimization of hair follicle spheroids for hair-on-a-chip

Currently, most models for hair follicle research have the limitation of not replicating some key features of the hair follicle microenvironment. To complement this, we transfected various factors for hair growth into dermal papilla cells (DPCs) by electroporation and cultured the spheroids with keratinocytes (KCs). We optimized the cell number and culture period for applying spheroids to hair-on-a-chip. Furthermore, we investigated the expression of hair growth factors in spheroids depending on the presence or absence of human umbilical vein endothelial cells (HUVECs) and transfection. In spheroids in which DPCs, KCs, and HUVECs were co-cultured for 21 days, the expression of lymphoid enhancer factor 1 (LEF1), T-cell factor 1 (TCF1), and keratin 25 (K25) in the center of the spheroid, the expression of keratin 17 (K17) on the outer surface of the spheroid, and the shape of hair extending outward from the spheroid surface were observed. From these results, it is expected that a hair-on-a-chip experiment in which short-term cultured TKH spheroids are injected into the dermis and co-cultured with KC will enable the production of full-thickness skin equivalents containing hair in vitro without transplantation into animals. We report spheroids prepared by injecting LEF1 and Wnt1 into DPCs via transfection and then adding KCs and HUVECs. Through SEM, we observed a part extending outward from the TK and TKH surfaces, as indicated by white arrows.