Tissue engineering ECM-enriched controllable vascularized human microtissue for hair regenerative medicine using a biomimetic developmental approach

INTRODUCTION: Regenerative medicine is a promising approach for hair loss; however, its primary challenge is the inductivity of human dermal papilla cells (DPCs), which rapidly lose hair growth-inducing properties in 2D culture. Despite extensive research efforts to construct DPCs, current 3D microenvironments fabricated to restore hair inductivity remain insufficient. OBJECTIVES: Here, we aimed to fabricate ECM-enriched controllable vascularized dermal papilla (DP) spheroids that highly mimic in vivo DPCs microenvironments to restore their hair inductivity. METHODS: We employed layer-by-layer (LbL) self-assembly using gelatin and alginate to construct nanoscale biomimetic ECM for DPCs, with Ca(2+) as a cross-linking agent to create controllable DP spheroids. DPCs were also co-cultured with human umbilical vein endothelial cells to construct vascularized DP spheroids. Immunofluorescence staining and angiography was used to detect angiogenesis in vitro and in vivo. RNA sequencing and in vivo implantation were employed to investigate DPCs signature. RESULTS: LbL technology enabled DPCs to aggregate into controllable DP spheroids of size and cell numbers similar to those of primary DP. Vascularization prevented hypoxia-induced necrosis and functioned in association with host vessels post-transplantation. Compared with traditional 3D culture, nanoscale ECM and vascularization were found to restore the transcriptional signature of DPCs and triple hair induction efficiency following engraftment. CONCLUSION: Our novel biomimetic developmental tissue engineering strategy is a crucial step toward the recovery of human DPC hair inductivity, which would enable the rapid clinical application of large-scale hair regeneration platforms.