Abstract
Hair follicle (HF) reconstruction in vitro is a promising field in alopecia treatment and human HF development research. Here, we combined postnatal human dermal papilla (DP) cells and skin epidermal keratinocytes (KCs) in a hanging drop culture to develop an artificial HF germ. The method is based on DP cell hair-inducing properties and KC self-organization. We evaluated two protocols of aggregate assembling. Mixed HF germ-like structures demonstrated the initiation of epithelial-mesenchymal interaction, including WNT pathway activation and expression of follicular markers. We analyzed the influence of possible DP cell niche components including soluble factors and extracellular matrix (ECM) molecules in the process of the organoid assembling and growth. Our results demonstrated that soluble factors had little impact on HF germ generation and Ki67+ cell score inside the organoids although BMP6 and VD3 maintained effectively the DP identity in the monolayer culture. Aggrecan, biglycan, fibronectin, and hyaluronic acid (HA) significantly stimulated cell proliferation in DP cell monolayer culture without any effect on DP cell identity. Most of ECM compounds prevented the formation of cell aggregates while HA promoted the formation of larger organoids. In conclusion, our model could be suitable to study cell-cell and cell-niche interactions during HF reconstruction in vitro.
Highlights
Hair loss or alopecia is a widespread pathology; men and women at any age could be affected by any of the different types of alopecia
We analyzed the expression of early hair follicle (HF) marker Lef1 and found that proliferative KCs at the site of epithelial-mesenchymal contact were positive for this marker (Figure 2(a))
The results provide evidence that the process of HF morphogenesis in mixed aggregates is induced by dermal papilla (DP) cells, and Lef1 and P cadherin are the specific markers of an onset of this process
Summary
Hair loss or alopecia is a widespread pathology; men and women at any age could be affected by any of the different types of alopecia. It was shown that HF germ keratinocytes express these molecules for the regulation of HF cycle [14] These factors support and stimulate hair-inducing abilities of DP cells in vitro [10,11,12,13]. Stem Cells International reproducing the initial steps of HF formation This process depends on ECM components, especially DP-specific protein versican [15]. DP niche could be reconstructed using KCs to provide all the necessary soluble factors and 3D culturing to stimulate DP aggregation processes. Soluble factors (BMP6, VD3, and VPA) maintained DP cell identity, while several ECM components (aggrecan, biglycan, fibronectin, and hyaluronic acid (HA)) significantly stimulated cell proliferation in 2D cultures. Our results may provide the new in vitro method of HF development, and the model could be suitable to study cell-cell and cell-niche interactions during HF reconstruction in vitro
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