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Description
INTRODUCTION
Skin possesses a complex structure with diverse components. The epidermis consists of tightly packed epithelial cells, while dermis contain fibroblasts, blood vessels, sensory neurons, immune cells, and hair follicles. Both in vitro and in vivo models serve as useful tools for studying skin biology and uncovering the cellular and molecular processes involved in degenerative skin conditions. [1] However, these models have significant limitations because they often lack key biological and structural components of native skin. The most notable drawback of current lab-grown skin models is the absence of critical skin appendages, such as hair follicles (HFs) which limit our understanding of proper micro-environmental factors that contribute to tissue organization. In this study, we aimed to integrate human primary hair follicle dermal papilla cell (HFDPC) spheroids (hair follicle germs) within a fully functionalized vascularized bi-layered construct to biofabricate skin substitute with natural mechanical strength and flexibility and an ability to develop hair follicles.
MATERIALS AND METHODS
A biofabrication platform was developed to incorporate human primary hair follicle dermal papilla cell (HFDPC) spheroids within three-dimensional melt-electro written (3D-MEW) scaffold embedding human umbilical vein endothelial cells (HUVECs) and human dermal fibroblasts (HDFs) within a gelatin-based hydrogel to construct a full skin substitute with hair follicles. Briefly, a functionalized Polycaprolactone (PCL) skin graft with a 0-90° architecture was fabricated using an advanced 3D-MEW technique. Following this, the dermal layer was engineered by embedding HUVECs and HDFs within a gelatin-based hydrogel to create a vascularized dermal construct. HFDPC spheroids were generated and then precisely incorporated into the scaffold using a novel bioprinting method for further evaluation. Subsequently, keratinocyte cells were seeded to form the epidermis layer for a full-thickness skin construct with hair follicles.
RESULTS AND DISCUSSION
The developed platform enabled the construction of a 3D biomimetic structure of a full skin substitute with hair follicle germs. Confocal microscopic images revealed that the HFDPC spheroids were located within the center of the PCL filaments in the dermis, with high cell viability throughout the 3D hybrid structure (Figure 1-I). The distinct spindle-shaped morphology clearly showed that HDFs extended along the PCL filaments and were directed toward the surroundings of the spheroids. This suggested that the cell-spheroid interaction could be effective in the hybrid structure and that the PCL scaffold geometry can have an important role in hair follicle development. After 21 days of in vitro incubation, hair follicle induction within the hybrid structure was observed (Figure 1-II).
CONCLUSIONS
The results demonstrated that the developed 3D platform effectively supported the construction of highly viable hybrid structure recapitulating structure of a full-thickness skin substitutes with hair follicles.
REFERENCES
[1] Randall, Matthew J., et al. "Advances in the Biofabrication of 3D Skin in vitro: Healthy and Pathological Models." Frontiers in bioengineering and biotechnology 6 (2018): 154.
[2] Hosseini, Motaharesadat, Karl R. Koehler, and Abbas Shafiee. "Biofabrication of human skin with its appendages." Advanced healthcare materials 11.22 (2022): 2201626.
ACKNOWLEDGEMENTS
This study is supported by the Scientific and Techno-logical Research Council of Turkey (221M539).
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