Cattier Bettina, Guignard Rina, Martel Israël, Martel Christian, Simard-Bisson Carolyne, Larouche Danielle, Guiraud Béatrice, Bessou-Touya Sandrine, Germain Lucie
Centre de Recherche en Organogénèse Expérimentale de l'Université Laval/LOEX, Quebec City, QC G1J 5B3, Canada.
Department of Surgery, Faculty of Medicine, Université Laval, Quebec City, QC G1V 0A6, Canada.
Int J Mol Sci. 2025 Feb 21;26(5):1852. doi: 10.3390/ijms26051852.
Hair follicle stem cells, located in the bulge region of the outer root sheath, are multipotent epithelial stem cells capable of differentiating into epidermal, sebaceous gland, and hair shaft cells. Efficient culturing of these cells is crucial for advancements in dermatology, regenerative medicine, and skin model development. This investigation aimed to develop a protocol for isolating enriched bulge-derived epithelial cells from scalp specimens to produce tissue-engineered substitutes. The epithelium, including hair follicles, was separated from the dermis using thermolysin, followed by microdissection of the bulge region. Epithelial stem cells were isolated using enzymatic dissociation to create a single-cell suspension and compared with the direct explant culture and a benchmark method which isolates cells from the epidermis and pilosebaceous units. After 8 days of culture, the enzymatic digestion of microdissected bulges yielded 5.3 times more epithelial cells compared to explant cultures and proliferated faster than the benchmark method. Cells cultured from all methods exhibited comparable morphology and growth rates. The fully stratified epidermis of tissue-engineered skin was similar, indicating comparable differentiation potential. This enzymatic digestion method improved early-stage cell recovery and expansion while maintaining keratinocyte functionality, offering an efficient hair bulge cell-extraction technique for tissue engineering and regenerative medicine applications.
毛囊干细胞位于外根鞘的隆突区，是一种多能上皮干细胞，能够分化为表皮细胞、皮脂腺细胞和毛干细胞。高效培养这些细胞对于皮肤病学、再生医学和皮肤模型开发的进展至关重要。本研究旨在开发一种从头皮标本中分离富集隆突来源上皮细胞的方案，以生产组织工程替代物。使用嗜热菌蛋白酶将包括毛囊在内的上皮组织与真皮分离，随后对隆突区进行显微解剖。通过酶解分离上皮干细胞以创建单细胞悬液，并与直接外植体培养以及从表皮和毛囊皮脂腺单位分离细胞的基准方法进行比较。培养8天后，显微解剖隆突的酶解消化产生的上皮细胞比外植体培养多5.3倍，且增殖速度比基准方法更快。所有方法培养的细胞表现出相似的形态和生长速率。组织工程皮肤的完全分层表皮相似，表明具有相当的分化潜能。这种酶解消化方法提高了早期细胞回收率和扩增能力，同时保持了角质形成细胞的功能，为组织工程和再生医学应用提供了一种高效的毛囊隆突细胞提取技术。
