毛囊单位促进组织工程皮肤形成的体外实验研究

目的 通过插入分离的毛囊单位,构建带有皮肤附属器的组织工程皮肤,探讨毛囊对组织工程皮肤形成的作用.方法 将头皮分离成毛囊单位,插入由Ⅰ型鼠尾胶原、成纤维细胞和角质形成细胞构建的组织工程皮肤模型中,浸没培养1周,气-液界面培养3周.通过镜下观察、HE染色和免疫组化检测组织工程皮肤和毛囊的生长状态.结果 相对于悬浮培养的毛囊,实验组毛囊体外生长期延长,结构更完整.HE染色显示,实验组组织工程皮肤可见毛囊和皮脂腺存在.免疫组化染色显示,带有毛囊的组织工程皮肤中可见反映基底膜完整性的Laminin和Ⅳ型胶原呈线状连续分布于表皮、真皮连接处;而反应表皮成熟度的CK4和CK10/13则呈阳性分布于基底上层非角化细胞.结论 复合毛囊单位可以促进组织工程皮肤表皮组织的分化和成熟.本方法为组织工程皮肤的构建和毛囊的体外培养提供了新的思路和方法.

Hair Follicle Units Improving the Construction of Tissue Engineered Skin in Vitro

Objective To construct tissue-engineered skin with skin appendages by inserting isolated hair follicle units and to explore the effect of hair follicle units on the construction of tissue-engineered skin. Methods Single undamaged hair follicle units were dissected by surgical blades. Then hair follicle units were inserted to the engineered dermis including Rat tail tendon collagen type I, fibroblasts and Keratinocytes. After 1 week of submerged culture and 3 weeks of air-liquid culture, tissue-engineered skin and the hair follicle were detected by inverted microscope, HE staining and immunohistochemical staining. Results Hair follicle morphology could maintain more than 21 days without hair papilla damage, which was longer than those of free-floating culture. After 4 weeks, skin appendages such as hair follicle and sebaceous glands were found in the tissue engineered skin by HE staining. A continuous basement membrane was observed and associated with the expression of laminin and collagen IV by immunohistochemical staining. Keratinocytes seeded on the tissue with hair follicle units formed a stratified and cornified epidermis and expressed typical markers of differentiation （keratin4, keratinl0/13）. Conclusion Inserting hair follicle units could promote the differentiation and mature of tissue- engineered skin, which could provide a new way to construct tissue-engineered skin.