Department of Tissue Engineering and Regeneration, School and Hospital of Stomatology, Cheeloo College of Medicine, Shandong University & Shandong Key Laboratory of Oral Tissue Regeneration & Shandong Engineering Laboratory for Dental Materials and Oral Tissue Regeneration, No.44-1 Wenhua Road West, Jinan, Shandong, China.
Shandong Provincial Key Laboratory of Animal Cells and Developmental Biology, School of Life Science, Shandong University, Qingdao, China.
Stem Cell Res Ther. 2020 Sep 18;11(1):407. doi: 10.1186/s13287-020-01924-z.
In recent years, significant progress has been made in developing highly complex tissue-engineered skin substitutes (TESSs) for wound healing. However, the lack of skin appendages, such as hair follicles and sweat glands, and the time required, are two major limitations that hinder its broad application in the clinic. Therefore, it is necessary to develop a competent TESS in a short time to meet the needs for clinical applications.
Adult scalp dermal progenitor cells and epidermal stem cells together with type I collagen as a scaffold material were used to reconstitute bilayer TESSs in vitro. TESSs at 4 different culture times (5, 9, 14, and 21 days) were collected and then grafted onto full-thickness wounds created in the dorsal skin of athymic nude/nude mice. The skin specimens formed from grafted TESSs were collected 4 and 8 weeks later and then evaluated for their structure, cell organization, differentiation status, vascularization, and formation of appendages by histological analysis, immunohistochemistry, and immunofluorescent staining.
Early-stage bilayer TESSs after transplantation had a better efficiency of grafting. A normal structure of stratified epidermis containing multiple differentiated layers of keratinocytes was formed in all grafts from both early-stage and late-stage TESSs, but higher levels of the proliferation marker Ki-67 and the epidermal progenitor marker p63 were found in the epidermis formed from early-stage TESSs. Interestingly, the transplantation of early-stage TESSs produced a thicker dermis that contained more vimentin- and CD31-positive cells, and importantly, hair follicle formation was only observed in the skin grafted from early-stage TESSs. Finally, early-stage TESSs expressed high levels of p63 but had low expression levels of genes involved in the activation of the apoptotic pathway compared to the late-stage TESSs in vitro.
Early-stage bilayer TESSs reconstituted from skin progenitor cells contained more competent cells with less activation of the apoptotic pathway and produced a better skin structure, including hair follicles associated with sebaceous glands, after transplantation, which should potentially provide better wound healing when applied in the clinic in the future.
近年来，在开发用于伤口愈合的高度复杂组织工程皮肤替代物（TESS）方面取得了重大进展。然而，缺乏皮肤附属物，如毛囊和汗腺，以及所需的时间，是阻碍其在临床上广泛应用的两个主要限制因素。因此，有必要在短时间内开发出有能力的 TESS，以满足临床应用的需求。
使用成人生发头皮真皮祖细胞和表皮干细胞以及 I 型胶原作为支架材料，在体外重建双层 TESS。收集培养 4 个不同时间（5、9、14 和 21 天）的 TESS，然后将其移植到无胸腺裸鼠背部全层皮肤创面。4 和 8 周后收集来自移植 TESS 的皮肤标本，然后通过组织学分析、免疫组织化学和免疫荧光染色评估其结构、细胞组织、分化状态、血管生成和附属物形成。
移植后早期双层 TESS 的移植效率更好。所有早期和晚期 TESS 移植的标本均形成了具有多个角化细胞分化层的正常分层表皮结构，但早期 TESS 形成的表皮中增殖标志物 Ki-67 和表皮祖细胞标志物 p63 的水平更高。有趣的是，早期 TESS 的移植产生了更厚的真皮，其中含有更多的波形蛋白和 CD31 阳性细胞，重要的是，仅在来自早期 TESS 的皮肤移植中观察到毛囊形成。最后，与体外晚期 TESS 相比，早期 TESS 表达高水平的 p63，但参与凋亡途径激活的基因表达水平较低。
由皮肤祖细胞重建的早期双层 TESS 含有更少激活凋亡途径的更有能力的细胞，并且在移植后产生更好的皮肤结构，包括与皮脂腺相关的毛囊，这在未来应用于临床时应该能提供更好的伤口愈合效果。
