论文部分内容阅读
目的探索一种简单而又高效的分离大鼠胎儿表皮干细胞方法,以及观察其在体内环境中的分化潜能。方法采集ED14、ED16、ED18及ED20不同发育阶段的SD大鼠胎儿皮肤,HE染色观察其结构。鼠尾胶原快速黏附法筛选胎鼠表皮细胞,在无血清培养基(K-SFM)中进行培养。免疫组织化学方法检测CK-19和1β-整合素表达情况。同时,也对细胞周期和细胞增殖能力进行检验。经荧光染料(PKH26)标记过的胎鼠表皮干细胞与多孔凝胶海绵结合,然后植入大鼠肾被膜下进行诱导分化。结果在ED18之前皮肤表皮和真皮尚未充分完成分化。使用快速黏附法能够有效的从ED18胎鼠皮肤分离到表皮干细胞。获得的细胞CK19和1β整合素呈阳性表达,细胞周期分析显示,细胞呈慢周期性,移植体在肾被膜下分化并展现典型的表皮样结构。结论表皮干细胞存在于早期发育皮肤中,但在ED18才能使用快速黏附法分离到表皮干细胞。此时的胎鼠表皮干细胞具有向表皮分化的能力。
Objective To explore a simple and efficient method for isolating rat fetal epidermal stem cells and to observe its differentiation potential in vivo. Methods Fetal skin of SD rats at different developmental stages of ED14, ED16, ED18 and ED20 were collected and their structures were observed by HE staining. Fetal rat epidermal cells were screened by rapid adhesion method and cultured in serum-free medium (K-SFM). Immunohistochemistry was used to detect the expression of CK-19 and 1β-integrin. At the same time, the cell cycle and cell proliferation ability is also tested. Fetal mouse epidermal stem cells labeled with fluorescent dye (PKH26) were combined with porous gel sponge and then implanted under rat renal capsule for differentiation induction. Results The epidermis and dermis of the skin had not yet been fully differentiated before ED18. Epidermal stem cells can be efficiently isolated from ED18 fetal rat skin using the rapid adhesion method. The obtained cells showed positive expression of CK19 and 1β integrin. Cell cycle analysis showed that the cells showed a slow cycle. The grafts differentiated under the renal capsule and showed the typical epidermal-like structure. Conclusion Epidermal stem cells exist in early developmental skin, but epidermal stem cells can be isolated by ED18. The fetal rat epidermal stem cells at this time have the ability to differentiate into the epidermis.