摘要
背景:研究报道用烧伤大鼠血清作为诱导剂可以诱导间充质干细胞分化为表皮细胞。目的:体外诱导骨髓间充质干细胞分化为表皮细胞,单独或与必要的诱导剂组合移植修复皮肤创面缺损与重建表皮。方法:无菌环境取大鼠骨髓,选取贴壁细胞用L-DMEM培养液培养至第4代,通过流式细胞术鉴定骨髓间充质干细胞。用含20%烧伤大鼠血清DMEM-F12培养液诱导骨髓间充质干细胞分化成表皮细胞,通过免疫组织化学鉴定。将Wistar大鼠制造全层皮肤缺损创面,分为3组,将BrdU标记的自体骨髓间充质干细胞及含有经诱导的骨髓间充质干细胞分别单次涂布到烧伤大鼠模型创面上,以未移植骨髓间充质干细胞做对照,观察再生皮肤创面收缩率及表皮层细胞与皮肤附件再生情况。结果与结论:分离、培养的骨髓间充质干细胞24 h后少部分细胞贴壁生长,形态长梭形,类似成纤维细胞,16 d时细胞贴满瓶底,呈鱼群样或网状排列,经流式细胞仪鉴定后加入20%烧伤大鼠血清的DMEM F-12培养基继续培养,细胞形态渐变为圆形或椭圆形细胞,经免疫组织细胞化学法和流式细胞术分析细胞角蛋白表达阳性,证实已分化为表皮细胞。动物实验结果表明无论是骨髓间充质干细胞单独移植还是与必要的诱导剂组合移植其皮肤的修复与再生情况均超过大鼠皮肤自然愈合,且不仅皮肤再生快,而且皮肤附件如毛囊等的再生也比对照组明显改善。初步推断间充质干细胞参与了表皮和皮肤附件毛囊的重建,从而改善皮肤愈合方式。
BACKGROUND：Burned rat serum has been reported as an inducer that can induce differentiation of mesenchymal stem cells into epidermal cells. OBJECTIVE：To induce in vitro differentiation of bone marrow mesenchymal stem cells into epidermal cells that were transplanted alone or combined with inducers for the repair of skin wound defect and epidermal reconstruction. METHODS：Under aseptic environment, rat bone marrow was harvested to culture adherent cells using low-glucose Dulbecco’s modified Eagle’s medium. Culture cells at passage 4 were confirmed as mesenchymal stem cells by flow cytometry. Bone marrow mesenchymal stem cells were induced by Dulbecco’s modified Eagle’s medium/Ham’s nutrient mixture F-12 containing 20%burned rat serum to differentiate into epidermal cells that were identified by immunohistochemistry. Wistar rat models of ful-thickness skin wounds were prepared and divided into three groups. The 5-bromo-2-deoxyuridine-labeled autologous bone marrow mesenchymal stem cells and induced bone marrow mesenchymal stem cells were coated singly onto a rat model of burn wounds, and rat models of burn wounds with no treatment served as controls. Wound contraction and regeneration of epidermal cells and skin appendages were observed. RESULTS AND CONCLUSION：After isolation and culture of cells for 24 hours, a few of adherent cells grew as fibroblast-like cells with fusiform shape. At 16 days, cells completely covered the bottom of bottle, exhibiting a fish＆amp;nbsp;or reticular arrangement. After detection by flow cytometry, cells were cultured in Dulbecco’s modified Eagle’s medium/Ham’s nutrient mixture F-12 containing 20%burned rat serum, and fusiform-shaped cells gradual y became round or oval cells. Flow cytometry analysis and immunocytochemistry results showed that cells expressed keratin, which were confirmed as epidermal cells. The results show that both the bone marrow mesenchymal stem cells transplantation alone or with necessary inductor is better for skin repair than natural h
出处
《中国组织工程研究》
CAS
CSCD
2014年第6期853-859,共7页
Chinese Journal of Tissue Engineering Research
