大鼠骨髓间充质干细胞在创面愈合中的作用

Contribution of rat bone marrow-derived cells to wound healing

背景:骨髓可能是体内所谓间质组织的干细胞库,一旦组织受损,信号可通过未知方式到达骨髓,促进体内的间充质干细胞向创伤部位迁移。目的:探讨骨髓间充质干细胞在创面愈合中的作用。设计、时间及地点:细胞学体内观察,于2006-07/2007-07在东南大学完成。材料:健康6～8周龄SD大鼠34只,由东南大学医学院实验动物中心提供。方法:取4只大鼠,采用全骨髓法体外分离培养骨髓间充质干细胞,取传至第5代细胞,移植前48h行BrdU标记。剩余30只大鼠麻醉后制作全层皮肤缺损模型,以对侧对称部位皮肤作为自身正常对照。造模后将标记的2×107个骨髓间充质干细胞经尾静脉注入体内。主要观察指标:于创伤后不同时间点采用大体观察、免疫荧光化学染色等手段,动态观察骨髓间充质干细胞参与创面修复情况。结果:大鼠造模后伤口有少量炎性渗出,细胞移植后7d创面均形成较硬的痂皮,14d创缘有新生表皮生成,21～28d大部分创面愈合,未出现炎症、化脓现象。细胞移植后第7,14,21,28天组织切片均可见BrdU染色阳性细胞,胞核呈绿色荧光,在创面边缘和创伤局部聚集。细胞移植后第14天组织切片可见BrdU染色阳性的细胞(胞核为绿色荧光)胞浆中有FⅧ表达,呈红色荧光,提示可能有部分骨髓间充质干细胞分化为血管内皮细胞参与创面愈合。对侧正常皮肤未见明显的骨髓间充质干细胞聚集和增殖分化。结论:创伤可能是一种刺激因素,招募自身或外源的骨髓间充质干细胞参与创面愈合。

BACKGROUND： Bone marrow may be the stem cell bank of so-called mesenchymal tissues. Once the tissues damaged, the signals can reach the bone marrow by unknown method, and promote the transportation of the mesenchymal stem cells （MSCs） in vivo to wound. OBJECTIVE： To investigate the effect of rat bone marrow-derived MSCs on wound healing. DESIGN, TIME AND SETTING： A cytological observation in vivo was performed in Southeast University from July 2006 to July 2007. MATERIALS： A total of 34 healthy SD rats, aged 4-6weeks, were provided by Experimental Animal Center of Southeast University. METHODS： The bone marrow-derived MSCs of four SD rats were isolated and cultured in vitro by the whole bone marrow culture method. The fifth passage cells were labeled with 5-BrdU at 48 hours before transplantation. Under anesthesia, full thickness dermal wound models were established on the back of the other thirty rats, and the opposite backs were as self-controls, The labeled MSCs of SD rats were transplanted into tail veins. MAIN OUTCOME MEASURES： The wound healing conditions at different time points after the transplantation were dynamically observed by general observation and immuno-tluorescence histochemical staining. RESULTS： A little inflammatory exudation was observed at wound after model establishment. Hard crusts were formed at wound surfaces at 7 days after transplantation, new epidermis were formed at 14 days, and most wounds healed at 21-28 days without inflammation and suppuration. At 7, 14, 21, and 28 days after transplantation, BrdU positive celts were observed in sections, green fluorescence was observed on nucleus. The positive cells aggregated at edge and local site of wound. At 14 days after transplantation, F Ⅷ showed red fluorescence on cytoplasm of BrdU positive cells on tissue sections, which suggested some of bone marrow-derived MSCs maybe differentiate into vascular endothelial cells and participant the wound healing. There was not obvious aggregation, proliferation and differentiation of bone marrow-derived MSCs in the normal skin. CONCLUSION： Wound may be a kind of stimulating factor, which can made the self and exogenous bone marrow-derived MSCs participant wound healing.