成体人牙髓干细胞的分离与鉴定

Identification and isolation of human dental pulp stem cells

目的从成体人牙髓组织中分离培养牙髓干细胞,初步探讨其分化潜能。方法选取年轻患者因正畸或阻生拔除的健康第三磨牙,取出牙髓,采用酶消化及过滤法得到单细胞悬液,有限稀释法原代培养。扩大培养细胞克隆,检测STRO1的表达。体外诱导分化后对各克隆从碱性磷酸酶(ALP)活性、矿化结节形成、牙本质涎蛋白(DSP)表达、OilRedO染色、PPARr2基因表达等方面进行检测。结果克隆来源细胞STRO1表达阳性。在矿化液诱导下,克隆细胞呈现明显高的ALP活性;能够形成矿化结节;可以分泌表达DSP,已向成牙本质细胞方向发生了分化。成脂肪诱导后,OilRedO染色阳性,可检测到PPARr2基因表达。结论从成体人牙髓组织中可以分离培养出干细胞,在体外能有效增殖并保持低分化状态。

Objective To isolate and cultivate human dental pulp stem cells(DPSCs). Methods Pulp tissuewas removed from healthy young human teeth extracted for orthodontic purposes. The pulp was digested by Type Ⅰcollagenase and dispase.Then single-cell suspensions were obtained by filter and cultivated. The clones were identified by expression of STRO-1. Under the conditions of inducement, clones were identified by activity of alkaline phosphatase(ALP), formation of mineralized nodule and expression of dentin sialoprotein(DSP), and by Oil Red-O dyeing and expressing of PPARr2. Results The clones had positive expression of STRO-1. When stimulated to differentiation, these cells took on dramatically high activity of ALP, had the ability of mineralization and expressed DSP. These cells also had ability to trans-differentiate into adipocytes. Conclusion There are stem cells in human dental pulp tissues ,which can be isolated and cultivated.