地塞米松诱导牙周膜干细胞的定向成骨分化及凋亡

Dexamethasone induces osteogenic differentiation and apoptosis of periodontal ligament stem cells

背景:牙周膜干细胞是一类起源于牙组织的成体干细胞,具有良好的成骨分化能力,有望在骨组织工程中得到应用。目的:观察成骨诱导液对牙周膜干细胞成骨分化能力及细胞早期凋亡的影响。方法:从原代牙周膜组织中分离得到牙周膜干细胞,以1×104/cm2浓度铺板后开始诱导。利用1,10,100 nmol/L地塞米松、β-磷酸甘油钠、维生素C为成骨诱导剂,以碱性磷酸酶活性检测、茜素红矿化结节染色、荧光定量PCR等方法对细胞成骨情况进行鉴定,采用AnnexinV/PI双染法检测细胞凋亡情况。结果与结论:地塞米松可有效诱导牙周膜干细胞成骨分化,可显著提高碱性磷酸酶活性,促进茜素红矿化结节形成,提高成骨相关基因骨粘连蛋白及Ⅰ型胶原表达。根据碱性磷酸酶活性和矿化结节实验结果,地塞米松的成骨诱导具有浓度梯度效应,其中100 nmol/L地塞米松具有最佳成骨诱导能力。细胞凋亡结果提示,地塞米松诱导的成骨分化具有一定促凋亡作用,可诱导牙周膜细胞的早期凋亡。

BACKGROUND：Periodontal ligment stem cells are adult stem cells derived from dental tissues, which possess the capability of osteogenic differentiation. They are promising“seed cells”in bone tissue engineering. OBJECTIVE：To investigate the effects of osteogenic induction medium on the osteogenic differentiation and early apoptosis of periodontal ligment stem cells. METHODS：Periodontal ligment stem cells were isolated from primary periodontal ligament tissues and seeded on the plate at a density of 1×104/cm2. Osteogenic induction medium containing 1, 10 and 100 nmol/L dexamethasone,β-glycerophosphate and ascorbic acid were used. Alkaline phosphatase activity, mineral deposition staining and real time PCR were performed to assess the osteogenic capacity. AnnexinV/PI staining was used to detected apoptosis. RESULTS AND CONCLUSION：Dexamethasone effectively promoted osteogenic differentiation of periodontal ligment stem cells, by up-regulating alkaline phosphatase activity, enhancing mineral deposition and increasing osteogenic related gene （osteonectin and type Ⅰ col agen） expression. Alkaline phosphatase activity and mineralized nodule detection showed that osteogenic effects of dexamethasone were gradient-dependent, and 100 nmol/L was the best concentration. Apoptosis detection results indicated that osteogenic differentiation of dexamethasone for periodontal ligment stem cells could also induce cellapoptosis to some extent.