人脐带间充质干细胞诱导为牙周韧带样细胞的实验研究

Experimental study of human umbilical cord mesenchymal stem cells differentiating into periodontal ligament-like cells

目的建立人牙周韧带细胞（human periodontal ligament cell，hPDLC）与人脐带间充质干细胞（human umbilical cord mesenchymal stem cell，hUCMSC）体外非接触式共培养模型，研究hUCMSC定向分化为hPDLC的可能性，探索新的可用于牙周组织工程的种子细胞。方法利用跨室培养装置（Transwell）培养板建立hPDLC与hUCMSC体外非接触式共培养模型，免疫组织化学方法检测其骨桥蛋白（osteopontin，OPN）、骨钙素（ostocalcin，OCN）及骨涎蛋白（bone sialoprotein，BSP）的表达情况，并采用蛋白质印迹法从蛋白水平定量分析诱导后hUCMSC在分子水平的改变。结果hUCMSC在非接触式共培养体系中可以被hPDLC诱导为多角形或梭形，蛋白质印迹法检测结果显示，共培养3、7、14及21d后的hUCMSC在蛋白水平OCN和OPN表达上调[OCN共培养前（0.88±0.21），共培养21d（1.42±0.17）；OPN共培养前（0.93±0.13），共培养21d（1.43±0.22）]；BSP表达逐渐下调[共培养前（1.60±0.09），培养21d（0.75±0.20）]，与共培养前相比差异均有统计学意义（P〈0.05）。结论hUCMSC在一定条件下可向hPDLC定向分化，并有望成为牙周组织工程的种子细胞。

Objective To investigate the possibility of human umbilical cord mesenchymal stem cells （hUCMSC） differentiating into human periodontal ligament cells （hPDLC） by establishing a coeulture model in vitro. Methods Indirect coculture model of hPDLC with hUCMSC was established on Transwell system. The protein expression of bone sialoprotein （ BSP）, ostocalcin （OCN） and osteopontin （OPN） was examined by immunohistochemical staining. The changes of hUCMSC on molecular level were analyzed by Western blotting. Results hUCMSC was in the form of polygon and rhombus after induction by hPDLC. Immunohistochemistry staining and Western blotting showed that OCN and OPN were up-regulated [OCN ： （0.88±0.21） vs （1.42±0.17 ） , OPN ： （0.93±0.13） vs （1.43±0.22 ） , P 〈 0.05] and BSP was down- regulated [（1.60±0.09） vs （ 0.75±0.20）, P 〈 0.05] after coculture. Conclusions hUCMSC can differentiate into hPDLC under certain conditions in vitro and hUCMSC may hopefully become the stem cells in periodontal tissue engineering.