骨髓间充质干细胞向神经元样细胞分化中褪黑素受体和多效蛋白的表达

目的体外诱导成人骨髓间充质干细胞（MSCs）向神经元样细胞分化，并探讨分化过程中多效蛋白（PTN）和褪黑素（Mel）受体亚型MT1、MT2 mRNA的表达，以探索MSCs向神经元样细胞分化的特性和机制。方法密度梯度离心加贴壁培养法分离成人MSCs，原代和传代培养。取第6代MSCs设对照和实验组进行诱导，诱导后30min至3d，观察细胞形态并计数。免疫细胞化学法和RT—PCR法测定分化后细胞神经细胞特异性表面标志神经元特异性烯醇化酶（NSE）、微管相关蛋白-2（MAP-2）、神经胶质纤维酸性蛋白（GFAP）和诱导前、诱导后12hPTN和MT1、MT2 mRNA的表达。结果接种24h后MSCs开始贴壁，呈圆形或椭圆形。3d后可见梭状细胞呈集落状生长，10d～14d融合。第5代～第6代时呈现较均一的成纤维细胞样形态。诱导后胞体向胞核收缩；出现双极及多极细胞。12h变形细胞增多，细长突起相互连接。24h后变形细胞增多不明显。诱导后12h大部分细胞表达NSE、MAP-2，未检测到GFAP的表达。实验组诱导后12h细胞有PTN和MT1 mRNA的表达。结论PTN和Me1信号传导可能参与调控了MSCs向神经元样细胞的分化。

he Expression of Melatonin Receptors and Pleiotrophin mRNA in Adult HUman Bone Marrow Mesenchymal Stem Cells Differentiation into Neural Like Cells ｜n vitro

Objective To study the expression of melatonin receptors （MT1,MT2） and pleiotrophin （PTN） mRNA on adult human bone marrow mesenchymal stem cells （MSCs） differentiated into neural like cells in vitro,and to investigate the mechanism of MSCs plasticity. Methods Human MSCs were isolated primarily from bone marrow of adult volunteers＇ lilac and purified sterilely with gradient centrifugation. The 6th passage of MSCs were planted into plastic culture flask and induced. The control group did not receive any induced treatment. After the induction for 12 hours,NSE,MAP- 2 and GFAP, which were neuron marker and astroeytic marker,were detected by immunocytochemistry method and real - time polymerase chain reaction（RT - PCR） assay. The express of MT1,MT2 and PTN mRNA was detected by RT - PCR assay before induction and at 12 h after induction. Results The MSCs were cultivated primarily and following passage successfully. The living behavior of MSCs from passage four to passage six were stable. After the induction for 12 hours,there was expression of NSE and MAP - 2,and there was not expression of GFAP. PTN and MT1 mRNA were detected to express after induction for 12 hours. Conclusion Pleiotrophin and melatonin might take part in MSCs differentiated into neural like cells.