中文摘要：

目的研究维甲酸（RA）和音猬因子（Shh）单独和联合作用对促进骨髓基质细胞（BMSCs）向运动神经元（MN）发育的作用。方法采用密度梯度离心法结合贴壁法原代培养BMSCs，应用神经干细胞培养基（专利号ZL02134314．4）培养细胞并分四组：（1）空白对照组；（2）RA组，加入0．5μg／mLRA；（3）Shh组，加入200ng／mL Shh因子；（4）RA＋Shh组，加入0．5μg／mL RA和200ng／mL Shh。细胞培养后第8、12、16、20天．采用SABC免疫细胞化学和间接免疫荧光法分别检测神经干细胞标记nestin和运动神经元标记Hb9的表达，选细胞密度适中视野计算阳性细胞数。对比各组细胞阳性表达比例。结果RA组nestin阳性表达率比其他三组都高（P〈0．05）；Shh组和空白组阳性率均很低。两者间无显著差别；RA＋Shh组和空白组比较有差别（P〈0．05）。四组中只有RA＋Shh组Hb9表达阳性．其他组均未检测到Hb9抗体。结论RA能显著促进BMSCs增殖并向骨髓基质源性神经干细胞（BMSC-D-NSCs）分化，然而单独使用Shh并没有明显效果。在RA＋Shh组，部分BMSC．D．NSCs在Sb_h诱导下表达了MN发育过程中的重要蛋白Hb9，提示BMSCs存在向MN发育的潜力．RA＋Shh可以促进该分化过程。

英文摘要：

Objective To study the singular and combined impact ofretinoic acid （RA） and sonic hedgehog （Shh） on the differentiation of the bone marrow stromal cells （BMSCs） into motor neurons. Methods By means of density gradient centrifugation and removing non-sticking-cells, BMSCs were primarily cultured in special neural stem cells medium （Patent number： ZL02134314）. Four groups were classified by level of RA and Shh： （1）control; （2）0.5 μg/mL RA in medium （RA）; （3）200 ng/mL Shh in medium （Shh）; （4）0.5 μg/mL RA and 200 ng/mL Shh in medium （RA＋Shh）. Nestin, the special mark of neural stem cells, and Hb9, the tag of motor neurons were detected on the 8th, 12th, 16th, 20th day after seeding via SABC immunocytochemistry and immunofluorescence. On the other hand, a reasonable sight was selected to count positive cells and compared the expression of each mark among four groups. Results The proportion of nestin positive in RA group was significantly higher than that in other groups （P〈0.05）. A low part ofnestin positive was showed in Shh group and control group, with no obvious difference between them;There＇s diffirence between RA＋Sb_h group and control group （P〈0.05）. In all four groups, only RA＋Shh group was Hb9 positive. Conclusion RA could greatly promote proliferation of BMSCs and differentiate them into BMSC-D-NSCs. However, solitary Shh could not make obvious effect on this process. In RA＋Shh group, BMSC-D-NSCs expressed Hb9, the key mark of neural development of MN, which indicates that the BMSCs possess the potential function to differentiate into MN, and RA＋Shh can promote this process.