大鼠骨髓间充质干细胞体外诱导向神经元样细胞分化的实验研究

目的研究大鼠骨髓间充质干细胞（BMSCs）离体分离和培养方法，探讨碱性成纤维细胞生长因子（bFGF）、表皮生长因子（EGF）、全反式维甲酸（RA）、神经营养因子（BDNF）在体外诱导BM—SCs向神经元样细胞分化的作用。方法采用出生3周SD大鼠的全骨髓细胞进行培养，传至第3代时，分为三组：A组，bFGF＋EGF＋RA进行诱导分化；B组，BDNF＋RA进行诱导分化；C组，RA诱导分化。在倒置显微镜下每日观察、记录BMSCs的诱导分化情况，并应用免疫细胞化学技术对细胞进行兔抗神经元特异性烯醇化酶（NSE）单抗鉴定。结果A组诱导5d后有大部分细胞具备神经元样细胞形态，胞体呈锥形或圆形，有较长单极或多极的突起，有NSE阳性细胞表达。而B组可有部分NSE阳性细胞、C组细胞有少量NSE阳性细胞。结论bFGF＋EGF＋RA、BDNF＋RA和RA均可在诱导BMSCs向神经样元细胞分化，bFGF＋EGF＋RA组更优于和BDNF＋RA组及RA组。

An experimental study on the inducible differentiation of rat bone marrow stromal cells to neuron-like cells

Objective To investigate the methods for isolating and culturing rat bone marrow stroreal cells （BMSCs） ii vitro and to determine whether adult BMSCs could be induced to differentiate into neurons by basic fibroblast growth factor （bFGF）, epidermal growth factor （EGF）, all--transretinoicacid （RA） and Brain--derived neurotrophic factor （BDNF）. Methods Adult marrow stromal cells isolated from bone marrow of four week--old SD rats were cultured, bFGF＋ EGF＋ RA （group A）, BDNF＋ RA （group B） and RA （group C） were used to induce the third generation BMSCs. Phase contrast microscope was used to observe the differentiation of BMSCs daily. Immunohistochemistry technique was used to identify the diferentiated BMSCs with neuron specific enolase （NSE）. Results Seven days after treatment, some BMSCs in grupe A appeared typical morphologicalneuronal traits. Cell bodies appeared refractile, cone--like or spherical with long processes. NSE was expressed in group A. The cells in group B and group C showed no NSE expression. Conclusions The neuron--induction efficacy in the combination with bFGF. EGF and RA in rBMSCs in vitro is significantly higher than that of the combination with BDNF and RA or using RA alone.