复方麝香注射液在人脐带间充质干细胞神经分化中的作用

Effects of compound musk injection on neurocyte-like differentiation of human umbilical cord-derived mesenchymal stem cells

目的 观察复方麝香注射液在体外对人脐带间充质干细胞（hUMSCs）神经分化的影响.方法 取足月妊娠剖宫产健康新牛儿脐带,获取间充质干细胞,采用流式细胞术进行细胞表型鉴定.以碱性成纤维细胞生长因子（bFGF）预诱导24 h,再以含5、10、15、20、25g/L复方麝香注射液的无血清培养基进行神经分化诱导,采用免疫细胞化学检测神经元特异性烯醇化酶（NSE）、微观相关蛋白-2（MAP-2）和胶质纤维酸性蛋白（GFAP）的表达.结果 流式细胞检测显示hUMSCs表达CD13、CD29、CD44、CD73、CD90、CD105、CD106、CD166和HLA-ABC抗原,不表达CD31、CD34、CD38、CD45、CD133和HLA-DR抗原.经不同浓度复方麝香注射液诱导5 d后,各诱导组中均出现神经样细胞.5、10、15、20、25 g/L组显微镜每随机视野中NSE阳性细胞数分别为19.00±3.61、66.60±5.37、60.20±10.06、44.80±9.44、47.80±10.45,MAP-2阳性细胞数分别为9.20±3.27、53.40±10.92、59.40±10.41、46.60±8.88、46.80±8.93.以10 g/L和15 g/L组中的NSE、MAP-2阳性细胞数为最多.各诱导组中仅极少数细胞呈GAFP弱阳性.结论 人脐带富含问充质干细胞（MSCs）,适量复方麝香注射液可有效诱导人脐带间充质干细胞分化为神经元细胞.

Objective To observe the effects of compound musk injection on neurocyte-like differentiation of human umbilical cord-derived mesenchymal stem cells （hUMSCs） in vitro. Methods The full-term umbilical cords from healthy neonates were washed with D-Hank＇ s,and MSCs were separated from them. After flow cytometry was performed,hUMSCs were pre-induced with 10 μg/L basic fibroblast growth factor （bFGF） ,then were induced to neural differentiation by serum-free culture medium containing different concentrations of compound musk injection including 5,10,15,20,25 g/L respectively. The expression of neuron specific enolase （NSE）, microtubule-associated protein 2 （MAP-2） and glial fibrillary acid protein （GFAP） was detected by immunocytochemistcy. Results Fluorescence activating cell sorter （FACS） showed that hUMSCs expressed CD13, CD29, CD44, CD73, CD90, CD105, CD106, CD166, HLA-ABC （HLA-I） ,and did not express CD31 ,CD34,CD38,CD45,CD133,HLA-DR （HLA-Ⅱ）. After induction for 5 days by DMEM/F12 medium containing different concentrations of compound musk injection, the hUMSCs of each group could differentiate into neuron-like cells. The number of NSE positive cells in 5,10,15,20,25 g/ L groups was 19.00 ±3.61,66.60 ±5.37,60.20 ＋ 10.06,44.80 ±9.44,47. 80 ± 10.45,and that of MAP-2 positive cells was 9.20 ± 3. 27,53.40 ± 10. 92,59.40 ± 10. 41,46. 60 ± 8. 88,46. 80 ± 8. 93, respectively. NSE and MAP-2 positive cells were most plentiful in 10 g/L and 15 g/L groups. But just few of GFAP weak positive cells were detected in all five groups. Conclusion Human umbilical cord is a rich source of MSCs. Adequate concentration of compound musk injection can effectively induce the differentiation of hUMSCs into neural-like cells.