摘要
目的探讨地黄多糖诱导大鼠骨髓间充质干细胞(BMSCs)分化为神经元样细胞的作用及对Notch1和Jagged1蛋白表达的影响。方法取大鼠P3-P5代BMSCs,随机分为正常对照组(CON组)、神经生长因子组(BDNF组,10μg/ml)和地黄多糖诱导A、B、C组(RGP组,浓度分别为50、100、200μg/ml),诱导后连续培养7d。观察细胞一般形态。采用Western blotting检测CON组与RGP C组神经标志物相关蛋白神经巢蛋白(Nestin)、βⅢ-微管蛋白(βⅢ-tubulin)、神经元特异性烯醇化酶(NSE)、胶质纤维酸性蛋白(GFAP)的表达,免疫荧光法和Western blotting分别检测各组诱导后0、1、3、7d时Notch1、Jagged1蛋白和Notch1胞内段NICD的表达。结果 CON组始终以梭形细胞为主,RGP C组第5天起出现神经元样细胞。Western blotting检测到RGP C组Nestin蛋白表达由强到弱,NSE和βⅢ-tubulin蛋白表达由弱到强,GFAP蛋白呈弱表达。免疫荧光结果显示,RGP各组各时间点Notch1蛋白阳性细胞率均显著低于CON组(P<0.05),且随时间延长Notch1蛋白表达阳性率逐渐降低;RGP C组中Jagged1阳性细胞率诱导后1d时具有明显的上升趋势,而后显著下降(P<0.05),BDNF组基本无Jagged1蛋白表达。Western blotting检测结果与免疫荧光检测结果类似。结论地黄多糖具有诱导BMSCs向神经元样细胞分化的作用,且可影响Notch1和Jagged1蛋白的表达。
Objective To explore the effect ofRehmannia glutinosa polysaccharide （RGP） on the differentiation of rat bone marrow mesenchymal stem cells （BMSCs） into neuron-like cells and the expression of Notch1 and Jagged 1 proteins. Methods P3-P5 BMSCs were randomly divided into control group, brain-derived neurotrophic factor （BDNF） group （10μg/ml）, and RGP group （50, 100, 200μg/ml subgroup, respectively）. Cells in each group were respectively induced and cultured for 7 days. The cell morphology was observed. The protein expressions of neuro-markers including nestin, β-tubulin, neuron-specific enolase （NSE） and gial fibrillary acid protein （GFAP） in both control group and 200μg/ml subgroup were determined by Western blotting. The expressions of Notch1, Jagged 1 and intracellular domain of Notch 1 （NICD） were determined by immunocytochemical assay and Western blotting 0, 1, 3, 7d after induction in all the groups. Results The spindle type cells were observed in control group all the time, while the neuron-like cells were found in 200μg/ml subgroup 5d after induction. Western blotting results showed that the expression of nestin decreased and the expressions of NSE and β 1H-tubulin increased gradually, and the GFAP was weakly expressed. Immunofluorescence assay showed that the percentage of Notch1-positive cells in RGP group was significantly lower than in control group at each time point （P〈0.05）, and decreased in time in each subgroup of RGP group. The percentage of Jagged 1-positive cells showed a significant ascending trend from d 0 to d 1 after induction, while a significant descending trend from ld to 7d （P〈0.05） in 200 μg/ml subgroup. Jagged i protein expression was hardly found in BDNF group. The result of Western blotting was similar to those of the immunofluorescence method. Conclusion Rehmannia glutinosa polysaccharide may induce differentiation of BMSCs into neuron-like cells, and may effect the expression of Notch i and Jagged i proteins.
出处
《解放军医学杂志》
CAS
CSCD
北大核心
2014年第6期448-453,共6页
Medical Journal of Chinese People's Liberation Army
基金
辽宁省科技厅自然科学基金(2013022059)
辽宁医学院青年科技启动基金(Y2011Z010)~~
