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目的:通过分析50Hz电磁场干预骨髓源神经祖细胞向神经细胞分化前后相关神经细胞标志物m RNA表达水平,探讨低频电磁场对骨髓源神经祖细胞诱导分化的作用。方法:利用全骨髓培养法获取骨髓间充质干细胞,将第2代骨髓间充质干细胞在无血清神经干细胞培养环境中悬浮诱导,获取骨髓源神经祖细胞。将第3代骨髓源神经祖细胞分为2组贴壁培养:电磁场组与对照组。电磁场组干预方法为正弦波磁场、频率50Hz、强度5m T,60min/d,共15天,对照组置于无磁场干预的同等环境中。骨髓源神经祖细胞诱导后,采用免疫细胞荧光检测巢蛋白(Nestin)与微管蛋白抗体(Tuj-1)的表达变化;采用实时定量基因扩增荧光检测系统(Q-PCR)检测Nestin,唾液酸-神经细胞粘附分子(PSA-NCAM)和β-微管蛋白-Ⅲ(β-Ⅲtubulin),乙酰胆碱酯酶(ACHE),5-羟色胺(5-HT),γ-氨基丁酸(GABA)m RNA表达水平变化。结果:骨髓间充质干细胞在无血清神经干细胞培养环境中可形成骨髓源神经祖细胞,表达Nestin阳性产物;两组诱导后的神经元样细胞免疫细胞荧光检测Tuj-1均呈阳性表达,Q-PCR结果示Nestin,PSA-NCAM,β-Ⅲtubulin,ACHE,5-HT,GABA m RNA表达水平与诱导前比较显著下降(P<0.01),电磁场组β-Ⅲtubulin m RNA表达水平显著高于对照组(P<0.05)。结论:50Hz电磁场可以促进骨髓源神经祖细胞向神经元分化。
OBJECTIVE: To investigate the effect of low frequency electromagnetic fields on the differentiation of bone marrow-derived neural progenitor cells (BMMNC) induced by 50Hz electromagnetic field (EMF) intervention in the expression of m RNA in neural cells before and after differentiation of neural progenitor cells. Methods: Bone marrow mesenchymal stem cells were obtained by whole bone marrow culture. The second generation of bone marrow mesenchymal stem cells were induced in suspension in the culture of serum-free neural stem cells to obtain bone marrow-derived neural progenitor cells. The third generation of bone marrow-derived neural progenitor cells were divided into two groups of adherent culture: electromagnetic field group and control group. Electromagnetic field intervention method for the sinusoidal magnetic field, frequency 50Hz, intensity 5m T, 60min / d, a total of 15 days, the control group placed in the same environment without magnetic field intervention. After induced by bone marrow-derived neural progenitor cells, the expression of Nestin and Tuj-1 was detected by immunofluorescence staining. The expression of Nestin and saliva was detected by real-time quantitative gene amplification fluorescence detection system (Q-PCR) (PSA-NCAM) and β-tubulin, acetylcholinesterase (ACHE), serotonin (5-HT) and γ- aminobutyric acid (GABA) m RNA expression level changes. Results: Bone marrow-derived mesenchymal stem cells could form bone marrow-derived neural progenitor cells in the culture of serum-free neural stem cells and express Nestin-positive products. The immunofluorescent staining of Tuj-1 neurons in both groups showed positive expression of QTL and Q -PCR results showed that the expression of Nestin, PSA-NCAM, β-Ⅲtubulin, ACHE, 5-HT and GABA mRNA were significantly decreased compared with that before induction (P <0.01) Group (P <0.05). Conclusion: 50Hz electromagnetic field can promote differentiation of bone marrow-derived neural progenitor cells into neurons.