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目的探讨人脐血间充质干细胞用于神经细胞再生的可行性与人脐血间充质干/祖细胞最佳的诱导培养条件。寻找一种合适的细胞来源进行移植以代替受损的神经元、星形胶质细胞和少突胶质细胞来修复神经损伤。方法将脐血单个核细胞置于低血清(2%)达氏修正依氏培养基中培养,生成贴壁细胞层,依传代方法,用相同培养条件进行扩增,扩增后的贴壁细胞置入细胞培养液(加入新生大鼠神经细胞分离培养上清液)中,并添加表皮生长因子(EGF)和碱性成纤维细胞生长因子(bFGF)进行诱导分化。采用流式细胞术、免疫组织化学法分别检测被诱导的神经样细胞的表面标志及细胞特异性标志、结构,并进行分析。结果脐血间充质干/祖细胞克隆在脐血单个核细胞中出现频率为0.5×10-6,在传20代时,可有效扩增1.5×106倍,诱导后,70%的脐血间充质干/祖细胞分化为神经样细胞。结论采用上述扩增与诱导条件,脐血间充质干/祖细胞可得到有效扩增,并可高效向神经样细胞分化,从而证实脐血是一种细胞替代治疗前景光明的细胞来源。
Objective To investigate the feasibility of using human umbilical cord blood mesenchymal stem cells for nerve cell regeneration and the optimal culture conditions of human umbilical cord blood mesenchymal stem / progenitor cells. Find a suitable source of cells for transplantation to replace damaged neurons, astrocytes and oligodendrocytes to repair nerve damage. Methods Umbilical cord blood mononuclear cells were cultured in low serum (2%) Dysosma to establish the adherent cell layer. According to the passage method, the adherent cells were expanded under the same culture conditions. The cells were cultured in the medium of neonatal rat neural cells and the supernatant was added into the culture medium. Epidermal growth factor (EGF) and basic fibroblast growth factor (bFGF) were added to induce differentiation. Flow cytometry and immunohistochemistry were used to detect the surface markers and cell-specific markers of induced neuron-like cells, and their structures were analyzed. Results The umbilical cord blood mesenchymal stem / progenitor cells cloned in cord blood mononuclear cells in the frequency of 0.5 × 10-6, in the passage of 20 generations can be effectively amplified 1.5 × 106 times, after induction, 70% of cord blood Mesenchymal stem / progenitor cells differentiate into neuron-like cells. Conclusion Using the above conditions of amplification and induction, umbilical cord blood mesenchymal stem / progenitor cells can be effectively expanded and differentiated into neuron-like cells with high efficiency, thus confirming that umbilical cord blood is a promising cell source for cell replacement therapy.