胶原凝胶三维支架对神经干细胞增殖和分化的影响

Effect of three-dimensional collagen gel scaffold on proliferation and differentiation of neural stem cells

目的：观察胶原凝胶三维支架对大鼠胚胎神经干细胞增殖和分化的影响。方法：将大鼠胚胎神经干细胞接种于胶原凝胶三维支架中，通过倒置显微镜观察胶原凝胶中神经干细胞生长和增殖的情况。以悬浮培养作为对照组，采用免疫荧光鉴定胶原凝胶中神经干细胞标志物，并检测神经干细胞分化成不同神经细胞的比例。以CCK-8方法检测不同胶原凝胶浓度对神经干细胞活力的影响。结果：胶原凝胶三维支架中的神经干细胞可以增殖并形成神经球，这些神经球表达神经干细胞标志物，并且可以分化形成神经元、星形胶质细胞和少突胶质细胞。与悬浮培养的神经球相比，三维培养中形成的神经球其分化为神经元、星形胶质细胞和少突胶质细胞的比例无差异。CCK-8检测表明胶原浓度为0．5mg／ml时最适合神经干细胞生长。结论：本实验表明浓度为0．5mg／ml的胶原凝胶三维支架适合神经干细胞的增殖和分化，三维培养的神经干细胞在脊髓损伤治疗中具有潜在的应用前景。

Objective： To observe the effect of three-dimensional （3D） collagen gel scaffold on proliferation and differentiation of embryonic rat neural stem cells （NSCs）. Methods~ The embryonic rat neural stem cells were seeded in 3D collagen gel to observe the survival and proliferation of neural stem cells under an inverted microscope. Compared with NSCs cultured in suspension, the NSCs marker and the proportion of differentiated cells in 3D collagen gel were identified by the immunofluorescence techniques. The cell viability of NSCs cultured in different concentration of collagen gel was detected by CCK-8 assay. Results： The collagen-trapped NSCs actively expanded and formed neurospheres. These neurospheres cultured in 3D collagen gel were positive for nestin and differentiated into the neurons, astrocytes and oligodendrocytes. Immunofluorescence showed that there was no significant difference in the proportion of differentiated cells between two groups. The CCK-8 testing showed that the concentration of 0. 5 mg/ml collagen gel was the most suitable for the growth of NSCs. Conclusion： The results suggest that the concentration of 0. 5 mg/ml collagen gel is suitable for the proliferation and differentiation of NSCs. The NSCs cultured in 3D collagen gel may have a potential application in the repair of spinal cord injury.