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目的:中风是一种严重危害人类健康的疾病,神经干细胞能够促进中枢神经系统功能的修复。神经干细胞的增殖、分化、迁移需要干细胞niche的调控,前期实验发现人参总皂苷可以显著增加中风后神经干细胞的数量。体外模拟神经干细胞niche内星形胶质细胞和脑微血管内皮细胞对神经干细胞的影响,以研究人参总皂苷是否能作用于星形胶质细胞和脑微血管内皮细胞而促进中风后神经干细胞的分化,修复脑损伤?将脑微血管内皮细胞、星形胶质细胞和神经干细胞共培养,观察模拟神经干细胞niche内复杂微环境条件下,人参总皂苷能否使脑微血管内皮细胞和星形胶质细胞分泌的VEGF增多,从而促进中风损伤的神经干细胞分化。方法:取1-3d的新生SD大鼠,分离培养星形胶质细胞和脑微血管内皮细胞。取孕16d SD大鼠,分离培养神经干细胞。利用Transwell装置,将神经干细胞、星形胶质细胞和脑微血管内皮细胞共培养。制备神经干细胞缺氧6h的脑中风模型。用含1μg/ml的人参总皂苷培养基作用1d,设置空白对照组。MAP-2标记成熟神经元,GFAP标记星形胶质细胞。用细胞免疫荧光化学染色检测缺氧6h神经干细胞分化后的MAP-2和GFAP阳性细胞比例,ELISA检测人参总皂苷对星形胶质细胞和脑微血管内皮细胞共培养上清中VEGF含量的影响。结果:与脑微血管内皮细胞、星形胶质细胞共培养条件下,与空白对照组比较,人参总皂苷可显著增加MAP-2阳性细胞比例(P<0.01);人参总皂苷作用于星形胶质细胞和脑微血管内皮细胞48h可显著上调VEGF表达(P<0.01)。结论:人参总皂苷作用于脑微血管内皮细胞和星形胶质细胞可增加中风后神经干细胞向神经元定向分化并促进其成熟,可能与人参总皂苷调控脑微血管内皮细胞和星形胶质细胞VEGF分泌,改善神经干细胞niche微环境有关。
Objective: Stroke is a serious disease that endangers human health. Neural stem cells can promote the repair of the central nervous system. Proliferation, differentiation and migration of neural stem cells require the regulation of stem cell niche. Previous studies found that total ginsenoside could significantly increase the number of neural stem cells after stroke. To investigate the effects of ginsenosides on neural stem cells in vitro and in vitro.METHODS The effects of ginseng total saponins on astrocytes and brain microvascular endothelial cells were observed in vitro to promote the differentiation of neural stem cells after stroke, Repair brain damage? Brain microvascular endothelial cells, astrocytes and neural stem cells were co-cultured to observe the simulated neural stem cell niche complex microenvironment conditions, ginseng total saponins can make brain microvascular endothelial cells and astrocytes secreted Increased VEGF contributes to stroke-induced neural stem cell differentiation. Methods: Newborn SD rats were used for 1-3 days to separate and culture astrocytes and brain microvascular endothelial cells. SD rats were taken for 16 days, and neural stem cells were isolated and cultured. Neural Stem cells, astrocytes and brain microvascular endothelial cells were co-cultured with Transwell apparatus. Preparation of Stroke Model of Neural Stem Cells Hypoxia for 6h. With 1μg / ml of total ginseng saponin medium role 1d, set the blank control group. MAP-2 labeled mature neurons, GFAP-labeled astrocytes. The ratio of MAP-2 and GFAP positive cells differentiated into NSCs at 6h after hypoxia was detected by immunofluorescence staining and the effect of total ginsenoside on VEGF in supernatants of astrocytes and brain microvascular endothelial cells was detected by ELISA. Results: Compared with the blank control group, the total ginsenosides significantly increased the proportion of MAP-2-positive cells (P <0.01) under the conditions of co-culture with cerebral microvascular endothelial cells and astrocytes; After 48h, the expression of VEGF increased significantly (P <0.01). Conclusion: The effect of ginseng total saponin on brain microvascular endothelial cells and astrocytes can increase the directional differentiation of neural stem cells into neurons after stroke and promote their maturation, which may be related to the regulation of brain microvascular endothelial cells and astrocytes by ginseng total saponins Secretion, improve neural stem cell niche microenvironment related.