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目的探讨在离体细胞水平下高压氧(HBO)对缺氧缺血性脑损伤(HIBD)时新生SD大鼠神经干细胞(NSCs)增殖分化的影响。方法采用Rice-Vannucci法制成HIBD模型,分别制备正常及HIBD时SD大鼠脑组织匀浆,将传至2～3代的SD大鼠NSCs根据处理的不同随机分为空白对照组(CON组)、与正常脑组织匀浆共培养组(NC+N组)、与HIBD脑组织匀浆共培养组(NC+HIBD组)、与HIBD脑组织匀浆共培养1h后给予HBO处理组(NC+HIBD+HBO组)和与正常脑组织匀浆共培养1h后给予HBO处理组(NC+N+HBO)5组。5组细胞在同一时间行免疫荧光染色,分别进行NSE、GFAP、O4染色,荧光显微镜下计数各组NSCs分化为NSE阳性细胞、GFAP阳性细胞和O4阳性细胞的百分率,并进行比较。结果与CON组相比,NC+N+HBO组、NC+N组、NC+HIBD组分化为神经元和少突胶质细胞的百分比明显增加(P<0.05),其中HIBD组增加最多,在此基础上再给予HBO处理,NSCs向神经元的分化会进一步增加,向少突胶质细胞的分化也相应增加;与CON组相比,NC+N组、NC+HIBD组、NC+HIBD+HBO组分化为星形胶质细胞的百分比明显减少(P<0.05)。结论在离体细胞水平,HBO可促进HIBD大鼠NSCs分化为神经元和少突胶质细胞而抑制其向星形胶质细胞分化。
Objective To investigate the effect of hyperbaric oxygen (HBO) on the proliferation and differentiation of neural stem cells (NSCs) of neonatal SD rats exposed to hypoxic-ischemic brain damage (HIBD) in vitro. Methods The HIBD model was made by the method of Rice-Vannucci, and the SD rat brain homogenates were prepared at normal and HIBD respectively. The SD rats NSCs transfected from 2 to 3 generations were randomly divided into blank control group (CON group) (NC + N group), HIBD brain homogenate co-culture group (NC + HIBD group) and HIBD brain homogenate co-culture group (NC + N group) HIBD + HBO group) and HBO group (NC + N + HBO group) were co-cultured with normal brain homogenate for 1 hour. Five groups of cells were immunofluorescently stained with NSE, GFAP and O4 respectively at the same time. The percentage of NSCs differentiated into NSE positive cells, GFAP positive cells and O4 positive cells in each group were counted under fluorescence microscope and compared. Results Compared with CON group, the percentages of neurons and oligodendrocytes in NC + N + HBO group, NC + N group and NC + HIBD group were significantly increased (P <0.05), and in HIBD group, On the basis of HBO treatment, the differentiation of NSCs to neurons was further increased and the differentiation to oligodendrocytes was also increased. Compared with CON group, NC + N group, NC + HIBD group, NC + HIBD + The percentage of HBO-differentiated astrocytes decreased significantly (P <0.05). Conclusion In vitro, HBO can promote the differentiation of NSCs into neurons and oligodendrocytes in HIBD rats and inhibit their differentiation into astrocytes.