高压氧影响离体神经干细胞分化的量效及时效关系

Effect of hyperbaric oxygen administered at different pressures and different exposure time on differentiation of neural stem cells in vitro

摘要:

目的：探讨不同压力和不同稳压时间下高压氧(HBO)对离体神经干细胞（NSCs）分化的影响，旨在找出HBO治疗的最佳压力与稳压时间。方法：取出生3日内新生Sprague-Dawley大鼠的脑皮质组织，在含B27、bFGF和EGF的DMEM/F12培养基中悬浮培养NSCs。将传至2~3代的NSCs随机分为不做处理的对照组（CON）和HBO处理组。HBO处理组根据所用压力和稳压时间分为6组：NSCs分别给予1个绝对大气压（ATA）、2 ATA、3 ATA 的HBO处理，每种压力再分别给予30和60 min两种稳压时间。HBO后继续培养24 h，7组细胞在同一时间行免疫荧光染色，荧光显微镜下观察NSCs分化为NSE阳性细胞的百分率。结果：NSCs分化为NSE阳性细胞的分化率从低到高依次为CON组（3.72±0.88%），1ATA-30min组（3.85±0.44%），1ATA-60min组（5.45±0.52%），3ATA-30min组（6.08±0.57%），2ATA-30min组（6.72±0.76%），3ATA-60min组（7.89±0.62%），2ATA-60min（9.17±0.50%）；除1ATA-30min组外，其余各HBO组与对照组比较，差异有统计学意义（P<0.01）；同一压力下60min组与30min组之间的差异及2ATA-60min组与其他各组之间的差异均有统计学意义（P<0.01）。结论：2 ATA压力下稳压时间为60 min的HBO促进NSCs向神经元分化的作用最强。［中国当代儿科杂志，2010，12（5）：368-372］

Abstract:

OBJECTIVE: To study the effect of hyperbaric oxygen (HBO) administered at different pressures and different exposure time on the differentiation of neural stem cells (NSCs) in vitro. METHODS: The cerebral cortices from newborn rats (0-3 days old) were sterilely collected, digested, and centrifuged. After removal of the supernatant, the cells were re-suspended with DMEM/F12 medium containing B27, bFGF and EGF. The NSCs of 2-3 passages were randomly divided into seven groups: a control (untreated) and 6 HBO treatment groups that NSCs were subjected to HBO treatment of different pressures (1, 2 or 3 ATA) and different exposure time (30 or 60 minutes). The differentiated NSCs were examined by neuron-specific enolase (NSE) immunocytochemistry 24 hrs later. Percentage of NSE positive cells differentiated from NSCs was assessed by fluorescent microscopy. RESULTS: The percentage of NSE positive cells differentiated from NSCs was the highest in the HBO 2ATA-60min group (9.17±0.50%) (P<0.01), followed by the HBO 3ATA-60min (7.89±0.62%), HBO 2ATA-30min (6.72±0.76%), HBO 3ATA-30min (6.08±0.57%), HBO 1ATA-60min (5.45±0.52%), HBO 1ATA30min (3.85±0.44%) and control groups (3.72±0.88%). In addition to the HBO 1ATA-30min group, the other HBO treatment groups had increased significantly percentage of NSE positive cells compared with the control group (P<0.01). Under the same pressure, the 60 min treatment groups had increased significantly percentage of NSE positive cells compared with the 30 min treatment groups (P<0.01). CONCLUSIONS: HBO treatment (2 ATA, 60 minutes) produces a best effect in the differentiation of NSCs into neurons.［Chin J Contemp Pediatr, 2010, 12 (5):368-372］