不同浓度臭氧对大鼠离体脑片氧糖剥夺及再灌注损伤模型的保护作用

Neuro-protective Effect of Different Concentration of Ozone on Cultured Rat Brain Slices Underwent Oxygen-glucose Deprivation and Reperfusion

目的探讨不同浓度臭氧对大鼠离体脑片氧糖剥夺及再灌注损伤模型的保护作用。方法制备SD大鼠脑片，室温下保存在人工脑脊液（ACSF）中，根据实验分组情况选择性经过下列程序：恢复期60min、平衡期30min、氧糖剥夺（OGD）期30min及再灌注期90min。对照组（CTRL）不经过OGD期；模型组不施加臭氧干预（OGD／R）；实验组于再灌注期分别给予10，20，30，40，50μg／mL的臭氧，并且根据ACSF中是否含有人血白蛋白（HSA）分成2个亚组。再灌注期结束后分别测定各组ACSF中谷氨酸（Glu）和乳酸脱氢酶（LDH）的含量。结果OGD 30min后给予90min的再灌注，模型组与对照组比较Glu和LDH释放增加（P〈0．01）。不同浓度的臭氧在不同程度上拮抗了Glu和LDH的释放，其中40μg／mL的臭氧效应最明显，低浓度（10～30μg／mL）或高浓度（50μg／mL）臭氧组与OGD／R组比较效应不明显（P〉0．05）；在含有HAS组与不合HAS组比较，给予臭氧（40μg／mL）干预能明显减少Glu和LDH的释放（P〈0．05）。结论40μg／mL臭氧气体对大鼠离体脑缺血再灌注损伤模型中的神经细胞具有保护作用，并且HSA能增强臭氧的保护作用。

Objective To evaluate the neuro-protective effect of different concentration of ozone on cultured rat brain slices which were subjected to oxygen-glucose deprivation and reperfusion. Methods In this study, brain slices of SD rats were cultured and subjected to oxygen-glucose depri- vation and reperfusion. Slices were maintained in ACSF for 1 hour at room temperature （ Recovery period） and then equilibrated for an additional pe- riod of 30 minutes （ Equilibration ）. Afterwards, the phase of oxygen/glucose deprivation （ OGD ） was carried out for 30 minutes. After the OGD peri- od, ischemic solution was replaced by 2 mL of fresh, oxygenated ACSF for an additional 90-minute period （ Reperfusion ）. Slices were incubated in ACSF for 120 minutes （control conditions, CTRL） or subjected to oxygen/glucose deprivation for 30 minutes followed by 90-minute immersion in normally oxygenated ACSF （OGD/R）. Increasing concentrations of ozone （ 10-50 μg/mL） were administrated to ACSF during the reperfusion. Ozone treated group were divided into two subgroups according to the absence or in presence of Human Serum Albumin HSA （ 150μg/mL） during the reperfusion phase. Neuronal damage was assessed quantitatively by measuring the amount of both glutamate （Glu） and lactate dehydregenase （LDH） released into the ACSF during 90-minutes-reperfusion period. Results As expected, 30 minutes of oxygen-glucose deprivation followed by 90 minutes of reperfusion （OGD/R） caused a highly significant release in LDH and Glu compared to the CTRL （P 〈 0.001 ）. Ozone （10-50 μ/mL） antagonized OGD/R-induced LDH and Glu release in different levels. In particular, 40 μg/ml proved to be the most effective concentration compared with those of CTRL. While lower （ 10- 30μg/mL） or higher （ 50 μg/mL） ozone concentrations had not statistically significant effects. The difference about the effects on LDH and Glu release was significance when comparing the groups with and without HAS （P 〈 0.01 ）. Conclusion Oxygen-ozone gaseous mixture in which the ozone＇ s concentration is 40 μg/mL appeared to be effective in reverting damage of rat brain slices which were subjected to oxygen-glucose deprivation ; moreover, the presence of HSA in ACSF improved the ozone＇ s effectiveness.