缺氧/复氧条件下星形胶质细胞水通道蛋白4和5表达变化的研究

The Expression of Aquaporin 4 and Aquaporin 5 in Astrocytes under the Condition of Hypoxia/ Reoxygenation

目的:观察缺氧/复氧条件下星形胶质细胞水通道蛋白(AQP)4和5表达的变化,探讨脑缺血再灌注后脑水肿与AQP4和AQP5的关系。方法:取新生24h内的SD大鼠皮质新鲜脑组织,进行原代和传代培养。以95%N2和5%CO2造成细胞缺氧,用倒置相差显微镜对细胞进行形态学观察,用锥虫蓝染色法测定缺氧及复氧后不同时间点星形胶质细胞的死亡数以反映星形胶质细胞的存活能力,应用细胞免疫化学技术测定星形胶质细胞缺氧及复氧后各个时间点AQP4、AQP5表达的变化。结果:缺氧4及8h后细胞形态变化不明显,随着复氧时间的延长出现细胞损伤的表现。与对照组比较,缺氧后4及8h有少量细胞死亡(P<0.05),随着复氧时间延长细胞死亡数亦逐渐增多(P<0.01);缺氧4及8h后,AQP4、AQP5阳性表达细胞数均减少(P<0.01),而复氧后AQP4、AQP5阳性表达细胞数逐渐升高并随时间延长呈增高趋势(P<0.01)。结论:星形胶质细胞对缺氧的耐受能力较强,但复氧时出现明显损伤;AQP4、AQP5表达的变化与缺血-再灌注损伤后脑水肿存在相关性。

To observe the changes of astrocytes on the expression of AQP4 and AQP5 under the condition of hypoxia/regoxygenation, and analyze the relation between the ischemia/reperfusion damage of brain and AQP4 and AQP5. Methods： The primary and subculture cells from the cortex of SD fetal rat （less than 24 hours） were divided into two groups：normal control group（C）, hypoxia/reoxygenation group（H/R）. Morphological observation was carried by inverted phase contrast microscope. Trypan blue staining was used to detect the survival rate of astrocytes at different time points after hypoxia/reoxygenation. Immunocytochemistry technology was used to examine the expression of AQP4, AQP5 in the astrocytes at the indicated time points after hypoxia/reoxygenation. Results： The cell morphology did not change significantly after hypoxia for 8 hours. However,reoxygenation made the damage obviously and the death of astrocytes increased, showing a clear time-effect relationship. The expression of AQP4-positive and AQP5-positive cells increasingly showed a tendency to decrease compared with the normal control group after 8 hours of hypoxia in 37℃, but expressed rising after reoxygenation and showed a rising trendwith time. Conclusion： Astrocytes have better tolerance to hypoxia. It suggested that there be an apparent correlation between the expression of AQP4 and AQP5 and brain edema after ischemical reperfusion injury.