The Effects Of Bumetanide On The Activity And The Expression Of AQP4mRNA Of Astrocytes After Hypoxia/Reoxygenation Injury

Objective: The objective of present work was to investigate the effects of administration of Bumetanide on the cell activity and the expression of AQP4 m RNA of Rat Astrocytes which experienced the injury of hypoxia/reoxygenaion in vitro.Methods:The Rat Astrocytes were abstracted from the newborn rats(less than 48 hours),and then primary cultured and subcultured. The Rat Astrocytes were identified after the forth passage. The oxygen-deprived environments was created by the mixture of 95%N2 and 5%CO2 The cell function and the expression of AQP4 of Astrocytes were tested The cell activity of Astrocytes: the cells were divided into four groups: control group, hypoxia/reoxygenation group(H/R group), bumetanide intervention group(50umol/L) and bumetanide intervention group(100umol/L). The degrading rate of MTT and release of lactate dehydrogenase(LDH) were estimated as the injury index of above astrocytes.The expression of AQP4: the cells were divided into three groups: control group, hypoxia/reoxygenation group(H/R group) and bumetanide intervention group(100umol/L).The transcription levels of AQP4 m RNA were detected by Q-PCR technique.Results:1.the MTT degradation rate of H/R group is obviously lower than that of control group(p<0.05);compared with H/R group, the MTT degradation rate of bumetanide intervention group(50umol/L)is significantly higher(p<0.05);compared with H/R group, the MTT degradation rate of bumetanide intervention group(100umol/L) group is significantly higher(p<0.05);There is no significant statistics difference between bumetanide intervention group(50umol/L)and bumetanide intervention group(100umol/L)(p>0.05).2.Compared with control group,the expression of LDH in H/R group is obviously higher(p<0.05);the expression of LDH in bumetanide intervention group(50umol/L)is lower than H/R group(p<0.05); the expression of LDH in bumetanide intervention group(100umol/L)is lower than H/R group(p<0.05);There is no significant statistics difference between bumetanide intervention group(50umol/L)and bumetanide intervention group(100umol/L)(p>0.05) about the expression of LDH.3.The transcription levels of AQP4 m RNA are gradually increase within 72 hours(0h,3h,24 h,72h) after the hypoxia/reoxygenation injury(p<0.01). Compared with control group, he expression of AQP4 m RNA in H/R group is significantly higher(p<0.01);in bumetanide intervention groups, the transcription levels of AQP4 m RNA are gradually increase within 72 hours(0h,3h,24 h,72h) after the hypoxia/reoxygenation injury(p<0.01); however, they are lower than H/P group(p<0.01).Conclusion: the result show that Bumetanide protect the astrocyte from the injury of hypoxia/reoxygenation;decrease the expression of AQP4 m RNA if the astrocyte has experienced the hypoxia/reoxygenation injury.