二氮嗪预处理对缺氧复氧后大鼠海马神经元凋亡及JNK表达的影响

目的探讨二氮嗪（DZ）预处理能否抑制C-Jun氨基末端激酶（JNK）表达而减轻缺氧复氧后大鼠海马神经元的凋亡。方法原代培养9～10d的SD大鼠海马神经元随机分为5组：对照组、DZ0，30，100μmol／L和DZ100μmol／L＋5-羟癸酸（5-HD）100μmol／L预处理组。除对照组外，其余4组神经元自缺氧前2d开始，每天实施以上对应浓度DZ预处理1h。连续3d。于体外缺氧4h复氧48h后，采用四唑蓝比色法测定海马神经元活力，AnnexinV—FITC流式细胞术测定凋亡率，Western blotting法检测JNK蛋白表达。结果与对照组比较，缺氧复氧后海马神经元的活力下降，凋亡率增大，JNK蛋白表达增强（P〈0．05）。与其他预处理组比较，DZ100μmol／L预处理组的神经元活力高，凋亡率低（P〈0．05），其他预处理组间比较无统计学意义：DZ预处理后，JNK蛋白表达减低，且DZ100μmol／L组表达弱于DZ30μmol／L组（P〈O．05）。同时给于5-HD预处理后，DZ未能抑制JNK蛋白表达。结论DZ预处理可能抑制JNK表达而减轻缺氧复氧后大鼠海马神经元的凋亡。

Effects of Diazoxide Preconditioning on JNK Expression and Anoxia-Reoxygenation-Induced Apoptosis of Hippocampal Neurons

Objective To study the effect of diazoxide （DZ） preconditioning on JNK protein expression and on anoxia-reoxygenation-induced apoptosis of hippocampal neurons. Methods Cultured for 9-10 d in vitro, the hippocampal neurons of SD rats were divided into the following 5 groups randomly： Control, DZ 0μmol/L, DZ 30 μmol/L, DZ 100 μmol/L, and DZ 100 μmol/Lq- 5-hydroxydeeanoate 100μmol/L （5-HD, a selective MitOKATP channel blocker）. Prior to oxygen deprivation, the hippocampal neurons except those in the control group were treated with the above concentrations of DZ or DZ＋5- HD for 1 h per day and for 3 d. Thereafter, neurons were subjected to anoxia for 4 h and followed by reoxygenation. After 48 h of reoxygenation, the neuronal vitality was measured by MTT method. The apoptotic rates were assayed by annexinV-FITC staining. The expression of JNK was detected and eval- uated by western blotting. Results Anoxia-reoxygenation injury reduced the neuronal vitality and increased the apoptotie rates significantly. In comparison with other anoxic groups, the DZ 100 μmol/L group showed an increased vitality but simultaneously decreased apoptotic rate（P〈0.05）. 5-HD could a- bolish the neuroprotection provided by 100μmol/L DZ. JNK in control normoxic neurons was expressed slightly while anoxia-reoxygenation led to its high expression. The administration of DZ reduced the expression of JNK significantly （P〈0.05）. Compared with DZ 30 μmol/L, DZ 100 μmol/L further decreased the expression of JNK protein （P〈0.05）. However, effects of DZ on the expression of JNK were reversed after the co-treatment with 5-HD. Conclusions DZ alleviates the anoxia-reoxygenation-in- duced apoptosis of hippoeampal neurons possibly through down-regulating the expression of JNK protein.