MK-801拮抗水杨酸钠致螺旋神经节细胞损伤的实验研究

Blocking NMDA Receptors for Reversal of Excitotoxicity Processes in Spiral Ganglion Neurons Induced by Sodium Salicylate

目的探讨非特异性阻断NMDA受体拮抗水杨酸钠对离体培养螺旋神经节细胞兴奋性损伤的作用。方法将培养的耳蜗螺旋神经节细胞随机分为3组,分别为正常对照组:培养液中仅加入1mM谷氨酸;水杨酸钠组:1mM谷氨酸+5mM水杨酸钠;MK-801组:50μM MK-801+1mM谷氨酸+5mM水杨酸钠。培养24h后加药物干预3h,收集细胞采用实时荧光定量PCR及免疫荧光技术检测BDNF exonⅣ,BDNF exonⅥ转录及Caspase-3mRNA转录和蛋白表达的改变,研究应用MK-801非特异性阻断NMDA受体拮抗水杨酸钠对离体培养螺旋神经节细胞兴奋性损伤的作用。结果水杨酸钠组较谷氨酸组及MK-801组螺旋神经节细胞中BDNF exonⅣ,BDNF exonⅥ和Caspase-3 mRNA的转录及Cas-pase-3蛋白的表达水平显著增高(P值均<0.05);BDNF exonⅣ的转录在MK-801组较谷氨酸组明显降低(P<0.05);BDNF exonⅥ的转录在MK-801组较谷氨酸组未发生具有统计学意义的改变;Caspase-3的转录及蛋白的表达在MK-801组较谷氨酸组显著提高(P<0.05)。结论非特异性阻断NMDA受体能拮抗水杨酸钠引起的离体培养螺旋神经节细胞中BDNFexonⅣ,BDNFexonⅥ及Caspase-3上调。

Objective To investigate the antagonism of non-specific blockage of NMDA receptors on the process of exci-totoxicity in spiral ganglion neurons induced by sodium salicylate. Methods Cultured spiral ganglion cells were divided into three groups on the basis of the drugs which were mixed into the culture medium：1 mM glutamic acid （the GLU group）, 1 mM glutamic acid ＋ 5 mM sodium salicylate （the salicylate group）, or 1 mM glutamic acid ＋ 5 mM sodium salicylate ＋ 50 μM MK-801 （the MK-801 group） . After 24 hours of culturing and 3 hours of reaction with the agents, cells were collected and test-ed for transcription and expression levels of BDNF exonⅣ, BDNF exonⅥand Caspase-3 using real-time PCR and immuno-fluorescence. Results Compared with the GLU and MK-801 groups, the transcription of BDNF exonⅣ, BDNF exonⅥand Caspase-3, as well as the expression of Caspase-3, in the salicylate group were upregulated （p〈0.05）. Compared with the GLU Group, the transcription of BDNF exon Ⅳin the MK-801 group decreased （p〈0.05）. Transcription of BDNF exon Ⅵshowed no difference between the MK-801 and GLU groups, although there was in increase in expression of Caspase-3 （p〈0.05）. Conclusions Non-specific blockage of NMDA receptor can reverse the process of excitotoxicity in spiral ganglion neu-rons induced by sodium salicylate.