摘要
目的观察不同浓度谷氨酸（Glu）损伤螺旋神经元中凋亡诱导因子（apoptosis inducing factor,AIF）的表达与分布变化,探讨利用谷氨酸损伤体外培养螺旋神经元是否与AIF凋亡途径相关。方法取40只出生后0~3天SD仔鼠螺旋神经元行体外培养,所得培养细胞平均分为正常组和10mM、20mM、40mM Glu组,正常组仅给予正常SGNs培养液培养,其余三组分别加入相应浓度谷氨酸和SGNs培养液,培养48h后,应用光学显微镜、免疫荧光染色及实时荧光定量PCR方法观察四组中SGN细胞形态、SGN中AIF的分布及AIF、钙蛋白酶（calpain）、半胱氨酸天冬氨酸3（caspase 3）的表达变化;用TUNEL法观察细胞凋亡。结果 20mM和40mM Glu组SGNs细胞形态改变和细胞凋亡明显,并伴有AIF核转位;不同浓度Glu组AIF和calpain基因表达明显高于正常组（P〈0.05）,但caspase 3表达无明显升高（P〉0.05）。结论谷氨酸损伤体外培养的螺旋神经元过程中,AIF凋亡途径参与了细胞凋亡,calpain的高表达也促进了兴奋性神经递质对SGNs的损伤,但caspase3途径无明显作用。
Objective The study aimed to explore the relationship between AIF related pathway and the inju-ring of cultured SGNs （spiral ganglion neurons）by glutamate toxicity,and to find AIF expression and distribution changes in SGNs.Methods SGNs of 40 newborn rats within 3 day were obtained and cultured in vitro.Cultured cells were divided into four groups：the normal control group,10 mM,20 mM and 40 mM glutamate injured group, separately.After 48 h hours culturing,optical microscopy,immune fluorescence staining and real-time fluores-cence quantitative PCR were used to observe the morphology,AIF distribution,and AIF,calpain,Caspase3 expres-sion changes in SGNs in vitro.TUNEL was used to verify the cell apoptosis.ResuIts Noticeable morphological chan-ges and cell apoptosis were occurred in 20 mM glutamate group,with AIF nuclear translocation.AIF gene expression was significantly higher than normal after glutamate administration （P〈0.05）;moreover,calpain gene expression increased（P〈0.05）;but caspase3 expression was not statistically significantly increased in all glutamate treated groups （P＆gt;0.05）. ConcIusion In the process of cultured SGNs injured by glutamate,AIF participated in the cell apoptosis.Noticeable cell apoptosis were occurred in 20 mM glutamate group with AIF nuclear translocation.Calpain up-expression also contributed to excitatory neurotransmitter injury on SGNs,but Caspase 3 had no obvious effects.
出处
《听力学及言语疾病杂志》
CAS
CSCD
北大核心
2014年第6期620-624,共5页
Journal of Audiology and Speech Pathology
基金
国家自然科学基金(81120108008
30930098
81271069)
国家"973"项目课题(2011CB504505)联合资助
