Necrostain-1对缺糖缺氧诱导的原代皮质神经元caspase非依赖性死亡的保护作用  

Protective effect of necrostain-I on oxygen-glucose deprivation induced caspase-independent cell death in primary cortical neurons

作　　者：樊红彬[1] 张翠翠[2] 陈巍巍[1] 程言博[1] 徐兴顺[1] 耿德勤[3] 

机构地区：[1]徐州医学院附属医院神经内科,徐州221002 [2]徐州医学院第二附属医院 [3]苏州大学第二附属医院

出　　处：《中华行为医学与脑科学杂志》2011年第12期1088-1091,共4页Chinese Journal of Behavioral Medicine and Brain Science

基　　金：国家自然科学基金（30700245）

摘　　要：目的研究低氧诱导因子（HIF）-1a是否介导缺糖缺氧诱导的原代皮质神经元caspase非依赖性细胞死亡，Necrostain-1（Nec-1）能否抑制其表达起到保护作用。方法（1）原代皮质神经元培养14d，予caspase抑制剂z—VAD．fmk（z—VAD）预保护30min，同时分别予Nec-10．1，1，5，10，25，50μmoL／L，缺糖缺氧（OGD）2h、再灌注12h，通过LDH测细胞死亡情况；（2）予z-VAD作用后30min后，缺糖缺氧（OGD）2h再灌注0，2，6，12，24，48h，Westernblot检测HIF-1α蛋白表达情况，RT—PCR测HIF-1αRNA表达；（3）予z-VAD和Nee-125μmo[／L作用30min，OGD2h、再灌注12h后检测HIF-10t蛋白及RNA表达情况。结果（1）予Nec-15μmoL／L（6．97±0．06）后与未加Nec-1组（14．23±0．08）比较LDH水平明显下降（P〈0．05），Nec-125μmol／L（2．21±0．05）时LDH降至正常水平与正常组（1．03±0．03）比较，P〉0．05）；（2）缺糖缺氧2h再灌注后HIF-10α蛋白表达增加，再灌注2h后（0．57±0．09）与正常组（0．24±0．01）相比差异有统计学意义（P〈0．05），再灌注12h（0．91±0．08）达高峰，HIF-10LRNA表达无变化（P〉0．05）；（3）与未予Nec．1组（0．83±0．03）相比，Nec-1组（0．32±0．04）HIF-1α蛋白表达明显降低（P〈0．05），HIF-1αRNA表达无变化（P〉0．05）。结论HIF一1仅介导了缺氧缺氧诱导的原代皮质神经元caspase非依赖性死亡，Nec-1能够抑制HIF-1α的表达起到保护作用。Objective To explore whether hypoxia-inducible factor-1α （HIF-1α ）is involved in oxygen- glucose deprivation （OGD）induced easpase-independent cell death in primary cortical neurons and whether their expression is infected by necrostatin-1 （ Nee-1 ）. Methods （ 1 ） Primary cerebrocortical neurons were cultured for 14 days. Pretred z-VAD. Fmk （z-VAD）and Nec-lwith 0.1,1,5,10,25 and 50 μmoL/L before the neurons were exposed to OGD for 2 hours and reoxygenated for 12 hours, then cell viability was determined by measure LDH lev- el. （2）Pretred z-VAD before the neurons were exposed to OGD for 2 hours , then reoxygenated for 0,2,6,12,24 and 48 hours. Then western blot analysis protein level of HIF-1 ct;rt-PCR check its RNA level . （3）Pretred z-VAD and Nee-1 with 251xmol/L before the neurons were exposed to OGD for 2 hours and reoxygenated for 12 hours. Then western blot analysis protein level of HIF-1α; rt-PCR check its RNA level. Result （ 1 ） When cells were pretread Nee-1 with 5 μmol/L（6.97± 0.06）, the level of LDH was lower than cells untreated （14.23 ± 0.08 ） （P 〈 0. 05 ） ; At 25 μmol/L（2.21 ±0. 05 ）, the level of LDH was essentially the same as that of the control （ 1.03 ± 0. 03 ） （P 〉 0.05 ）. （2）The protein level of HIF-1 etwas different from normal （0.24 ± 0.01 ） when exposed to OGD for 2 hours and reoxygenated for 2 hours （ 0. 57 ± 0.09 ） and was highest after cells were exposed to OGD for 2 hours and reoxygenated for 12 hours （0.91 ± 0. 08 ） （P 〈 0.05 ）. The RNA level of HIF-1α when cells were exposed to OGD was not deferent from normal （P 〉 0.05 ）. （ 3 ） When cells were pretread with Nec-1 （ 0.32 ± 0.04）, the protein level of HIF-1αwere lower than untreated （0.83 ± 0. 03 ） （P 〈 0.05 ）, but the RNA level of HIF-1 ct had no deference（P〉0.05）. Conclusion HIF-1α was involved in cell＇ s caspase- independent cell death;Nec-1 can protect neurons through inhibiting the expression of HIF-1α

关 键 词：低氧诱导因子 Necrostain-1 缺糖缺氧 caspase非依赖性死亡 

分 类 号：R285.5[医药卫生—中药学]