摘要
背景:大量研究显示高热可诱导机体细胞发生广泛凋亡,但对于高热如何介导神经元细胞凋亡并没有深入研究报道。目的:检测热打击细胞模型中核因子κB信号通路对活性氧诱导细胞凋亡的影响。方法:使用细胞培养箱建立细胞热打击模型,热打击组分别将细胞置于39,41,43℃培养箱中进行热打击2 h,对照组(37℃组)将细胞置于标准37℃、体积分数5%CO2细胞培养箱。使用Annexin V-FITC/PI双染色方法检测不同温度热打击下神经元细胞凋亡率,Westen blot检测caspase-3及抗-核因子КB65蛋白表达,DCFH法检测细胞内活性氧含量,同时检测活性氧抑制剂MnTMPyP及核因子κB抑制剂PDTC对热打击细胞凋亡的影响。结果与结论:39℃热打击对细胞凋亡无影响,41℃热打击诱导细胞少量凋亡(10.19%),43℃热打击诱导诱导细胞大量凋亡(43.02%)。caspase-3和抗-核因子κB65蛋白的表达于热打击温度依赖的方式增加。MnTMPyP及PDTC均可有效阻断热打击引起的caspase-3、抗-核因子κB65蛋白的表达和细胞凋亡。结果证实热打击后细胞内活性氧增加诱导神经元细胞凋亡,提示核因子κB可能作为中间信号通路参与了热打击引起的细胞凋亡。
BACKGROUND：Hyperpyrexia can induce a wide range of cel apoptosis in organisms, but no study has introduced the mechanism of heat stress-induced neuronal apoptosis. OBJECTIVE：To observe the effect of nuclear factor kappa B （NF-κB） signal pathway on heat stress-induced neuronal apoptosis through reactive oxygen species.METHODS：Heat stress model was established in the cel incubator. Heat stress group of cel s were incubated at 39,41,43℃for2hours,whilecontrolgroupofcelswereincubatedat37 ℃in5% CO2 for 2 hours. Apoptosis was analyzed by flow cytometry using Annexin V-FITC/PI staining. The expression levels of caspase-3 and p-NF-κB65 were determined by western blot analysis. The amounts of intracel ular reactive oxygen species were assayed by DCFH staining. In addition, the effect of MnTMPyP and PTDC on heat stress-induced apoptosis was also studied.RESULTS AND CONCLUSION：39 ℃ heat stress had no impact on the apoptosis, 41 ℃ heat stress induced a smal amount of apoptosis （10.19%）, and 43 ℃ heat stress triggered a large amount of apoptosis （43.02%）. The expression of caspase-3 and p-NF-κB65 was increased, in a temperature-dependent manner. In addition, both MnTMPyP and PTDC significantly decreased the heat stress-induced apoptosis and expression of caspase-3 and p-NF-κB65. Experimental findings indicate that, the increase of intracel ular reactive oxygen species may induce neuronal apoptosis, and NF-κB participates in the heat stress-induced neuronal apoptosis as the intermedial signal pathway.
出处
《中国组织工程研究》
CAS
CSCD
2014年第11期1641-1646,共6页
Chinese Journal of Tissue Engineering Research
基金
国家自然科学基金(81071529
81101467)
全军十二.五重点课题(BWS12J018)~~
