Clemens J A, Stephenson D T, Smalstig E B, Dixon E P, Little S P
Eli Lilly and Company, CNS Division, Lilly Research Laboratories, Indianapolis, Ind. 46285, USA.
Stroke. 1997 May;28(5):1073-80; discussion 1080-1. doi: 10.1161/01.str.28.5.1073.
After global ischemia, brain levels of hydrogen peroxide, oxygen radicals, and the cytokines tumor necrosis factor-alpha (TNF-alpha) and interleukin-1 beta (IL-1 beta) are increased. Oxygen radicals, TNF-alpha, and IL-1 beta are known to activate nuclear factor-kappa B (NF-kappa B) in vitro. The present study was performed to determine whether NF-kappa B was activated in vivo by global ischemia in hippocampal CA1 neurons.
Adult male rats were subjected to 30 minutes of four-vessel occlusion and killed 72 hours later. Levels of NF-kappa B p50 and p65 subunits in hippocampus were determined by immunocytochemistry, Western blot, and gel-shift analysis. Specific labeling of DNA strand breaks was demonstrated by means of an Apoptag apoptosis detection kit.
Labeling of DNA strand breaks was present at 72 hours. Chromatin compaction and segregation, a characteristic of apoptosis, was observed in sections stained with hematoxylin and eosin. NF-kappa B p50 and p65 immunoreactivity localized only to nuclei of CA1 neurons at 72 hours after reperfusion. Induction of the activated p50 and p65 subunits was confirmed by Western blot and electromobility shift analysis. The results demonstrate that NF-kappa B is activated selectively in hippocampal CA1 neurons at 72 hours after four-vessel occlusion, which is at the approximate time of CA1 neuronal cell death.
Transient forebrain ischemia resulted in a marked activation of nuclear NF-kappa B in the highly vulnerable CA1 sector. Intense nuclear localization of NF-kappa B was associated only with dying neurons; regions of the hippocampus that were not vulnerable to four-vessel occlusion did not exhibit nuclear NF-kappa B localization. The elevation of NF-kappa B in degenerating CA1 neurons may be associated mechanistically with apoptotic or necrotic cell death.
全脑缺血后，脑内过氧化氢、氧自由基以及细胞因子肿瘤坏死因子-α（TNF-α）和白细胞介素-1β（IL-1β）的水平会升高。已知氧自由基、TNF-α和IL-1β在体外可激活核因子-κB（NF-κB）。本研究旨在确定全脑缺血是否会在体内激活海马CA1神经元中的NF-κB。
成年雄性大鼠接受30分钟的四血管闭塞，72小时后处死。通过免疫细胞化学、蛋白质印迹法和凝胶迁移分析测定海马中NF-κB p50和p65亚基的水平。使用Apoptag凋亡检测试剂盒检测DNA链断裂的特异性标记。
72小时时出现DNA链断裂标记。在苏木精和伊红染色的切片中观察到染色质浓缩和分离，这是凋亡的特征。再灌注72小时后，NF-κB p50和p65免疫反应性仅定位于CA1神经元的细胞核。蛋白质印迹法和电泳迁移率变动分析证实了活化的p50和p65亚基的诱导。结果表明，四血管闭塞72小时后，NF-κB在海马CA1神经元中被选择性激活，这大约是CA1神经元细胞死亡的时间。
短暂性全脑缺血导致高度易损的CA1区核NF-κB显著激活。NF-κB的强烈核定位仅与濒死神经元相关；海马中不易受四血管闭塞影响的区域未表现出核NF-κB定位。退变的CA1神经元中NF-κB的升高可能在机制上与凋亡或坏死性细胞死亡有关。
