Yan Jun-hao, Yang Xiao-mei, Chen Chun-hua, Hu Qin, Zhao Jing, Shi Xian-zhong, Luan Li-ju, Yang Lei, Qin Li-hua, Zhou Chang-man
Department of Anatomy and Embryology, Peking University Health Science Centre, Beijing 100083, China.
Chin Med J (Engl). 2008 Mar 5;121(5):414-9.
The mechanism of cerebral vasospasm following subarachnoid haemorrhage (SAH) is not understood. Here, we hypothesized that apoptosis of endothelial cells induced by p53 and its target gene em dash p53 upregulated modulator of apoptosis (PUMA) played an important role in development of cerebral vasospasm. We also observed the effects of a p53 inhibitor, pifithrin-alpha (PFT-alpha), on reducing the expression of p53 and PUMA, consequently decreasing the apoptosis of endothelial cells and alleviating cerebral vasospasm.
Male Sprague-Dawley rats weighing 300-350 g were randomly divided into five groups: a control group (sham surgery), a SAH group, a SAH+dimethyl sulfoxide (DMSO) group, a SAH + PFT-alpha (0.2 mg/kg) group and a SAH + PFT-alpha (2.0 mg/kg) group. PFT-alpha was injected intraperitoneally immediately after SAH. Rats were sacrificed 24 hours after SAH. Western blot and immunohistochemical staining were used to detect the levels of p53, PUMA and caspase-3 protein. In addition, mortality and neurological scores were assessed for each group. Statistical significance was assured by analysis of variance performed in one way ANOVA followed by the Tukey test. The neurological and mortality scores were analyzed by Dunn's method and Fisher exact test, respectively.
After SAH, Western blot and immunohistochemical staining showed the levels of p53, PUMA and caspase-3 in the endothelial cells and the numbers of TdT mediated dUTP nick end labelling (TUNEL) positive endothelial cells were all significantly increased in the basilar arteries (P<0.05), but significantly reduced by PFT-alpha (P<0.05). These changes were accompanied by increasing diameters and declining wall thickness of basilar arteries (P<0.05), as well as reduced mortality and neurological deficits of the rats (P<0.05).
PFT-alpha could protect cerebral vessels from development of vasospasm and improve neurological outcome as well as reduce the mortality via suppressing apoptosis induced by p53 in the endothelial cells of cerebral vessels.
蛛网膜下腔出血（SAH）后脑血管痉挛的机制尚不清楚。在此，我们假设由p53及其靶基因——p53上调凋亡调节因子（PUMA）诱导的内皮细胞凋亡在脑血管痉挛的发生中起重要作用。我们还观察了p53抑制剂pifithrin-α（PFT-α）对降低p53和PUMA表达、从而减少内皮细胞凋亡及减轻脑血管痉挛的作用。
将体重300 - 350 g的雄性Sprague-Dawley大鼠随机分为五组：对照组（假手术）、SAH组、SAH + 二甲基亚砜（DMSO）组、SAH + PFT-α（0.2 mg/kg）组和SAH + PFT-α（2.0 mg/kg）组。SAH后立即腹腔注射PFT-α。SAH后24小时处死大鼠。采用蛋白质免疫印迹法和免疫组织化学染色检测p53、PUMA和半胱天冬酶-3蛋白水平。此外，评估每组的死亡率和神经功能评分。通过单因素方差分析及Tukey检验确保统计学意义。神经功能和死亡率评分分别采用Dunn法和Fisher确切概率法进行分析。
SAH后，蛋白质免疫印迹法和免疫组织化学染色显示基底动脉内皮细胞中p53、PUMA和半胱天冬酶-3水平以及末端脱氧核苷酸转移酶介导的dUTP缺口末端标记（TUNEL）阳性内皮细胞数量均显著增加（P<0.05），但PFT-α可使其显著降低（P<0.05）。这些变化伴随着基底动脉直径增加和管壁厚度减小（P<0.05），以及大鼠死亡率降低和神经功能缺损减轻（P<0.05）。
PFT-α可通过抑制脑血管内皮细胞中p53诱导的凋亡来保护脑血管免受血管痉挛的发生，改善神经功能结局并降低死亡率。
