神经生长因子预处理对沙土鼠全脑缺血/再灌注损伤脑神经细胞凋亡及Bcl-2和Bax蛋白表达的影响

目的探讨神经生长因子(NGF)预处理对沙土鼠全脑缺血/再灌注(I/R)损伤的脑保护作用的可能机制及最佳给药时间窗。方法采用夹闭沙土鼠双侧颈总动脉造成全脑I/R损伤模型。采用NGF侧脑室注射法进行预处理。沙土鼠30只随机分为5组,每组6只:假手术组(A组)、I/R损伤组(B组)、NGF预处理12、24和48h组(C、D、E组),除A组外各组分别于脑缺血20min、再灌注72h后处死取标本。用末端脱氧核苷酸转移酶介导的dUTP缺口末端标记法(TUNEL)检测沙土鼠全脑I/R损伤后脑皮质及海马CA1区凋亡神经细胞,用免疫组化法检测凋亡相关调控基因Bcl-2、Bax蛋白的表达。结果与B组比较,NGF预处理各组可显著减少沙土鼠全脑I/R损伤后脑皮质及海马CA1区神经细胞凋亡数目(P均<0.05),诱导Bcl-2蛋白及抑制Bax蛋白的表达(P均<0.05),其中以NGF预处理48h时脑皮质及海马CA1区细胞凋亡指数和Bax蛋白表达的阳性细胞指数最低,Bcl-2蛋白表达的阳性细胞指数最高。结论NGF预处理能明显减轻沙土鼠全脑I/R损伤引起的神经细胞凋亡,而以NGF预处理48h对脑保护效果最好;其抑制神经细胞凋亡的机制可能是通过调节凋亡相关调控基因Bcl-2及Bax的不同表达来发挥作用。

Effects of nerve growth factor pretreatment on apoptosis of neurons and expression of Bcl-2, Bax protein in brain tissue following cerebral ischemia/reperfusion injury in gerbils

Objective To study the effect of nerve growth factor (NGF) pretreatment on apoptosis of neurons and the expression of Bcl-2 and Bax protein in cerebral cortex and hippocampus CA1 zone following global cerebral ischemia/reperfusion (I/R) injury in gerbils and to explore the mechanism of protection and the best time window of NGF pretreatment. Methods Global cerebral I/R injury model was induced by occlusion of bilateral carotid arteries. NGF was injected into the lateral ventricle. Thirty gerbils were randomly divided into five groups, with six animals in each: sham operation group (A group), I/R injury group (B group), NGF pretreatment 12, 24 and 48 hours groups (C, D and E group). Gerbils in all groups were sacrificed after being subjected to 20 minutes of cerebral ischemia followed by 72 hours reperfusion, except A group. Neural apoptosis was identified by terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling (TUNEL), and immunohistochemistry was used to detect the expression of Bcl-2 and Bax protein in cerebral cortex and hippocampus CA1 zone. Results Compared with B group, the number of apoptotic neurons and the expression of Bax positive cells in NGF pretreatment groups were decreased significantly (all P<0.05), while the expression of Bcl-2 positive cells was increased significantly (all P<0.05). The apoptotic rate in cerebral cortex and hippocampus CA1 zone and expression rate of Bax protein positive cells were the lowest, but the expression rate of Bcl-2 protein positive cells was the highest at 48 hours. Conclusion NGF pretreatment can significantly decrease the neuronal apoptosis of the cerebral I/R injury in gerbils, and the best time window of NGF pretreatment is 48 hours. The mechanism of protection may be related to induction of Bcl-2 protein expression and inhibition of Bax protein expression by NGF pretreatment, thereby preventing neuronal apoptosis.