甘草酸对癫痫大鼠海马神经元损伤的保护作用及其机制

Protective effect and mechanism of glycyrrhizic acid on the hippocampal neuron injury in epileptic rat models

目的探讨甘草酸对癫痫大鼠海马神经元损伤的保护作用及相关机制。方法采用氯化锂-匹鲁卡品点燃制作大鼠癫痫模型,造模成功后大鼠随机分为癫痫组及甘草酸组,另外将不做任何处理的大鼠作为正常对照组,每组12只。利用Nissl法和TUNEL法检测大鼠海马神经元损伤及凋亡情况;JC-1法检测大鼠海马神经元线粒体膜电位;比色法检测天冬氨酸蛋白水解酶3(caspase 3)和caspase 9的活性;Western blot法检测大鼠海马组织中cleaved-caspase 3、9,B淋巴细胞瘤-2(Bcl-2),Bcl-2相关X蛋白(Bax),细胞色素C(CytC),凋亡蛋白酶激活因子(Apaf-1)蛋白表达。结果与正常对照组相比,癫痫组神经元细胞数减少(P<0.01),TUNEL阳性细胞数增加(P<0.01),线粒体膜电位降低(P<0.01),caspase 3、9活性提高(P<0.01),Bcl-2及线粒体中CytC表达量下调(P<0.01),Bax及细胞质中CytC、Apaf-1表达量上调(P<0.01)。与癫痫组比较,甘草酸组神经元细胞数增加(P<0.01),TUNEL阳性细胞数减少(P<0.01),线粒体膜电位提高(P<0.01),caspase 3、9活性降低(P<0.01),Bcl-2及线粒体中CytC表达量上调(P<0.01),Bax及细胞质中CytC、Apaf-1表达量下调(P<0.01)。结论通过阻断线粒体途径,甘草酸能够抑制癫痫大鼠的海马神经元损伤。

Objective To explore the protective effect and mechanism of glycyrrhizic acid on hippocampal neuroninjury in epileptic rat models. Methods The epileptic rat models were established by lithium and pilocarpine kindling.The successful models were randomly divided into epilepsy group and glycyrrhizic acid group. In addition, the rats withoutany treatment were used as the normal control group, with 12 rats in each group. Hippocampal neuron injury and apoptosiswere detected by Nissl and TUNEL staining. Mitochondrial membrane potential in the hippocampal neurons of rats wasdetected by JC-1 method. The activity of aspartic acid protein hydrolase 3 （ caspase 3） and caspase 9 was detected bycolorimetric assay. The expressions of cleaved-caspase 3, 9, B lymphocyte tumor-2 （ Bcl-2）, Bcl-2 related X protein（Bax）, cytochrome C （CytC） and apoptotic protease-activating factor （Apaf-1） protein in the hippocampal tissues of rats were detected by western blot assay. Results Compared with the normal group, the number of neurons was reduced （P 〈0.01）, the number of TUNEL-positive cells was increased （P 〈 0.01）, mitochondrial membrane potential was decreased（P 〈 0.01）, caspase 3 and 9 activity was increased （P 〈 0.01）, the expressions of Bcl-2 and mitochondrial CytC weredown-regulated （P 〈 0.01）, the expressions of Bax, cytoplasm CytC and Apaf-1 were up-regulated （P 〈 0.01） in theepilepsy group. Compared with the epilepsy group, the number of neurons was increased （ P 〈 0.01）, the number ofTUNEL-positive cells was reduced （P 〈 0.01）, mitochondrial membrane potential was increased （P 〈 0.01）, caspase 3and 9 activity was decreased （ P 〈 0.01）, the expressions of Bcl-2 and mitochondrial CytC were up-regulated （ P 〈0.01）, the expressions of Bax, cytoplasm CytC and Apaf-1 were down-regulated （ P 〈 0.01） in the glycyrrhizic acidgroup. Conclusions These results suggest that glycyrrhizic acid can inhibit the hippocampal neuron injury in epileptic ratsby blocking the mitochondrial pathway.