甘草酸苷对幼鼠癫痫持续状态模型的保护机制研究

Protective effect of glycyrrhizin in young rats with status epilepticus

目的研究甘草酸苷对幼鼠癫痫持续状态(SE)模型的保护机制。方法年幼SD大鼠采用腹腔注射氯化锂和匹罗卡品制作SE大鼠模型,分为5组:对照组、SE组、SE+低剂量甘草酸苷组、SE+中剂量甘草酸苷组和SE+高剂量甘草酸苷组,腹腔内注入不同剂量的甘草酸苷(20 mg/kg、40 mg/kg和60 mg/kg)。注射后6 h、12 h和24 h眶内采血,检测各组大鼠血清和(或)海马组织中肿瘤坏死因子(TNF)-α、白细胞介素(IL)-1β、IL-6、高迁移率族蛋白B1(HMGB1)、晚期糖基化终末产物受体(RAGE)、磷酸化细胞外信号调节激酶(p-ERK)、细胞外信号调节激酶(ERK)、磷酸化c-Jun氨基酸末端激酶(p-JNK)和c-Jun氨基末端激酶(JNK)的表达水平;流式细胞仪测定海马组织中细胞内钙离子水平。结果甘草酸苷可以有效减少SE发作后血清和海马组织中TNF-α、IL-1β和IL-6的释放水平,注射甘草酸苷后,与SE组相比,低剂量组、中剂量组和高剂量组血清炎症因子表达水平均有不同程度的下调,差异有统计学意义(P<0.01);高剂量组TNF-α表达水平随时间的推移有下降趋势,但差异无统计学意义(P=0.070);中剂量组注射甘草酸苷后6 h、12 h和24 h,血清IL-1β和IL-6水平随时间推移明显下降(P=0.030,0.012);而在低剂量组,随时间的推移TNF-α和IL-6水平呈升高趋势,但差异无统计学意义(P=0.235,0.229),IL-1β呈明显升高趋势(P=0.034)。同时甘草酸苷还可抑制海马组织细胞内Ca^(2+)的上调(P<0.05);中、高剂量甘草酸苷还能有效下调SE导致的海马组织中HMGB、RAGE、p-ERK/ERK和p-JNK/JNK比值异常升高(P<0.05)。结论甘草酸苷可以通过抑制HMGB1/RAGE、ERK和JNK信号通路对SE大鼠模型发挥保护作用。

Objective To clarify the protective effect of glycyrrhizin in young rats with status epilepticus(SE).Methods Young Sprague-Dawley rats were exposed to lithium chloride and pilocarpine to induce SE in vivo and then were injected with different doses of glycyrrhizin(20,40 and 60 mg/kg).Blood was collected intraorbitally at 6 h,12 h,24 h after injection.The levels of tumor necrosis factor(TNF)-α,interleukin(IL)-1β,IL-6,high mobility group box-1 protein(HMGB 1),receptor for advanced-glycation end products(RAGE),phosphorylated extracellular signal-regulated kinase(p-ERK),extracellular signal-regulated kinase(ERK),phosphorylated c-Jun N-terminal kinase(p-JNK)and c-Jun N-terminal kinases(JNK)in hippocampal tissues and/or in serum were determined.Intracellular Ca^(2+)levels were measured by flow cytometry using Fluo 3-AM dye.Results Glycyrrhizin injection significantly decreased SE-induced elevations of TNF-α,IL-1βand IL-6 levels in serum and hippocampal tissues.After injection of glycyrrhizin,the expression levels of inflammatory factors in the low-dose,medium-dose and high-dose groups were down-regulated to varying degrees compared with the SE group,and the difference was statistically significant(P<0.01).The expression level of TNF-αin the high-dose group showed a downward trend over time,but the difference was not statistically significant(P=0.070).In the medium-dose group,the serum levels of IL-1βand IL-6 decreased significantly over time at 6,12,and 24 hours after injection of glycyrrhizin(P=0.030,0.012).In the low-dose group,the levels of TNF-αand IL-6 showed an increasing trend over time,but the difference was not statistically significant(P=0.235,0.229),and IL-1βshowed a significant increasing trend(P=0.034).Additionally,SE-induced mitochondrial damages and increases in intracellular Ca^(2+),levels were ameliorated by glycyrrhizin.Furthermore,the middle and high dose of glycyrrhizin injection blocked SE-induced increases in HMGB1,RAGE,p-ERK/ERK and p-JNK/JNK ratios(P<0.05).Conclusion Glycyrrhizin protected young rats against SE by inactivating the HMGB1/RAGE,ERK and JNK signaling pathways.