黄芩素衍生物对大鼠急性肝性脑病模型的干预

Study on the intervention effect of baicalein derivative on acute hepatic encephalopathy in rats

关键词:  肝性脑病  黄芩素衍生物  血氨  神经递质  PI3K/AKT信号通路

Key words:  hepatic encephalopathy  baicalein derivative  blood ammonia  neurotransmitter  PI3K/AKT signaling pathway

摘要:

[目的] 通过构建硫代乙酰胺（TAA）诱导的大鼠急性肝性脑病模型，研究黄芩素衍生物二磺酸基黄芩素（DB）对急性肝性脑病模型大鼠的保护作用。[方法] 选]择Wistar大鼠为研究对象，将其随机分为对照组、模型组、DB低、中、高剂量（6.25、25、100 mg/kg）组和阳性药（乳果糖，6 g/kg）组。各组大鼠灌胃给予相应剂量药物，对照组与模型组给予等量生理盐水。实验过程中通过Morris水迷宫实验检测大鼠行为学变化。DB干预7 d后处死大鼠，收集结肠内容物、肝脏及脑组织。通过快速血氨测定仪检测大鼠血氨含量；采用pH计法测定结肠内容物pH；通过全自动生化仪检测血清中天冬氨酸氨基转移酶（AST）、丙氨酸氨基转移酶（ALT）、碱性磷酸酶（ALP）、总胆红素（TBiL）的浓度；通过苏木精-伊红（HE）染色观察肝组织病理变化；采用酶联免疫吸附法（ELISA）测定血清中肿瘤坏死因子-α（TNF-α）和白介素-6（IL-6）水平，前额叶皮质中γ-氨基丁酸（GABA）和谷氨酸（Glu）含量；通过免疫印迹法检测前额叶皮质中磷脂酰肌醇3激酶（PI3K）和磷酸化蛋白激酶B（p-AKT）的蛋白表达。[结果] 与模型组相比，DB低、中、高剂量组可降低肝性脑病大鼠死亡率；提高大鼠学习记忆能力；改善大鼠肝组织病理学损伤和纤维化；DB中、高剂量组可显著降低血氨和结肠内容物pH（P<0.01）；显著降低大鼠血清ALT、AST、ALP、TBiL、TNF-α、IL-6水平（P<0.05或P<0.01）；显著降低前额叶皮质中GABA含量（P<0.01），升高前额叶皮质中Glu的含量。DB中剂量组可显著抑制前额叶皮质中PI3K、p-AKT蛋白表达（P<0.01）。[结论] DB能够改善TAA诱导的急性肝性脑病大鼠的学习记忆能力和肝功能，降低体内氨水平，抑制炎症因子表达，调节前额叶皮质中神经递质（GABA和Glu）水平，其作用机制可能与调控PI3K/AKT信号通路有关。

Abstract:

[Objective] The protective effect of baicalein derivative (disulfonate baicalein,DB) on acute hepatic encephalopathy by constructing a thioacetamide (TAA) induced acute hepatic encephalopathy rats was studied. [Methods] Wistar rats,as the research objects,were randomly divided into control group,model group,DB groups (6.25,25,100 mg/kg) and positive drug group (lactulose,6 g/kg). The rats in each group were intragastric the corresponding dose of drugs,while the rats in the control group and the model group were given the same amount of normal saline. During this period,the behavioral changes of rats were detected by the morris water maze test. The rats were sacrificed after 7 days of administration and colon contents,hepatic and brain tissue were collected. The content of blood ammonia was detected by rapid ammonia analyzer. The pH of colon contents was measured by pH meter. The concentrations of aspartate aminotransferase (AST),alanine aminotransferase (ALT),alkaline phosphatase (ALP),and total bilirubin (TBiL) in serum were determined by automatic biochemical analyzer. HE staining was performed for hepatic tissues. The levels of tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) in serum and the contents of gamma aminobutyric acid (GABA) and glutamic acid (Glu) in prefrontal cortex were measured by enzyme-linked immunosorbent assay (ELISA). The expressions of phosphatidylinositol 3 kinase (PI3K) and phosphorylated protein kinase B (p-AKT) in prefrontal cortex were detected by Western bloting. [Results] Compared with the model group,DB low,middle and high dose groups reduced the mortality of rats with hepatic encephalopathy,enhanced rat's learning and memory ability,and improved the pathological damage and fibrosis of liver tissue. The middle and high dose groups significantly reduced blood ammonia and pH of colon contents (P<0.01),also decreased the serum levels of ALT,AST,ALP,TBiL,TNF-α and IL-6 (P<0.05, 0.01),decreased GABA content in prefrontal cortex (P<0.01),increased Glu content in prefrontal cortex. The middle dose group could decrease the protein expression levels of PI3K and p-AKT in prefrontal cortex (P<0.01). [Conclusion] DB can improve the learning and memory ability and liver function of rats with TAA induced acute hepatic encephalopathy,reduce ammonia levels,inhibit the expression of inflammatory factors,and regulate neurotransmitters (GABA and Glu) levels in prefrontal cortex. Its mechanism may be related to the regulation of PI3K/AKT signaling pathway.