氨和谷氨酸培养星形胶质细胞AQP4表达及干预其表达的实验研究
发布时间：2019-02-16 11:36
【摘要】： 目的:观察氨、谷氨酸培养的SD乳鼠脑星形胶质(astrocytes,AS)细胞水通道蛋白4(aquaporin-4,AQP4)的表达,探讨其和肝性脑病脑水肿发生的关系。以及分别应用地塞米松、二甲基亚砜(Dimethyl sulfoxide,DMSO)和丝裂原活化蛋白激酶(mitogen activated protein kinases,MAPKs)信号转导通路抑制剂U0126观察对其表达的影响,寻求预防和治疗肝性脑病脑水肿的药物。方法:通过氨和谷氨酸培养SD乳鼠脑星形胶质细胞,用倒置相差显微镜和电镜观察细胞形态及超微结构、Fluo-3/Am荧光探针观察细胞内钙离子变化、自动生化分析仪测定LDH漏出量、采用免疫组织化学及逆转录聚合酶链式反应检测AQP4在蛋白和基因水平的表达及其变化,以及分别加用U0126、地塞米松、二甲基亚砜后观察上述指标的变化。结果:氨和谷氨酸可致培养的星形胶质细胞死亡,星形胶质细胞LDH漏出量增加,细胞内钙离子浓度增高,AQP4 mRNA和AQP4蛋白的表达增加。U0126、地塞米松、DMSO可使氨和谷氨酸培养星形胶质细胞LDH漏出明显减少,抑制细胞内钙离子的增加,同时使AQP4 mRNA和AQP4蛋白的表达减少。结论:1、氨和谷氨酸可致培养的星形胶质细胞AQP4表达增加,且随着氨浓度的升高AQP4表达也相应增加。2、U0126、地塞米松、DMSO可以减少星形胶质细胞AQP4的表达,具有细胞保护作用。
[Abstract]:Aim: to investigate the expression of aquaporin 4 (aquaporin-4,AQP4) in astrocytes (astrocytes,AS) of SD rats cultured with ammonia and glutamate, and to explore the relationship between the expression of aquaporin 4 (aquaporin-4,AQP4) and the occurrence of cerebral edema in hepatic encephalopathy. The effects of Dexamethasone, dimethyl sulfoxide (Dimethyl sulfoxide,DMSO) and mitogen activated protein kinase (mitogen activated protein kinases,MAPKs) signal transduction pathway inhibitor U0126 on its expression were observed. To seek drugs for the prevention and treatment of cerebral edema in hepatic encephalopathy. Methods: the astrocytes of SD were cultured with ammonia and glutamate. The morphology and ultrastructure of the cells were observed by inverted phase contrast microscope and electron microscope. The changes of intracellular calcium were observed by Fluo-3/Am fluorescence probe. The leakage of LDH was measured by automatic biochemical analyzer, the expression and changes of AQP4 at protein and gene levels were detected by immunohistochemistry and reverse transcriptase polymerase chain reaction, and U0126 and dexamethasone were added respectively. The changes of above indexes were observed after dimethyl sulfoxide. Results: ammonia and glutamate could induce the death of cultured astrocytes, increase the amount of LDH leakage, increase the intracellular calcium concentration, and increase the expression of AQP4 mRNA and AQP4 protein, U0126, dexamethasone, U0126, dexamethasone, DMSO could significantly reduce the leakage of LDH from astrocytes cultured with ammonia and glutamate, inhibit the increase of intracellular calcium ion, and decrease the expression of AQP4 mRNA and AQP4 protein at the same time. Conclusion: 1. Ammonia and glutamate can induce the increase of AQP4 expression in cultured astrocytes, and the expression of AQP4 increases with the increase of ammonia concentration. 2Dexamethasone and DMSO can reduce the expression of AQP4 in astrocytes. It has cytoprotective effect.
【学位授予单位】：昆明医学院
【学位级别】：硕士
【学位授予年份】：2010
【分类号】：R329
本文编号：2424405
[Abstract]:Aim: to investigate the expression of aquaporin 4 (aquaporin-4,AQP4) in astrocytes (astrocytes,AS) of SD rats cultured with ammonia and glutamate, and to explore the relationship between the expression of aquaporin 4 (aquaporin-4,AQP4) and the occurrence of cerebral edema in hepatic encephalopathy. The effects of Dexamethasone, dimethyl sulfoxide (Dimethyl sulfoxide,DMSO) and mitogen activated protein kinase (mitogen activated protein kinases,MAPKs) signal transduction pathway inhibitor U0126 on its expression were observed. To seek drugs for the prevention and treatment of cerebral edema in hepatic encephalopathy. Methods: the astrocytes of SD were cultured with ammonia and glutamate. The morphology and ultrastructure of the cells were observed by inverted phase contrast microscope and electron microscope. The changes of intracellular calcium were observed by Fluo-3/Am fluorescence probe. The leakage of LDH was measured by automatic biochemical analyzer, the expression and changes of AQP4 at protein and gene levels were detected by immunohistochemistry and reverse transcriptase polymerase chain reaction, and U0126 and dexamethasone were added respectively. The changes of above indexes were observed after dimethyl sulfoxide. Results: ammonia and glutamate could induce the death of cultured astrocytes, increase the amount of LDH leakage, increase the intracellular calcium concentration, and increase the expression of AQP4 mRNA and AQP4 protein, U0126, dexamethasone, U0126, dexamethasone, DMSO could significantly reduce the leakage of LDH from astrocytes cultured with ammonia and glutamate, inhibit the increase of intracellular calcium ion, and decrease the expression of AQP4 mRNA and AQP4 protein at the same time. Conclusion: 1. Ammonia and glutamate can induce the increase of AQP4 expression in cultured astrocytes, and the expression of AQP4 increases with the increase of ammonia concentration. 2Dexamethasone and DMSO can reduce the expression of AQP4 in astrocytes. It has cytoprotective effect.
【学位授予单位】：昆明医学院
【学位级别】：硕士
【学位授予年份】：2010
【分类号】：R329
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