临床儿科杂志 ›› 2016, Vol. 34 ›› Issue (7): 542-.doi: 10.3969 j.issn.1000-3606.2016.07.016

• 实验研究 • 上一篇    下一篇

癫痫持续状态大鼠海马内Notch 通路对HIF-1α调控位点的研究

庄亚飞1, 张林1, 袁宝强1, 戴园园2, 李蕊2   

  1. 徐州医学院附属医院 1. 儿童神经与康复科,2. 儿科(江苏徐州 221002)
  • 收稿日期:2016-07-15 出版日期:2016-07-15 发布日期:2016-07-15
  • 通讯作者: 袁宝强 E-mail:yuanbqiang@hotmail.com

Exploration of the regulation site of HIF-1α in Notch signaling pathway in the hippocampus of rats with status epilepticus

ZHUANG Yafei1, ZHANG Lin1, YUAN Baoqiang1, DAI Yuanyuan2, LI Rui2   

  1. 1. Department of Child Neurology and Rehabilitation, 2. Department of Pediatrics, Xuzhou Medical College Affiliated Hospital, Xuzhou 221006, Jiangsu, China
  • Received:2016-07-15 Online:2016-07-15 Published:2016-07-15

摘要: 目的 研究低氧诱导因子-1α(HIF-1α) 及Notch 信号通路下游基因HES1在发育期癫痫持续状态(SE) 大鼠海马内的表达变化,探寻Notch信号通路对于HIF-1α的调控位点。方法 176只21日龄SD大鼠随机分为生理盐水对照组(NS组)、戊四氮(PTZ) 致SE模型组(PTZ 组)、Notch 信号通路特异性抑制剂(DAPT) 干预组(DAPT组)。PTZ组大鼠腹腔注射PTZ溶液制作SE模型,NS组注射等剂量的生理盐水作为对照,予安定腹腔注射终止SE发作。造模成功后分别于0.5、1、2、4、8 h 断头取脑分离双侧海马组织,采用RT-PCR法检测HES1、HIF-1α mRNA的表达;于2、4、8、12、24 h 断头取海马,采用Western blot 法检测蛋白的表达。DAPT组在开始造模前30 min 腹腔注射DAPT溶液,分别于造模成功后2 h、8 h 断头取海马组织,采用RT-PCR法检测2 h 时HES1、HIF-1α mRNA的表达,用Western blot 法检测8 h 时蛋白的表达。结果 SE后的各个时间点,大鼠海马内HES1、HIF-1α mRNA和蛋白表达是增高的,与同一时间的NS组相比差异有统计学意义(P < 0.05);与同一时间点的PTZ组比较,DAPT组的HES1、HIF-1α mRNA和蛋白表达明显降低,差异有统计学意义(P < 0.05)。结论 在发育期大鼠SE发作后的海马内,HES1基因可能是Notch信号通路中对HIF-1α表达的调控位点。

Abstract: Objective To study the expression of hypoxia inducible factor-1 (HIF-1) and Notch signaling pathway downstream gene HES1 in the hippocampus of pubertal rats with status epilepsy (SE), and to explore the regulation site of HIF-1α in Notch signaling pathway. Methods One hundred and seventy-six 21-day-old SD rats were randomly divided into control group (NS group), pentetrazole (PTZ)-induced SE group (PTZ group), and Notch signaling pathway specific inhibitor (DAPT) intervention group (DAPT group). In PTZ group PTZ was intraperitoneally injected to build SE model and in NS group normal saline was injected as control. The intraperitoneal injection of diazepam was used to terminate SE seizures. After successful modeling, the bilateral hippocampuses were isolated after the rats were sacrificed at 0.5, 1, 2, 4 and 8 h, respectively, and RT-PCR was performed to detect the mRNA expression of HES1 and HIF-1α. The Western Blot was performed to detect protein expression in hippocampuses which were collected at 2, 4, 8, 12, and 24 h after successful modeling. DAPT group received intraperitoneal injection of DAPT 30 min before the start of molding, then the hippocampuses were isolated at 2 and 8 h after successful modeling. RT-PCR was performed to detect the mRNA expression of HES1 and HIF-1α at 2 h, and Western blot was performed to detect protein expression at 8 h. Results At each time point after SE, the expression of mRNA of HES1 and HIF-1α and the expression of protein were higher than the same time point of NS group (P < 0.05). Compared with the same time point of PTZ group, the mRNA expression of HES1 and HIF-1α and the expression of protein of DAPT group were obviously reduced (P < 0.05). Conclusion HES1 gene may be the regulatory site of HIF-1 expression in Notch signaling pathway in the hippocampus of puberty rats with SE.