PI3K／Akt信号通路在发育期大鼠癫痫持续状态后海马内HIF一1α表达中的作用

PI3K/Akt pathway involves in regulating of hypoxia-inducible factor-lot expression in the hippocampus of developing rats after status epilepticus

目的探讨发育期大鼠癫痫持续状态后海马内低氧诱导因子-1α（HIF-1α）表达与磷脂酰肌醇．3激酶（P13K1／丝氨酸一苏氨酸蛋白激酶（Akt）信号通路之间的关系。方法21d龄SD大鼠54只按随机数字表法分为生理盐水组（n=24）、模型组（n=24）和渥曼青霉素干预组（n=6），其中模型组腹腔注射戊四氮（PTZ）诱导建立癫痫持续状态模型，生理盐水组腹腔注射等量生理盐水，渥曼青霉素干预组于癫痫持续状态模型建立前30min腹腔注射0．5mg/kg渥曼青霉素。分别于建模后1、4、8、24h处死大鼠取出海马，应用免疫组化染色检测HIF-1α、Akt阳性细胞表达，Westernblottiong检测HIF-1α、Akt及磷酸化Akt（p-Akt）蛋白含量。结果模型组海马内HIF-1α阳性细胞及蛋白于建模后lh即开始表达，4h明显升高，8h达高峰，24h后下降；Akt阳性细胞及蛋白表达在各时间点间无明显变化；P-Akt蛋白于建模后lh即明显升高，4h达高峰，8h开始下降。生理盐水组各时间点HIF—let阳性细胞及蛋白、P-Akt蛋白仅有极少量表达，其中模型组各时间点HIF-let阳性细胞及蛋白表达均明显高于生理盐水组，差异有统计学意义fP〈O．05）；Akt阳性细胞与蛋白表达与生理盐水组比较差异无统计学意义（pO．05）；建模后1、4、8hP．Akt蛋白表达明显高于生理盐水组，差异有统计学意义（氏0．05）。渥曼青霉素干预组HIF-1α、p-Akt蛋白表达于建模后4h有明显下降，与模型组比较差异均有统计学意义（p＜0．05）。结论发育期大鼠癫痫持续状态后可激活P13K／Akt信号通路，并参与调控海马内HIF—1α的表达。

Objective To investigate the correlation between hypoxia-inducible factor-lα （HIF-lαexpression and activation of phosphatidyl inositol 3-kinase and serine/threonine kinase signal pathway in the hippocampus of developing rats after status epilepticus （SE）. Methods Fifty-four SD rats aged 21 days were randomly divided into control group （n=24）, SE group （n=24）, wortmannin treatment group （n=6）; SE rat models of the SE group were induced by intraperitoneal injection of 1% 1, 5-Pentamethylenetetrazole （PTZ）; rats of the control group received injection of normal saline （NS）; for wortmannin treatmnet group, the rats received intraperitoneal injection ofwortmannin 30 minutes before the inducement; the brain tissues were harvested from the rats at 1, 4, 8 and 24 h after the inducement, but only at 4 h in the wortmannin treatment group. The HIF-la and Akt positive cells were detected with immunohistochemistry method. HIF-1α, Akt and p-Akt protein expressions were measured by Western blotting. Results In SE group, the HIF-1α expression began to occur at 1 h, significantly increased at 4 h after inducement, reached the peak level at 8 h, and began to decrease at 24 h; Akt protein positive cells showed no significant difference between each two time points; the p-Akt protein was significantlyincreased at 1 h, reached the peak level at 4 h and began to decrease at 8 h. However, the expression levels of HIF-lot and p-Akt protein in the control group were extremely low at each time point. So, the HIF-Ia expression level in the SE vehicle group was significantly higher than that in the control group （P〈0.05）; the p-Akt protein expression in SE group at 1, 4 and 8 h was significantly higher than that in the control group （/〈0.05）, The changes of Akt protein in the SE group were not time-dependent, and no significant difference was evident when it was compared with that of the control group （P〉0.05）. Using wortmannin, the PI3K/Akt specific inhibitor, HIF-1 a protein expression was significantly decreased when it was compared with the SE vehicle group （/〈0.05）. Conclusion After status epilepticus in the developing rats, the PI3K/Akt signaling pathway is activated and the pathway involves in regulating the HIF- 1expression.