淀粉样β蛋白片段25~35下调大鼠海马PI3K/Akt/p70S6K信号传导通路(英文)

目的探讨阿尔茨海默病(AD)的淀粉样β蛋白(Aβ)的沉积是否损害神经细胞存活的信号传导通路。方法实验分为生理盐水对照组;Aβ25-35组;Aβ25-35+布洛芬组;Aβ25-35+布洛芬+LY294002组;Aβ25-35+LY294002组。大鼠分别灌胃给予布洛芬7.5或15 mg.kg-1,每日1次,连续3周后,左侧脑室内注射Aβ25-35(10 μL, 1 mmol.L-1),之后继续灌胃给予布洛芬1周。PI3K特异性阻断剂LY294002 (5 μL, 4 mmol.L-1)在注射Aβ25 -35前1 h左侧脑室内注射。注射Aβ25-35后1周,取海马CA1区,Western免疫印迹法观察P53,Bax, FasL, Bcl-2, Akt和p70S6K的蛋白表达水平。应用半胱氨酸天冬氨酸蛋白酶(caspase)3活性测定试剂盒分析caspase 3活性变化,RT-PCR方法观察p70s6k mRNA表达水平。结果脑室内注射Aβ25-35可引起大鼠海马CA1区磷酸化Akt/PKB和磷酸化p70S6K表达明显降低,分别从对照组1.32±0.14和0.769±0.028下降到0.69±0.08和0.479±0.032。同时,海马CA1区促凋亡蛋白P53, Bax和FasL表达及caspase 3活性明显增加,抗凋亡蛋白Bcl-2表达明显降低。预先注射LY294002可使caspase 3活性较单独注射Aβ25-35组进一步增加。给Aβ25-35前后连续给予布洛芬4周可明显对抗Aβ25-35引起的上述变化。LY294002可明显减弱布洛芬上调磷酸化Akt/PKB和磷酸化p70S6K表达的作用。结论 Aβ25-35引起抗凋亡通路PI3K/Akt/p70S6K下调可能参与AD的神经元损伤。布洛芬具有较好的对抗作用,这可能与上调PI3K/Akt/p70S6K通路中的一些蛋白有关。

Amyloid β-protein fragment 25-35 down-regulates PI3K/Akt /p70S6K pathway in rat hippocampus in vivo

AIM To investigate whether Aβ deposit in Alzheimer disease(AD) impairs signal transduction pathway responsible for neuronal survival. METHODSThe rats were randomly divided into six groups: control group and Aβ25-35 group, Aβ25-35+ibuprofen groups (7.5 and 15 mg·kg-1, respectively), Aβ25-35+ibuprofen+LY294002 group, and Aβ25-35+LY294002 group. Rats were given ibuprofen (7.5 and 15 mg·kg-1 daily, ig) for 3 weeks prior to and 1 week after icv single dose of Aβ25-35 (10 μL, 1 mmol·L-1). LY294002 was injected icv 1 h before the injection of Aβ25-35. Seven days after Aβ25-35 injection, the hippocampal expressions of P53, Bax, Fas ligand (FasL), Bcl-2 proteins, phospho-Akt/PKB, and phosphorylated 70 ku ribosomal protein S6 kinase (p70S6K) and caspase 3 were determined in the brain tissue preparations from CA1 area with Western blot. The activity of caspase 3 was measured using a caspase 3 colorimetric activity assay kit. RT-PCR was used to show the change of p70s6k mRNA level. RESULTS Aβ25-35 icv injection significantly down-regulated phosphorylated Akt/PKB from 1.32±0.14 to 0.69±0.08 and p70S6K from 0.769±0.028 to 0.479±0.032 in hippocampal CA1 region. These changes were accompanied by increased expressions of the proapoptotic proteins P53, Bax, and FasL and decreased expression of the anti-apoptotic protein Bcl-2 in rat hippocampus. In addition, caspase 3 activity was significantly enhanced in hippocampal CA1 region in Aβ25-35-treated rats compared with control rats. Ibuprofen can reverse these Aβ25-35-induced changes. CONCLUSION Down-regulated anti-apoptotic PI3K/Akt/p70S6K signalingpathway induced by Aβ25-35 in rat hippocampus may contribute to the neuronal damage in AD. Ibuprofen prevents Aβ25-35-induced down-regulation of PI3K/Akt/p70S6K signaling pathway.