S14G-humanin对APP/PS1双转基因小鼠的神经保护作用及其机制研究

Neuroprotective role and mechanism of S14G-humanin in amyloid precursor protein/presenolin-1 double transgenic mice

目的探讨S14G-humanin(HNG)对淀粉样前体蛋白(APP)/早老蛋白-1(PS1)双转基因小鼠空间学习记忆能力和海马神经元自噬功能的影响。方法将16只APP/PS1双转基因小鼠按随机数字表法分为模型组、HNG治疗组,每组8只;另取8只C57BL/6J小鼠作为对照组。对照组和模型组小鼠每天腹腔注射0.5 mL双蒸水,HNG治疗组小鼠每天腹腔注射0.5 mL HNG(50μg/kg)。连续注射4周后采用Morris水迷宫实验检测各组小鼠的空间学习记忆能力,采用Western blotting、RT-PCR实验分别检测各组小鼠海马泛素化激酶1(ULK1)、P62、兔抗微管相关蛋白1轻链3(LC3)Ⅱ、LC3Ⅰ及组织蛋白酶D蛋白和mRNA的表达,采用免疫组化染色检测各组小鼠海马神经元β淀粉样蛋白(Aβ)的沉积。结果Morris水迷宫实验显示对照组、HNG治疗组、模型组小鼠的逃避潜伏期依次延长,穿梭原平台所在象限次数依次减少,差异均有统计学意义(P<0.05)。Western blotting、RT-PCR检测显示对照组、HNG治疗组、模型组小鼠海马ULK1蛋白和mRNA的表达依次减少,P62蛋白和mRNA的表达依次增加、LC3Ⅱ蛋白/LC3Ⅰ蛋白和LC3ⅡmRNA/LC3ⅠmRNA值依次增加、组织蛋白酶D蛋白和mRNA的表达依次减少,差异均有统计学意义(P<0.05)。免疫组化染色检测显示对照组、HNG治疗组、模型组小鼠海马神经元Aβ阳性表达依次增加,差异均有统计学意义(P<0.05)。结论HNG可提高APP/PS1双转基因小鼠海马神经元的自噬活性,减少脑内Aβ的沉积,从而改善小鼠的空间学习记忆能力。

Objective To investigate the effect of S14G-humanin(HNG)on spatial learning and memory abilities and hippocampal neuron autophagy in amyloid precursor protein(APP)/presenolin-1(PS1)double-transgenic mice.Methods The APP/PSl transgenic mice were randomly divided into model group and HNG treatment group(n=8).Another 8 C57BL/6J mice were chosen as controls;0.5 mL double-steamed water was intraperitoneally injected into mice in the control group and model group every day,and 0.5 mL of HNG(50μg/kg)was intraperitoneally injected into mice in the HNG treatment group every day;after 4 weeks of continuous injection,Morris water maze test was used to test the spatial learning and memory abilities of mice in each group.Western blotting and reverse transcription-PCR were used to detect the protein and mRNA expressions of ubiquitin like kinase1(ULK1),P62,anti-microtubule associated protein 1 light chain 3(LC3)II/LC3 I and cathepsin D in the hippocampus.Immunohistochemical staining was used to detect the deposition of amyloidβprotein(Aβ)in the hippocampus of mice in each group.Results Morris water maze test showed that the escape latency of mice in the control group,HNG treatment group and model group was statistically prolonged in turn,and the number of times traversing through the quadrant of the original platform was significantly smaller in turn(P<0.05).Western blotting and reverse transcription-PCR showed that the ULK1 protein and mRNA expressions in the hippocampus decreased successively,P62 protein and mRNA expressions increased successively,LC3 II protein/LC3 I protein and LC3 II mRNA/LC3 I mRNA ratio increased successively,and cathepsin D protein and mRNA expressions decreased successively in the control group,HNG treatment group and model group,with significant differences(P<0.05).Immunohistochemical staining showed that the Aβpositive expressions in the hippocampal neurons of the control group,HNG treatment group and model group increased successively,with statistical differences(P<0.05).Conclusion HNG treatment can improve the spatial learning and memory abilities,which may be attributed to ameliorate autophagic network dysfunction and reduce Aβplaques in the hippocampi of APP/PSl transgenic mice.