APP17肽对D-半乳糖老化小鼠海马神经元IRS和PI3K表达的影响

Effect of APP 17-mer peptide on IRS-1 and IRS-2 of D-galactose induced aging murine brain

目的通过对胰岛素受体底物-1（IRS-1）、胰岛素受体底物-2（IRS-2）、磷脂酰肌醇-3激酶（PI3K）在海马表达的观察，研究APP17肽对D-半乳糖老化小鼠海马神经元信号转导途径上游步骤的影响。方法小鼠随机分为3组，每组22只：正常对照组（C组）、D-半乳糖模型组（D组）、APP17肽治疗组（D＋P组）。D、D＋P 2组每日皮下注射半乳糖100mg·kg^-1。D＋P组每周一、二、五皮下给药APP17肽0．35μg／只。4个月后，小鼠灌注固定，行脑冰冻切片，用PI3K、IRS-1、IRS-2抗体进行免疫组化染色。Western blot应用IRS—1抗体染色。结果免疫组织化学染色方法检测海马神经元胰岛素受体底物-1、胰岛素受体底物-2、PI3K的表达，结果显示3种抗体C组深染的阳性细胞排列整齐。染色深，突起深染，可见2—3级分支，细胞计数与D组差异有显著性；D组海马阳性反应细胞着色浅，少有突起，细胞数目少，分别为正常对照的67．5％、63．68％和48．6％。APP17肽治疗组（D＋P组）染色结果与C组相似，阳性细胞数目接近C组，与半乳糖老化组比较差异均有显著性，P〈0．05，但阳性细胞突起少于C组，排列亦不如C组整齐。Western blot结果亦显示D组海马表达IRS-1减少，应用17肽后IRS-1表达上调，但不如C组数值高。结论D-半乳糖老化小鼠海马神经元IRS—1、IRS-2、PI3K的表达明显下降；APP17肽能上调D-半乳糖老化小鼠海马IRS—1、IRS-2和PI3K的表达。

Aim In order to explore the impact of APP17-mer peptide on the hippoeampus of D-galaetose induced aging of murine （DGAM）, authors examined expressions of insulin receptor substrate-1 （IRS-1）,insulin receptor substrate-2 （ IRS-2）, and phosphatidyl inoditide-3′ OH kinase （PI3K） in the hippocampus. Methods Mice were randomized into three groups： control （ group C ） , D-galactose （ group D ） , and APP17-mer peptide treatment groups （ group D ＋ P）. Each had 22 subjects. In the D and D ＋ P groups, a dose of 100 mg · kg^-1 of D-galactose was injected subcutaneously daily to each subject. In addition, on Monday, Wednesday, Friday, each mouse in the D ＋ P group was given a dose of 0. 35μg APP17- mer peptide as the treatment. Sixteen weeks later, mice＇s brains were cryostat sectioned and stained immunohis-tochemically with IRS-1, IRS-2 and PI3K antibodies. The results of the immunohistochemical stain were then recorded. Results In the DGAM group, the number of neurons stained positively by IRS-1,2 and PI3K antibodies was significantly lower than their counterparts in the control group （P 〈 0. 05 ）. In terms of the high power magnification, cells were stained lightly in the D group as compared to the C group, where cells were stained darkly. In the control group, numerous primary level dendrites and some secondary to tertiary level branches were clearly observed. Results of the APP17- mer peptide treatment group and the control group were the similar. However, neurons stained in the APP17- mer peptide treatment group were significantly different from those of the DGAM group （ P 〈 0. 05 ）. Conclusion Amounts of expressions of insulin receptor substrate-1, insulin receptor substrate-2, and phosphatidyl inoditide-3′ OH kinase decrease dramatically in the hippocampal neurons of aging mice. A treatment with APP17-mer peptide will increase expressions of IRS-1, IRS-2, and PI3K.