叶酸对AD模型小鼠脑内Aβ沉积及BACE1表达影响

Effects of folic acid on Aβ deposition and BACE1 expression in brain of APP/PS1 transgenic mouse

目的探讨叶酸对阿尔茨海默病(AD)小鼠脑内β–淀粉样蛋白(Aβ)沉积影响及机制。方法将6月龄雄性APP/PS1模型小鼠随机分为4组:叶酸缺乏组、叶酸对照组、叶酸低、高剂量组(120、600μg/kg);同月龄野生型小鼠作为阴性对照组。叶酸缺乏组小鼠饲以叶酸缺乏饲料,其他各组饲以普通饲料,干预60 d。采用实时荧光定量PCR法(RT-PCR)检测β位淀粉样前体裂解酶–1(BACE1)mRNA水平;免疫印迹法(WB)检测脑组织BACE1蛋白表达;免疫组化法检测脑组织Aβ表达;酶联免疫吸附法检测脑组织中和含量。结果与叶酸对照组比较,叶酸高剂量组小鼠脑组织中Aβ蛋白表达降低,叶酸缺乏组Aβ蛋白表达升高(P<0.05);与叶酸对照组(0.295±0.034)比较,叶酸高剂量组小鼠脑组织含量(0.209±0.041)降低,叶酸缺乏组含量(0.396±0.043)升高(P<0.05);与叶酸对照组(15.99±1.434)比较,叶酸缺乏组小鼠脑组织BACE1 mRNA水平(21.97±4.396)升高,叶酸低、高剂量组小鼠脑组织中BACE1 mRNA表达水平[分别为(9.932±1.903)、(10.53±2.750)]均降低(均P<0.05);与叶酸对照组(1.371±0.213)比较,叶酸缺乏组小鼠脑组织中BACE1蛋白表达水平(1.655±0.241)升高,叶酸低、高剂量组小鼠脑组织中BACE1蛋白表达水平[分别为(1.003±0.271)、(0.906±0.188)]均降低,差异均有统计学意义(均P<0.05)。结论补充叶酸可减少AD小鼠脑组织中Aβ沉积及含量,其机制可能与叶酸抑制小鼠脑组织中淀粉样蛋白生成途径的分泌酶BACE1表达有关。

Objective To explore the effect and mechanisms of folic acid on deposition of amyloid β-protein （Aβ） in brain tissues of mice with Alzheimer＇s disease. Methods Forty 6-month-old male APP/PS1 mice were randomly assigned into four groups：a folate deficiency group （AD ＋ FA-D） with folic acid deficient feed and daily gavage of water, a folate control group （AD ＋ FA-N） with normal feed and daily gavage with water, and low- and high-dose folate groups （AD ＋ FA-L and AD ＋ FA-H） with normal feed and daily gavage of 120 and 600 μg/kg folic acid; ten 6-month-old male wild type mice with normal feed and daily gavage of water （WT ＋ FA-N） were used as negative controls. The treatments were performed consecutively for 60 days. By the end of the treatments, the mRNA expression of β-site amyloid precursor protein cleaving enzyme-1 （BACE1） in brain tissues was quantified by real-time PCR. The protein expression level of β-site amyloid precursor protein cleaving enzyme-1 （BACE1） in brain tissues was detected with Western blot; the expression of Aβ in brain tissues was measured with immunohistochemistry; the expression levels of Aβ1–40 and Aβ1–42 in brain tissues were measured with enzyme-linked immunosorbent assay （ELISA）.Results The expression of Aβ decreased in AD＋FA-H group and increased in AD＋FA-D group significantly compared to that in AD＋FA-N group （P 〈 0.05）; the content of Aβ1–42 （0.209 ±0.041） decreased in AD ＋ FA-H group and increased （0.396 ±0.043） in AD ＋ FA-D group significantly compared to that （0.295 ±0.034） in AD ＋ FA-N group （P 〈 0.05）; the expression of BACE1 mRNA increased in AD ＋ FA-D group （21.97 ±4.396） but decreased in AD ＋ FA-L and AD ＋ FA-H groups （9.932 ±1.903 and 10.53 ±2.750） significantly compared to that （15.99 ±1.434） in AD ＋ FA-N group （P 〈 0.05 for all）; the expression of BACE1 protein increased in AD ＋ FA-D group （1.655 ±0.241） but decreased in AD ＋ FA-L and AD ＋ FA-H groups （1.003 ±0.271 and 0.906 ±0.188） significantly compared to that （1.371 ±0.213） in AD ＋ FA-N group （P 〈 0.05 for all）.Conclusion The study results show that folic acid supplementation can reduce brain Aβ deposition and Aβ1–42 content and the mechanism of the effects may be related to folic acid-induced inhibition of BACE1 expression associated with Aβ formation in brain tissue of mice.