端粒缩短和p53-p21相关通路蛋白在苯并[a]芘致小鼠学习记忆损伤中的作用  

Role of telomere shortening and p53-p21-related pathway protein in learning and memory impairment induced by benzo[a]pyrene in mice

作　　者：张瑾琰 魏建宏[1] ZHANG Jin-yan;WEI Jian-hong(Department of Anatomy,Fenyang College of Shanxi Medical University,Shanxi Fenyang 032200,China)

机构地区：[1]山西医科大学汾阳学院解剖学教研室,山西汾阳032200

出　　处：《解剖学报》2020年第2期178-183,共6页Acta Anatomica Sinica

基　　金：山西省自然科学基金(201801D121264)。

摘　　要：目的通过观察不同时间和不同剂量染毒小鼠端粒长度和p53、p21蛋白表达水平的改变,探讨端粒缩短和p53-p21相关通路蛋白在苯并[a]芘致学习记忆损伤中的作用。方法取健康雄性C57小鼠90只,每批30只,分别染毒1个月、2个月和3个月,每批均设空白对照组、溶剂对照组、1.00 mg/kg、2.50 mg/kg和6.25 mg/kg苯并[a]芘染毒组,每组6只,隔日腹腔注射染毒。测试小鼠学习记忆能力,ELISA检测小鼠血浆中B[a]P-7,8-二氢二醇9,10-环氧化物-DNA加合物(BPDE-DNA)含量,Western blotting检测小鼠海马p53-p21相关通路蛋白及神经干细胞增殖、分化相关蛋白巢蛋白(Nestin)、βⅢ-微管蛋白(βⅢ-tubulin)表达水平,Real-time PCR方法检测端粒相对长度。结果各染毒组小鼠逃避潜伏期明显延长,穿越平台次数明显减少;非染毒组中未检测到BPDE-DNA,染毒组小鼠血浆中BPDE-DNA含量呈上升趋势(P<0.05);染毒1月时,染毒组与非染毒组小鼠外周血相对端粒长度无明显区别;染毒2月和3月组小鼠,染毒组与非染毒组外周血相对端粒长度相比明显缩短(P<0.05),各染毒组小鼠血浆中BPDE-DNA含量与相对端粒长度成负相关;与非染毒组相比,染毒组p53、p21蛋白表达水平随染毒剂量的加大而增加(P<0.05);1.0 mg/kg剂量染毒1月时,Nestin表达水平未见明显增高,余各染毒组Nestin表达水平均增高(P<0.05);染毒6.25 mg/kg剂量3月时,Nestin表达水平较1.0 mg/kg和2.5 mg/kg组有所降低(P<0.05);染毒1月时,染毒组与非染毒组βⅢ-tubulin表达量无明显区别,染毒2月和3月时,染毒组βⅢ-tubulin表达量下降(P<0.05)。结论苯并[a]芘腹腔注射染毒可诱导BPDE-DNA形成。造成DNA损伤和端粒缩短,从而引起p53-p21相关信号通路激活。端粒缩短和p53-p21信号通路活化介导的DNA损伤应答反应可能是抑制海马神经元增殖和分化的重要机制。Objective To observe the changes of telomere length and p53 and p21 protein expression levels in mice exposed to different time and doses,and to explore the role of related pathways protein in benzo[a]pyrene in learning and memory impairment.Methods Ninety healthy male C57 mice,30 in each batch,were exposed for 1 month,2 months and 3 months respectively.Each batch was set with blank control group,solvent control group,1.00 mg/kg,2.50 mg/kg and 6.25 mg/kg benzo[a]pyrene-treated groups,6 in each group,inoculated intraperitoneally every other day.Learning and memory ability was detected by Morris water maze test,ELISA was used to detect the content of benzo[a]pyrene-7,8-dihydrodiol-9,10-epoxide-DNA(BPDE-DNA)adduct in mice plasma.There was no significant difference in the relative telomere length of the mice at 1 mouth of each group.The length in mice after treated for 2 and 3 months were significantly shorter than that of the non-infected group(P<0.05).And the plasma BPDE-DNA adduct content in each exposed group was negatively correlated with the relative telomere length.Western blotting was used to detect the expression of p53-p21 related pathway protein and neural stem cell proliferation and differentiation related proteins Nestin andβⅢ-tubulin in mouse hippocampus.Relative telomere length was detected by Real-time PCR.Results The escape latency of mice in each exposed group was significantly prolonged,and the times of crossing platforms was significantly reduced.BPDE-DNA adduct was not detected in the non-dose group and in the exposed group showed an upward trend(P<0.05).Compared with the non-dose group,the expression levels of p53 and p21 in the exposed group increase with the dose of the drug(P<0.05).When the dose was 1.0 mg/kg,the expression level of Nestin did not significantly increase.The expression level of Nestin increased in the remaining exposure groups(P<0.05),and the expression level of Nestin was lower than that in the 1.0 mg/kg and 2.5 mg/kg groups at the dose of 6.25 mg/kg for 3 months(P<0.05).A

关 键 词：苯并[A]芘 端粒 海马 学习 记忆 免疫印迹法 小鼠 

分 类 号：R322[医药卫生—人体解剖和组织胚胎学]