长期铅暴露导致大鼠神经退行性改变研究

发布时间：2018-04-23 11:41

本文选题：铅暴露 + 神经退行性病变　；参考：《南昌大学》2013年硕士论文

【摘要】：目的：研究表明铅不仅具有神经发育毒性,而且可能是神经退行性疾病的一个重要的危险因素,本文旨在探讨长期铅暴露致大鼠神经退行性改变及其相关的基因蛋白表达研究。方法：将成年雌性Sprague-Dawley (SD)大鼠随机分为3个组：空白组、低铅暴露组(LLG)和高铅暴露组(HLG),分别给予0,800,1500mg/L乙酸铅水溶液,染毒10天后与自由饮用去离子水的雄性大鼠合笼。在观察到阴栓后21天,仔鼠降生,选择雄性仔鼠分别饲养至断乳(Postnatal week 3, PNW3)、中年(Postnatal week 41, PNW41)、老年(Postnatal week 70, PNW70)。雄性仔鼠在三个组分别给予0,300,900mg/L乙酸铅水溶液。染毒末,通过活体磁共振成像(Magnetic resonance imaging, MRI)技术测量老年大鼠大脑及海马容积；用离体电感耦合等离子体原子发射光谱(Inductively coupled plasma atomic emission spectrum-etry, ICP-AES)方法测定大鼠三个年龄段血铅和皮层、海马铅含量；硫堇染色观察三个年龄段皮层、海马神经元密度情况；采用透射电子显微镜观察老年大鼠海马神经元超微结构损伤情况；采用酶联免疫吸附试验(Enzyme-linked immunosorbent assay, ELIS A)方法测量大脑DNA氧化损伤指标8。羟基脱氧鸟苷(8-hydroxy2'deoxyguanosine,8-OHdG)含量；采用实时荧光定量PCR (Quantitative Real Time PCR, qRT-PCR)和Western blot方法进行淀粉样前体蛋白(Amyloid precursor protein, APP)、tau、葡萄糖反应蛋白78(Glucose-regulated protein 78, GRP78)、葡萄糖反应蛋白94 (Glucose-regulated protein 94, GRP94)的表达量分析。结果：(1)各年龄段血铅及脑组织铅含量均随着铅暴露剂量的增加而增加,呈现剂量-反应关系,且均有统计学差异；各组在不同年龄段的血铅无显著性差异,但皮层、海马中铅在中年与老年期均比断乳期高,均有显著性差异。(2)铅暴露大鼠形态学改变：活体MRI结果显示,铅暴露老年大鼠大脑容积下降(pD.D5),海马容积呈下降趋势,但无统计学意义；离体硫堇染色结果可知,在断乳期,铅暴露组海马CA1区神经元密度下降,在老年期,铅暴露组大鼠皮层顶区和海马CA1区神经元密度明显下降,且与空白组比较有统计学差异,海马DG区呈下降趋势；电镜结果提示,随着染铅剂量增加,胞质、胞核、内质网结构和突触结构均遭到不同程度的破坏,出现早期神经元细胞凋亡现象。(3)在断乳和老年期,铅暴露使大脑皮层、海马APP表达增加,在中年有上升趋势,但差异无统计学意义；在断乳和老年期,铅暴露使海马tau及磷酸化tau (Ser396)蛋白表达增加,但在大鼠皮层tau表达无差异。(4)铅暴露增加了断乳和老年大鼠大脑8-OHdG的含量,且呈剂量-反应关系；但在中年变化不明显。(5)在老年期,铅暴露使大脑皮层、海马GRP78表达上升,在断乳和中年期变化均不明显；在断乳和老年期,海马各组GRP94表达呈上升趋势,但在皮层变化不明显,中年变化均不明显。结论：(1)大鼠形态学结果和阿尔茨海默病特征性基因上调提示铅暴露大鼠出现了神经退行性改变。(2)铅暴露导致的神经退行性改变可能与增加了内质网应激反应有关,也可能与增加了APP/tau的表达有关。(3)长期铅暴露大鼠各指标变化具有年龄阶段性。
[Abstract]:Objective: the study shows that lead not only has neurodevelopmental toxicity, but also may be an important risk factor for neurodegenerative disease. This paper aims to investigate the neurodegenerative changes and related gene protein expression in rats induced by long-term lead exposure. Methods: adult female Sprague-Dawley (SD) rats were randomly divided into 3 groups: empty In the white group, the low lead exposure group (LLG) and the high lead exposure group (HLG) were given 08001500mg/L lead acetic acid solution respectively, and the male rats who drank the deionized water for 10 days were caged. The offspring were born 21 days after the observation of the suppository, and the male offspring were fed to the weaning (Postnatal week 3, PNW3), middle age (Postnatal week 41, PNW41), old age, and old. (Postnatal week 70, PNW70). Male offspring rats were given 0300900mg/L acetate aqueous solution in three groups. The volume of brain and hippocampus in old rats was measured by living magnetic resonance imaging (Magnetic resonance imaging, MRI), and the isolated atomic emission spectrum (Inductively coupled plasma atom) was used as an isolated inductor coupling. IC emission spectrum-etry, ICP-AES) method was used to determine the blood lead and cortex and hippocampus lead content in three age rats. The density of hippocampal neurons in three age groups was observed by thio staining. The ultrastructural damage of hippocampal neurons in the aged rats was observed by transmission electron microscope, and the enzyme linked immunosorbent assay (Enzyme-li) was used. Nked immunosorbent assay, ELIS A) method was used to measure the content of 8. hydroxyl deoxy guanosine (8-hydroxy2'deoxyguanosine, 8-OHdG) of DNA oxidative damage in the brain. The expression of protein 78 (Glucose-regulated protein 78, GRP78) and glucose reactive protein 94 (Glucose-regulated protein 94, GRP94) was analyzed. Results: (1) the lead content of blood and brain tissue in all age groups increased with the increase of lead exposure dose, showing a dose response relationship, and there were statistical differences; each group was in different ages. There was no significant difference in blood lead in the segment, but in the cortex and the hippocampus, lead in the hippocampus was higher in the middle age and the aged than in the weaning period. (2) the morphological changes in the exposed rats showed that the volume of the brain in the aged rats decreased (pD.D5), the volume of the hippocampus decreased, but the volume of the hippocampus decreased, but the result of the isolated thio staining could be found. In the weaning period, the density of neurons in the hippocampal CA1 area decreased in the lead exposed group. In the elderly, the density of neurons in the cortex and hippocampus CA1 area of the lead exposed rats decreased significantly, and there was a statistical difference between the group and the blank group. The hippocampal DG area decreased. The electron microscope results suggested that the cytoplasm, the nucleus, the endoplasmic reticulum structure and the process of the endoplasmic reticulum were increased with the increase of the dose of lead. The contact structure was damaged in different degrees and appeared early neuronal apoptosis. (3) in weaning and old age, lead exposure increased the expression of APP in the cerebral cortex and hippocampus, and increased in middle age, but the difference was not statistically significant; in weaning and old age, the expression of tau and phosphorylated tau (Ser396) protein in the hippocampus increased, but in the weaning and old age, the expression of the protein in the hippocampus and phosphorylated tau (Ser396) was increased, but in the weaning and the old age, the expression of the protein was increased. There was no difference in the expression of tau in the cortex of rats. (4) lead exposure increased the content of 8-OHdG in the brain of weaning and old rats, and showed a dose response relationship, but the change in middle age was not obvious. (5) the expression of lead in the cerebral cortex and hippocampal GRP78 increased in the old age, and the changes in the weaning and middle age were not obvious; in the weaning and the old age, the hippocampus was GRP94 The expression showed an upward trend, but the changes in the cortex were not obvious, and the changes in middle age were not obvious. Conclusion: (1) the morphological results of the rats and the up-regulated genes of Alzheimer's disease suggest that the lead exposed rats have neurodegenerative changes. (2) the neurodegenerative changes caused by lead exposure may be related to the increase of endoplasmic reticulum stress response, and may also be associated with the increase of the endoplasmic reticulum stress response. It was related to the increase of APP/tau expression. (3) the changes of each indicator in the long-term lead exposed rats were of age.

【学位授予单位】：南昌大学
【学位级别】：硕士
【学位授予年份】：2013
【分类号】：R114

【参考文献】