發(fā)布時(shí)間：2018-06-01 06:32

  本文選題：三氧化二砷 + 神經(jīng)毒性�。� 參考：《大連醫(yī)科大學(xué)》2012年碩士論文

【摘要】：研究背景：砷（Arsenic, As）是一種廣泛分布于土壤、巖石和水環(huán)境中的有毒類(lèi)金屬元素。As在自然界中主要以化合物的形式存在，對(duì)健康具有多方面的危害，可引發(fā)多器官和多系統(tǒng)的形態(tài)學(xué)和功能上的異常改變。As對(duì)神經(jīng)系統(tǒng)的影響主要表現(xiàn)為頭痛、嗜睡、煩躁、記憶力減退、定向力障礙，驚厥甚至昏迷等亞臨床的神經(jīng)損傷。兒童由于正處于生理發(fā)育階段對(duì)外源化合物的毒作用較為敏感。大量流行病學(xué)調(diào)查、動(dòng)物實(shí)驗(yàn)表明，As暴露兒童的中樞神經(jīng)系統(tǒng)損傷主要表現(xiàn)為學(xué)習(xí)記憶能力的損害，已廣泛引起社會(huì)的高度關(guān)注，但目前，關(guān)于砷致學(xué)習(xí)記憶能力損害的分子作用機(jī)制尚不清楚。本課題組的前期研究發(fā)現(xiàn)，砷暴露可顯著抑制腦組織中鈣離子/鈣調(diào)蛋白依賴(lài)性蛋白激酶Ⅳ(calcium/calmodulin-dependent protein kinaseⅣ, Camk4)基因、蛋白的表達(dá)及c-Fos、JunB等與學(xué)習(xí)記憶相關(guān)基因、蛋白的表達(dá)，破壞腦組織中長(zhǎng)時(shí)程記憶（LTM）的形成。據(jù)文獻(xiàn)報(bào)道，腦組織Camk4基因的轉(zhuǎn)錄和翻譯依賴(lài)于兩種核受體即甲狀腺激素受體（TR）和視黃醇X受體(RXR)的存在。我們的前期研究已經(jīng)發(fā)現(xiàn)，砷暴露小鼠大、小腦組織TR受體亞型之一TRβ在基因、蛋白水平均表達(dá)顯著下調(diào)，，但砷是否也影響RXR基因表達(dá)國(guó)內(nèi)外未見(jiàn)相關(guān)報(bào)道。 目的：觀察砷對(duì)小鼠大、小腦組織中RXR基因和蛋白的表達(dá)影響，為闡明砷的神經(jīng)毒作用機(jī)制以及防治砷的神經(jīng)毒性危害提供靶基因依據(jù)。 方法：SPF級(jí)小鼠40只，按體重將小鼠隨機(jī)分為飲用水對(duì)照組、1ppm As_2O_3染毒組、2ppm As_2O_3染毒組、4ppm As_2O_3染毒組、4ppm As_2O_3+150mg/kg�；撬岜Ｗo(hù)組。通過(guò)自然飲用含不同濃度As_2O_3蒸餾水的方式使小鼠染砷，保護(hù)劑是以灌胃方式給予，連續(xù)染毒60天后取腦組織。用基因芯片技術(shù)和PCR技術(shù)檢測(cè)小鼠腦組織視黃醇X受體(RXR)的差異表達(dá)，并進(jìn)一步用Western blot和免疫組化的方法證實(shí)RXR蛋白水平表達(dá)和組織分布的改變。采用SPSS11.5統(tǒng)計(jì)軟件，用單因素方差分析（ANOVA），比較各染砷組與對(duì)照組間的統(tǒng)計(jì)學(xué)差異，兩組間比較用LSD法分析，以P0.05表示統(tǒng)計(jì)學(xué)差異顯著。 結(jié)果：基因芯片結(jié)果顯示，與對(duì)照組比較，各砷暴露組大腦組織RXR基因表達(dá)均顯著上調(diào)，尤其4ppm組上調(diào)更明顯(P 0.05)，而各組之間，小鼠小腦組織RXR的基因表達(dá)差異無(wú)統(tǒng)計(jì)學(xué)意義(P0.05)。Real Time PCR定量檢測(cè)結(jié)果顯示，砷暴露組大腦中RXR基因表達(dá)顯著上調(diào)，差異有統(tǒng)計(jì)學(xué)意義(P 0.05)，與基因芯片結(jié)果一致。同時(shí)，Western blotting檢測(cè)結(jié)果顯示，砷暴露導(dǎo)致小鼠大腦RXR蛋白表達(dá)上調(diào)，差異有統(tǒng)計(jì)學(xué)意義(P 0.05)，且呈劑量-反應(yīng)關(guān)系。免疫組化顯示RXR在砷染毒小鼠大腦4ppm組高表達(dá)。另外，我們發(fā)現(xiàn)，在砷暴露小鼠大腦組織，抗氧化劑�；撬釋�(duì)砷致2ppm和4ppm劑量組的RXR基因和蛋白表達(dá)上調(diào)有拮抗作用(P 0.05)。 結(jié)論：在亞慢性砷暴露可上調(diào)小鼠大腦組織中RXR基因表達(dá)水平，但在小腦組織中，RXR基因表達(dá)水平并無(wú)顯著差異變化。�；撬釋�(duì)亞慢性砷暴露使小鼠大腦組織中RXR基因和蛋白的表達(dá)上調(diào)有拮抗作用。提示，亞慢性砷暴露導(dǎo)致腦組織中RXR基因異常表達(dá)可能與砷誘導(dǎo)的氧化應(yīng)激作用有關(guān)。
[Abstract]:Research background: Arsenic (As) is a kind of toxic metal element, which is widely distributed in soil, rock and water environment,.As exists mainly in the form of compound in nature, and has many harmful effects on health. It can cause the abnormal changes of morphology and energy of multiple organs and multiple systems and the influence of.As on the nervous system. It is the subclinical nerve injury of headache, drowsiness, irritability, memory loss, disorder of orienteering, convulsion and even coma. Children are more sensitive to the toxic effects of exogenous compounds at the stage of physiological development. A large number of epidemiological investigations have shown that the central nervous system injury in children exposed to As is mainly learning and memory. The damage of capacity has aroused great concern in society, but at present, the molecular mechanism of arsenic induced impairment of learning and memory is not clear. The previous study in our group found that arsenic exposure could significantly inhibit the calcium ion / calmodulin dependent protein kinase IV (calcium/calmodulin-dependent protein kinase IV, C) in brain tissue. Amk4) gene, expression of protein and c-Fos, JunB, and learning and memory related genes, protein expression, destroying the formation of long history memory (LTM) in the brain tissue. It is reported that the transcription and translation of the Camk4 gene of the brain is dependent on the existence of two nuclear receptors, the thyroid hormone receptor (TR) and the retinol X receptor (RXR). Our preliminary study has already been reported. It was found that arsenic exposure in mice was large, and TR beta, one of the TR receptor subtypes of cerebellar tissue, decreased significantly in gene and protein levels, but whether arsenic also affects the expression of RXR gene has not been reported at home and abroad.
Objective: To observe the effect of arsenic on the expression of RXR gene and protein in the large and cerebellar tissues of mice, and to provide the target gene for elucidating the mechanism of arsenic neurotoxicity and the prevention and treatment of arsenic neurotoxicity.
Methods: 40 mice of grade SPF were randomly divided into drinking water control group, 1ppm As_2O_3 dye group, 2ppm As_2O_3 dye group, 4ppm As_2O_3 poisoning group, 4ppm As_2O_3+150mg/kg taurine protection group. The mice were exposed to arsenic by natural drinking with different concentrations of As_2O_3 distilled water, and the protective agent was given by gavage and continuous dyeing. The brain tissue was extracted after 60 days. The differential expression of retinol X receptor (RXR) in mouse brain tissue was detected by gene chip technology and PCR technique. The expression of RXR protein and the change of tissue distribution were confirmed by Western blot and immunohistochemistry. The SPSS11.5 statistics software was used to compare the arsenic staining groups with the single factor variance analysis (ANOVA). The statistical difference between the two groups was analyzed by LSD. P0.05 showed significant difference.
Results: the results of gene chip showed that the expression of RXR gene in the brain tissue of each arsenic exposure group was significantly up-regulated compared with the control group, especially in the 4ppm group (P 0.05), but there was no significant difference in the gene expression of RXR in the cerebellar tissues of the mice (P0.05).Real Time PCR quantitative detection results showed that the RXR base in the arsenic exposed group was in the brain. The difference was statistically significant (P 0.05), which was consistent with the results of gene chip. At the same time, the results of Western blotting detection showed that arsenic exposure led to the up regulation of RXR protein expression in the brain of mice (P 0.05), and showed a dose response relationship. Immunohistochemistry showed that RXR was highly expressed in the 4ppm group of arsenic exposed mice. In addition, we found that the antioxidant taurine had an antagonistic effect on the up regulation of RXR gene and protein expression in arsenic induced 2ppm and 4ppm doses in arsenic exposed mice brain tissue (P 0.05).
Conclusion: subchronic arsenic exposure can increase the level of RXR gene expression in the brain tissue of mice, but there is no significant difference in the expression of RXR gene in the cerebellar tissues. Taurine has antagonistic effect on the up regulation of RXR gene and protein expression in the brain tissue of mice. Abnormal expression of genes may be related to arsenic induced oxidative stress.
【學(xué)位授予單位】：大連醫(yī)科大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2012
【分類(lèi)號(hào)】：R114

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