發(fā)布時(shí)間：2018-01-15 13:02

  本文關(guān)鍵詞：喹乙醇誘導(dǎo)胰島細(xì)胞DNA損傷及其機(jī)制探討　出處：《大連醫(yī)科大學(xué)》2013年碩士論文　論文類型：學(xué)位論文

  更多相關(guān)文章： 喹乙醇 DNA損傷 氧化應(yīng)激 溶酶體膜穩(wěn)定性 Western Blot

【摘要】：目的：喹乙醇(olaquindox),是一種抗菌促生長(zhǎng)劑，被廣泛應(yīng)用在禽畜和水產(chǎn)養(yǎng)殖業(yè)，，已有實(shí)驗(yàn)表明喹乙醇對(duì)肝腎均有損害，但尚未有關(guān)于喹乙醇對(duì)胰島損害的研究，本實(shí)驗(yàn)通過彗星實(shí)驗(yàn)觀察喹乙醇是否會(huì)對(duì)大鼠胰島細(xì)胞的DNA有損傷并探討其損害可能的機(jī)制，通過western blot實(shí)驗(yàn)觀察喹乙醇對(duì)p53蛋白表達(dá)的影響，從而探討由持續(xù)DNA損傷可能引起的與癌有關(guān)基因表達(dá)的增加，為更全面的評(píng)估喹乙醇多臟器的損害提供依據(jù)。 方法：喹乙醇引起的DNA損傷是通過單細(xì)胞凝膠電泳實(shí)驗(yàn)檢測(cè)，用2，7，-二氫二氯熒光素（DCFH）檢測(cè)細(xì)胞內(nèi)活性氧（ROS）含量，用吖啶橙（AO）檢測(cè)細(xì)胞內(nèi)溶酶體膜穩(wěn)定性，通過辣根過氧化物酶標(biāo)記的熒光指示劑來檢測(cè)westernblot實(shí)驗(yàn)中P53蛋白表達(dá)量，通過堿性保護(hù)劑NH4Cl和NAC來干預(yù)彗星實(shí)驗(yàn)及分別干預(yù)溶酶體實(shí)驗(yàn)和ROS實(shí)驗(yàn)。 結(jié)果：喹乙醇（0~1600ug/ml）作用于INS-1細(xì)胞1h后，彗星實(shí)驗(yàn)中尾長(zhǎng)、尾距、尾DNA％含量均增加，并呈劑量依賴，在對(duì)800、1600ug/ml劑量組分別加NH4Cl和NAC干預(yù)后均有保護(hù)作用。喹乙醇同樣作用于INS-1細(xì)胞1h，可使細(xì)胞內(nèi)溶酶體膜穩(wěn)定性下降、ROS含量增加，分別用NH4Cl和NAC干預(yù)后均有保護(hù)作用。喹乙醇分別作用于INS-1細(xì)胞24h和48h后，P53蛋白表達(dá)量增加。 結(jié)論：喹乙醇可以誘導(dǎo)大鼠胰島細(xì)胞DNA損傷，對(duì)大鼠胰島細(xì)胞有遺傳毒性， NAC和NH4Cl均可對(duì)喹乙醇所致的DNA損傷起到保護(hù)作用，說明喹乙醇DNA損傷的途徑是氧化應(yīng)激和溶酶體途徑。喹乙醇可使p53蛋白表達(dá)量增加，表明喹乙醇可使與癌有關(guān)基因表達(dá)增加，其機(jī)制可能是DNA的持續(xù)損傷。
[Abstract]:Objective: olaquindox (olaquindox), is a kind of antibacterial auxogen, is widely used in livestock and aquaculture, previous studies have demonstrated that the damage to the liver and kidney of olaquindox, but not yet a study of olaquindox on islet lesions, the experimental observation of the comet will ethanol on rat islet cell damage and DNA to explore the possible mechanism of the damage effect of Western blot through the experimental observation of olaquindox on expression of p53 protein increased, so as to explore the possible damage caused by sustained DNA cancer related gene expression, and provide the basis for a more comprehensive assessment of olaquindox multiple organ damage.
Methods: DNA damage caused by olaquindox by single cell gel electrophoresis assay, 2,7, two - two hydrogen chloride fluorescein (DCFH) detection of intracellular reactive oxygen species (ROS) content, using acridine orange (AO) to detect the intracellular lysosomal membrane stability by horseradish peroxidase labeled fluorescent indicator to detect the expression of P53 in mind the amount of protein in the Westernblot experiment, the protective agent NH4Cl and NAC by alkaline comet assay and intervention intervention respectively lysosomes and ROS experiment.
Results: olaquindox (0~1600ug/ml) in INS-1 cells after 1h, the comet tail length, tail length, the tail DNA% content were increased in a dose-dependent manner, the protective effect of 8001600ug/ml was NH4Cl and NAC respectively with the dose group after the intervention. The same effect on INS-1 cells of olaquindox 1H, the lysosomal membrane stability decreased, ROS content increased by NH4Cl and NAC, respectively. After the intervention has a significant protective effect of olaquindox were acting in 24h and 48h INS-1 cells, the expression of P53 protein was increased.
Conclusion: olaquindox can islet cell injury in rats induced by DNA, genetic toxicity on rat islet cells, NAC and NH4Cl on DNA damage induced by olaquindox can play a protective role, how olaquindox DNA injury is oxidative stress and lysosomal pathway. Olaquindox can make the expression of p53 protein was increased, and that of olaquindox can make cancer the increased gene expression, its mechanism may be sustained damage in DNA.

【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】：2013
【分類號(hào)】：R114

【參考文獻(xiàn)】

相關(guān)期刊論文 前10條

1 鄒家杰;張婷;湯樹生;陳倩;靳溪;陳開跑;肖希龍;;喹乙醇誘導(dǎo)HepG2細(xì)胞線粒體的氧化損傷[J];癌變.畸變.突變;2009年05期

2 張婷;陳倩;湯樹生;靳溪;鄒家杰;劉鳳英;張q

本文編號(hào)：1428444