殺草強(qiáng)對(duì)人甲狀腺細(xì)胞系Nthy-ori-3-1細(xì)胞全基因表達(dá)譜的影響
發(fā)布時(shí)間：2018-10-16 14:27
【摘要】：目的甲狀腺是人體最大的內(nèi)分泌腺,其分泌的甲狀腺激素對(duì)人體的生長(zhǎng)發(fā)育和新陳代謝有很重要的作用。甲狀腺干擾物對(duì)人及動(dòng)物的甲狀腺有很顯著的影響,能夠干擾甲狀腺功能、誘發(fā)自身免疫性甲狀腺疾病、還有潛在的致甲狀腺腫瘤的作用。在已知的甲狀腺干擾物中,農(nóng)藥占有很大的比例。殺草強(qiáng)作為一種非選擇性除草劑,被證實(shí)能夠干擾甲狀腺激素的合成以及導(dǎo)致甲狀腺增生和甲狀腺腫瘤,但是對(duì)其作用機(jī)制的研究報(bào)道很少。本研究以人甲狀腺細(xì)胞系Nthy-ori-3-1細(xì)胞為受試對(duì)象,在排除細(xì)胞毒性作用的劑量下,從基因水平探索殺草強(qiáng)致甲狀腺腫瘤相關(guān)的作用機(jī)制。方法實(shí)驗(yàn)設(shè)計(jì)了殺草強(qiáng)0μg/ml、μg/ml.10pg/ml.100μg/ml四個(gè)劑量組,用含相應(yīng)受試物的培養(yǎng)基培養(yǎng)人甲狀腺細(xì)胞系Nthy-ori-3-1細(xì)胞24小時(shí)后,經(jīng)MTT法檢測(cè)細(xì)胞吸光度值,并計(jì)算細(xì)胞存活率。采用Agilent單標(biāo)表達(dá)譜基因芯片對(duì)差異表達(dá)基因進(jìn)行了篩選,并應(yīng)用實(shí)時(shí)定量PCR技術(shù)隨機(jī)選擇了五個(gè)差異基因驗(yàn)證基因芯片結(jié)果的可靠性。結(jié)果MTT實(shí)驗(yàn)結(jié)果顯示,殺草強(qiáng)劑量在1～100-g/ml時(shí),Nthy-ori-3-1細(xì)胞的形態(tài)以及增殖同樣無(wú)顯著的變化,表明在此劑量下,殺草強(qiáng)對(duì)Nthy-ori-3-1細(xì)胞的影響與細(xì)胞毒性無(wú)關(guān)�；蛐酒Y選結(jié)果顯示,有90個(gè)基因的表達(dá)發(fā)生了顯著的變化,其中55個(gè)基因的表達(dá)上調(diào),35個(gè)基因表達(dá)下調(diào)；差異基因共影響了45個(gè)信號(hào)通路,其中大部分與腫瘤發(fā)生發(fā)展有關(guān)。實(shí)時(shí)定量PCR技術(shù)驗(yàn)證差異基因表達(dá),AGT、CA11基因顯著上調(diào)；MUC12、RAPH1、POLR1A基因顯著下調(diào),與基因芯片結(jié)果一致。結(jié)論殺草強(qiáng)劑量在1-100μg/ml時(shí),其對(duì)Nthy-ori-3-1細(xì)胞的影響與細(xì)胞毒性無(wú)關(guān)。AGT、CAll、MUC12基因在甲狀腺腫瘤的發(fā)生發(fā)展中起到很重要的作用；RAPH1、POLR1A基因能夠影響細(xì)胞的內(nèi)部結(jié)構(gòu),包括蛋白質(zhì)的合成、修飾和代謝以及細(xì)胞骨架系統(tǒng)的裝配,實(shí)時(shí)定量PCR驗(yàn)證結(jié)果與基因芯片結(jié)果一致。
[Abstract]:Objective thyroid gland is the largest endocrine gland in human body. Thyroid hormone secreted by thyroid gland plays an important role in growth, development and metabolism of human body. Thyroid disrupters have a significant effect on the thyroid gland of human and animals, which can interfere with thyroid function, induce autoimmune thyroid diseases, and have potential thyroid tumor effects. Pesticides account for a large proportion of known thyroid disruptions. As a non-selective herbicide, it has been proved to interfere with the synthesis of thyroid hormones and lead to thyroid hyperplasia and thyroid neoplasms. In this study, the human thyroid cell line Nthy-ori-3-1 cells were selected as subjects, and the mechanism related to the thyroid neoplasms induced by chlorpropoxide was explored at the gene level at the dose excluding cytotoxicity. Methods four dose groups of 0 渭 g / ml and 渭 g/ml.10pg/ml.100 渭 g/ml were designed. Human thyroid cell line Nthy-ori-3-1 cells were cultured on the culture medium containing the corresponding test material for 24 hours. The cell absorbance was measured by MTT method and the cell survival rate was calculated. The differentially expressed genes were screened by Agilent single standard gene microarray, and five differentially expressed genes were randomly selected by real-time quantitative PCR to verify the reliability of the results. Results the results of MTT test showed that the morphology and proliferation of Nthy-ori-3-1 cells had no significant change at the dose of 1~100-g/ml, which indicated that the effect of Xizuoqiang on Nthy-ori-3-1 cells was not related to the cytotoxicity. The results of gene chip screening showed that there were significant changes in the expression of 90 genes, of which 55 genes were up-regulated and 35 genes were down-regulated. Most of them are related to tumorigenesis and development. The differential gene expression was confirmed by real-time quantitative PCR, the AGT,CA11 gene was up-regulated and the MUC12,RAPH1,POLR1A gene was down-regulated, which was consistent with the results of gene chip. Conclusion the cytotoxicity of Nthy-ori-3-1 cells is not related to its effect on Nthy-ori-3-1 cells at a dose of 1-100 渭 g/ml. AGT,CAll,MUC12 gene plays an important role in the development of thyroid neoplasms, and RAPH1,POLR1A gene can affect the internal structure of the cells, including the synthesis of proteins. The results of modification and metabolism, assembly of cytoskeleton system, and real-time quantitative PCR were consistent with those of gene chip.
【學(xué)位授予單位】：昆明醫(yī)科大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2015
【分類號(hào)】：R114
本文編號(hào)：2274652
[Abstract]:Objective thyroid gland is the largest endocrine gland in human body. Thyroid hormone secreted by thyroid gland plays an important role in growth, development and metabolism of human body. Thyroid disrupters have a significant effect on the thyroid gland of human and animals, which can interfere with thyroid function, induce autoimmune thyroid diseases, and have potential thyroid tumor effects. Pesticides account for a large proportion of known thyroid disruptions. As a non-selective herbicide, it has been proved to interfere with the synthesis of thyroid hormones and lead to thyroid hyperplasia and thyroid neoplasms. In this study, the human thyroid cell line Nthy-ori-3-1 cells were selected as subjects, and the mechanism related to the thyroid neoplasms induced by chlorpropoxide was explored at the gene level at the dose excluding cytotoxicity. Methods four dose groups of 0 渭 g / ml and 渭 g/ml.10pg/ml.100 渭 g/ml were designed. Human thyroid cell line Nthy-ori-3-1 cells were cultured on the culture medium containing the corresponding test material for 24 hours. The cell absorbance was measured by MTT method and the cell survival rate was calculated. The differentially expressed genes were screened by Agilent single standard gene microarray, and five differentially expressed genes were randomly selected by real-time quantitative PCR to verify the reliability of the results. Results the results of MTT test showed that the morphology and proliferation of Nthy-ori-3-1 cells had no significant change at the dose of 1~100-g/ml, which indicated that the effect of Xizuoqiang on Nthy-ori-3-1 cells was not related to the cytotoxicity. The results of gene chip screening showed that there were significant changes in the expression of 90 genes, of which 55 genes were up-regulated and 35 genes were down-regulated. Most of them are related to tumorigenesis and development. The differential gene expression was confirmed by real-time quantitative PCR, the AGT,CA11 gene was up-regulated and the MUC12,RAPH1,POLR1A gene was down-regulated, which was consistent with the results of gene chip. Conclusion the cytotoxicity of Nthy-ori-3-1 cells is not related to its effect on Nthy-ori-3-1 cells at a dose of 1-100 渭 g/ml. AGT,CAll,MUC12 gene plays an important role in the development of thyroid neoplasms, and RAPH1,POLR1A gene can affect the internal structure of the cells, including the synthesis of proteins. The results of modification and metabolism, assembly of cytoskeleton system, and real-time quantitative PCR were consistent with those of gene chip.
【學(xué)位授予單位】：昆明醫(yī)科大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2015
【分類號(hào)】：R114
【參考文獻(xiàn)】
相關(guān)期刊論文 前1條
1 張?jiān)谙?脫碘酶[J];衛(wèi)生研究;1998年05期
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