双酚A、对枯基苯酚和己烯雌酚对MCF-7细胞的单一和联合效应研究
发布时间：2019-02-15 02:58
【摘要】：随着人类社会农业、工业、经济和科学的快速发展,大量化合物被合成并通过各种迁移转化途径进入环境中。外源雌激素(Xenoestrogens, XEs)就是其中的一大类物质,它可以通过模拟或者干扰内源雌激素的作用方式而发挥雌激素效应。不仅仅在水体,大气和土壤等环境介质中能检测到XEs,它也广泛存在于各种动物体内甚至是人体中,人类时时刻刻都进行着多种XEs的联合暴露。与此同时,大量研究证实了XEs暴露与乳腺癌、子宫内膜癌、卵巢癌、宫颈癌以及前列腺癌等癌症的发生发展有着密切关系。本论文以双酚A (BPA),己烯雌酚(DES)和对枯基苯酚(4-CP)三种结构类似的XEs为目标化合物,以雌激素受体阳性的人类乳腺癌MCF-7细胞为受试对象,通过细胞增殖实验研究了三种目标化合物和人体浓度的雌二醇(E2)单独作用和联合作用的雌激素效应。采用蛋白质免疫印迹法(Western blot)检测了雌激素单独和联合作用时雌激素受体α (ERα)、雌激素相关受体γ (ERRγ)和抗凋亡蛋白Bcl-2的表达水平,推测引起细胞增殖可能的机制。得到以下的研究结果：(1) BPA、4-CP、DES在诱导细胞增殖时具有时间依赖性和低剂量、非线性效应特征。三种XEs诱导细胞增殖能力为DESBPA4-CP,而高浓度时的细胞毒性为DES4-CPBPA。(2)三种目标化合物都可以通过与ERa结合,激活基因组通路,从而影响细胞的生长和增殖。四种雌激素与ERa结合能力为：E2 DES 4-CP BPA。 ERRγ在三种外源雌激素引起的增值效应中也具有重要作用,BPA和4-CP对ERRγ的亲和力大于DES,而DES大于E2。在单一效应实验中,Bcl-2蛋白的表达与各实验组的细胞增殖率之间有比较好的对应关系,提示：三种XEs也可以通过作用于细胞凋亡蛋白家族,从而对细胞增殖和凋亡产生影响。(3)三种外源雌激素两两混合时,BPA和DES可以通过混合作用相互促进,发挥出更强的雌激素效应。这可能是由于掺入BPA后,混合物的ERRγ的表达水平相对于DES单独作用时更高；而掺入DES后,混合物结合ERa的水平相对于DES单独作用时更强。但对于三种外源雌激素形成的三元混合物,其增值效应有一定减弱,高浓度的4-CP会使混合物发挥抑制增殖的效应,而相应的Western blot结果显示XEs三元混合对ERα、 ERRγ和Bcl-2的表达均有不同程度的影响。(4)生理浓度的E2可以使单一XEs对MCF-7细胞的促增殖能力显著提高。产生这一现象的原因可能是由于掺入XEs后,混合物的ERRγ的表达水平相对于E2单独作用时更高；而掺入E2后,混合物结合ERa的水平相对于XEs单独作用时更强。相反地,当E2与两种或三种XEs混合时,E2会弱化XEs混合物产生的雌激素效应,很多实验组雌激素活性基本丧失。且其对ERα、 ERRγ和Bcl-2表达的影响并不具有一般规律性。
[Abstract]:With the rapid development of agriculture, industry, economy and science, a large number of compounds have been synthesized and entered into the environment through various migration and transformation pathways. Exogenous estrogen (Xenoestrogens, XEs) is one of the major substances. It can play an estrogen effect by simulating or interfering with the action of endogenous estrogen. Not only can XEs, be detected in environmental media such as water body, atmosphere and soil, but it also exists in various animals and even in human body. Human beings are exposed to a variety of XEs all the time. At the same time, numerous studies have confirmed that XEs exposure is closely related to the occurrence and development of breast, endometrial, ovarian, cervical and prostate cancers. In this study, three similar XEs compounds, bisphenol A (BPA), diethylstilbestrol (DES) and p-withered phenol (4-CP), were used as target compounds, and MCF-7 cells with positive estrogen receptor were used as subjects. The estrogenic effects of three target compounds and estradiol (E _ 2) were studied by cell proliferation experiments. The expression levels of estrogen receptor 伪 (ER 伪), estrogen associated receptor 纬 (ERR 纬) and anti-apoptotic protein (Bcl-2) were detected by Western blot (Western blot). We speculate on the possible mechanism of cell proliferation. The results are as follows: (1) BPA,4-CP,DES has the characteristics of time-dependent, low-dose and nonlinear effects in the induction of cell proliferation. Three kinds of XEs can induce cell proliferation ability to be DESBPA4-CP, but at high concentration the cytotoxicity is DES4-CPBPA. (2). All three target compounds can activate genomic pathway by binding with ERa, thus affecting cell growth and proliferation. The binding ability of four estrogens to ERa is E2 DES 4-CP BPA.. ERR 纬 also plays an important role in the three exogenous estrogen-induced value-added effects. The affinity of BPA and 4-CP to ERR 纬 is greater than that of DES, and DES is greater than E2. In the single effect experiment, the expression of Bcl-2 protein had a good correspondence with the cell proliferation rate of each experimental group, which suggested that three kinds of XEs could also act on the apoptotic protein family. Thus, the proliferation and apoptosis of cells were affected. (3) when the three exogenous estrogens were mixed, BPA and DES could promote each other and play a stronger estrogen effect. This may be due to the higher expression level of ERR 纬 in the mixture after incorporation of BPA than that in the DES alone, while the level of ERa binding in the mixture with DES is stronger than that with DES alone. However, for the ternary mixture of three exogenous estrogens, the increment effect was weakened, and the high concentration of 4-CP could inhibit the proliferation of the mixture, and the corresponding Western blot results showed that the XEs ternary mixture could inhibit the proliferation of ER 伪. The expression of ERR 纬 and Bcl-2 were affected in different degrees. (4) the physiological concentration of E2 could significantly increase the proliferation of MCF-7 cells induced by single XEs. The reason for this phenomenon may be that the expression level of ERR 纬 in mixture is higher than that in E2 alone, and the level of ERa binding in mixture is stronger than that in XEs alone. On the contrary, when E2 was mixed with two or three kinds of XEs, E2 weakened the estrogenic effect of XEs mixture, and the estrogenic activity of many experimental groups was basically lost. Moreover, its effect on the expression of ER 伪, ERR 纬 and Bcl-2 has no general regularity.
【学位授予单位】：昆明理工大学
【学位级别】：硕士
【学位授予年份】：2016
【分类号】：X503
本文编号：2422890
[Abstract]:With the rapid development of agriculture, industry, economy and science, a large number of compounds have been synthesized and entered into the environment through various migration and transformation pathways. Exogenous estrogen (Xenoestrogens, XEs) is one of the major substances. It can play an estrogen effect by simulating or interfering with the action of endogenous estrogen. Not only can XEs, be detected in environmental media such as water body, atmosphere and soil, but it also exists in various animals and even in human body. Human beings are exposed to a variety of XEs all the time. At the same time, numerous studies have confirmed that XEs exposure is closely related to the occurrence and development of breast, endometrial, ovarian, cervical and prostate cancers. In this study, three similar XEs compounds, bisphenol A (BPA), diethylstilbestrol (DES) and p-withered phenol (4-CP), were used as target compounds, and MCF-7 cells with positive estrogen receptor were used as subjects. The estrogenic effects of three target compounds and estradiol (E _ 2) were studied by cell proliferation experiments. The expression levels of estrogen receptor 伪 (ER 伪), estrogen associated receptor 纬 (ERR 纬) and anti-apoptotic protein (Bcl-2) were detected by Western blot (Western blot). We speculate on the possible mechanism of cell proliferation. The results are as follows: (1) BPA,4-CP,DES has the characteristics of time-dependent, low-dose and nonlinear effects in the induction of cell proliferation. Three kinds of XEs can induce cell proliferation ability to be DESBPA4-CP, but at high concentration the cytotoxicity is DES4-CPBPA. (2). All three target compounds can activate genomic pathway by binding with ERa, thus affecting cell growth and proliferation. The binding ability of four estrogens to ERa is E2 DES 4-CP BPA.. ERR 纬 also plays an important role in the three exogenous estrogen-induced value-added effects. The affinity of BPA and 4-CP to ERR 纬 is greater than that of DES, and DES is greater than E2. In the single effect experiment, the expression of Bcl-2 protein had a good correspondence with the cell proliferation rate of each experimental group, which suggested that three kinds of XEs could also act on the apoptotic protein family. Thus, the proliferation and apoptosis of cells were affected. (3) when the three exogenous estrogens were mixed, BPA and DES could promote each other and play a stronger estrogen effect. This may be due to the higher expression level of ERR 纬 in the mixture after incorporation of BPA than that in the DES alone, while the level of ERa binding in the mixture with DES is stronger than that with DES alone. However, for the ternary mixture of three exogenous estrogens, the increment effect was weakened, and the high concentration of 4-CP could inhibit the proliferation of the mixture, and the corresponding Western blot results showed that the XEs ternary mixture could inhibit the proliferation of ER 伪. The expression of ERR 纬 and Bcl-2 were affected in different degrees. (4) the physiological concentration of E2 could significantly increase the proliferation of MCF-7 cells induced by single XEs. The reason for this phenomenon may be that the expression level of ERR 纬 in mixture is higher than that in E2 alone, and the level of ERa binding in mixture is stronger than that in XEs alone. On the contrary, when E2 was mixed with two or three kinds of XEs, E2 weakened the estrogenic effect of XEs mixture, and the estrogenic activity of many experimental groups was basically lost. Moreover, its effect on the expression of ER 伪, ERR 纬 and Bcl-2 has no general regularity.
【学位授予单位】：昆明理工大学
【学位级别】：硕士
【学位授予年份】：2016
【分类号】：X503
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