The mechanism of exocrine gland disorder through estrogen and/or endocrine disruptors

为了研究雌激素和内分泌干扰物对EBY再激活的可能性，我们分析了BZLF1启动子试验和ZEBRA蛋白的免疫印迹。雌二醇抑制TPA诱导的B95-8和Akata细胞中ZEBRA的表达。我们建立了转染pZp221-Luc和pZp552-Luc的稳定的B95-8细胞系。在这些细胞中，雌二醇也抑制TPA激活的荧光素酶活性。在Hela、HSY和HSG细胞中，雌二醇抑制转染ZEBRA或TPA刺激诱导的Zp活性。雌激素的作用在30分钟内显示出来，因此我们推测这些作用的一部分可能是通过膜ER或GPR30发生的非基因组事件。本文对雌激素影响Zp活性的机制进行了详细的探讨。接下来，我们使用BPA、3-MC、B[a]P和TCDD以相同的方法分析了内分泌干扰物的影响。这些化学物质均未改变B94-8和Akata细胞中ZEBRA的表达或Zp活性。与B细胞的作用相反，3-MC和TCDD在Hela、HSG和HSY细胞中增强了Zp活性。当我们转染AhR/Arnt和ER时，与单独转染AhR/Arnt相比，Zp在Hela细胞中的活性降低，而在HSG和HSY细胞中活性增强。结果表明，内质网对二恶英诱导的Zp活性具有不同类型的调节作用。在HSY细胞中，虽然没有AhR/Arnt的二恶英没有改变Zp的ZEBRA转激活，但当ZEBRA和AhR/Arnt共转染时，Zp的活性较高。我们推测二恶英可能是eb病毒在上皮细胞中再激活的增强剂。为了确定AhR/Arnt和ZEBRA之间的相互作用，我们正在使用免疫沉淀、GST下拉实验、凝胶移位实验和脚印分析进行研究。

To study the possibility of EBY reactivation by estrogen and endocrine disruptors, we analyzed BZLF1 promoter assay and immunoblotting for ZEBRA protein. Estradiol inhibited ZEBRA expression induced by TPA in B95-8 and Akata cells. We established stable B95-8 cell lines which transfected pZp221-Luc and pZp552-Luc. In these cells estradiol also inhibited luciferase activity activated by TPA. In Hela, HSY and HSG cells, estradiol inhibited Zp activity induced by both ZEBRA transfection or TPA stimulation. The effect of estrogen was revealed in 30 minutes, thus we speculated a part of these effects might be a non-genomic event through membrane ER or GPR30. The mechanisms of estrogen on Zp activity are being searched in detail. Next, we analyzed the effects of endocrine disruptors with the same procedure using BPA,3-MC,B[a]P and TCDD. None of these chemicals changed the ZEBRA expression or Zp activity in B94-8 and Akata cells. In contrast to these effects on B cells, Zp activity was enhanced by 3-MC and TCDD in Hela, HSG and HSY cells. When we transfected AhR/Arnt and ER, the activity of Zp was decreased in Hela cells whereas enhanced in HSG and HSY cells compared with AhR/Arnt alone. These results indicated that ER could regulate the Zp activity induced by dioxins depending on the cell types. In HSY cells, although ZEBRA transactivation of Zp was not changed by dioxins without AhR/Arnt, high activity of Zp was observed when ZEBRA and AhR/Arnt were cotransfected. We speculated that dioxins might be an enhancer of EBV reactivation in epithelial cells. To determine the interaction between AhR/Arnt and ZEBRA, we are investigating using immunoprecipitation, GST pull-down assay, Gel shift assay and foot printing.