Determining the Role of Atrazine and Atrazine-Induced Estrogen in Increasing CD4+

说明(申请人提供)：阿特拉津是美国使用最广泛的除草剂之一。由于作物施用后的农业径流，它是美国最常见的地下水污染物(：//water.usgs.gov/nawqa/)。阿特拉津被归类为内分泌干扰物，因为它诱导了CYP19(芳香酶)，其催化活性导致雌激素产生增加。虽然已经对与阿特拉津暴露相关的潜在健康风险进行了大量研究，但大多数研究都集中在与雌激素升高有关的生殖异常上。少数研究考察了阿特拉津对哺乳动物免疫系统的影响，但对免疫系统的影响在很大程度上仍不清楚。此前没有一项研究关注免疫细胞的一个关键亚群--CD4T辅助细胞的激活和功能。这些细胞有助于形成对病原体和癌症的保护性免疫反应，并保持自身的耐受性。CD4T细胞数量或效应器活动的紊乱可能会对个人的健康造成可怕的后果。例如，调节性T细胞(Treg)的增加可能会导致免疫抑制，从而增加感染或癌症的易感性。在初步的体外研究中，观察到阿特拉津暴露显著抑制了常规T细胞的增殖、激活和细胞因子的产生。此外，在阿特拉津暴露的细胞中，Foxp3 Treg的频率显著增加。在这项建议中，我们将研究阿特拉津介导的抑制CD4T细胞的机制。这一建议的假设是，阿特拉津提高雌激素水平，通过直接抑制CD4T细胞和诱导Treg细胞显着增加来抑制CD4T细胞的激活。它将通过两个特定的目标进行测试：第一个目标是在体外和体内研究阿特拉津诱导Treg频率的机制。使用来自Foxp3gfp OT-II小鼠的TCR转基因T细胞，我们将确定阿特拉津是诱导nTreg细胞的扩张，还是将传统T细胞转化为诱导的Treg(ITreg)细胞。我们还将研究在缺乏Treg的情况下，阿特拉津暴露对常规T细胞表型的直接影响。在第二个目标中，使用药物抑制剂和抗体阻断的组合，将检查阿特拉津介导的雌激素产生对Treg增加的作用。

DESCRIPTION (provided by applicant): Atrazine is one of the most widely used herbicide in the United States. As a result of agricultural runoff after crop applications, it is the most commo ground water contaminant in the US (://water.usgs.gov/nawqa/). Atrazine is classified as an Endocrine-Disrupting Compound due its induction of CYP19 (aromatase), whose catalytic activity leads to increased estrogen production. While a significant amount of research has been done looking into the potential health risks associated with atrazine exposure, most have focused on reproduction abnormalities related to elevated estrogen. A small number of studies have examined the effect of atrazine on the mammalian immune system, but the effects on the immune system remain largely unknown. None of the previous studies have focused on the activation and functions of a critical subset of immune cells, the CD4+ T helper cells. These cells help shape the protective immune response to pathogens and cancer as well as maintaining tolerance of self. Perturbations of CD4+ T cell numbers or effector activity can have dire consequences for the health of the individual. For example, an increase in regulatory T cells (Treg), can lead to immune suppression and resulting in increased susceptibility to infection or cancer. In preliminary in vitro studies, it was observed that atrazine exposure significantly inhibits the proliferation, activation and cytokine production by conventional T cells. Further, there is a significant increase in the frequency of Foxp3+ Treg in atrazine-exposed cells. In this proposal we will examine the mechanism of atrazine-mediated inhibition of CD4+ T cells. The hypothesis of this proposal is that atrazine increases estrogen levels which suppresses CD4+ T cell activation by both directly inhibiting these cells and by inducing a significant increase of Treg cells. It will be tested using 2 specific aims: In the first aim, the mechanism of atrazine-induced Treg frequency will be examined in vitro and in vivo. Using TCR transgenic T cells from Foxp3gfp+ OT-II mice, we will determine whether atrazine is inducing expansion of nTreg or conversion of conventional T cells to induced Treg (iTreg) cells. We will also examine the direct effects of atrazine exposure on the phenotype of conventional T cells in the absence of Treg. In the second aim, using a combination of pharmacological inhibitors and antibody blockade, the role of atrazine-mediated estrogen production on the Treg increase will be examined.