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

描述（由申请人提供）：阿特拉津是美国使用最广泛的除草剂之一。作为作物施用后农业径流的结果，它是美国最常见的地下水污染物（://water.usgs.gov/nawqa/）。阿特拉津因其诱导CYP19（芳香化酶）而被归类为内分泌干扰化合物，其催化活性导致雌激素产生增加。虽然已经有大量的研究调查了与阿特拉津接触有关的潜在健康风险，但大多数研究都集中在与雌激素升高有关的生殖异常上。少数研究调查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然未知。之前的研究都没有关注免疫细胞的一个关键亚群——CD4+ T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，以及维持自我耐受性。CD4+ T细胞数量或效应活性的扰动可能对个体的健康产生可怕的后果。例如，调节性T细胞（Treg）的增加可导致免疫抑制并导致对感染或癌症的易感性增加。在初步的体外研究中，观察到阿特拉津暴露显著抑制常规T细胞的增殖、活化和细胞因子的产生。此外，暴露于阿特拉津的细胞中Foxp3+ Treg的频率显著增加。在这个提议中，我们将研究阿特拉津介导的CD4+ T细胞抑制的机制。该建议的假设是，阿特拉津增加雌激素水平，通过直接抑制CD4+ T细胞和诱导Treg细胞显著增加来抑制CD4+ T细胞的激活。试验将采用两个特定目的：第一个目的是在体外和体内研究阿特拉津诱导Treg频率的机制。利用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扩增还是诱导常规T细胞向诱导Treg （iTreg）细胞转化。我们还将研究在Treg缺失的情况下，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，阿特拉津介导的雌激素产生对Treg增加的作用将被检查。

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.