How environmental chemicals impair immunity

描述(由PI提供)：这些研究的目标是确定环境化学物质损害免疫细胞发育的分子机制。虽然许多研究表明，环境中普遍存在的多环芳烃(PAH)会抑制成熟的免疫系统，但相对较少的研究评估了它们对发育中的免疫系统的影响。为了弥补这一差距，我们利用B细胞发育模型来确定PAH如何影响骨髓B细胞。我们证明PAH通过间接诱导PRO-B和Pre-B细胞的凋亡来抑制B淋巴细胞的生成，这些细胞是通过构成造血微环境的基质细胞间接介导的。这些研究导致了一种假设，即发育中的B细胞室对环境化学物质非常敏感，环境化学物质不适当地激活了B细胞死亡程序。 我们最近的研究已经开始阐明执行B细胞死亡程序的一连串信号。建立在这些研究基础上的工作模型提出了至少两条可能重叠的途径，一条涉及caspase-8，另一条涉及线粒体激活。一个独特的资源组合，包括凋亡基因缺陷的、未转化的早期B细胞系和稳定的基因敲除小鼠，将被用来测试和完善这个模型。我们提出了三个具体的目标：1)明确PAH诱导的、caspase-8依赖的、骨髓B细胞中的凋亡信号通路(D.1)：我们将：a)确定死亡受体在caspase-8激活中的作用，b)评估JNK、PKR和ASK1激酶在caspase-8依赖的细胞凋亡中的作用，以及c)用质谱仪(MALDI-TOF)鉴定激活的caspase-8复合体的成分。2)评估线粒体依赖的细胞凋亡信号的影响因素。(D.2)：将绘制出多环芳烃诱导的线粒体凋亡放大途径的要素。具体地说，我们将：a)确定caspase-2和-8在Bid介导的线粒体激活中的作用，b)评估JNK、PKR和P53在caspase非依赖性线粒体激活中的作用，以及c)确定核因子-kappaB下调与caspase信号转导的关系。3)对体外绘制的细胞凋亡途径进行体内相关性研究。(D.3)：为了建立在体外获得的结果，我们将使用基因敲除小鼠：a)在体内确定骨髓B细胞亚群对AhR依赖的凋亡信号的相对敏感性，以及b)在体内研究PAH诱导的骨髓B细胞亚群中caspase依赖的死亡信号。这些研究不仅有助于我们理解多环芳烃对B细胞发育的分子作用，而且也将验证一个易于在分子水平上适用于其他免疫毒性环境化学物质和其他造血细胞亚群研究的平台。

DESCRIPTION (provided by PI): The goal of these studies is to determine the molecular mechanisms by which environmental chemicals impair immune cell development. While many studies demonstrate that environmentally ubiquitous polycyclic aromatic hydrocarbons (PAH) suppress the mature immune system, relatively few have evaluated their effects on the developing immune system. To bridge this gap, we exploited B cell development models to define how PAH affect bone marrow B cells. We demonstrated that PAH suppress B lymphopoiesis by inducing pro-B and pre-B cell apoptosis mediated indirectly through stromal cells constituting the hematopoietic microenvironment. These studies led to the hypothesis that the developing B cell compartment is exquisitely sensitive to environmental chemicals, which inappropriately activate B cell death programs. Our recent studies have begun to elucidate the cascade of signals that executes the B cell death program. The working model built on these studies proposes at least two potentially overlapping pathways, one involving caspase-8 and another involving mitochondrial activation. A combination of unique resources, including apoptosis gene-defective, nontransformed early B cell lines and a stable of knockout mice will be exploited to test and to refine this model. Three specific aims are proposed: 1) Define PAH-induced, caspase-8-dependent, apoptosis signaling pathways in bone marrow B cells (D. 1): We will: a) determine the role of death receptors in caspase-8 activation, b) evaluate the contribution of JNK, PKR, and ASK1 kinases to caspase-8-dependent apoptosis, and c) identify components of activated caspase-8 complexes by mass spectrometry (MALDI-TOF). 2) Evaluate factors contributing to mitochondria-dependent apoptosis signaling. (D.2): The elements of a PAH-induced mitochondrial apoptosis amplification pathway will be mapped out. Specifically, we will: a) determine the roles of caspases-2 and -8 in mitochondrial activation mediated by Bid, b) assess the contribution of JNK, PKR and p53 in caspase-independent mitochondrial activation, and c) define where NF-kappaB down-regulation, a prerequisite to PAH-induced pro/pre-B cell death, interfaces with caspase signaling. 3) Investigate in vivo correlates of the apoptosis pathway mapped out in vitro. (D.3): To build on results obtained in vitro, we will use gene-knockout mice: a) to determine the relative sensitivities of bone marrow B cell subsets to AhR-dependent apoptosis signaling in vivo and b) to investigate PAH-induced, caspase-dependent death signaling in bone marrow B cell subsets in vivo. These studies will not only contribute greatly to our understanding of the molecular effects of PAH on developing B cells, but they also will validate a platform that is easily applicable to the study of other immunotoxic environmental chemicals and other hematopoietic cell subsets on the molecular level.