Choroid Plexus a Target in Metal-Induced Neurotoxicity

描述(申请人提供)：最近添加的一种锰 含(Mn)抗爆化合物甲基环戊二烯基锰三羰基(MMT) 给美国汽油供应带来的健康风险引起了极大的关注 与环境中锰水平的潜在增加有关。两者都有 环境和职业接触(锰)会导致神经退行性变 症状类似帕金森氏症。然而，MN背后的机制 神经毒性仍不清楚。我们最近的结果表明，在体内和体外 暴露于锰会改变全身和亚细胞铁的状态。而前者 促进铁从血液循环流入脑脊液 体液(CSF)，后者促进细胞铁超载。我们还发现， 脉络丛中锰的积聚，脉络丛是血-脑脊液屏障的组织 驻留，提高转铁蛋白受体(TFR)mRNA的密度，它具有3‘ 结合IRE(铁响应元件)环接受[4Fe-4S] 含簇铁调节蛋白-1(IRP-1，或胞浆乌头酸酶)。 这些发现使我们提出了锰引起的机制 铁代谢异常可能是通过与IRP-1和 随之而来的是TFR过表达。这些事件反过来又加速了铁的运输 脑屏障系统和加重神经细胞中铁的蓄积。 因此，我们在这个建议中假设，脉络膜中锰的积累 神经丛改变血-脑脊液屏障中的铁调节机制，从而 干扰脑脊液中铁的动态平衡，这可能是锰诱导的 神经退行性帕金森综合症。我们的研究目标是更好地了解 锰致帕金森病的机制及其识别和预防 神经退行性疾病的环境原因。我们的具体目标是(1) 为了检验锰暴露改变铁调节的工作假说 脉络丛中的铁机制，导致脑脊液中铁的状态扭曲。 我们将定义锰暴露和铁暴露的剂量和时间效应关系 脑脊液、血液和脉络丛，检测IRP-1的活性，并确定 转铁蛋白受体在血-脑脊液屏障和特定脑区的表达； 检验锰促进铁在土壤中运输的工作假设 血-脑脊液屏障指向脑脊液，在数量上更多 比通过血脑屏障运输更重要。我们将在体外使用 确定两个点铁通量方向和大小的输送模型 主要的脑屏障，并调查是否阻止细胞转运 TFR拮抗锰增强的铁的单向转运；以及(3)测试 细胞铁调节中锰的变化作用假说 放在mRNA表达水平，而不是转录水平 基因组DNA的调控。我们将阻止锰对RNA结合的影响 IRP-1和TFR相关表达的能力，检测锰是否抑制 TFR mRNA的降解，并研究两种屏障中细胞铁蛋白的状态。 此外，我们还将研究锰对细菌RNA结合能力的影响。 新发现的IRP，它调节[Fe-S]亚基的合成 Complex-I。

DESCRIPTION (provided by applicant): The recent addition of a manganese (Mn)-containing antiknock compound methylcyclo-pentadienyl Mn tricarbonyl (MMT) to the US gasoline supply has raised a great concern about the health risks associated with a potential increase in the environmental levels of Mn. Both environmental and occupational exposures to (Mn) result in neurodegenerative symptoms resembling Parkinson's disease. However, the mechanisms underlying Mn neurotoxicity remain unknown. Our recent results show that in vivo and in vitro exposures to Mn alter both systemic and subcellular Fe status. While the former facilitates influx of Fe from the blood circulation to the cerebral spinal fluid (CSF), the latter promotes cellular Fe overload. We also found that accumulation of Mn in the choroid plexus, a tissue where blood-CSF barrier resides, enhances the density of transferrin receptor (TfR) mRNA, which has 3' binding IRE (iron responsive element) loops receptive to [4Fe-4S] cluster-containing iron regulatory protein-1 (IRP-1, or cytosolic aconitase). These findings have led us to propose that the mechanism by which Mn causes abnormal Fe metabolism is likely via its interaction with IRP-1 and the subsequent overexpression of TfR. The events, in turn, expedite Fe transport at the brain barrier systems and aggravate Fe accumulation in neuronal cells. Thus, we hypothesize in this proposal that accumulation of Mn in the choroid plexus alters Fe regulatory mechanisms in the blood-CSF barrier and thereby disturbs Fe homeostasis in the CSF, which may contribute to Mn-induced neurodegenerative Parkinsonism. Our research goals are to better understand the mechanism of Mn-induced Parkinsonism and in so doing identify and prevent environmental causes of neurodegenerative diseases. Our specific aims are (1) to test the working hypothesis that Mn exposure alters the Fe regulatory mechanism in the choroid plexus, leading to a distorted Fe status in the CSF. We will define the dose and time response relationship of Mn exposure and Fe in CSF, blood, and choroid plexus, examine the activity of IRP-1, and determine the expression of TfR in blood-CSF barrier and selected brain areas; (2) to test the working hypothesis that Mn-facilitated transport of Fe at the blood-CSF barrier is directed toward the CSF and quantitatively is more significant than transport by the blood-brain barrier. We will use in vitro transport models to determine the direction and magnitude of Fe fluxes at two major brain barriers and to investigate if blocking of cellular trafficking of TfR antagonizes Mn-augmented unidirectional transport of Fe; and (3) to test the working hypothesis that alteration by Mn of cellular Fe regulation takes place at the level of mRNA expression, but not at the level of transcriptional modulation of genomic DNA. We will deterrnine the effect of Mn on RNA binding capability of IRP-1 and pertinent expression of TfR, examine if Mn inhibits degradation of TfR mRNA, and study cellular ferritin status in both barriers. In addition, we will study the effect of Mn on the RNA binding capability of a newly discovered IRP which regulates the synthesis of an [Fe-S] subunit of Complex-I.