Choroid Plexus a Target in Metal-Induced Neurotoxicity

描述（由申请人提供）：最近添加了锰 含(Mn)抗爆化合物甲基环戊二烯基三羰基锰(MMT) 美国汽油供应引发了人们对健康风险的高度关注 与环境中锰含量的潜在增加有关。两者都 环境和职业接触（Mn）会导致神经退行性疾病 类似帕金森病的症状。然而，锰的潜在机制 神经毒性仍未知。我们最近的结果表明，体内和体外 接触锰会改变全身和亚细胞的铁状态。而前者 促进铁从血液循环流入脑脊髓 液（CSF），后者会促进细胞铁超载。我们还发现 脉络丛中锰的积累，脉络丛是血脑脊液屏障的组织 驻留，增强转铁蛋白受体 (TfR) mRNA 的密度，该受体具有 3' 结合 IRE（铁反应元件）环，可接受 [4Fe-4S] 含有簇的铁调节蛋白-1（IRP-1，或胞质乌头酸酶）。 这些发现使我们提出，锰引起的机制 Fe 代谢异常可能是通过其与 IRP-1 和 随后 TfR 过度表达。这些事件反过来又加速了铁的运输 脑屏障系统并加剧神经元细胞中铁的积累。 因此，我们在这个提议中假设脉络膜中锰的积累 丛改变血脑脊液屏障中的铁调节机制，从而 扰乱 CSF 中的 Fe 稳态，这可能有助于 Mn 诱导 神经退行性帕金森症。我们的研究目标是更好地了解 锰诱发帕金森病的机制，并在此过程中识别和预防 神经退行性疾病的环境原因。我们的具体目标是（1） 检验锰暴露改变铁调节的工作假设 脉络丛中的机制，导致脑脊液中 Fe 状态的扭曲。 我们将定义 Mn 暴露和 Fe 的剂量和时间响应关系 脑脊液、血液和脉络丛，检查IRP-1的活性，并确定 血脑脊液屏障和选定脑区中 TfR 的表达； (2) 至 检验以下工作假设：Mn 促进 Fe 在 血脑脊液屏障指向脑脊液，数量上更多 比通过血脑屏障的运输更重要。我们将在体外使用 输运模型确定两点处 Fe 通量的方向和大小 主要的脑屏障并研究是否阻断细胞运输 TfR 拮抗 Mn 增强的 Fe 单向传输； (3) 测试 Mn 改变细胞 Fe 调节的工作假设 位于mRNA表达水平，但不位于转录水平 基因组 DNA 的调节。我们将确定 Mn 对 RNA 结合的影响 IRP-1 的能力和 TfR 的相关表达，检查 Mn 是否抑制 TfR mRNA 的降解，并研究两种屏障中的细胞铁蛋白状态。 此外，我们将研究Mn对RNA结合能力的影响 新发现的IRP调节[Fe-S]亚基的合成 复合体-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.