Iron regulation at the CNS vascular barriers and aging

说明(申请人提供)：虽然铁(Fe)对正常的大脑功能是必不可少的，但它在老化的大脑中的积累与神经退化有关。积聚的机制仍不清楚。我们最近的研究表明，转铁蛋白(Tf)结合的和游离的铁进入中枢神经系统是由(I)脑毛细血管(BBB)和脉络丛(CP)最初的快速隔离，以及(Ii)随后从血管结构向脑组织液(ISF)和脑脊液(CSF)的受控和缓慢释放所决定的。此外，我们的试验数据表明，铁主要通过转铁蛋白药物机制从大脑迅速清除到血液中，进入血脑屏障。根据这些初步数据，我们假设铁跨血脑屏障和CP是一个两步过程，需要Tf受体(TFR)和DMT1(二价金属转运体1)，其中DMT1是一个限速步骤，而从CNS外流是一个依赖于Tf途径的一步过程，需要血脑屏障上的TFR。我们进一步假设，铁中枢神经系统的内流和外流运输机制之间的失衡，主要是由于血管中枢神经系统屏障的清除减少导致其在衰老的大脑中积累。有两个目标。1.探讨转铁蛋白(Tf)介导和非转铁蛋白(Non-Tf)途径在选择性增龄大鼠中枢神经系统铁外流中的作用及铁在中枢神经系统中的滞留机制。2.测定Tf和Non-Tf介导的铁跨血脑屏障和CP在选择年龄的野生型和贝尔格莱德大鼠体内的动力学。这些目的将通过使用脑灌流和清除技术以及贝尔格莱德大鼠(b/b)来实现，因为它是DNT1的生理敲除。在b/b/和对照组大鼠中测定Tf和非Tf结合的铁的动力学参数，以确定DMT1在血脑屏障和CP中铁的流入和流出中的作用。在选定年龄的对照组大鼠中进行的类似研究将确定与年龄相关的变化。TFR和DMT1在血脑屏障和CP的相对水平将通过标准分子技术来确定，以补充运输研究。这项研究将有助于我们了解DMT1和TFR在BBB和CP中与脑内铁积累相关的神经退行性变化的作用，并确定未来开发脑内铁降低策略的潜在靶点。

DESCRIPTION (provided by applicant): While iron (Fe) is essential for normal brain function its accumulation in the aging brain is associated with neurodegeneration. The mechanism of accumulation is still unclear. Our recent studies indicated that entry of transferrin (Tf)-bound and free iron into the CNS is determined by (i) initial rapid sequestration by brain capillaries (BBB) and choroid plexus (CP), and (ii) subsequent controlled and slow release from vascular structures into brain interstitial fluid (ISF) and cerebrospinal fluid (CSF). In addition, our pilot data suggest that iron is rapidly cleared from brain into blood across the blood-brain barrier mainly by a Tf-medicated mechanism. Based on these preliminary data, we hypothesize that influx of iron across the blood-brain barrier and CP is a two-step process that requires both Tf receptor (TfR) and DMT1 (divalent metal transporter 1) with DMT1 being a rate limiting step, while efflux of Fe from the CNS is a one-step process that depends on Tf route and requires TfR at the blood-brain barrier. We further hypothesize that imbalance between iron CNS influx and efflux transport mechanisms, due mainly to reduced clearance at the vascular CNS barriers leads to its accumulation in the aging brain. There are two Aims. 1. To determine the role of Tf-mediated and non-Tf route for Fe efflux from CNS and the retention mechanism for Fe in CNS in bold type and DMT1 mutant Belgrade rats at selective ages. 2. To determine the kinetics of Tf and non-Tf mediated Fe influx across blood-brain barrier and CP in wild type and Belgrade rats at selective ages. These Aims will be achieved by using the brain perfusion and clearance techniques and the Belgrade rat (b/b) since it is a physiological knock out for DNT1. The kinetic parameters of Tf and non Tf bound Fe will be determined in b/b/ and control rats to establish the role of DMT1 in the influx and efflux of Fe at the blood-brain barrier and CP. Similar studies in control rats at selected ages will determine age related changes. Relative levels of TfR and DMT1 at blood-brain barrier and CP will be determined by standard molecular techniques to complement the transport studies. This study will help us understand the roles of DMT1 and TfR at the BBB and CP with respect to iron accumulation in the brain, associated neurodegenerative effects, and identify potential targets for future development of brain iron lowering strategies.