A Mechanistic Study of Methamphetamine Neurotoxicity: Involvement of HO-1 & MnSO

项目概述：本研究的目的是帮助弥合甲基苯丙胺（MA）神经毒性与有效治疗干预之间的差距。使用缺乏DA转运体的D1多巴胺（DA）受体表达的神经上皮瘤细胞系（SK-N-MC），我们开发了一种体外系统，可以模拟MA毒性直接靶向的突触后神经元。利用这一模型，我们将研究ma诱导神经毒性的新分子机制，特别是血红素加氧酶-1 （HO-1）、活性氮物种（RNS）和锰超氧化物歧化酶（MnSOD）的作用。我们假设DA刺激通过HO-1介导的过程，通过MnSOD的硝化作用导致RNS的产生和线粒体介导的细胞凋亡。我们的假设将在以下三个特定目的中得到验证：1)测试HO-1表达是否参与RNS的产生，2)测试HO-1抑制是否减弱da诱导的线粒体介导的细胞凋亡，以及3)确定MnSOD是否被硝化，以及用一般抗氧化剂或模拟物治疗是否减弱da诱导的硝化和/或线粒体介导的细胞凋亡。我们将通过研究旨在控制HO-1和MnSOD活性以限制RNS的产生和随后的凋亡来研究降低MA神经毒性的方法。这将使用创新的方法，如sirna介导的HO-1表达的敲除，或通过药理学模拟物增加MnSOD的催化活性，以及利用先前在其他疾病状态中证明具有临床相关性的化合物，如Sn （IV）介卟啉IX。这些发现将有助于开发新的治疗方法，当作为一种预处理时，将最大限度地减少对那些与MA成瘾疾病作斗争的人的进一步神经损伤。

DESCRIPTION (provided by applicant): Project Summary: The goal of this research is to help bridge the gap between methamphetamine (MA) neurotoxicity and effective therapeutic intervention. Using a D1 dopamine (DA) receptor-expressing neuroepithelioma cell line (SK-N-MC) that lacks DA transporters, we have developed an in vitro system that models post-synaptic neurons directly targeted by MA toxicity. By utilizing this model we will study novel molecular mechanisms involved in MA-induced neurotoxicity, in particular the contribution of heme oxygenase-1 (HO-1), reactive nitrogen species (RNS) and manganese superoxide dismutase (MnSOD). We hypothesize that DA stimulation, via a process mediated by HO-1, leads to generation of RNS and mitochondrial-mediated apoptosis through nitration of MnSOD. Our hypothesis will be tested in the following three specific aims: 1) Test whether HO-1 expression is involved in RNS production, 2) Test whether inhibition of HO-1 attenuates DA-induced mitochondrial-mediated apoptosis, and 3) Determine if MnSOD is nitrated and if treatment with a general antioxidant or mimetic attenuates DA-induced nitration and/or mitochondrial-mediated apoptosis. We will investigate ways to decrease MA neurotoxicity through studies aimed at manipulation of HO-1 and MnSOD activity in order to limit RNS production and subsequent apoptosis. This will be done using innovative methods such as siRNA-mediated knock-down of HO-1 expression, or increased MnSOD catalytic activity via pharmacological mimetics as well as utilizing compounds that have previously demonstrated clinical relevance in other disease states, such as Sn (IV) mesoporphyrin IX. These findings will enable the development of new therapeutic approaches that, when given as a pre-treatment, will minimize further neurological damage to those struggling with the disease of MA addiction. PUBLIC HEALTH RELEVANCE: Methamphetamine is a particularly devastating drug as its abuse has been shown to cause localized brain damage equivalent to that seen in patients with early dementia and greater than that seen in those with schizophrenia. In order to develop effective therapeutics to combat the brain damage seen in methamphetamine addicts, it is essential to first understand how the brain damage occurs. This research proposal is aimed at determining, on a molecular level, what happens inside brain cells to cause them to die after being exposed to methamphetamine. In light of this new-found information, the scientific community will be closer to developing new and effective treatments for methamphetamine toxicity.