ROLE OF GLYCERALDEHYDE-3-PHOSPHATE DEHYDROGENASE IN DEPRESSION & ALCOHOLISM

这个子项目是许多研究子项目中利用 资源由NIH/NCRR资助的中心拨款提供。子项目和 调查员(PI)可能从NIH的另一个来源获得了主要资金， 并因此可以在其他清晰的条目中表示。列出的机构是 该中心不一定是调查人员的机构。 抑郁症和酗酒被越来越多地理解为衰弱的，往往是致命的疾病，并且偶然地比预期更常见地同时发生。在严重抑郁障碍和酒精中毒中，导致观察到的神经细胞和神经胶质细胞丢失的分子机制尚不清楚。甘油醛-3-磷酸脱氢酶(GAPDH)是一种传统的糖酵解酶，近年来的研究主要集中在它在细胞凋亡中的作用。几个研究小组已经报道，当细胞受到压力时，GAPDH会移位到细胞核中。我们的初步数据显示，细胞应激源或乙醇增加了 GAPDH在脑源性细胞系中被激活，并增加了GAPDH与早期生长反应-1(Egr-1)的结合，导致GAPDH/Egr-1复合体的核转位。同时用细胞应激源和乙醇处理脑源性细胞系比单独用细胞应激源或乙醇处理更能增加GAPDH的表达和GAPDH/Egr-1的相互作用。我们的初步数据还表明，与正常对照组相比，患有抑郁症和酗酒的受试者前额叶皮质中的GAPDH水平比 在患有抑郁症或酗酒的受试者中。在目前的应用中，我们提出了一种新的GAPDH介导的 可能涉及GAPDH与Egr-1结合的神经元应激途径。一旦进入细胞核，GAPDHEgr-1复合体就会分离，Egr-1靶向单胺氧化酶B(MAO B)基因，从而增加MAO B的表达，然后MAO B酶促降解一些神经递质，产生有毒的活性氧(H2O2)，导致神经细胞应激，促进细胞死亡或凋亡。 我们推测，GAPDH-Egr-1-MAO-B介导的神经元应激通路参与了抑郁症和酒精中毒的发病机制，并且在抑郁症和酒精中毒的共病中比单独抑郁症或酒精中毒更明显。我们进一步假设，抑制GAPDH-Egr-1-MAO-B介导的神经细胞应激通路将保护脑细胞免受有害应激和乙醇诱导的影响。 我们的具体目标是：(1)通过检测应激或乙醇处理的大鼠前额叶皮质中的凋亡标志物(片段DNA)及其下游靶标Egr-1和MAO-B的水平(与未处理的对照组相比)，来表征GAPDH-Egr-1-MAO-B介导的神经元应激通路中的活性成分；(2)检测抑郁症酒精依赖者死后脑组织中凋亡标志物(片段DNA)、GAPDH及其下游靶标的表达，并与抑郁症组、酒精依赖组和正常对照组进行比较；以及(3)阻断GAPDH-Egr-1-MAO-B介导的神经元应激途径(使用siRNA)，以检验抑制GAPDH将减少细胞应激源和乙醇的有害影响的假设。 这些研究与其他CPN项目相关，因为据报道，过多的谷氨酸信号(子项目2)会触发GAPDH介导的神经细胞死亡级联反应。GAPDH-Egr-1-MAO-B通路可能参与了导致重度抑郁症脑细胞丢失的死亡级联反应(子项目1)。此外，GAPDH还影响MAO的遗传调节，MAO是降解5-羟色胺的关键酶。因此，阐明GAPDH-Egr-1-MAO-B介导的神经元应激/细胞死亡通路可能为了解5-羟色胺系统功能障碍(子项目3和4)提供新的见解，并可能导致治疗抑郁症和酒精中毒的新策略。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Depression and alcoholism are increasingly understood as debilitating and oftentimes fatal disorders, and co-occur more commonly than expected by chance. The molecular mechanisms contributing to observed neuronal and glial cell loss in both major depressive disorder and alcoholism remain unclear. Recent studies have focused on the apoptotic role of glyceraldehyde-3-phosphate dehydrogenase (GAPDH), a traditional glycolytic enzyme. Several groups have reported that GAPDH translocates into the cell nucleus when cells are stressed. Our preliminary data show that cell stressors or ethanol increase the expression of GAPDH in brain-derived cell lines and also increase the binding of GAPDH with early growth response-1 (Egr-1), resulting in nuclear translocation of the GAPDH/Egr-1 complex. Treatment of brain-derived cell lines with both cell stressors and ethanol increases GAPDH expression and the interaction of GAPDH/Egr-1 more than that of treatment with cell stressors or ethanol alone. Our pilot data also demonstrate that when compared to normal control subjects, GAPDH levels are more elevated in the prefrontal cortex of subjects with co-morbid depression plus alcoholism than in subjects with either depression or alcoholism alone. In the current application, we propose a novel GAPDHmediated neuronal stress pathway that may involve GAPDH binding to Egr-1. Once inside the nucleus, the GAPDHEgr-1 complex separates, Egr-1 targets the monoamine oxidase B (MAO B) gene, thus increasing the expression of MAO B. MAO B then enzymatically degrades a number of neurotransmitters, producing toxic reactive oxygen (H2O2) and resulting in neuronal cell stress which promotes cell death or apoptosis. We hypothesize that the GAPDH-Egr-1-MAO-B-mediated neuronal stress pathway contributes to the pathogenesis of depression and alcoholism and is more evident in co-morbid depression and alcoholism than in either depression or alcoholism alone. We further hypothesize that the inhibition of the GAPDH-Egr-1-MAO-Bmediated neuronal stress pathway will protect brain cells from harmful stress- and ethanol-induced effects. Our Specific Aims are: (1) to characterize the components active in the GAPDH-Egr-1-MAO-B -mediated neuronal stress pathway by measuring the apoptotic marker (fragmented DNA) and the levels of Egr-1 and MAO B (the downstream targets of GAPDH) in the prefrontal cortex from stressed or ethanol-treated rats as compared to untreated controls; (2) to examine the expression of apoptotic markers (fragmented DNA), GAPDH and its downstream targets in postmortem brain tissue from depressed alcoholic subjects as compared to depressed subjects, alcohol-dependent subjects and normal control subjects; and (3) to disrupt the GAPDH-Egr-1-MAO-B -mediated neuronal stress pathway (using the siRNA) to test the hypothesis that inhibition of GAPDH will reduce the harmful effects of cell stressors and ethanol. These studies are relevant to other CPN projects, because excessive glutamate signaling (Subproject 2) has been reported to trigger the GAPDH-mediated neuronal cell death cascade. The GAPDH-Egr-1-MAO-B pathway may contribute to this death cascade which causes brain cell loss in major depression (Subproject 1). Furthermore, GAPDH affects the genetic regulation of MAO, a key enzyme in degradation of serotonin. Thus, elucidating the GAPDH-Egr- 1-MAO-B -mediated neuronal stress/cell death pathway may provide new insights into serotonin system dysfunction (Subprojects 3 and 4), and might lead to novel therapeutic strategies for co-morbid depression and alcoholism.