Mediators of Lithium Action in Olfactory Epithelium

描述（由申请人提供）：双相情感障碍（BD）是一种慢性衰弱性疾病，具有巨大的社会负担。我们开发BD改进疗法的能力的主要障碍是我们对其病理生理学的理解不完整。用于治疗BD的锂改变了大量基因的稳态mRNA水平;特别是GSK-32、肌醇单磷酸酶（IMPase）、MIP合酶、Bcl-2、Akt、蛋白激酶C、MARCKS、胰岛素样生长因子结合蛋白-2（IGFBP-2）和ERK。但是，这些分子变化如何引起临床反应（如果有的话）仍然是未知的。然而，研究锂在活的BD患者的中枢神经（CNS）组织中的细胞和分子效应是不可行的。虽然死后的大脑已被用于生物标志物的研究，他们是不够的捕获状态依赖的分子特征的BD。血细胞可能不能充分反映神经元细胞中的分子分布。在这项研究中，我们建议使用嗅觉上皮（OE）通过鼻活检，结合新的激光捕获显微切割，以阐明前瞻性确定的临床反应锂的分子和细胞标志物。随后将确定从相同受试者获得的淋巴母细胞中这些分子模式的平行分布，以用于血液标志物的可能临床和预后效用。OE是CNS的一个独特部分，在整个生命过程中不断再生并分化为成熟的嗅觉受体神经元（ORN）。这提出了一种途径，通过检查的密度变化的增殖（p75 NGRF+）的神经元前体和成熟（嗅觉标志物蛋白+）的神经元（替代标记物的神经发生和神经元存活的OE）检查BD的神经发育假说。同样特别有趣的是，与锂作用相关的基因失调，因此BD可能反映在嗅觉功能的缺陷中。因此，结合嗅觉组织情绪稳定的分子机制和嗅觉功能的检测，可以加深我们对BD病理生理的理解，并为BD的其他病因学研究提供新的数据来源。此外，在淋巴母细胞（LB）中验证基因（来自OE研究）的平行研究，通过确认蛋白质研究证实，可能会抢占未来血液标志物的临床和预后效用。在本项目的具体目标1中，我们建议使用我们成功招募的BD受试者方法，确定40例未用药、不吸烟的BD急性发作受试者和20例性别、年龄和种族匹配的不吸烟正常对照（CTL）。在具体目标2中，我们将：a）比较BD和CTL在通过RT-PCR测量的（上述）9个锂作用候选基因的mRNA水平上的差异，和B）检查BD受试者中mRNA水平的锂相关变化对临床应答（LR）的影响。为了进一步阐明特定目标3中情绪稳定的细胞机制，我们将采用免疫组织化学技术来检测针对增殖的低亲和力神经生长因子受体（p75 NGFR）、有丝分裂后未成熟神经元和神经元的细胞类型特异性抗体的密度。（生长相关蛋白43 [GAP 43]）和嗅觉标记蛋白（OMP），以确定增殖神经元前体和成熟神经元（分别为神经发生和神经元存活的替代标志物）的密度增加是否是锂相关分子改变对LR的影响的介导物。在其他研究中，我们将：1）检查BD和CTL之间嗅觉的差异; 2）确定基线基因活性和嗅觉功能之间的关联，以及性别对这些关联的影响; 3）确定基线嗅觉功能和锂相关嗅觉变化作为BD严重性和情绪稳定性的替代标志物的效用;（4）确定OE和LB在锂作用的分子标记上的平行配置。这些目标的实现将促进我们对BD病理生理学的理解，促进对情绪稳定具有特定预期效果的新型和改进的治疗方法的开发，并为科学家团队进一步研究BD的其他病理学产生新的数据资源。

DESCRIPTION (provided by applicant): Bipolar Disorder (BD) is a chronic debilitating illness with substantial societal burden. A major impediment in our ability to develop improved therapeutics for BD is the fact that our understanding of its pathophysiology is incomplete. Lithium, used for treatment of BD, alters steady-state mRNA levels of a large number of genes; notably GSK-32, inositol monophosphatase (IMPase), MIP Synthase, Bcl-2, Akt, Protein Kinase C, MARCKS, insulin-like growth factor binding protein-2 (IGFBP-2), and ERK. But it remains unknown, how any of these molecular changes bring about clinical response, if, at all. Hitherto, studying cellular and molecular effects of lithium in the central nervous (CNS) tissues of living BD patients has been infeasible. Although postmortem brains have been useful for studies of biomarkers, they are insufficient to capture state-dependent molecular signature of BD. Blood cells may not reflect adequately, molecular disposition in neuronal cells. In this study, we propose use of olfactory epithelium (OE) via nasal biopsy, combined with novel laser-captured microdissection to elucidate molecular and cellular markers of prospectively-determined clinical response to lithium. This will be followed by determination of parallel dispositions of these molecular patterns in lymphoblasts obtained from same subjects, for possible clinical and prognostic utility of blood markers. OE is a unique part of the CNS that continually regenerates and differentiates into mature olfactory receptor neurons (ORNs) throughout life. This presents an avenue to examine the neurodevelopmental hypotheses of BD through examination of changes in the densities of proliferating (p75NGRF+) neuronal precursors and mature (olfactory marker protein+) neurons (surrogate markers of neurogenesis and neuronal survival in OE). Also of particular interest, dysregulation of genes relevant to lithium action, and therefore BD may be reflected in deficits in olfactory functions. Therefore, combined study of molecular mechanisms underlying mood stabilization in olfactory tissue and tests of olfactory function could advance our understanding of the pathophysiology of BD and generate new data resource to study other pathoetiologies of BD. Furthermore, parallel studies of validated genes (from OE studies) in lymphoblasts (LB), confirmed by confirmed protein studies, may preempt future clinical and prognostic utility of blood markers. In specific aim 1 of this project, we propose to use our successful recruitment approach of BD subjects, to ascertain 40 non-medicated, non-smoking subjects with acute episodes of BD and 20 gender-, age-, and race- matched non-smoking normal controls (CTL) for this study. In specific aim 2, we will: a) compare BD and CTL on mRNA levels of (above-stated) 9 candidate genes of lithium action measured via RT-PCR, and b) examine the effect of lithium-associated changes in mRNA levels on clinical response (LR) in the BD subjects. To further illuminate cellular mechanisms of mood stabilization in specific aim 3, we will employ immunohistochemical techniques to examine densities of cell type-specific antibodies directed at the proliferating low-affinity nerve growth factor receptor (p75NGFR), postmitotic immature neurons (growth- associated protein 43 [GAP43]), and olfactory marker protein (OMP) in OE samples of all participants pre- and post-treatment, to determine if increased density of proliferating neuronal precursors and mature neurons (surrogate markers of neurogenesis and neuronal survival, respectively) are mediators of the effect of lithium- associated molecular alterations on LR. In other studies, we will: 1) examine differences in olfaction between BD and CTLs; 2) determine the association between baseline gene activity and olfactory function, and the impact of gender on these associations; 3) determine the utility of baseline olfactory function and lithium- associated changes in olfaction as surrogate markers of BD severity and mood stabilization; and 4) determine parallel dispositions between OE and LB on molecular markers of lithium action. Accomplishment of these aims will advance our understanding of the pathophysiology of BD, facilitate development of novel and improved therapeutics with specific desired effect on mood stabilization, and generate new data resource for team of scientists to further investigate other pathoetiologies of BD.