Mechanisms of TNFa Enhancement of Nicotinic Receptor Upregulation

描述(由申请人提供)：尼古丁通过与神经元烟碱型乙酰胆碱受体(NAChR)相互作用而对细胞功能产生影响。其中一些受体，特别是那些由？4？2亚基组成的受体，通过高亲和力尼古丁结合位点密度显著增加所定义的上调过程，对持续的配体暴露做出反应。上调的生理后果与许多不同的过程有关，从成瘾到导致阿尔茨海默病早期的病理生理过程。在与促炎细胞因子的相互作用方面，nAChRs正在发挥一个令人兴奋的、相对较新的作用。事实上，尼古丁的抗炎特性，通过各种nAChRs发挥作用，现在被广泛报道影响许多炎症病因学的疾病。然而，对于控制炎症的机制：nAChR相互作用，人们知之甚少。这项建议的总体目标是定义细胞内机制，以调节关键的促炎细胞因子和确定的nAChR亚型组合之间的相互作用。特异性目的1.假设：肿瘤坏死因子？-激活p38MAPK通路和/或IL-1？PI3K/Akt信号通路可拮抗PKA通路的激活，可增强β4？2的表达。在这个目标中，我们将继续定义由肿瘤坏死因子？和/或IL-1？为了增强尼古丁介导的nAChR？4？2.特异性目的2.假设：？4的直接磷酸化和其他nAChR亚基的存在改变了促炎细胞因子信号，介导了？4？2的上调。这一目的将决定以下影响：1)修改？4中的PKA磷酸化位点；2)引入(或删除)其他nAChR(特别是？7或？5)；以及3)确定肿瘤坏死因子的作用？还是IL-1？细胞内信号导致培养神经元增强？4？2上调。具体目的3.假设：中枢神经系统细胞如神经元和小胶质细胞(Mg)之间的相互作用直接影响整个炎症反应的结果：nAChR反应。将神经元和小胶质细胞以不同的比例共培养，分析旁分泌、自分泌或旁分泌相互作用在调节肿瘤坏死因子？IL-1？介导的混合细胞系统中？4？2-上调。 公共卫生相关性：我们有兴趣了解尼古丁和炎症之间的相互作用。我们将确定细胞因子用来促进或抑制尼古丁受体上调的信号转导途径。两种主要的细胞因子，肿瘤坏死因子α(TNF？)那白介素1呢？(IL-1？)增强尼古丁诱导的高亲和力尼古丁受体nAChR？4？2的上调。上调反应与成瘾行为和多种脑部病理如阿尔茨海默病有关。了解炎症系统和对尼古丁的反应是如何相互作用的，这将影响我们如何使用治疗剂来处理这些问题。

DESCRIPTION (provided by applicant): Nicotine imparts its effects on cellular function through interaction with neuronal nicotinic acetylcholine receptors (nAChR). Some of these receptors, especially those composed of ?4?2 subunits, respond to sustained ligand exposure through the process of upregulation as defined by a substantial increase in the density of high affinity nicotine binding sites. The physiological consequences of upregulation have been linked to many diverse processes ranging from addiction to pathophysiological processes contributing to early phases of Alzheimer's disease. An exciting and relatively new role for nAChRs is emerging with regard to the interaction with pro-inflammatory cytokines. In fact, the anti-inflammatory properties of nicotine, acting through various nAChRs, is now widely reported to influence many diseases of inflammatory etiology. However, relatively little is known about the mechanisms controlling the inflammatory:nAChR interaction. The overall goal of this proposal is to define intracellular mechanisms that regulate the interactions between key pro-inflammatory cytokines and defined combinations of nAChR subtypes. SPECIFIC AIM 1. Hypothesis: TNF?-activation of the p38MAPK pathway and/or IL-1? activation of PKA pathways enhance ?4?2 expression and these are antagonized by pathways involving PI3K/Akt signaling. In this Aim we will pursue the definition of intracellular pathways initiated by TNF? and/or IL-1? to enhance nicotine-mediated upregulation of nAChR??4?2. SPECIFIC AIM 2. Hypothesis: Direct phosphorylation of ?4 and the presence of other nAChR subunits modify the pro-inflammatory cytokine signals mediating enhanced upregulation of ?4?2. This Aim will determine the impact of: 1) modifying PKA phosphorylation sites in ?4; 2) introducing (or deleting) other nAChRs (specifically ?7 or ?5); and 3) determining effects of TNF? or IL-1? intracellular signaling leading to enhanced ?4?2 upregulation in cultured neurons. SPECIFIC AIM 3. Hypothesis: Interactions between cells of the CNS such as neurons and microglia (Mg) direct the outcome of the overall inflammatory:nAChR response. Neurons and microglia will be co-cultured in different ratios to dissect the relative contribution of paracrine, autocrine or juxtacrine interactions in modulating the TNF? and IL-1?-mediated enhancement of ?4?2-upregulation in a mixed cell system. PUBLIC HEALTH RELEVANCE: We are interested in understanding the interaction between nicotine and inflammation. We will determine the signal transduction pathways cytokines use to promote or inhibit enhanced upregulation of nicotinic receptors. Two major cytokines, tumor necrosis factor alpha (TNF?) and interleukin-1? (IL-1?) enhance nicotine-induced upregulation of the high affinity nicotine receptor termed nAChR?4?2. The upregulation response is correlated with addiction behaviors and multiple pathologies of the brain such as Alzheimer's disease. Understanding how the inflammatory system and response to nicotine interact will impact upon how we approach these problems with therapeutic agents.