Nicotine and nicotinic receptors in lung transitional remodeling

描述（由申请人提供）： 烟草和酒精是世界上使用和滥用最多的两种药物。它们加在一起，每年造成全世界数百万人死亡。尽管进行了大量的研究来了解这些药物如何影响肺部，但导致临床重大进展的新信息令人失望。由于退伍军人中烟草和酒精暴露的高患病率，罗马实验室正在探索这些药物影响肺部的机制。我们的研究表明，酒精和尼古丁，烟草中存在的生物碱，促进肺基质组成的改变，我们称之为“过渡性”重塑，一个过程的特点是微妙的改变，在相对组成的肺细胞外基质。虽然这些事件与肺部疾病的确切机制仍不完全清楚，但我们认为，过渡性重塑使宿主易患肺部疾病，这一观点得到了体外和体内已发表研究的支持。考虑到上述情况，我们着手确定负责这些事件的细胞/分子机制。使用培养的原代肺成纤维细胞，我们发现，尼古丁诱导细胞增殖和基质的表达，通过激活特定的7烟碱乙酰胆碱受体（7 nAChR）。这些受体的重要性在体内得到证实，表明产前暴露于尼古丁会影响肺发育，并诱导野生型动物的气道增厚和高反应性，但在7 nAChR敲除小鼠中则不然。使用相同的培养模型，我们发现酒精通过另一组nAChR（4个nAChR）作用于肺成纤维细胞。在啮齿类动物的肺部，这些受体的表达增加慢性酒精暴露，这反过来又容易导致急性肺损伤。这项工作表明，胆碱能系统在肺内稳态和对损伤的反应中起着重要作用，并指出nAChRs作为干预的潜在靶点。最近，我们发现nAChRs通过配体依赖性和非依赖性途径激活。具体而言，虽然尼古丁以传统的配体依赖性方式作用于7种nAChR，但酒精至少部分地通过诱导特定形式的氧化应激（半胱氨酸/胱氨酸氧化还原电位或Eh Cys/CySS的氧化）起作用，该氧化应激与策略性地位于某些nAChR五聚体亚基上的半胱氨酸残基一起，通过配体非依赖性作用机制激活nAChR信号传导。重要的是，我们已经产生了初步的数据，表明在老化损伤的易感性可能会通过类似的途径激活nAChR，从而促进过渡性重塑和增加肺损伤的易感性。考虑到上述情况，我们提出衰老（如尼古丁和乙醇暴露所观察到的）与通过不同的nAChR进行的肺过渡性重塑相关。nAChR可以通过配体依赖性和非依赖性途径激活，后者由Eh Cys/CySS氧化诱导的氧化应激触发。具体而言，氧化应激影响策略性地位于nAChR的β亚基中的半胱氨酸，这反过来触发信号转导并促进肺过渡性重塑，并增加肺损伤后对破损的易感性。为了验证这一假设，我们提出了实验设计：目的1。研究特定亚基半胱氨酸在肺细胞和体内nAChR活化中的作用。目标2.确定氧化应激的作用，特别是，在体外和体内nAChR激活的Eh Cys/CySS的氧化。目标3.检查nAChR激活在与衰老相关的肺部疾病中的作用，以及烟草和酒精暴露如何影响这一点。

DESCRIPTION (provided by applicant): Tobacco and alcohol are the two most used and abused drugs in the world. Together, they are responsible for millions of deaths each year worldwide. Despite the enormous amount of research carried out to understand how these agents affect the lung, new information leading to significant advances at the clinic has been disappointing. Because of the high prevalence of tobacco and alcohol exposure in veterans, the Roman lab is exploring the mechanisms by which these agents affect the lung. Our research revealed that both alcohol and nicotine, an alkaloid present in tobacco, promote alterations in lung matrix composition that we have termed 'transitional' remodeling, a process characterized by subtle alterations in the relatively composition of the lung extracellular matrix. Although the exact mechanisms linking these events to lung disease remain incompletely understood, we believe that transitional remodeling renders the host susceptible to lung disease, and this belief is supported by published research performed both in vitro and in vivo. Considering the above, we set out to identify the cellular/molecular mechanisms responsible for these events. Using cultured primary lung fibroblasts, we found that nicotine induces cellular proliferation and matrix expression through the activation of specific 7 nicotinic acetylcholine receptors (7 nAChRs). The importance of these receptors was confirmed in vivo showing that prenatal exposure to nicotine affects lung development and induces airway thickening and hyperreactivity in wildtype animals, but not in 7 nAChR knockout mice. Using the same culture model, we discovered that alcohol acts on lung fibroblasts via another set of nAChRs, 4 nAChRs. In rodent lungs, the expression of these receptors is increased by chronic alcohol exposure which, in turn, predisposes to acute lung injury. This work suggests that the cholinergic system plays an important role in lung homeostasis and response to injury, and points to nAChRs as potential targets for intervention. More recently, we found that nAChRs are activated through both ligand-dependent and -independent pathways. Specifically, while nicotine acts on 7 nAChRs in a traditional ligand-dependent fashion, alcohol works, at least in part, through induction of a particular form of oxidant stress (oxidation of the cysteine/cystine redox potential or Eh Cys/CySS) which, together with cysteine residues strategically located on the subunits of certain nAChR pentamers, activates nAChR signaling via ligand-independent mechanisms of action. Importantly, we have generated preliminary data to suggest that susceptibility to injury in aging might activate nAChRs through similar pathways thereby promoting transitional remodeling and increased susceptibility to lung injury. Considering the above, we propose that aging (as observed for nicotine and ethanol exposure) is associated with lung transitional remodeling via distinct nAChRs. nAChRs can be activated via ligand-dependent and -independent pathways, the latter triggered by oxidant stress induced by oxidation of the Eh Cys/CySS. Specifically, oxidant stress affects cysteines strategically located in the � subunits of nAChRs which, in turn, triggers signal transduction and promotes lung transitional remodeling and increase susceptibility to disrepair after lung injury. To test this hypothesis, we propose experiments designed to: Aim 1. Investigate the role of specific subunit cysteines in nAChR activation in lung cells and in vivo. Aim 2. Determine the role of oxidant stress, especially, oxidation of the Eh Cys/CySS in nAChR activation both in vitro and in vivo. Aim 3. Examine the role of nAChR activation in lung disease related to aging and how this is affected by tobacco and alcohol exposure.