INTERLEUKIN 13 STIMULATION OF HUMAN MONOCYTES

单核细胞是白细胞的一个子集，对正常的免疫功能至关重要。它们是炎症反应的重要组成部分，被认为是动脉粥样硬化病变病理的重要组成部分。在动脉粥样硬化病变中，单核细胞/巨噬细胞表达一种脂质氧化酶，15-脂氧合酶（15-LO），可产生有效的炎症介质。这种酶的独特产物是氧化亚油酸，我们已经证明它是动脉粥样硬化病变中主要的氧化脂肪酸。正常单核细胞不表达15-LO，但暴露于细胞因子IL-13会诱导15-LO表达，该酶的表达被认为与动脉粥样硬化动物模型中的疾病进展有关。本申请的研究将探索IL-13在原代人单核细胞中触发的信号转导通路，并将其与15-LO的下游表达联系起来。Aim I的研究将表征人单核细胞中的IL-13受体复合物。接下来，在Aim 2中，将评估Jaks与这些受体成分的关联和激活，并确定激活状态。将评估Ser/Thr激酶的激活（在Aim 3中），扩展我们的初步数据，即MAP激酶被IL-13激活。Jaks在这一激活过程中的作用将被研究。Aim 4的研究将探讨Ser/Thr磷酸化在调控Jak/Stat磷酸化及其功能中的作用。每种激活途径与15-脂氧合酶表达的生物学相关性以及单核细胞介导的LDL氧化的相关增强是Aim 5中描述的研究要研究的主题。在这一最终目标中，将进行进一步的研究，以确定已确定的IL-13反应途径对一般单核细胞功能的作用。识别选择性参与15-LO表达的信号通路，但不需要其他必要的单核细胞功能，如吞噬和超氧阴离子的产生。这些研究的新颖之处在于，由于单核细胞非常有利于反义寡脱氧核糖核苷酸对特定蛋白质表达的操纵，因此采用了独特的方法来专门剖析这些通路在完整的人类单核细胞中的作用。这些研究将大大提高我们对单核细胞暴露于IL-13诱导的信号机制的理解，并可能提出调节这种反应的特异性和选择性方法。

Monocytes are a subset of white blood cells critical for normal immune function. They are important components of inflammatory responses and are believed to significantly contribute to atherosclerotic lesion pathology. In atherosclerotic lesions, monocyte/macrophages have been shown to express a lipid oxidizing enzyme, 15-lipoxygenase (15-LO), that produces potent inflammatory mediators. Unique among the products of this enzyme are oxidized linoleate which we have shown to be the predominant oxidized fatty acid in atherosclerotic lesions. Normal monocytes do not express 15-LO, but are induced to do so by exposure to the cytokine IL-13 and expression of this enzyme is believed to correlate with disease progression in animal models of atherosclerosis. The studies proposed in this application will explore the IL-13 triggered signal transduction pathways in primary human monocytes and relate them to the downstream expression of 15-LO. Studies in Aim I will characterize the IL-13 receptor complex in human monocytes. Next, in Aim 2, the association and activation of Jaks with these receptor components will be assessed and activated Stats will be identified. The activation of Ser/Thr kinases will be evaluated (in Aim 3), expanding on our preliminary data that MAP kinases are activated by IL-13. The role of Jaks in this activation process will be investigated. Studies in Aim 4 will explore the role of Ser/Thr phosphorylation in regulating Jak/Stat phosphorylation and function. The biologic relevance of each of the activated pathways to the expression of 15-lipoxygenase and the related enhancement of monocyte-mediated LDL oxidation is the topic to be examined in studies described in Aim 5. In this final aim, additional investigations will be conducted to determine the role of the identified, IL-13 responsive pathways to general monocyte function with the goal.of identifying signaling pathways that are selectively involved in 15-LO expression but not required for other essential monocyte functions such as phagocytosis and superoxide anion production. The novelty of these studies is derived from the unique approaches to be employed to specifically dissect the roles of these pathways in intact human monocytes by virtue of the fact that monocytes are very conducive to antisense oligodeoxyribonucleotide manipulation of specific protein expression. These studies will substantially enhance our understanding of the signaling mechanisms induced upon exposure of monocytes to IL-13 and may suggest specific and selective means for modulating this response.