Glucocorticoid/cytokine mechanisms in endotoxemia.

描述（由申请方提供）：本提案中待检验的假设是糖皮质激素（GC）对脂多糖（LPS）诱导的单核细胞/巨噬细胞（MOMac）激活程序的关键组分的控制发挥双相作用。PI和Co-I提出了一个模型，以协调中等皮质醇水平的刺激（“允许”）作用与高皮质醇水平的抑制作用。该模型预测皮质醇对炎症介质的影响呈钟形曲线，峰值位于或接近皮质醇昼夜变化的上限。由于GC的允许作用不如抑制作用那么好理解，它们提供了我们提出的MOMac激活的介质和调节剂的研究重点。具体目标是：（1）验证GC对LPS诱导的细胞因子产生和MOMac表型产生双相影响的假设;（2）阐明GC对MOMac激活程序的容许和抑制作用的分子机制。我们建议解决这些问题，采用在体外和人体在体内系统。最初，我们将使用淘洗MO的培养物，其将在皮质醇维持在预测为亚容许、容许或抑制的水平的条件下暴露于LPS。我们将使用ELISA和流式细胞仪分析来确定皮质醇和LPS对MOMac激活程序的两个关键组成部分-IL-6和CD 163的相互作用。使用LPS和皮质醇的剂量和时间，首先确定导致CD 163和IL-6的最大增强或抑制，我们将检查其他细胞因子，信号分子和受体，已被证明是由LPS诱导或抑制。我们还将使用一个良好控制的人类实验性内毒素血症的范例来阐明导致GC在体内对内毒素的反应过程中抑制或增强推定的促炎和抗炎分子的产生和作用的生理和分子机制。我们相信，这些研究将产生重要的见解GC可以增强和抑制免疫和炎症功能的机制，导致更有效的方法，他们在炎症的临床环境中使用。

DESCRIPTION (provided by applicant): The hypothesis to be tested in this proposal is that glucocorticoids (GC) exert a biphasic effect on the control of key components of the lipopolysaccharride (LPS)-induced monocyte/ macrophage (MOMac) activation program. A model was proposed by the PI and Co-I to reconcile the stimulatory ("permissive") actions of moderate cortisol levels with the suppressive actions of high cortisol levels. This model predicts a bell-shaped curve for the effects of cortisol on inflammatory mediators that peaks at or near the upper levels of diurnal cortisol variation. As the permissive effects of GC are less well understood than the inhibitory actions, they provide the focus of our proposed studies on mediators and regulators of MOMac activation. The Specific Aims are: (1) To test the hypothesis that GC exert a biphasic influence on cytokine production and MOMac phenotype in response to LPS, and (2) To elucidate the molecular mechanisms of permissive and suppressive effects of GC on the MOMac activation program. We propose to address these questions employing both in vitro and human in vivo systems. Initially, we will use cultures of elutriated MO that will be exposed to LPS under conditions in which cortisol is maintained at levels predicted to be sub-permissive, permissive or suppressive. We will use ELISA and flow cytometric analyses to determine the interactive effects of cortisol and LPS on two key components of the MOMac activation program--IL-6 and CD163. Using LPS and cortisol at dosages and times that are first determined to result in maximal enhancement or inhibition of CD 163 and IL-6, we will examine additional cytokines, signaling molecules and receptors that have been shown to be induced or suppressed by LPS. We will also use a well-controlled paradigm of human experimental endotoxemia to elucidate the physiologic and molecular mechanisms that lead GC to either suppress or enhance the production and actions of putative pro- and anti-inflammatory molecules during the response to endotoxin in vivo. We believe that these studies will yield important insights into the mechanisms by which GC can both enhance and suppress immune and inflammatory functions, leading to more effective approaches to their use in clinical settings of inflammation.