REGULATION OF MACROPHAGE FUNCTION BY PROTEIN KINASES

巨噬细胞可以被刺激到高度增强的状态 功能上具有杀菌和杀瘤活性 定义为激活。细胞内事件，通过它 巨噬细胞的激活是否受到调控尚未有明确的定义。 我们建议系统地定义 与刺激有关的蛋白质磷酸化 巨噬细胞，并鉴定参与这些过程的蛋白激酶 流程。在这些研究中将使用两种不同的刺激； 集落刺激因子与细菌脂多糖 (LP)。脂多糖和脑脊液-1可引起多种常见的功能改变 巨噬细胞，但每一个也诱导独特的功能。为 例如，只有内毒素才能诱导杀瘤活性。为了 确定CSF-1和内毒素如何改变巨噬细胞的活性 将采用药理学和生化方法。这个 药理学方法将利用可能模拟或 抑制内毒素或CSF-1的活性以帮助识别第二种 可能介导这些活动的信使/蛋白激酶系统 脑脊液-1和内毒素水平。生化方法将决定 各自诱导的蛋白质磷酸化的快速变化 刺激物。在初步的药理和生化研究之后 给出了可能的蛋白激酶介导的特定的 行动，涉及的蛋白激酶将被识别。这将是 通过比较体内的磷酸化模式来完成 与通过刺激特定基因获得的磷酸化模式 体外细胞制剂中的蛋白激酶。蛋白激酶 将被刺激的将根据结果选择 药理和磷酸化研究。比较将是 在体外和体内的磷酸化之间 确定磷酸化是否在相同的磷蛋白上 就在那个磷蛋白的同一位置。这些结果将 使我们能够确定磷酸化反应是否被调节 由所研究的特定蛋白激酶组成。希望这将是 帮助定义巨噬细胞功能的机制(S) 受监管的。

Macrophages can be stimulated to states of heightened microbicidal and tumoricidal activity that are functionally defined as activated. The intracellular events by which macrophage activation is regulated have not yet been defined. We propose to systematically define the rapid alterations in protein phosphorylation that are associated with stimulation of macrophages and identify the protein kinases involved in these processes. Two different stimuli will be used in these studies; colony stimulating factor (CSF-1) and bacterial lipopolysaccharide (LPS). LPS and CSF-1 induce many common functional changes in macrophages, but each also induces unique functions. For example, only LPS induces tumoricidal activity. In order to determine how CSF-1 and LPS alter macrophage activity both a pharmacologic and biochemical approach will be employed. The pharmacologic approach will utilize agents that may mimic or inhibit activities of LPS or CSF-1 to help identify the second messenger/protein kinase systems that might mediate the actions of CSF-1 and LPS. The biochemical approach will determine the rapid alterations in protein phosphorylation induced by each stimuli. After initial pharmacologic and biochemical studies have given an indication of possible protein kinases mediating specific actions, the protein kinases involved will be identified. This will be accomplished by comparing the in vivo phosphorylation pattern with the phosphorylation pattern obtained by stimulating specific protein kinases in cell preparations in vitro. The protein kinases that will be stimulated will be chosen based on the results of the pharmacologic and phosphorylation studies. Comparisons will be made between the in vitro and in vivo phosphorylations to determine if the phosphorylations are on the same phosphoprotein and on the same site of that phosphoprotein. These results will allow us to determine if the phosphorylation reaction is mediated by the specific protein kinase studied. It is hoped that this will help define the mechanism(s) by which macrophage function is regulated.