POSTTRAUMATIC SEPSIS--REGULATION OF LPS-BINDING PROTEIN

革兰氏(-)败血症仍然是发病率和死亡率的主要原因 在创伤之后。我们假设革兰氏(-)败血症是在 内毒素与肝细胞合成的一种蛋白质结合 称为内毒素结合蛋白(LBP)。我们还提出，由此产生的 LBP-内毒素复合体与巨噬细胞上的受体结合，然后依赖于 在受体类型上，该复合体要么激活巨噬细胞， 分泌细胞因子(CD14受体)或促进摄取和 内毒素(乙酰化低密度脂蛋白受体)的降解。自肝枯否 已知细胞是循环内毒素的主要部位 摄取，我们认为肝细胞释放的LBP促进摄取 相邻Kupffer细胞对内毒素的影响。很可能细胞因子 由Kupffer细胞释放，然后进一步上调肝细胞LBP 合成创造了一个重要的反馈回路和LBP在 肝脏。目的I：测定细胞外和细胞内 肝细胞内脂多糖结合蛋白的调节我们将从 定义外部信号(例如，细胞因子、糖皮质激素等) 在体外调节肝细胞LBP的合成。我们预测LBP 将以类似的方式对几个描述良好的急性阶段做出反应 反应物。接下来，我们将确定Kupffer细胞在 在体外和体内提供这些信号。除了我们的 关于细胞外信号的研究，我们将研究细胞内信号 LBP的调节机制。我们将确定LBP的合成是否 是转录调控的。如果是这样的话，我们将分离LBP基因 推动者，这将使我们能够详细研究和比较LBP 对上调的其他分泌型肝细胞蛋白的调节 在败血症(急性期反应物)中。目的II：确定功能 内毒素结合蛋白在内毒素与CD14相互作用中的作用 肝细胞上的乙酰化低密度脂蛋白受体。研究伦敦银行间同业拆借利率在 肝脏对内毒素的处理，我们首先需要检查 脂多糖-LBP复合体受体的表达。因此，我们将 肝细胞CD14和乙酰化低密度脂蛋白受体的表达 无论是在静息状态下，还是在脓毒症和炎症条件下。 如果受体的表达在体内受到调节，我们将研究 LBP和其他肝细胞衍生因子在这一调节中。最后， 我们将确定LBP在这些受体摄取内毒素中的作用 在分离的库普弗细胞和整个器官中。我们将确定 LBP-内毒素结合的功能后果(激活与非激活) 到库普弗细胞。在我们的研究完成后，我们将拥有 表征了LBP合成的调节和功能作用 LBP在肝脏内清除内毒素。此信息将提供 对宿主-内毒素相互作用机制的重要见解和 败血症反应的诱导。

Gram (-) sepsis remains a major cause of morbidity and mortality following trauma. We hypothesize that gram (-)sepsis is initiated when lipopolysaccharide (LPS) is bound by a protein synthesized by hepatocytes known as LPS binding protein (LBP). We also propose that the resultant LBP-LPS complex binds to receptors on macrophages which then, depending on the receptor type, the complex either activates the macrophage to secrete cytokines (CD14 receptor) or facilitates the uptake and degradation of LPS (acetylated-LDL receptor). Since the liver Kupffer cells are already known to be the principal site of circulating LPS uptake, we propose that LBP released by hepatocytes promote the uptake of LPS by the adjacent Kupffer cell. It is likely that cytokines released by the Kupffer cells then further upregulates hepatocyte LBP synthesis creating an important feedback loop and function of LBP in the liver. AIM I: TO DETERMINE THE EXTRACELLULAR AND INTRACELLULAR REGULATION OF LPS BINDING PROTEIN IN HEPATOCYTES. We will begin by defining the external signals (e.g., cytokines, glucocorticoids, etc.) which regulate hepatocyte LBP synthesis in vitro. We predict that LBP will respond in a similar manner to several well-described acute phase reactants. Next, we will determine the role of Kupffer cells in providing these signals both in vitro and in vivo. In addition to our studies on the extracellular signals, we will study the intracellular mechanisms of LBP regulation. We will determine if the synthesis of LBP is transcriptionally regulated. If so, we will isolate the LBP gene promoter, which will allow us to study and compare, in detail, LBP regulation with other secreted hepatocyte proteins that are upregulated in sepsis (acute phase reactants). AIM II: TO DETERMINE THE FUNCTIONAL ROLE OF LPS BINDING PROTEIN IN THE INTERACTION OF LPS WITH CD14 AND THE ACETYLATED-LDL RECEPTOR ON LIVER CELLS. To study the role of LBP in the processing of LPS by the liver, we will first need to examine the expression of the receptors for LPS-LBP complexes. Therefore, we will define the expression of CD14 and acetylated-LDL receptors on liver cells both in the resting state and under septic and inflammatory conditions. If receptor expression is regulated in vivo, we will study the role of LBP and other hepatocyte-derived factors in this regulation. Finally, we will determine the role of LBP in the uptake of LPS by these receptors in isolated Kupffer cells and the whole organ. We will determine the functional consequence (activation vs no activation) of LBP-LPS binding to Kupffer cells. At the completion of our studies we will have characterized the regulation of LBP synthesis and the functional role of LBP in LPS clearance within the liver. This information will provide important insights into the mechanisms of host-LPS interaction and the induction of the septic response.