PROCESSING OF ENDOTOXINS BY LIVER MACROPHAGES

革兰氏阴性菌感染是一个主要的临床问题， 循环中的细菌内毒素导致危及生命的全身性 反应。内毒素也参与了许多 包括肝硬变在内的肝病。肝是黄褐斑的主要部位 肠源性内毒素的代谢。这些细胞负责 从门静脉血中清除内毒素的是肝巨噬细胞 (Kupffer细胞)，并与肝细胞合作解毒。 由于持续长期接触低水平的内毒素， 库普弗细胞对内毒素的反应与其他巨噬细胞不同 单核细胞与内毒素摄取和反应的分子细节 Kupffer细胞尚不清楚。为了调查这一点，我们将：1)描述 内毒素与枯否细胞结合所需的结构 细胞。我们已经鉴定了两个新的内毒素结合蛋白31和 34kD的大鼠枯否细胞上的内部蛋白似乎是 参与传递细胞因子反应的信号。 实验的目的是提纯这些蛋白质并测定氨基酸。 酸序列。我们将生产针对31和 使用多抗原肽(MAP)的34kD结合蛋白 技术这些抗体将被用于研究功能性 这些结合蛋白的意义和分布。2)我们会 鉴定、克隆和测序编码31kD和34kD基因 利用多肽序列合成内毒素结合蛋白 特定的寡核苷酸探针。序列数据将用于 为结合蛋白创建潜在的结构域模型并检查 它们与其他蛋白质的结构关系。克隆的基因将 也可用于转染真核细胞并提供一种 模型来研究这些新的内毒素结合蛋白的生物学作用。 3)我们将确定31和34 kD结合蛋白在 与内毒素与Kupffer细胞相互作用相关的信号事件 Kupffer细胞对内毒素的反应中分泌细胞因子。至 研究Mn/++和其他过渡金属的保护作用 抗内毒素及其对内毒素与31和34结合的抑制作用 KD蛋白。这些研究将增加我们对 内毒素的生物学效应及其新的治疗方法 中毒性休克。

Infection due to gram negative bacteria is a major clinical problem and bacterial endotoxins in the circulation cause life threatening systemic reactions. Endotoxin is also involved in the pathology of a number of liver diseases including cirrhosis. The liver is the major site of metabolism for gut derived endotoxins. The cells responsible for endotoxin clearance from the portal blood is the liver macrophage (Kupffer cell) and co-operates with hepatocytes in detoxification. Because of continuous chronic exposure to low levels of endotoxin the Kupffer cell response to endotoxin is different to other macrophages and monocytes and the molecular details of endotoxin uptake and response in Kupffer cells in unclear. To investigate this we will: 1) Characterize the structures necessary for the binding of endotoxin to the Kupffer cells. We have identified two novel endotoxin binding proteins of 31 and 34 kD on rat Kupffer cells that are internal proteins that appear to be involved in transducing the signal for the cytokine response. Experiments are designed to purify these proteins and to determine amino acid sequences. We will produce monoclonal antibodies to both the 31 and 34 kD binding proteins using multiple antigenic peptide (MAP) technology. These antibodies will be used to study the functional significance and distribution of these binding proteins. 2) We will identify and clone and sequence the genes encoding the 31 and 34 kD endotoxin binding proteins using peptide sequences to synthesize specific oligonucleotide probes. The sequence data will be used to create a potential domain model for the binding proteins and to examine their structural relationships to other proteins. The cloned genes will also be available for transfection into eukaryotic cells and provide a model to study the biological role of these novel LPS binding proteins. 3) We will determine the role of the 31 and 34 kD binding proteins in signaling events associated with LPS interaction with Kupffer cells and in the secretion of cytokines by Kupffer cells in response to LPS. To examine the protective effects of Mn/++ and other transition metals against endotoxin and their inhibition of LPS binding to the 31 and 34 kD proteins. These studies will increase our understanding of the biological effects of endotoxins and lead to novel therapies against toxic shock.