A MACROPHAGE ENDOGENOUS FACTOR THAT SUPPRESSES ANABOLISM

我们正在对分子事件进行调查， 引发感染的生化后遗症。其中一位临床医生 患有慢性感染或肿瘤的动物的特征是 存在分解代谢状态，可发展为恶病质， 震惊和死亡。这种现象的生化基础是 不被理解，但可能一旦被触发，就是普遍的 大自然。为了深入了解这一现象的机制 过程中，我们选择内毒素血症作为模型系统 已经将细胞培养技术应用于它的研究。 巨噬细胞系的条件培养液，RAW 264.7， 在暴露于内毒素后是一种介体的来源 当添加到3T3-L1脂肪细胞时，显着抑制 合成水平上的脂蛋白脂酶活性。这 纯化至均一的中介物称为cachectin和 被证明与肿瘤坏死因子(TNF)相同。它是 我们的目标是了解肿瘤坏死因子 调节细胞新陈代谢。要做到这一点，首先 肿瘤坏死因子对脂蛋白脂酶基因表达的影响 级别、该消息的半衰期以及 抄写。我们还将确定肿瘤坏死因子是否会影响 脂肪酶的分泌。我们观察到，肿瘤坏死因子能刺激 3T3-L1成纤维细胞对葡萄糖的摄取增加250%，而在完全... 分化的脂肪细胞的己糖摄取被抑制了50%。 葡萄糖进入细胞的这种调节将被描述为 通过在运动学上量化转运蛋白水平，以及通过 平衡结合和光亲和标记。我们还将 检测对mRNA水平和转录速率的影响。 我们已经定义了第二个单核细胞在 刺激原始264.7细胞的条件培养液 L6大鼠糖原耗竭和随后的己糖摄取 培养中的肌肉肌管。我们计划将这一单核细胞纯化为 均一性和量化其对：己糖摄取，细胞 糖原水平，乳酸的产生，并检查 结合和光亲和力对葡萄糖转运蛋白水平的影响 标记研究。我们还将确定数字是否发生变化 转运者的数量是由于转运者水平的变化 M RNA和转录速率。这些研究将有助于我们 了解这些单核细胞是如何在 免疫系统暴露在侵入性刺激下，能够 调节储能组织的新陈代谢。

We are pursuing an investigation of the molecular events that trigger the biochemical sequelae of infection. One of the clinical hallmarks of animals with chronic infections or tumors is the presence of a catabolic state which can proceed to cachexia, shock, and death. The biochemical basis for this phenomenon is not understood but presumably once triggered is of a universal nature. In order to gain insight into the mechanism of this process, we have selected endotoxemia as a model system and have applied cell culture techniques to its investigation. conditioned medium from the macrophage cell line, RAW 264.7, after exposure to endotoxin was the source of a mediator that when added to 3T3-L1 adipocytes markedly suppressed the activity of lipoprotein lipase at the level of synthesis. This mediator purified to homogeneity was termed cachectin and demonstrated to be identical to tumor necrosis factor (TNF). It is our objective to understand the mechanism by which TNF regulates cellular metabolism. To do this it is critical to first characterize the effect of TNF on lipoprotein lipase mRNA levels, the half-life of that message as well as rates of transcription. We will also determine if TNF affects the rate of secretion of the lipase. We have observed that TNF stimulates glucose uptake by 250% in 3T3-L1 fibroblasts while in the fully- differentiated adipocytes hexose uptake is suppressed by 50%. This regulation of glucose entry into the cell will be characterized by quantifying levels of transporter kinetically, as well as by equilibrium binding and photoaffinity labeling. We will also examine the effects on mRNA levels and rates of transcription. we have defined the presence of a second monokine in the conditioned medium of the RAW 264.7 cells that stimulates glycogen depletion and subsequently hexose uptake in L6 rat muscle myotubes in culture. We plan to purify this monokine to homogeneity and quantify its effects on: hexose uptake, cellular glycogen levels, lactate production, and examine changes in the levels of the glucose transporter by binding and photoaffinity labeling studies. We will also determine if changes in the number of transporters are due to changes in the levels of transporter mRNA and rates of transcription. These studies will aid in our understanding of how these monokines, produced when the immune system is exposed to invasive stimuli, are capable of regulating metabolism in energy storage tissues.