A MACROPHAGE ENDOGENOUS FACTOR THAT SUPPRESSES ANABOLISM

我们正在调查分子事件， 引发感染的生化后遗症 一种临床 患有慢性感染或肿瘤的动物的特征是 存在可发展为恶病质的分解代谢状态， 震惊和死亡 这种现象的生物化学基础是 不被理解，但据推测一旦触发是一个普遍的 自然 为了深入了解这一机制， 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其研究。 从巨噬细胞系RAW 264.7， 是一种介质的来源， 当加入3 T3-L1脂肪细胞时， 脂蛋白脂酶在合成水平上的活性。 这 纯化至均一的介质被称为恶病质素， 证实与肿瘤坏死因子（TNF）相同。 是 我们的目标是了解TNF 调节细胞代谢。 要做到这一点，关键是首先 表征TNF对脂蛋白脂酶mRNA的影响 水平，该消息的半衰期以及 转录。 我们还将确定TNF是否影响 脂肪酶的分泌。 我们已经观察到TNF刺激 3 T3-L1成纤维细胞的葡萄糖摄取率为250%，而在完全- 分化的脂肪细胞己糖摄取被抑制50%。 葡萄糖进入细胞的这种调节将被表征 通过定量转运蛋白的动力学水平，以及 平衡结合和光亲和标记。 我们还将 检查对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因子的存在， RAW 264.7细胞的条件培养基， L 6大鼠糖原消耗和随后的己糖摄取 培养中的肌管。 我们计划纯化这种单核细胞因子， 并量化其对以下方面的影响：己糖摄取、细胞 糖原水平，乳酸盐的产生，并检查 通过结合和光亲和性测定葡萄糖转运蛋白的水平 标签研究。 我们还将确定，如果数量的变化， 是由于运输水平的变化， mRNA和转录速率。 这些研究将有助于我们 了解这些单核因子是如何产生的 免疫系统暴露于侵入性刺激，能够 调节能量储存组织的新陈代谢。

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.