Zechner R, Moser R, Newman T C, Fried S K, Breslow J L
Institute of Medical Biochemistry, University of Graz, Austria.
J Biol Chem. 1991 Jun 5;266(16):10583-8.
Apolipoprotein E (apoE) is an important constituent of plasma lipoproteins and a ligand for several lipoprotein receptors. It is produced mainly in the liver but also in several peripheral tissues like brain, adrenal glands, kidney, and macrophages. Some of these tissues also coexpress lipoprotein lipase (LPL), an important enzyme in the metabolism of lipids and lipoproteins. This suggested a possible coordinate expression of these genes and led us to analyze whether adipocytes, a major source of LPL, could also synthesize apoE. Northern blotting experiments showed that apoE mRNA is found in differentiated mouse 3T3-L1 adipocytes as well as biopsies of human adipose tissue maintained in organ culture but not in undifferentiated 3T3-L1 preadipocytes. [35S]Methionine pulse-labeling experiments revealed that apoE protein is produced in human adipose tissue and differentiated mouse 3T3-L1 adipocytes but not in preadipocytes. In biosynthetic labeling experiments, most apoE was found to be cell associated even after prolonged chase periods. Heparin treatment of the cultured cells did not enhance apoE secretion. During differentiation of 3T3-L1 cells, the onset of apoE gene expression was later than that of LPL. The apoE mRNA and intracellular apoE protein concentrations increased linearly with time of differentiation, at least through day 11, whereas LPL showed highest expression at day 7 and then declined. The increase in apoE mRNA correlated with the cellular lipid content. Inhibition of lipid accumulation in differentiated cells by biotin deprivation decreased apoE expression. Cholesterol-loading experiments suggested that apoE mRNA expression is regulated by the intracellular free cholesterol content of 3T3-L1 adipocytes. In contrast, the LPL mRNA level was not influenced by biotin deprivation or cholesterol loading. Human recombinant tumor necrosis factor, a potent inhibitor of LPL gene transcription, had no effect on adipocyte apoE mRNA levels. Therefore, although apoE and LPL are both expressed in adipocytes in a differentiation-dependent manner, the time course of their expression differs as do their responses to cellular lipid content and tumor necrosis factor. We conclude that these genes are not coordinately regulated in adipocytes.
载脂蛋白E（apoE）是血浆脂蛋白的重要组成部分，也是几种脂蛋白受体的配体。它主要在肝脏中产生，但也在一些外周组织中产生，如脑、肾上腺、肾脏和巨噬细胞。这些组织中的一些也共同表达脂蛋白脂肪酶（LPL），这是脂质和脂蛋白代谢中的一种重要酶。这表明这些基因可能存在协同表达，从而促使我们分析脂肪细胞（LPL的主要来源）是否也能合成apoE。Northern印迹实验表明，在分化的小鼠3T3-L1脂肪细胞以及器官培养中保存的人脂肪组织活检样本中发现了apoE mRNA，但在未分化的3T3-L1前脂肪细胞中未发现。[35S]甲硫氨酸脉冲标记实验表明，人脂肪组织和分化的小鼠3T3-L1脂肪细胞中产生了apoE蛋白，但前脂肪细胞中未产生。在生物合成标记实验中，即使经过长时间的追踪期，大多数apoE仍与细胞相关。用肝素处理培养细胞并未增强apoE的分泌。在3T3-L1细胞分化过程中，apoE基因表达的开始时间晚于LPL。apoE mRNA和细胞内apoE蛋白浓度随分化时间呈线性增加，至少在第11天之前是这样，而LPL在第7天表达最高，然后下降。apoE mRNA的增加与细胞脂质含量相关。通过生物素剥夺抑制分化细胞中的脂质积累会降低apoE表达。胆固醇加载实验表明，apoE mRNA表达受3T3-L1脂肪细胞内游离胆固醇含量的调节。相比之下，生物素剥夺或胆固醇加载对LPL mRNA水平没有影响。人重组肿瘤坏死因子是LPL基因转录的有效抑制剂，对脂肪细胞apoE mRNA水平没有影响。因此，尽管apoE和LPL都以分化依赖的方式在脂肪细胞中表达，但它们表达的时间进程不同，对细胞脂质含量和肿瘤坏死因子的反应也不同。我们得出结论，这些基因在脂肪细胞中不是协同调节的。
