Nagaoka I, Trapnell B C, Crystal R G
Pulmonary Branch, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, Maryland 20892.
J Clin Invest. 1990 Feb;85(2):448-55. doi: 10.1172/JCI114458.
Activated macrophages release tissue forms of insulin-like growth factor I (IGF-I), 20-25-kD products of the IGF-I gene, thus providing an extracellular growth and differentiation signal at sites of inflammation. To examine the control of IGF-I gene expression in mononuclear phagocytes, the human macrophage-like cell line U937 was evaluated at rest and after surface activation with phorbol myristate acetate (PMA) or Ca2+ ionophore. Northern analysis and RNAse protection analysis with 32P-labeled IGF-I-specific probes demonstrated that the IGF-I mRNA transcripts of resting U937 cells were similar in size and amount to those of resting human alveolar macrophages, mononuclear phagocytes known to express the IGF-I gene. Nuclear run-off assays demonstrated that surface activation of U937 cells increased the transcription rate of the IGF-I gene four- to fivefold, a process that was inhibited by cycloheximide, suggesting that active protein synthesis was involved in the activation pathway. Despite this, cytoplasmic IGF-I mRNA levels after surface activation declined markedly, a process blocked by a protein kinase C inhibitor (for PMA activation) or a calmodulin antagonist (for Ca2+ ionophore activation). Like the increased transcription of the IGF-I gene, modulation of IGF-I mRNA transcript levels required active protein synthesis; in the presence of cycloheximide constitutive IGF-I mRNA levels increased and surface activation no longer caused a decrease in transcript number. Interestingly, surface activation caused a rapid release of IGF-I, even in the presence of a protein synthesis inhibitor, suggesting that mononuclear phagocytes have a preformed, stored, releasable pool of IGF-I. Together these observations demonstrate that IGF-I gene expression is complex and probably involves control of transcription rate, cytoplasmic mRNA levels possibly mediated through protein kinase C, calcium influx and calmodulin, and finally, release of preformed IGF-I from a storage pool.
活化的巨噬细胞释放胰岛素样生长因子I（IGF-I）的组织形式，即IGF-I基因的20 - 25kD产物，从而在炎症部位提供细胞外生长和分化信号。为了研究单核吞噬细胞中IGF-I基因表达的调控，对人巨噬细胞样细胞系U937在静息状态以及用佛波酯（PMA）或钙离子载体进行表面活化后进行了评估。用32P标记的IGF-I特异性探针进行的Northern分析和RNA酶保护分析表明，静息U937细胞的IGF-I mRNA转录本在大小和数量上与静息人肺泡巨噬细胞（已知表达IGF-I基因的单核吞噬细胞）的相似。核转录分析表明，U937细胞的表面活化使IGF-I基因的转录速率提高了4至5倍，这一过程被放线菌酮抑制，表明活性蛋白质合成参与了活化途径。尽管如此，表面活化后细胞质IGF-I mRNA水平显著下降，这一过程被蛋白激酶C抑制剂（用于PMA活化）或钙调蛋白拮抗剂（用于钙离子载体活化）所阻断。与IGF-I基因转录增加一样，IGF-I mRNA转录本水平的调节也需要活性蛋白质合成；在放线菌酮存在的情况下，组成型IGF-I mRNA水平升高，表面活化不再导致转录本数量减少。有趣的是，即使存在蛋白质合成抑制剂，表面活化仍会导致IGF-I快速释放，这表明单核吞噬细胞有一个预先形成的、储存的、可释放的IGF-I池。这些观察结果共同表明，IGF-I基因表达是复杂的，可能涉及转录速率的控制、可能通过蛋白激酶C、钙内流和钙调蛋白介导的细胞质mRNA水平，以及最终从储存池中释放预先形成的IGF-I。
