Division of Pulmonary and Critical Care Medicine, Department of Medicine, University of Michigan Health Systems, Ann Arbor, Michigan, United States of America.
PLoS One. 2010 Nov 11;5(11):e13962. doi: 10.1371/journal.pone.0013962.
Cyclic AMP (cAMP) regulates many biological processes and cellular functions. The importance of spatially localized intracellular gradients of cAMP is increasingly appreciated. Previous work in macrophages has shown that cAMP is produced during phagocytosis and that elevated cAMP levels suppress host defense functions, including generation of proinflammatory mediators, phagocytosis and killing. However, the spatial and kinetic characteristics of cAMP generation in phagocytosing macrophages have yet to be examined. Using a Förster resonance energy transfer (FRET)-based cAMP biosensor, we measured the generation of cAMP in live macrophages. We detected no difference in bulk intracellular cAMP levels between resting cells and cells actively phagocytosing IgG-opsonized particles. However, analysis with the biosensor revealed a rapid decrease in FRET signal corresponding to a transient burst of cAMP production localized to the forming phagosome. cAMP levels returned to baseline after the particle was internalized. These studies indicate that localized increases in cAMP accompany phagosome formation and provide a framework for a more complete understanding of how cAMP regulates macrophage host defense functions.
环磷酸腺苷（cAMP）调节许多生物过程和细胞功能。细胞内 cAMP 局部浓度梯度的重要性越来越受到重视。巨噬细胞中的先前研究表明，cAMP 在吞噬作用过程中产生，并且升高的 cAMP 水平抑制宿主防御功能，包括促炎介质的产生、吞噬作用和杀伤作用。然而，吞噬作用的巨噬细胞中 cAMP 生成的空间和动力学特征尚未被检测到。使用基于Förster 共振能量转移（FRET）的 cAMP 生物传感器，我们测量了活巨噬细胞中 cAMP 的生成。我们在静止细胞和主动吞噬 IgG 包被颗粒的细胞之间未检测到细胞内 cAMP 水平的差异。然而，生物传感器的分析显示，FRET 信号的快速下降对应于形成的吞噬体中 cAMP 产生的短暂爆发。在颗粒被内化后，cAMP 水平恢复到基线。这些研究表明，cAMP 的局部增加伴随着吞噬体的形成，并为更全面地了解 cAMP 如何调节巨噬细胞宿主防御功能提供了框架。
