SUMMARY In advanced atherosclerosis, macrophage apoptosis coupled with defective phagocytic clearance of the apoptotic cells (efferocytosis) promotes plaque necrosis, which precipitates acute atherothrombotic cardiovascular events. Oxidative and endoplasmic reticulum (ER) stress in macrophages are important causes of advanced lesional macrophage apoptosis. We now show that pro-apoptotic oxidative/ER stress inducers trigger another stress reaction in macrophages, autophagy. Inhibition of autophagy by silencing ATG5 or other autophagy mediators enhances apoptosis and NADPH oxidase-mediated oxidative stress, while at the same time rendering the apoptotic cells less well recognized by efferocytes. Most importantly, macrophage ATG5 deficiency in fat-fed Ldlr−/− mice increases apoptosis and oxidative stress in advanced lesional macrophages, promotes plaque necrosis, and worsens lesional efferocytosis. These findings reveal a protective process in oxidatively stressed macrophages relevant to plaque necrosis, suggesting a mechanism-based strategy to therapeutically suppress atherosclerosis progression and its clinical sequelae.

Tissue-resident macrophages of barrier organs constitute the first line of defense against pathogens at the systemic interface with the ambient environment. In lung, resident alveolar macrophages (AMs) provide sentinel function against inhaled pathogens1. Bacterial constituents ligate toll-like receptors (TLRs) on AMs2, causing AMs to secrete proinflammatory cytokines3 that activate alveolar epithelial receptors4, leading to recruitment of neutrophils that engulf pathogens5,6. However, since the AM-induced immune response could itself cause tissue injury, it is unclear how AMs modulate the response to prevent injury. Here, through real-time alveolar imaging in situ, we show that a subset of AMs attached to the alveolar wall, formed connexin 43 (Cx43)-containing gap junctional channels (GJCs) with the epithelium. During lipopolysaccharide (LPS)-induced inflammation, the AMs remained alveolus-attached and sessile, and they established intercommunication through synchronized Ca2+ waves, using the epithelium as the conducting pathway. The intercommunication was immunosuppressive, involving Ca2+ dependent activation of Akt, since AM-specific knockout of Cx43 enhanced alveolar neutrophil recruitment and secretion of proinflammatory cytokines in the bronchoalveolar lavage (BAL). The picture emerges of a novel immunomodulatory process in which a subset of alveolus-attached AMs intercommunicates immunosuppressive signals to reduce endotoxin-induced lung inflammation.

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SUMMARY Clearance of apoptotic cells (ACs) by phagocytes (efferocytosis) prevents post-apoptotic necrosis and dampens inflammation. Defective efferocytosis drives important diseases, including atherosclerosis. For efficient efferocytosis, phagocytes must be able to internalize multiple ACs. We show here that uptake of multiple ACs by macrophages requires dynamin-related protein 1 (Drp1)-mediated mitochondrial fission, which is triggered by AC uptake. When mitochondrial fission is disabled, AC-induced increase in cytosolic calcium is blunted owing to mitochondrial calcium sequestration, and calcium-dependent phagosome formation around secondarily encountered ACs is impaired. These defects can be corrected by silencing the mitochondrial calcium uniporter (MCU). Mice lacking myeloid Drp1 showed defective efferocytosis and its pathologic consequences in the thymus after dexamethasone treatment and in advanced atherosclerotic lesions in fat-fed Ldlr−/ − mice. Thus, mitochondrial fission in response to AC uptake is a critical process that enables macrophages to clear multiple ACs and to avoid the pathologic consequences of defective efferocytosis in vivo.