Abstract
The purpose of physiological cell death is the noninflammatory clearance of cells that have become inappropriate or nonfunctional. Consistent with this function, the recognition of apoptotic cells by professional phagocytes, including macrophages and dendritic cells, triggers a set of potent anti-inflammatory responses manifest on multiple levels. The immediate-early inhibition of proinflammatory cytokine gene transcription in the phagocyte is a proximate consequence of recognition of the apoptotic corpse, independent of subsequent engulfment and soluble factor involvement. Here, we show that recognition is linked to a characteristic signature of responses, including MAPK signaling events and the ablation of proinflammatory transcription and cytokine secretion. Specific recognition and response occurs without regard to the origin (species, tissue type, or suicidal stimulus) of the apoptotic cell and does not involve Toll-like receptor signaling. These features mark this as an innate immunity fundamentally distinct from the discrimination of "self" versus "other" considered to be the hallmark of conventional immunity. This profound unconventional innate immune discrimination of effete from live cells is as ubiquitous as apoptotic cell death itself, manifest by professional and nonprofessional phagocytes and nonphagocytic cell types alike. Innate apoptotic immunity provides an intrinsic anti-inflammatory circuit that attenuates proinflammatory responses dynamically and may act systemically as a powerful physiological regulator of immunity.
Highlights
The purpose of physiological cell death is the non-inflammatory clearance of cells that have become inappropriate or non-functional
Specific recognition and response to apoptotic cells is not limited to macrophages
The antiinflammatory response triggered in macrophages by their specific recognition of apoptotic cells is exerted on the level of cytokine gene transcription
Summary
Cells and Death Induction – RAW 264.7 murine macrophages, DO11.10 murine T hybridoma cells, Jurkat human acute T leukemia cells, Ramos (RA1) human Burkitt's B lymphoma cells, and PLB985 human myelomonoblastic leukemia cells (generously provided by Dr Peter Henson, National Jewish Medical and Research Center) were cultured at 37oC in a humidified, 5% (v / v) CO2 atmosphere in RPMI 1640 medium (Mediatech; Herndon, VA) supplemented with heat inactivated fetal bovine serum (10% v / v), 2 mM L-glutamine, and 50 μM 2-mercaptoethanol. Target cells (viable, apoptotic, and necrotic) were washed twice and resuspended in the medium of the responder cells to be tested. 400 μl of binding buffer was added per sample and cells were analyzed cytofluorimetrically. Transfections and Luciferase Assays – Apoptotic modulation of specific transcription (e.g. dependent on NFκB or the IL-8 promoter) was assessed in various cell types following transfection of relevant transcriptional reporter constructs, using a dual luciferase strategy, as described previously (13). Cell extracts were prepared after further incubation as indicated, and luciferase activities were measured by the Dual Luciferase Reporter. Cellular Extract Preparation and Immunoblot Analysis – Activation of Akt and inhibition of ERK1/2 were assessed in 3T3 cells cultured overnight in serum-free medium and left unstimulated or stimulated for 15 min.
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