Cell Death and Differentiation (2010) 17, 381–397 & 2010 Macmillan Publishers Limited All rights reserved 1350-9047/10 $32.00 www.nature.com/cdd Review Molecular mechanisms of late apoptotic/necrotic cell clearance IKH Poon1,2, MD Hulett*,1,2,3 and CR Parish*,1,3 Phagocytosis serves as one of the key processes involved in development, maintenance of tissue homeostasis, as well as in eliminating pathogens from an organism. Under normal physiological conditions, dying cells (e.g., apoptotic and necrotic cells) and pathogens (e.g., bacteria and fungi) are rapidly detected and removed by professional phagocytes such as macrophages and dendritic cells (DCs). In most cases, specific receptors and opsonins are used by phagocytes to recognize and bind their target cells, which can trigger the intracellular signalling events required for phagocytosis. Depending on the type of target cell, phagocytes may also release both immunomodulatory molecules and growth factors to orchestrate a subsequent immune response and wound healing process. In recent years, evidence is growing that opsonins and receptors involved in the removal of pathogens can also aid the disposal of dying cells at all stages of cell death, in particular plasma membrane-damaged cells such as late apoptotic and necrotic cells. This review provides an overview of the molecular mechanisms and the immunological outcomes of late apoptotic/necrotic cell removal and highlights the striking similarities between late apoptotic/necrotic cell and pathogen clearance. Cell Death and Differentiation (2010) 17, 381–397; doi:10.1038/cdd.2009.195; published online 18 December 2009 The immune system is constantly under pressure to accu- phagocytosis.1,2 Not surprisingly, impairment of phagocytosis rately distinguish foreign materials or pathogens (non-self) due to a deficiency in key phagocytic components such as from normal healthy tissues (self), and make an appropriate C1q, one of the first components of the classical complement immune response to non-self molecules through a range of cascade, has been implicated in an increased susceptibility effector mechanisms. It is equally important for the immune to bacterial infection3 as well as in the development of auto- system to distinguish healthy viable cells (self) from dying immune diseases such as systemic lupus erythematosus cells (altered-self) during the course of tissue remodelling or (SLE).4 Therefore, understanding the molecular mechanisms tissue injury to prevent the release of intracellular molecules of phagocytosis will provide new insights into numerous that may damage neighbouring cells or stimulate an immuno- physiological and pathological processes. genic response against self (i.e., an autoimmune response). Under normal physiological conditions, cells of a multi- Professional phagocytes of the innate immune system use cellular organism predominantly die through the well-defined a broad range of germline-encoded receptors and opsonins process known as apoptosis or programmed cell death (see to discriminate viable cells from pathogens and dying cells, Elmore5 for an extensive review). During different stages of and aid the removal of non-self and altered-self through apoptotic cell death, a precise set of morphological and 1John Curtin School of Medical Research, Australian National University, Canberra, 2601, Australia and 2Department of Biochemistry, La Trobe University, Melbourne, 3086, Australia *Corresponding authors: MD Hulett, Department of Biochemistry, La Trobe University, Physical Sciences 4 Building, Kingsbury Drive, Bundoora, Melbourne, 3086, Australia. Tel: þ 61 3 9479 6567; Fax: þ 61 3 9479 2467; E-mail: [email protected] or C Parish, The John Curtin School of Medical Research, The Australian National University, Building 131, Garran Road, Acton, ACT, 0200, Australia. Tel: þ 61 2 6125 2604; Fax: þ 61 2 6125 2337; E-mail: [email protected] 3These authors contributed equally to this work. Keywords: phagocytosis; necrotic cell; apoptotic cell; pathogen Abbreviations: ACAMP, apoptotic cell-associated molecular pattern; b2GPI, b2-glycoprotein I; BAI1, brain-specific angiogenesis inhibitor 1; C4BP, C4b-binding protein; CFHR4, human complement factor H-related protein 4; CR, complement receptors; CRP, C-reactive protein; DAMP, danger-associated molecular pattern; DC, dendritic cell; DNA, deoxyribonucleic acid; ds, double-stranded; EMAP II, endothelial monocyte-activating polypeptide II; FcgR, Fcg receptor; FPR, formyl peptide receptor; GAG, glycosaminoglycan; GlcNAc, N-acetyl-D-glucosamine; HERDS, heterogeneous ectopic ribonucleoprotein-derived structures; HMDDC, human monocyte-derived dendritic cell; HMDM, human monocyte-derived macrophage; HMGB1, high mobility group box 1 protein; HRG, histidine-rich glycoprotein; HS, heparan sulphate; HSP, heat shock protein; HSPG, heparan sulphate proteoglycans; IFN-g, interferon-g; Ig, immunoglobulin; IL, interleukin; ITIM, immunoreceptor tyrosine-based inhibitor motif; LBP, LPS-binding protein; LPC, lysophosphatidylcholine; LPS, lipopolysaccharide; LTA, lipoteichoic acid; MAC, membrane-attack complex; MASP, MBL-associated serine protease; MBL, mannose-binding lectin; MFG-E8, milk fat globular-EGF factor 8 protein; MIP2, macrophage-inflammatory protein-2; MHC, major histocompatibility complex; MSU, monosodium urate; NOD-like, nucleotide-binding oligomerization domain; PAMP, pathogen-associated molecular pattern; PBL, peripheral blood lymphocyte; PCh, phosphocholine; PRM, pattern recognition molecule; PS, phosphatidylserine; PTX3, pentraxin-3; RAGE, receptor for advanced glycation end-products; RIG-like, retinoic acid-inducible gene-I-like; RNA, ribonucleic acid; SAMP, self-associated molecular pattern; SAP, serum amyloid protein; SAP130, spliceosome-associated protein 130; SLE, systemic lupus erythematosus; TCR, T-cell receptor; TFAM, mitochondrial transcription factor A; TGF-b, transforming growth factor-b; TIM, T-cell immunoglobulin mucin; TLR, Toll-like receptor; TNF-a, tumour necrosis factor-a; Trem, triggering receptor expressed on myeloid cells; TSP-1, thrombospondin-1; VEGF, vascular endothelial growth factor Received 18.6.09; revised 30.10.09; accepted 02.11.09; Edited by P Vandenabeele; published online 18.12.09 Mechanisms of late apoptotic/necrotic cell clearance IKH Poon et al 382 biochemical changes occur within the dying cell.5 Dying cells soluble molecules such as lysophosphatidylcholine (LPC),16 can be characterized as early apoptotic cells, in which the endothelial monocyte-activating polypeptide II (EMAP II)17 plasma membrane remains intact but exposes phosphatidyl- and fractalkine/CX3CL118 by early apoptotic cells has also serine (PS) on the cell surface to mediate its recognition by been suggested to promote the recruitment of macrophages phagocytes. Early apoptotic cells can become late apoptotic and monocytes towards cells undergoing apoptosis. Interest- cells, also known as secondary necrotic cells, when the ingly, apoptotic leukocytes (HL-60 cells) generated by heat plasma membrane becomes permeabilized.2 Alternatively, treatment can secret both S19 ribosomal homodimer and a direct exposure of healthy viable cells to trauma, such as serine protease to promote and subsequently retard mono- extreme temperature, mechanical, and chemical insults, can cyte migration, respectively, during the early and later stages lead to the generation of membrane-permeabilized necrotic of apoptotic cell death.19 Besides facilitating the chemotaxis of cells (i.e., primary necrotic cells).6 Importantly, depending on mononuclear phagocytes, induction of apoptosis can also the type of dying cells (e.g., early apoptotic versus necrotic trigger the release of lactoferrin, an anti-inflammatory glyco- cells), recognition and internalization by phagocytes can protein that selectively inhibits the migration of neutrophils result in an anti- or a pro-inflammatory response.1,7 In towards early apoptotic cells, possibly to prevent any aberrant addition, uptake of apoptotic cells by macrophages can inflammation at sites of physiological forms of cell death.20 also promote cell growth8 and wound healing9 through the Although such an elegant phagocyte recruitment system may release of vascular endothelial growth factor (VEGF) and be necessary in mediating efficient clearance of apoptotic transforming growth factor-b (TGF-b), respectively. Thus, the cells at certain tissues, it should be noted that non-profes- process of phagocytosis in mammals not only serves as sional phagocytes, such as fibroblasts, endothelial and an effector mechanism to clear infectious agents, dying cells, epithelial cells, can also contribute to removing adjacent cells or unwanted materials, but also has a fundamental role in undergoing apoptosis (see Majai et al.21 for an extensive determining the subsequent adaptive immune response review).21 towards the phagocytosed materials as well as orchestrating When a phagocyte comes into close proximity to a target the processes of regeneration and angiogenesis at sites cell, the target cell must expose a sufficient level of ‘eat-me’ of tissue injury. Furthermore, phagocytosis of dying cells, signals to trigger phagocytosis. For phagocytes to discrimi- especially early apoptotic cells in an anti-inflammatory nate pathogens from host cells, Janeway22 proposed that a context, has a vital role in maintaining immunological limited number of germline-encoded pattern recognition tolerance against cell-associated antigens.10 molecules (PRMs) are used by the innate immune