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From Regulation of Dying Cell Engulfment to Development of Anti-Cancer Therapy

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From Regulation of Dying Cell Engulfment to Development of Anti-Cancer Therapy Cell Death and Differentiation (2008) 15, 29–38 & 2008 Nature Publishing Group All rights reserved 1350-9047/08 $30.00 www.nature.com/cdd Review From regulation of dying cell engulfment to development of anti-cancer therapy DV Krysko1,2 and P Vandenabeele*,1,2 Cell death and efficient engulfment of dying cells ensure tissue homeostasis and is involved in pathogenesis. Clearance of dying cells is a complex and dynamic process coordinated by interplay between ligands on dying cell, bridging molecules, and receptors on engulfing cells. In this review, we will discuss recent advances and significance of molecular changes on the surface of dying cells implicated in their recognition and clearance as well as factors released by dying cells that attract macrophages to the site of cell death. It is now becoming apparent that phagocytes use a specific set of mechanisms to discriminate between live and dead cells, and this phenomenon will be illustrated here. Next, we will discuss potential mechanisms by which removal of dying cells could modulate immune responses of phagocytes, in particular of macrophages. Finally, we will address possible strategies for manipulating the immunogenicity of dying cells in experimental cancer therapies. Cell Death and Differentiation (2008) 15, 29–38; doi:10.1038/sj.cdd.4402271; published online 16 November 2007 ‘While I thought that I was learning how to live, I have been and Zhou1) that converge at a common effector, ced-10 learning how to die.’ (mammalian homolog Rac-1). One pathway is composed of Leonardo da Vinci the proteins ced-2, ced-5, and ced-12 (mammalian homo- logues are CrkII, Dock180, and ELMO, respectively), and its Activation of the innate immune system is the first line of upstream components are GTPase, RhoG/MIG-2, and TRIO/ defense against infection and the effects of localized injury or UNC-73. The second pathway includes membrane proteins trauma. This activation is aimed at eliminating the infectious ced-1, ced-7, ced-6, an adaptor protein, and dynamin 1.1,2 agent and restoring homeostasis. However, in the human Clearance of dying cells is a complex process in which body close to 500 billion cells die each day, and they are either many surface molecules, adaptors, and chemotactic mole- shed off directly from body surfaces or continuously removed cules are involved, and it is controlled at multiple levels. This by a remarkably efficient phagocytic system without causing review covers recent advances in understanding the mole- inflammation or scars. In the late 19th century, the immuno- cular interactions involved in recognition and clearance of logist Ilya Metchnikoff first observed the process of phago- stressed/dying/dead cells, and the effect of their interaction on cytosis and made a link between cellular immunity and cells the immune system. We also discuss repelling signals that ‘eating’ other cells. Since then it has been established that protect living cells from being engulfed. We will briefly phagocytosis has a key role in immunological processes. consider how exposure of certain molecules on the surface Rapid recognition and clearance of dying cells by phagocytes of dying cells, such as phosphatidylserine (PS) and calreticulin, plays pivotal roles in development, maintenance of tissue can be used for raising immunogenicity of tumor cells,3,4 and homeostasis, control of immune responses, and resolution of how these findings could be applied for designing novel inflammation. Dying cells can be cleared by professional experimental anti-cancer treatments. phagocytes, such as macrophages and immature dendritic cells (DCs), or by nonprofessional phagocytes, such as endothelial cells, fibroblasts, smooth cells, and epithelial Positive Regulators of uptake Dying Cell cells. Important insight into the genetic pathways implicated in In order to guarantee instant recognition and uptake by clearance of dying cells came from studies on Caenorhabditis phagocytes, apoptotic cells undergo early membrane modi- elegans. The engulfment genes fall into two partially fications. The best characterized change in the surface of redundant pathways (reviewed in detail in Mangahas apoptotic cells that facilitates their recognition is the loss of 1Molecular Signalling and Cell Death Unit, Department for Molecular Biomedical Research, VIB, Technologiepark 927, Ghent, Belgium and 2Department of Molecular Biology, Ghent University, Technologiepark 927, Ghent, Belgium *Corresponding author: P Vandenabeele, Department for Molecular Biomedical Research, VIB, Ghent University, Technologiepark 927, Zwijnaarde-Gent B-9052, Belgium. Tel: þ 32 9 3313760; Fax: þ 32 9 3313609; E-mail: [email protected] Keywords: phagocytosis; apoptosis; necrosis; autophagy; macrophages; macropinocytosis Abbreviations: CR, complement receptor; DC, dendritic cell; eIF2a, eukaryotic initiation factors 2-a; ER, endoplasmic reticulum; Gas6, growth arrest-specific 6; HMGB- 1, high mobility group box 1; IFN-g, interferon-g; IL, interleukin; JNK, c-Jun N-terminal kinases; LPC, lysophosphatidylcholine; LPS, lipopolysaccharide; MAPK, mitogen- activated protein kinase; Mer, myeloid epithelial reproductive tyrosine kinase; MFG-E8, milk fat globule protein E8; NF-kB, nuclear factor-kB; PAF, platelet activating factor; PP1, protein phosphatase; PS, phosphatidylserine; ROS, reactive oxygen species; SIRPa, signal regulator protein-a; SOCS, suppressors of cytokine signaling; STAT1, signal transducer and activator of transcription 1; TGF-b, transforming growth factor-b; TLR, Toll-like receptor; TNF, tumor necrosis factor Received 22.8.07; revised 05.10.07; accepted 05.10.07; Edited by G Kroemer; published online 16.11.07 Dying cell engulfment and anti-cancer therapy DV Krysko and P Vandenabeele 30 phospholipid asymmetry and translocation of PS from the homeostatic function for PS exposed on apoptotic cells has inner to the outer leaflet of the lipid bilayer, which occurs very recently been proposed. Exposure of PS on UV-induced early during the apoptotic process.5 Although the precise apoptotic Jurkat T lymphocytes was necessary and sufficient mechanisms involved in initiating the movement of PS to the to stimulate the efflux of cholesterol from J774 murine external surface of the membrane are still unclear, some macrophages and human blood monocyte-derived macro- possibilities have been suggested, including a coordinate phages.11 Failure to stimulate cholesterol efflux by the increase in phospholipid flip-flop (flip refers to inward and flop macrophages during the uptake of necrotic cells (but not to outward movement) due to inactivation of the aminophos- during apoptotic cell uptake) could lead to progression of pholipid translocase.6 Recently, a novel molecular mechanism atherosclerosis during late-stage atherosclerotic lesions, of PS exposure has been discovered in C. elegans, by using which contain apoptotic cells and a core of necrotic cells.11 an RNAi reverse genetic approach and a transgenic strain A new alternative treatment for atherosclerosis in the future expressing a green fluorescent proteinHAnnexin-V reporter. may be the addition of either artificial targets that mimic Zullig et al.7 identified a tat-1 gene as a possible aminopho- apoptotic cells, such as PS-containing liposomes, or the spholipd transporter gene, because knocking down this gene triggering of specific engulfment receptors on phagocytes, abrogated PS exposure on apoptotic cells and reduced promoting cholesterol efflux from phagocytes.11 In addition, it clearance of dead cell corpses. Recognition of PS is mediated has been reported that statins, potent cholesterol-lowering by a broad range of extracellular bridge molecules8 (summar- agents, could increase clearance of apoptotic cells in vitro and ized in Figure 1), such as b2-glycoprotein, milk fat globule in vivo, supporting the notion that efficient clearance of protein E8 (MFG-E8), protein S, growth arrest-specific 6 apoptotic cells in the atherosclerotic lesion is an important (Gas6), and thrombospondin. Oxidation of proteins and lipids regulatory mechanism.12 at the apoptotic cell surface is also important for engulfment. PS on the surface of apoptotic cells also colocalizes with Oxidized PS has higher specificity than non-oxidized PS for other molecules, including calreticulin13 (Figure 2) and bridging molecules such as MFG-E8.9 Moreover, clearance of annexin-I.14 However, it is not yet clear whether this apoptotic cells in vivo also occurs through interaction of CD36, colocalization is just a topological issue or involves a a heavily glycosylated multi-ligand receptor belonging to the functional interaction providing cooperative stimuli for engulf- class B scavenger receptor group, with membrane-asso- ment. Calreticulin was identified in 1974 as a soluble protein ciated oxidized PS.10 A new and possibly clinically relevant from the lumen of the endoplasmic reticulum (ER). The multifunctional calreticulin binds (buffers) Ca2 þ in the lumen of the ER with high affinity, participates in the folding of newly synthesized glycoproteins, preventing their aggregation,15 and also performs other cellular functions outside the ER, such as cellular adhesion, and gene expression.15 Calreticulin is also present on the surface of most cell types, and its level on the surface increases during apoptosis. The precise mechanism initiating externalization of calreticulin to the surface of the plasma membrane is still unclear. It is possible that the chaperone calreticulin is transported together with membrane proteins, as in the case of the
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