microglia are macrophage - like resident immune cells in the central nervous system ( cns ) and possess both neurotoxic and neuroprotective function . microglia accumulate in the lesions of a variety of neurodegenerative disorders , such as alzheimer 's disease ( ad ) , parkinson 's disease , and multiple sclerosis , and are thought to play both toxic and protective functions for neuronal survival . microglia are considered to be a first line defense and respond quickly to various stimuli . when activated , microglia undergo morphological changes to ameboid , proliferate , migrate toward injured areas , and release many soluble factors and phagocytosis of foreign substances or unwanted self - debris . appropriate migration of microglia to damaged area is controlled by chemokines and nucleotide atp [ 2 , 3 ] . phagocytosis seems to be important to prevent the senile plaque expansion in ad by removing amyloid  ( a ) deposit . microglia not only engulf the a protein but also phagocytose apoptotic cells and degenerated neuronal debris . phagocytosis of apoptotic or degenerated neuronal debris is crucial to reduce inflammation and maintain healthy neuronal networks . another type of phagocytosis , phagocytosis with inflammation , occurs in chronic inflammatory - related neurodegenerative disorders including alzheimer disease [ 57 ] . degenerated neurons releases several signaling molecules , including nucleotides , cytokines , and chemokines , to recruit microglia and enhance their activities [ 8 , 9 ] . the phenomenon are now termed as find - me , eat - me , and help - me signals . in this paper , we focused on find - me , and eat - me signals from degenerated neurons to microglia . most distinguished and examined eat - me signal is phosphatidylserine ( ps ) , which is a component of cellular membrane and is everted on apoptotic cell membrane [ 10 , 11 ] . nucleotides are also considered as the eat - me signal lately ; microglia expresse various p2x and p2y receptors , nucleotide receptors , which regulate not only chemotaxis but also phagocytosis [ 8 , 12 ] . microglia express many other surface receptors , which have direct interaction with the target to initiate phagocytosis , including ps receptor , lipopolysaccharide ( lps ) receptor cd14 , the scavenger receptor cd36 , the purine receptor p2y6 , and the toll - like receptors ( tlrs )   ( table 1 , figure 1 ) . another surface receptor , the cx3c chemokine fractalkine receptor cx3cr1 , is almost exclusively expressed in microglia throughout the cns , which is involved in progression of neurodegenerative disease by altering microglial activities [ 16 , 17 ] ( figure 2 ) . deletion of cx3cr1 expression in microglia results in progressive neuronal cell death in an animal model of neurodegenerative disease , by inducing microglial dysfunction . it has been identified recently that neurons themselves produce cytokine and chemokine , such as fractalkine . as shown in figure 2 , we previously reported that interleukin-34 ( il-34 ) , a newly discovered cytokine , is produced by neurons , and that its receptor , colony - stimulating factor 1 receptor , is primarily expressed on microglia . fractalkine and il-34 might be important mediator between neurons and microglia , and it is important to clarify this cellular crosstalk signaling pathways for seeking future therapeutic target of neurodegenerative diseases including ad . in the following sections , we will discuss about recent advances of microglial chemotaxis and phagocytosis and their implications for ad therapy . the expression levels were increased under pathological conditions , which seem to facilitate the recruitment and trafficking of glial cells to the damaged area . microglia constitutively express several chemokine receptors ( table 1 ) , which are implicated in the recruitment and accumulation microglia in ad lesions . when exposed to a , microglia are induced to produce several chemokines , such as ccl2 , ccl4 , and cxcl12 . ccr2 , the receptor of ccl2 , deficiency resulted in reduction of microglial accumulation and higher brain a levels in mouse model of ad , which might be mediated via suppression of anti - inflammatory molecule , transforming growth factor  ( tgf- ) . however , there is a conflicting report showing increased tgf- signaling in microglia surrounding a plaques in ccr2 knockout in ad model mouse ( appswe / ps1/ccr2/ ) . this ad model has been shown to have accumulation of oligomeric a and memory impairment . ccl2 expression level is also related to another neurodegenerative disorder , multiple sclerosis ( ms ) ; ccl2 level is downregulated in cerebrospinal fluid from ms patients . ccl21 triggers chemotaxis of microglia through cxcr3 , but not ccr7 which implicated in peripheral lymphoid organs . neuronal ccl21 upregulates p2x4 receptor , the nucleotide receptor , expressed in microglia . this cascade is implicated in pathophysiology of tactile allodynia to cause chronic neuropathic pain . a-induced microglial activation is mediated through p2x7 receptor that is reported as proinflammatory response conductive receptor . atp predominantly induced microglial migration among nucleotides through p2y receptors , especially p2y12 [ 2 , 37 ] . following cns injury , expression of p2y12 in microglia drastically reduced after microglial activation , suggesting that p2y12 is a primary and temporary receptor to induce microglial chemotaxis at early stages of the local cns injury . the other nucleotide udp increases mainly microglial phagocytosis ( uptake of microspheres ) via p2y6 receptor . in the condition brain damage by kainic acid administration , the cx3c chemokine cx3cl1 ( fractalkine , also called as neurotactin ) , which has been identified as two forms , soluble or membrane - anchored forms , plays a pivotal role in signaling between degenerating neurons and microglia . cx3cl1 and its receptor cx3cr1 are highly expressed in brain tissue , particularly in neurons and microglia [ 4042 ] . cx3cl1 directly induces various microglial functions including migration , proliferation , inhibition of fas - ligand - induced cell death   and glutamate - induced neurotoxicity [ 45 , 46 ] , and inhibition of proinflammatory cytokine production [ 42 , 47 ] . recently , we have shown that soluble form of cx3cl1 also directly enhances microglial clearance of degenerated neuronal debris , which is mediated through phosphatidylserine ( ps ) receptor and production of milk fat globule - egf factor 8 protein ( mfg - e8 )   ( figure 2 ) . the membrane - anchored cx3cl1 is cleaved by several proteases including a disintegrin and metalloprotease ( adam ) family , adam-10 and adam-17 [ 4850 ] , and cathepsin s . when neurons are injured or exposed to glutamate , shedding of cx3cl1 occurs immediately [ 41 , 52 ] . however , little is known about direct connection with a-induced neuronal toxicity and the cx3cl1-shedding . another important function of adam family enzyme , except for cx3cl1 shedding , is an -secretase . it cleaves app in the centre of the a domain , and generated -app is considered to have neurotrophic function [ 5356 ] . cathepsin s is expressed predominantly in microglia and implicated in microglial activation of neuropathic pain . therefore , these cx3cl1-shedding proteases may regulate the microglial phagocytosis directly or indirectly through cx3cl1 expression , and may also play a role on pathogenesis of ad . there are some reports showing that ccl2 activates cx3cr1 expression , and the induction of cx3cr1 expression by ccl2-ccr2 axis is mediated through p38 mapk activation . cx3cr1 deficiency increases susceptibility to neurotoxicity in mouse models of parkinson 's disease , amyotrophic lateral sclerosis , and systemic lps administration . cx3cl1-induced neuroprotection in a rat model of parkinson 's disease has also been reported recently . in addition , there are some reports showing that pathologic features of ad mouse model are worsened by knockout of cx3cr1 [ 17 , 61 ] . the receptor leading to inflammatory status includes cd14 , cd36 , the receptor for advanced glycation end products ( rages ) , and toll - like receptor ( tlr ) 1 , tlr2 , tlr4 , and tlr6 . the receptor leading to anti - inflammatory status includes triggering receptor expressed on myeloid cells 2 ( trem2 ) and ps receptor ( psr ) , mfg - e8 . microglia also express many other phagocytosis - related receptors which are not yet unclear to the correlation of inflammatory status . cr3microglia express classical phagocytosis - related receptor , the  
 m 
  
 2 integrin complement - receptor-3 ( cr3 ; mac-1 ; cd11b / cd18 ) , and scavenger - receptor ( sr)-ai / ii ( sr - ai / ii ; cd204 ) . cr3 synergistically activated sr - ai / ii - mediated myelin phagocytosis by microglia [ 63 , 64 ] . cr3 is implicated in clearance of bacteria through induction of major histocompatibility complex class ii ( mhc ii ) antigens . cr3 is involved in endocytosis in normal conditions and mhc ii antigens in an inflammatory state . microglia express sr - ai / ii and sr - bi during neonatal period , while in adult they lack the expression . microglia express classical phagocytosis - related receptor , the  
 m 
  
 2 integrin complement - receptor-3 ( cr3 ; mac-1 ; cd11b / cd18 ) , and scavenger - receptor ( sr)-ai / ii ( sr - ai / ii ; cd204 ) . cr3 synergistically activated sr - ai / ii - mediated myelin phagocytosis by microglia [ 63 , 64 ] . cr3 is implicated in clearance of bacteria through induction of major histocompatibility complex class ii ( mhc ii ) antigens . cr3 is involved in endocytosis in normal conditions and mhc ii antigens in an inflammatory state . microglia express sr - ai / ii and sr - bi during neonatal period , while in adult they lack the expression . cd14cd14 is the lps receptor which is also considered as classical phagocytosis - related receptor in macrophage and microglia . cd14-mediated phagocytosis does not require expression of ps receptor and possibly induces inflammatory conditions through activation of cd14 signaling . the authors suggested that it might be due to reduction of insoluble , but not soluble , a . cd14 is the lps receptor which is also considered as classical phagocytosis - related receptor in macrophage and microglia . cd14-mediated phagocytosis does not require expression of ps receptor and possibly induces inflammatory conditions through activation of cd14 signaling . the authors suggested that it might be due to reduction of insoluble , but not soluble , a . fcreceptorexposure of fibrillar a to microglia induces phagocytosis through the receptors distinct from those used by the classical phagocytosis : immunoglobulin receptors ( fcri and fcriii ) or complement receptors . exposure of fibrillar a to microglia induces phagocytosis through the receptors distinct from those used by the classical phagocytosis : immunoglobulin receptors ( fcri and fcriii ) or complement receptors . cd36 and cd47a directly interacted with microglial cell surface receptor complex comprising the class b scavenger receptor cd36 ,  
 6 
  
 1 integrin , and integrin - associated protein cd47 , all of them involved in migration and phagocytosis of microglia [ 7072 ] . cd36 is required for fibrillar a-induced chemotaxis and proinflammatory molecules including reactive oxygen species ( ros ) , tnf , il-1 , and several chemokines in microglia . a activates microglial recruitment to amyloid deposition site through cd36-dependent signaling cascade involving the src kinase family members , lyn and fyn , and the erk1/2 . cd47 is a membrane glycoprotein , broadly expressed in the various cell types in the cns , including neurons and microglia . signal regulatory protein- ( sirp ; cd172a ) is a receptor binding to cd47 , which is also expressed on neurons and myeloid cells . sirp is an inhibitory molecule of cd47 that downregulates mapk phosphorylation which is a downstream pathway of cd47 . sirp interacts with the protein tyrosine phosphatases shp-1 and shp-2 , which are predominantly expressed in neurons , dendritic cells , and macrophages . therefore , cd47 functions normally as a marker of  self  to protect intact body component [ 76 , 77 ] . a directly interacted with microglial cell surface receptor complex comprising the class b scavenger receptor cd36 ,  
 6 
  
 1 integrin , and integrin - associated protein cd47 , all of them involved in migration and phagocytosis of microglia [ 7072 ] . cd36 is required for fibrillar a-induced chemotaxis and proinflammatory molecules including reactive oxygen species ( ros ) , tnf , il-1 , and several chemokines in microglia . a activates microglial recruitment to amyloid deposition site through cd36-dependent signaling cascade involving the src kinase family members , lyn and fyn , and the erk1/2 . cd47 is a membrane glycoprotein , broadly expressed in the various cell types in the cns , including neurons and microglia . signal regulatory protein- ( sirp ; cd172a ) is a receptor binding to cd47 , which is also expressed on neurons and myeloid cells . sirp is an inhibitory molecule of cd47 that downregulates mapk phosphorylation which is a downstream pathway of cd47 . sirp interacts with the protein tyrosine phosphatases shp-1 and shp-2 , which are predominantly expressed in neurons , dendritic cells , and macrophages . therefore , cd47 functions normally as a marker of  self  to protect intact body component [ 76 , 77 ] . ragein has been reported that the direct interaction of a peptide with the receptor for rage is important in ad pathophysiology . in ad brains , rage expression is increased , and rage directly induces neurotoxicity by production of oxidative stressors and indirectly by activating microglia . rage increases macrophage - colony stimulating factor ( m - csf ) from neurons via nuclear - factor-b- ( nf-b- ) dependent pathway and released m - csf induced interaction of cognate receptor c - fms on microglia which prolongs survival of microglia . rage recognizes multiple ligands other than a peptide , such as advanced glycation end products ( ages ) , ps , and high - mobility group box 1 protein ( hmgb1 ) [ 80 , 81 ] . these molecules act as an agonist of rage on microglia , by inducing proinflammatory molecules , such as no , tnf- , il-1 , and il-6 , via map - kinase - kinase ( mek ) and phosphatidylinositol 3-kinase ( pi3k ) pathways . activation of rage leaded to nf-b and mapk - mediated signaling to propagate and perpetuate inflammation status . rage also mediates the transport of peripheral a into the brain across the blood - brain barrier ( bbb ) . 
  in has been reported that the direct interaction of a peptide with the receptor for rage is important in ad pathophysiology . in ad brains , rage expression is increased , and rage directly induces neurotoxicity by production of oxidative stressors and indirectly by activating microglia . rage increases macrophage - colony stimulating factor ( m - csf ) from neurons via nuclear - factor-b- ( nf-b- ) dependent pathway and released m - csf induced interaction of cognate receptor c - fms on microglia which prolongs survival of microglia . rage recognizes multiple ligands other than a peptide , such as advanced glycation end products ( ages ) , ps , and high - mobility group box 1 protein ( hmgb1 ) [ 80 , 81 ] . these molecules act as an agonist of rage on microglia , by inducing proinflammatory molecules , such as no , tnf- , il-1 , and il-6 , via map - kinase - kinase ( mek ) and phosphatidylinositol 3-kinase ( pi3k ) pathways . activation of rage leaded to nf-b and mapk - mediated signaling to propagate and perpetuate inflammation status . rage also mediates the transport of peripheral a into the brain across the blood - brain barrier ( bbb ) . cd200rcd200 is a transmembrane glycoprotein and is expressed on many different cell types including neurons , endothelial cells , lymphocytes , and dendritic cells . the receptor of cd200 , cd200r expression , is predominant in myeloid cells , macrophages , and microglia [ 84 , 85 ] . as in the case of sirp-cd47 , cd200 exerts inhibitory effect on cd200r , so that cd200-cd200r interaction can downregulate activity of microglia . in retina , it has been shown that activation of cd200r in microglia does not show direct effect on migration , but cd200-cd200r signaling restores lps / ifn-induced loss of migration . cd200 knockout leads to an expansion of the myeloid population in several tissues and increased expression of the activation markers in microglia , including the signaling adaptor protein dnax - activating protein of 12  kda ( dap12 ) , cd11b , cd45 , cd68 , and inducible no synthase ( inos ) . these observations suggest that cd200-cd200r signaling leads to anti - inflammatory state and protection against neurotoxic stimuli . cd200 and cd200r expression levels ( neurons and microglia , resp . ) are decreased in ad hippocampus and inferior temporal gyrus , indicating that inhibition of cd200-cd200r axis contributes to ad pathology . cd200 is a transmembrane glycoprotein and is expressed on many different cell types including neurons , endothelial cells , lymphocytes , and dendritic cells . the receptor of cd200 , cd200r expression , is predominant in myeloid cells , macrophages , and microglia [ 84 , 85 ] . as in the case of sirp-cd47 , cd200 exerts inhibitory effect on cd200r , so that cd200-cd200r interaction can downregulate activity of microglia . in retina , it has been shown that activation of cd200r in microglia does not show direct effect on migration , but cd200-cd200r signaling restores lps / ifn-induced loss of migration . cd200 knockout leads to an expansion of the myeloid population in several tissues and increased expression of the activation markers in microglia , including the signaling adaptor protein dnax - activating protein of 12  kda ( dap12 ) , cd11b , cd45 , cd68 , and inducible no synthase ( inos ) . these observations suggest that cd200-cd200r signaling leads to anti - inflammatory state and protection against neurotoxic stimuli . cd200 and cd200r expression levels ( neurons and microglia , resp . ) are decreased in ad hippocampus and inferior temporal gyrus , indicating that inhibition of cd200-cd200r axis contributes to ad pathology . trem2trem2 , a recently identified innate immune receptor , and its adaptor protein dap12 are expressed on microglia and cortical neurons , but not on hippocampal neurons , astrocytes , and oligodendrocytes . their expression correlates with clearance of apoptotic neurons by microglia without inflammation [ 9092 ] . however , endogenous ligand or specific agonist of trem2 had not been identified until recently . heat shock protein 60 ( hsp60 ) is a mitochondrial chaperone , which interacts with trem2 directly . hsp60-induced phagocytosis is only found in microglia which have robust expression of trem2 . in ad model mouse , trem2 expression was highest in the outer zone in a plaques , and the expression level correlated with the size of a plaque . forced expression of trem2 positively regulated microglial phagocytosis , the ability of microglia to stimulate cd4 t - cell proliferation , tnf- and ccl2 production , but not ifn production . trem2 , a recently identified innate immune receptor , and its adaptor protein dap12 are expressed on microglia and cortical neurons , but not on hippocampal neurons , astrocytes , and oligodendrocytes . their expression correlates with clearance of apoptotic neurons by microglia without inflammation [ 9092 ] . however , endogenous ligand or specific agonist of trem2 had not been identified until recently . heat shock protein 60 ( hsp60 ) is a mitochondrial chaperone , which interacts with trem2 directly . hsp60-induced phagocytosis is only found in microglia which have robust expression of trem2 . in ad model mouse , trem2 expression was highest in the outer zone in a plaques , and the expression level correlated with the size of a plaque . forced expression of trem2 positively regulated microglial phagocytosis , the ability of microglia to stimulate cd4 t - cell proliferation , tnf- and ccl2 production , but not ifn production . tlrs are class of pattern - recognition receptors in the innate immune system to induce inflammatory responses . 13 tlrs have been identified in human and mouse to date , except for tlr10 which is expressed only in human [ 95 , 96 ] . cd11b , a marker of macrophages and microglia , has been shown to interact with tlr signaling . cd11b knockout mouse exhibited enhanced tlr - mediated responses and subsequent more susceptibility to endotoxin shock . among the tlrs , lps receptor tlr4 potently activates microglia in various aspects , such as a phagocytosis and proinflammatory molecules production [ 98 , 99 ] . activation of tlr1 , tlr2 , tlr3 , and tlr9 by each selective agonist also increased phagocytosis and several cytokines and chemokine production [ 98 , 100 ] . the response of microglia to fibrillar a is mediated via cd14 , which act together with tlr4 and tlr2 to bind fibrillar a and induced microglial activation through p38 mapk . deficiency of cd200 induces the expression of tlr2 and tlr4 in glial cells and proliferation of cd11b / mhcii / cd40 activated microglia . these mice show memory impairment , suggesting that dual activation of tlr2 and tlr4 may induce an inflammatory phenotype of microglia which negatively regulate synaptic plasticity in the ad model . a triggers the inflammatory status in microglia via heterodimer of tlr4 and tlr6 , which is regulated by cd36 . therefore , cd14-tlr2-tlr4 and cd36-tlr4-tlr6 signaling are crucial to a-induced inflammatory response , and also in microglial phagocytosis . bacterial dna containing motifs of unmethylated cpg dinucleotides ( cpg - dna ) is a ligand of tlr9 , which is initially identified to activate microglia and strongly induces tnf- and il-12 production . however , we have shown that cpg activated microglia to produce neuroprotective molecule , such as hemeoxygenase-1 ( ho-1 ) and matrix metalloproteinase 9 ( mmp-9 ) without producing neurotoxic molecules , such as tnf- , glutamate and nitric oxide ( no ) , and enhanced a clearance to protect memory disturbance in vivo . there are the discrepancies about cpg effect between aforementioned two reports despite using the same origin cells ( mouse primary microglia ) . it may be due to concentrations of this tlr9 ligand used : higher concentration ( 10  m ) in former report , whereas lower concentrations ( 1 to 100  nm ) in latter report . moreover , the latter report revealed the difference on neuroprotective effect of cpg among synthetic oligodeoxynucleotides ( odns ) classes ( a to c ) . class a cpg did not activate microglia , but classes b and c cpgs increased microglial neuroprotective effect through induction of clearance of a and production of neuroprotectant . cpg increased chemokine ccl9 and its receptor ccr1 expression in macrophages and microglia via tlr9/myd88 signaling involving erk , p38 mapk , and pi3k pathways . phagocytotic cells recognize apoptotic cells by several mechanisms , including recognition of ps expressed on the cells . the receptors of ps ( psrs ) had not been clarified for a long time but have uncovered in recent years . these include mfg - e8 ( the lactadherin homolog in humans ) and t - cell immunoglobulin mucin domain 4 ( tim4 ) [ 107 , 108 ] . these act as a bridge between ps - expressing apoptotic cells and psr expressing phagocytes and trigger engulfment of cellular debris . mfg - e8 is expressed on various macrophage subsets in the periphery and on microglia in the cns . recently , we have shown that cx3cl1 induces mfg - e8 expression in primary mouse microglia to lead to the microglial clearance of degenerated neuronal debris . others also reported the induction of mfg - e8 by cx3cl1 in macrophages and rat microglia [ 109 , 110 ] . mfg - e8 bridges ps and integrins  
 v 
  
 3 or  
 v 
  
 5 on the surface of phagocytes [ 107 , 111 ] . high - mobility group box 1 protein ( hmgb1 ) is an intracellular protein that activates transcriptional factors , including p53 and nf-b . hmgb1 reportedly suppresses the interaction between mfg - e8 and  
 v 
  
 3 integrin in macrophage and inhibits the phagocytosis of apoptotic cells through erk - mediated signaling pathway . mfg - e8 may also be involved in a phagocytosis , since its expression is reduced in ad . therefore , mfg - e8 may possibly lead to targeted clearance of unwanted molecules , such as a , without inflammation . the other well - studied psr , tim4 , is expressed in mac-1 cells in various mouse tissues , including spleen , lymph nodes , and fetal liver . among the other tim family members , tim1 , but neither tim2 nor tim3 , also specifically binds to ps . it has been shown recently that rage also recognizes ps and induces apoptotic cell clearance . however as mentioned previously , rage - guided intracellular signaling pathway induces prolonged inflammatory status . -secretase is a protein complex of four essential membrane proteins : aph-1 , pen-2 , nicastrin , and presenilin . a recent study suggests that presenilin increases microglial phagocytosis of a , and this -secretase has dual role for ad pathogenesis : one is cleavage of amyloid precursor protein ( app ) to produce pathologic a , and the other is reduction of microglial phagocytosis of a by -secretase inhibitor . vertebrates have two presenilin genes , psen1 ( located on chromosome 14 in humans ; encoded presenilin 1 ) and psen2 ( located on chromosome 1 ; encoded presenilin 2 ) . there is a report that presenilin 2 is identified as the predominant -secretase in mouse microglia ( but not presenilin 1 ) , which repressed microglial activation via its function as a -secretase , and its expression is increased by inflammatory stimuli ( ifn- ) . in order to explore the detailed mechanism how -secretase regulates microglial activity , further studies are needed since -secretase is a therapeutic target for ad . in traumatic injury of the brain , presenilin and nicastrin expressions -secretase mainly cleaves app to lead to accumulation of a 
 1 - 42 , which then results in aggregation of a protein to worsen ad pathology . microglia express a wide array of receptors characteristic to immune cell , such as cd molecules , integrins , chemokine receptors , and psr ( figure 1 ) . chemokine receptors not only induce migration of microglia but also contribute directly to ad pathogenesis through regulation of phagocytosis and neuroprotective activity . ps acts as eat - me signal , and psr - mediated phagocytosis so far is regarded as inducing anti - inflammatory responses . however , according to some recent reports , rage interacts with psr and facilitates phagocytosis with robust inflammation status [ 80 , 120 ] . therefore , if the other phagocytosis - related receptors including tlrs interact with psr , microglia would be activated to outbreak excessive phagocytosis with robust inflammation . microglia from old app / ps1 mouse , but not from younger ones , show the reduction of sra , cd36 , rage , and the a-degrading enzymes including insulysin , neprilysin , and mmp9 . thus , it is important to consider the microglial status depending on the disease stage , to treat ad effectively . as shown in figure 2 , fkn and il-34 may be an intrinsic neuroprotectant for damaged but still surviving neurons through activation of microglia .