Central nervous system diseases related to pathological microglial phagocytosis

Microglia are important phagocytes of the central nervous system (CNS). They play an important role in protecting the CNS by clearing necrotic tissue and apoptotic cells in many CNS diseases. However, recent studies have found that microglia can phagocytose parts of neurons excessively, such as the neuronal cell body, synapse, or myelin sheaths, before or after the onset of CNS diseases, leading to aggravated injury and impaired tissue repair. Meanwhile, reduced phagocytosis of synapses and myelin results in abnormal circuit connections and inhibition of remyelination, respectively. Previous studies focused primarily on the positive effects of microglia phagocytosis, whereas only a few studies have focused on the negative effects. In this review, we use the term "pathological microglial phagocytosis" to refer to excessive or reduced phagocytosis by microglia that leads to structural or functional abnormalities in target cells and brain tissue. The classification of pathological microglial phagocytosis, the composition, and activation of related signaling pathways, as well as the process of pathological phagocytosis in various kinds of CNS diseases, are described in this review. We hypothesize that pathological microglial phagocytosis leads to aggravation of tissue damage and negative functional outcome. For example, excessive microglial phagocytosis of synapses can be observed in Alzheimer's disease and schizophrenia, leading to significant synapse loss and memory impairment. In Parkinson's disease, ischemic stroke, and traumatic brain injury, excessive microglial phagocytosis of neuronal cell bodies causes impaired gray matter recovery and sensory dysfunction. We therefore believe that more studies should focus on the mechanism of pathological microglial phagocytosis and activation to uncover potential targets of therapeutic intervention.     

Macrophage and microglial responses to cytokines in vitro: Phagocytic activity,proteolytic enzyme release,and free radical production

Certain cytokines are believed to play a key role in the development of autoimmune demyelinating diseases. Little is known, however, about the effects of these cytokines in the regulation of the key event in myelin destruction, the phagocytosis of myelin by phagocytic cells. We investigated the effects of certain cytokines and growth factors on cultured peritoneal macrophages and microglia in respect to their various functions, phagocytosis, secreted proteolytic activity, and oxidative activity. Interferon-γ (IFN-γ), tumor necrosis factor-α (TNF-α), and lipopolysaccharide (LPS), all proinflammatory factors, actually decreased (IFN-γ and LPS), or had no effect (TNF-α) on myelin phagocytosis by macrophages, but substantially increased phagocytic activity by microglia. Surprisingly, interleukins 4 and 10 (IL-4 and IL-10), considered to be downregulating cytokines, increased phagocytic activity by macrophages, while with microglia, IL-4 had no effect, but IL-10 almost doubled myelin phagocytosis. Transforming growth factor-β (TGF-β) had no significant effect on either cell. These cytokines did not affect proteolytic secretion in microglia, while IFN-γ and LPS induced a doubling of the secreted proteases. This proteolytic activity was almost completely suppressed by calpain inhibitors, although some gelatinase appeared to be present. Microglia exerted much more oxidative activity on the membranes than macrophages, and granulocyte-macrophage colony stimulating factor (GM-CSF) and interleukin 1β (IL-1β) significantly increased microglial oxidative activity. The pattern of responses of macrophages and microglia to the cytokine types indicate that in cytokine-driven autoimmune demyelinating disease, microglia may be the more aggressive cell in causing tissue injury by phagocytosis and oxidative injury, while infiltrating macrophages may produce most of the proteolytic activity thought to contribute to myelin destruction. J. Neurosci. Res. 54:68–78, 1998. © 1998 Wiley-Liss, Inc. This article is a US Government work and, as such, is in the public domain in the United States of America.     

Normal polyclonal immunoglobulins ('IVIg') inhibit microglial phagocytosis in vitro

Phagocytosis removes pathogens and tissue debris during inflammatory reactions, but also plays an important role in autoimmune reactions. The main phagocytes in the central nervous system (CNS) are microglial cells that are activated during CNS inflammation. In the treatment of inflammatory demyelinating diseases like multiple sclerosis (MS), administration of intravenous immunoglobulins (IVIg) has become a promising immunomodulatory therapy. Although a large number of potential mechanisms for the effects of IVIg has been suggested, the precise mode of action in CNS inflammation is unknown. We assessed the influence of IVIg on phagocytosis and endocytosis in microglia in vitro. IVIg had little effect on non-specific phagocytosis of latex particles in untreated microglia, while there was a dose-dependent inhibition in microglia activated with LPS and IFNgamma. Endocytosis of soluble myelin basic protein (MBP) was downregulated by IVIg in both untreated and activated microglia. The effect was mediated by an F(ab')(2) preparation of immunoglobulins, suggesting that Fc receptor-mediated phagocytosis is not involved. Intact IVIg, but not F(ab')(2) fragments also suppressed Fc receptor-mediated phagocytosis of opsonised erythrocytes in both untreated and activated microglia. These results show that IVIg can inhibit the phagocytic activity of microglia via different mechanisms. Such an effect could contribute to the immunomodulatory capacity of IVIg in inflammatory CNS diseases.     

Distinct inflammatory stimuli induce different patterns of myelin phagocytosis and degradation in recruited macrophages

Injury and demyelinating diseases result in the disruption of the myelin sheath that surrounds axons in the nervous system. The removal of degenerating myelin by macrophages and microglia is central to repair mechanisms that follow. The efficiency of myelin removal depends on magnitudes and rates of myelin phagocytosis and degradation. In the present study we test whether environmental conditions within a tissue can control patterns of myelin removal. We document that macrophages that are recruited to the same tissue but by distinct inflammatory stimuli differ in their ability to phagocytose and degrade myelin. These observations may apply to the nervous system where different pathological conditions that involve distinct inflammatory stimuli may induce different functional states in microglia and macrophages.     

Astrocytes modulate macrophage phagocytosis of myelin in vitro

Previous work from this laboratory has shown that both macrophages and microglia phagocytize relatively little myelin in vitro under basal conditions. In an effort to better simulate the conditions within the central nervous system (CNS), we have co-cultured these cells with astrocytes, the most numerous of the neural cells in the CNS, and have compared myelin phagocytosis in the co-cultures with that in cells cultured alone. Both macrophages and microglia in company with astrocytes phagocytized about three times as much myelin as controls, as measured by the formation of cholesterol ester, while astrocytes alone showed little evidence of myelin phagocytosis. Astrocyte-conditioned medium increased phagocytic activity in macrophages by 2.3-fold, and by 3.5-fold in microglia. A number of adhesion molecules and extracellular matrices were tested for their effects on myelin phagocytosis. Matrigel was most effective in activating the macrophages, and in the presence of conditioned medium, stimulated these cells to phagocytize as much myelin as when co-cultured with astrocytes. On the other hand, Matrigel inhibited myelin phagocytosis in microglia. These results indicate that activation of macrophages by astrocytes may be due to an adhesion component, as well as to soluble factors secreted by the astrocytes. While microglia were also stimulated by conditioned medium, adhesion to astrocytes or Matrigel induced a downregulation in phagocytic activity.     

Myelin phagocytosis by peritoneal macrophages in organ cultures of mouse peripheral nerve. A new model for studying myelin phagocytosis in vitro

Organ cultures of degenerating nerve fascicles were exposed to cultured macrophages obtained by peritoneal lavage. Invasion of the nerve fascicle by phagocytes was shown by prelabeling with carbon and with electron microscopy. There was massive active phagocytosis of degenerating myelin sheaths. The invading phagocytic cell population was identified as Fc receptor-positive, Mac-1-positive macrophages by immunocytochemistry. The Schwann cell population persisted without significant myelin phagocytosis. The vitality of the Schwann cell population was shown by subsequent reimplantation of the organ cultures into host animals. The reimplants had retained their ability to remyelinate regenerating axon sprouts. These observations were made in cultures exposed to cytostatic agents. If cytostatic agents were omitted, there was proliferation of endogenous phagocytes in the nerve fascicles without added peritoneal cells. These endogenous phagocytes were identified as proliferating resident monocytes and were positive for the Fc receptor and Mac-1 markers. This model allows studies on how monocytes recognize and digest degenerating myelin apart from surviving Schwann cells.     

Phagocytosis of apoptotic inflammatory cells by microglia and modulation by different cytokines: mechanism for removal of apoptotic cells in the inflamed nervous system

Apoptosis of autoaggressive T cells in the central nervous system (CNS) is an effective, nonphlogistic mechanism for the termination of autoimmune inflammation in experimental autoimmune encephalomyelitis (EAE). The clearance of apoptotic leukocytes by tissue-specific phagocytes is a critical event in the resolution of the inflammatory attack. To investigate the role of microglia in the removal of apoptotic cells and potential regulatory mechanisms of microglial phagocytosis, an in vitro phagocytosis assay was established, using Lewis rat microglia. Microglia exhibited a high capacity for the uptake of apoptotic autologous thymocytes, as well as apoptotic encephalitogenic myelin basic protein (MBP)-specific T cells, in contrast to nonapoptotic target cells. Pretreatment of microglia with interferon-gamma (IFN-gamma) raised the proportion of microglia capable of phagocytosing apoptotic cells to 75% above the untreated controls. The increased phagocytic activity was selective for apoptotic target cells and was not dependent on phosphatidylserine-mediated recognition mechanisms. In contrast, preincubation of microglia with interleukin-4 (IL-4) inhibited the uptake of apoptotic cells, whereas tumor-necrosis factor-alpha (TNF-alpha) and transforming growth factor-beta (TGF-beta) did not alter phagocytosis. Phagocytic clearance of apoptotic inflammatory cells by microglia may be an important mechanism for the termination of autoimmune inflammation in the CNS. Augmentation of microglial phagocytosis by the Th-1-type cytokine IFN-gamma suggests a feedback mechanism for the accelerated clearance of the inflammatory infiltrate in the CNS.     

Galectin-3/MAC-2 in experimental allergic encephalomyelitis

The removal of degenerating myelin by phagocytosis is central to pathogenesis and repair in traumatized and diseased nervous system. Galectin-3/MAC-2 is a differentiation and activation marker of murine and human monocytes/macrophages/microglia. Galectin-3/MAC-2, along with MAC-1 that mediates myelin phagocytosis, marks an in vivo activation state in macrophages, which are involved in myelin degeneration and phagocytosis in injured mouse peripheral nerves. In contrast, high levels of MAC-1 but extremely low levels of Galectin-3/MAC-2 are expressed in vivo in injured CNS where myelin degeneration and phagocytosis progress extremely slowly. The present study was aimed at testing whether an activation state marked by Galectin-3/MAC-2 is present in vivo in the CNS of EAE mice concomitant with autoimmune induced myelin degeneration and phagocytosis. EAE was inflicted by mouse spinal cord homogenate. Demyelination was assessed by light microscopy and Galectin-3/MAC-2, MAC-1, and F4/80 expression by immunocytochemistry. We presently document that Galectin-3/MAC-2 expression is up regulated, along with MAC-1 and F4/80, in spinal cords and optic nerves of EAE mice in areas of demyelination and myelin degeneration, in myelin phagocytosing microglia and macrophages. Copolymer 1 (Glatiramer acetate) suppresses EAE, demyelination, and Galectin-3/MAC-2 expression. EAE pathogenesis thus involves a state of activation in microglia and macrophages characterized by the expression Galectin-3/MAC-2 along with MAC-1. Furthermore, the in vivo responses to injury and autoimmune challenge in the CNS differ in the activation pattern of microglia and macrophages with regard to Galectin-3/MAC-2 expression and the corresponding occurrence of myelin degeneration and phagocytosis.     

Predominant phagocytic activity of resident microglia over hematogenous macrophages following transient focal cerebral ischemia: an investigation using green fluorescent protein transgenic bone marrow chimeric mice

Activated microglia and hematogenous macrophages are known to be involved in infarct development after cerebral ischemia. Traditionally, hematogenic macrophages are thought to be the primary cells to remove the ischemic cell debris. However, phagocytosis is a well known property also of activated microglia. Due to a lack of discriminating cellular markers, the cellular origin of phagocytes and the temporal course of phagocytosis by these two cell types are largely unknown. In this study, we used green fluorescent protein (GFP) transgenic bone marrow chimeric mice and semithin serial sections after methyl methacrylate embedding of the brains to dissect in detail the proportion of identified activated resident microglial cells and infiltrating hematogenous macrophages in phagocytosing neuronal cell debris after 30 min of transient focal cerebral ischemia. Already at day one after reperfusion, we found a rapid decrease of neurons in the ischemic tissue reaching minimum numbers at day seven. Resident GFP-negative microglial cells rapidly became activated at day one and started to phagocytose neuronal material. By contrast, hematogenous macrophages incorporating neuronal cell debris were observed in the ischemic area not earlier than on day four. Quantitative analysis showed maximum numbers of phagocytes of local origin within 2 days and of blood-borne macrophages on day four. The majority of phagocytes in the infarct area were derived from local microglia, preceding and predominating over phagocytes of hematogenous origin. This recruitment reveals a remarkable predominance of local defense mechanisms for tissue clearance over immune cells arriving from the blood after ischemic damage.     

Differential regulation of myelin phagocytosis by macrophages/microglia, involvement of target myelin, Fc receptors and activation by intravenous immunoglobulins.

Macrophages/microglia are the key effector cells in myelin removal. Differences exist in the amount and time course of myelin uptake in the central (CNS) and peripheral nervous system (PNS), the basis of this difference, however, is not yet clarified. In the present experiments we studied the phagocytosis rate of CNS or PNS myelin by macrophages and microglia in vitro. Additionally, the effects of intravenous immunoglobulins (IVIg) on this process were investigated. In the PNS experiments, sciatic nerves were cocultured with peritoneal macrophages. Optic nerve fragments were used to characterize the myelin-removing properties of microglia. Cocultures with peritoneal macrophages aimed at investigating the differences in phagocytosis between resident microglia and added macrophages. The myelin phagocytosis in sciatic nerve fragments was higher than in optic nerves, indicating differences in the myelin uptake rate between peripheral macrophages and microglia. IVIg increased the phagocytosis of PNS myelin by macrophages, but not by microglia in optic nerves. The addition of peritoneal macrophages to optic nerve fragments did not lead to an increase in the phagocytosis of CNS myelin either. The IVIg induced phagocytosis of PNS myelin by peripheral macrophages was associated with an increased expression of macrophage Fc receptors measured by FACS. Blocking of Fc receptors by anti-Fc receptor antibody reduced the IVIg induced PNS myelin phagocytosis to basic levels, indicating that the induced but not the basic myelin uptake by macrophages is Fc receptor dependent. In contrast to peripheral macrophages, IVIg did not increase Fc receptor density on microglia. These data indicate that phagocytosis of PNS and CNS myelin by macrophages or microglia is differentially regulated. Local factors within the CNS or PNS may affect this process by modulating the surface receptor profile and activation state of the phagocytic cell or the structure of the myelin sheath.     

Microglial-neuronal interactions in synaptic damage and recovery.

An understanding of the role of microglial cells in synaptic signaling is still elusive, but the neuron-microglia relationship may have important ramifications for brain plasticity and injury. This review summarizes current knowledge and theories concerning microglial-neuronal signaling, both in terms of neuron-to-microglia signals that cause activation and microglia-to-neuron signals that affect neuronal response to injury. Microglial activation in the brain involves a stereotypical pattern of changes including proliferation and migration to sites of neuronal activity or injury, increased or de novo expression of immunomodulators including cytokines and growth factors, and the full transformation into brain-resident phagocytes capable of clearing damaged cells and debris. The factors released from neurons that elicit such phenotypical and functional alterations are not well known but may include cytokines, oxidized lipids, and/or neurotransmitters. Once activated, microglia can promote neuronal injury through the release of low-molecular-weight neurotoxins and support neuronal recovery through the release of growth factors and the isolation/removal of damaged neurons and myelin debris. Because microglia respond quickly to neuronal damage and have robust effects on neurons, astrocytes, and oligodendrocytes, microglial cells could play potentially key roles in orchestrating the multicell cascade that follows synaptic plasticity and damage.     

Galectin-3/MAC-2, Ras and PI3K activate complement receptor-3 and scavenger receptor-AI/II mediated myelin phagocytosis in microglia

The removal of degenerated myelin is essential for repair in Wallerian degeneration that follows traumatic injury to axons and in autoimmune demyelinating diseases (e.g., multiple sclerosis). Microglia can remove degenerated myelin through phosphatidylinositol-3-kinase (PI3K)-dependent phagocytosis mediated by complement receptor-3 (CR3/MAC-1) and scavenger receptor-AI/II (SRAI/II). Paradoxically, these receptors are expressed in microglia after injury but myelin is not phagocytosed. Additionally, Galectin-3/MAC-2 is expressed in microglia that phagocytose but not in microglia that do not phagocytose, suggesting that Galectin-3/MAC-2 is instrumental in activating phagocytosis. S-trans, trans-farnesylthiosalicylic (FTS), which inhibits Galectin-3/MAC-2 dependent activation of PI3K through Ras, inhibited phagocytosis. K-Ras-GTP levels and PI3K activity increased during normal phagocytosis and decreased during FTS-inhibited phagocytosis. Galectin-3/MAC-2, which binds and stabilizes active Ras, coimmunoprecipitated with Ras and levels of the coimmunoprecipitate increased during normal phagocytosis. A role for Galectin-3/MAC-2 dependent activation of PI3K through Ras, mostly K-Ras, is thus suggested. An explanation may thus be offered for deficient phagocytosis by microglia that express CR3/MAC-1 and SRAI/II without Galectin-3/MAC-2 and efficient phagocytosis when CR3/MAC-1 and SRAI/II are co-expressed with Galectin-3/MAC-2.     

Microglia and macrophage activation and the regulation of complement-receptor-3 (CR3/MAC-1)-mediated myelin phagocytosis in injury and disease

Microglia and macrophages play critical roles in the response of the central and peripheral nervous systems (CNS and PNS, respectively) to injury and disease, one of which is the removal of degenerated myelin by phagocytosis. Myelin removal is efficient during Wallerian degeneration, which follows injury to PNS axons, and in CNS autoimmune demyelinating diseases (e.g., multiple sclerosis) but is inefficient after injury to CNS axons. We suggest that inefficient myelin removal results from deficient microglia activation, reflected by the failure to up-regulate Galectin-3/MAC-2 expression, which marks a state of activation correlated with efficient myelin phagocytosis. Surprisingly, whether or not executing myelin phagocytosis, CNS microglia express the alphaM/beta2 integrin complement receptor-3 (CR3/MAC-1), which has the potential of mediating efficient myelin phagocytosis. We hypothesize that CR3/MAC-1 might be present in distinct inactive and active states that determine, respectively, efficient and inefficient CR3/MAC-1-mediated myelin phagocytosis. We present evidence that CR3/MAC-1-mediated myelin phagocytosis is regulated in microglia and macrophages. First, CR3/MAC-1- mediated myelin phagocytosis has complement-dependent and -independent components. Second, an active complement system augments CR3/MAC-1-mediated myelin phagocytosis. Third, anti-alphaM monoclonal antibodies (MAbs) inhibit and anti-beta2 MAbs augment CR3/MAC-1-mediated myelin phagocytosis in the presence and absence of an active complement system. Fourth, an active complement system modulates MAb-induced regulation of CR3/MAC-1-mediated myelin phagocytosis. Overall, MAb-induced phagocytosis regulation might range three- to sevenfold from inefficient to efficient. We suggest that one of the mechanisms underlying MAb-induced phagocytosis regulation is the induction/stabilization of inactive and active conformational changes. Monoclonal antibody-induced phagocytosis regulation must reveal a mechanism by which native extracellular molecules bind to and regulate CR3/MAC-1-mediated myelin phagocytosis in microglia and macrophages.     

Phagocytosis of apoptotic inflammatory cells by microglia and its therapeutic implications: termination of CNS autoimmune inflammation and modulation by interferon-beta

Apoptosis of autoaggressive T-cells in the CNS is an effective, noninflammatory mechanism for the resolution of T-cell infiltrates, contributing to clinical recovery in T-cell-mediated neuroinflammatory diseases. The clearance of apoptotic leukocytes by tissue-specific phagocytes is critical in the resolution of the inflammatory infiltrate and leads to a profound downregulation of phagocyte immune functions. Adult human microglia from surgically removed normal brain tissue was used in a standardized, light-microscopic in vitro phagocytosis assay of apoptotic autologous peripheral blood-derived mononuclear cells (MNCs). Microglia from five different patients had a high capacity for the uptake of apoptotic MNCs in contrast to nonapoptotic target cells with the phagocytosis rate for nonapoptotic MNCs amounting to only 61.6% of the apoptotic MNCs. A newly described phosphatidylserine receptor, critical in the phagocytosis of apoptotic cells by macrophages, is also expressed at similar levels on human microglia. The effects of the therapeutically used immunomodulatory agent interferon-beta (IFNbeta) were investigated using Lewis rat microglia and apoptotic, encephalitogenic, myelin basic protein-specific autologous T-cells. Also, rat microglia had a high capacity to phagocytose apoptotic T-cells specifically. IFNbeta increased the phagocytosis of apoptotic T-cells to 36.8% above the untreated controls. The enhanced phagocytic activity was selective for apoptotic T-cells and was not mediated by increased IL-10 secretion. Apoptotic inflammatory cells may be efficiently and rapidly removed by microglial cells in the autoimmune-inflamed human CNS. The in vitro increase of phagocytosis by IFNbeta merits further investigations whether this mechanism could also be therapeutically exploited.     

Inhibition of UDP/P2Y6 purinergic signaling prevents phagocytosis of viable neurons by activated microglia in vitro and in vivo

Microglia activated through Toll‐like receptor (TLR)‐2 or ‐4 can cause neuronal death by phagocytosing otherwise‐viable neurons—a form of cell death called “phagoptosis.” UDP release from neurons has been shown to provoke microglial phagocytosis of neurons via microglial P2Y₆ receptors, but whether inhibition of this process affects neuronal survival is unknown. We tested here whether inhibition of P2Y₆ signaling could prevent neuronal death in inflammatory conditions, and whether UDP signaling can induce phagoptosis of stressed but viable neurons. We find that delayed neuronal loss and death in mixed neuronal/glial cultures induced by the TLR ligands lipopolysaccharide (LPS) or lipoteichoic acid was prevented by: apyrase (to degrade nucleotides), Reactive Blue 2 (to inhibit purinergic signaling), or MRS2578 (to specifically block P2Y₆ receptors). In each case, inflammatory activation of microglia was not affected, and the rescued neurons remained viable for at least 7 days. Blocking P2Y₆ receptors with MRS2578 also prevented phagoptosis of neurons induced by 250 nM amyloid beta 1–42, 5 μM peroxynitrite, or 50 μM 3‐morpholinosydnonimine (which releases reactive oxygen and nitrogen species). Furthermore, the P2Y₆ receptor agonist UDP by itself was sufficient to stimulate microglial phagocytosis and to induce rapid neuronal loss that was prevented by eliminating microglia or inhibiting phagocytosis. In vivo, injection of LPS into rat striatum induced microglial activation and delayed neuronal loss and blocking P2Y₆ receptors with MRS2578 prevented this neuronal loss. Thus, blocking UDP/P2Y₆ signaling is sufficient to prevent neuronal loss and death induced by a wide range of stimuli that activate microglial phagocytosis of neurons. GLIA 2014;62:1463–1475     

Phagocytosis of neuronal debris by microglia is associated with neuronal damage in multiple sclerosis

Neuroaxonal degeneration is a pathological hallmark of multiple sclerosis (MS) contributing to irreversible neurological disability. Pathological mechanisms leading to axonal damage include autoimmunity to neuronal antigens. In actively demyelinating lesions, myelin is phagocytosed by microglia and blood-borne macrophages, whereas the fate of degenerating or damaged axons is unclear. Phagocytosis is essential for clearing neuronal debris to allow repair and regeneration. However, phagocytosis may lead to antigen presentation and autoimmunity, as has been described for neuroaxonal antigens. Despite this notion, it is unknown whether phagocytosis of neuronal antigens occurs in MS. Here, we show using novel, well-characterized antibodies to axonal antigens, that axonal damage is associated with HLA-DR expressing microglia/macrophages engulfing axonal bulbs, indicative of axonal damage. Neuronal proteins were frequently observed inside HLA-DR(+) cells in areas of axonal damage. In vitro, phagocytosis of neurofilament light (NF-L), present in white and gray matter, was observed in human microglia. The number of NF-L or myelin basic protein (MBP) positive cells was quantified using the mouse macrophage cell line J774.2. Intracellular colocalization of NF-L with the lysosomal membrane protein LAMP1 was observed using confocal microscopy confirming that NF-L is taken up and degraded by the cell. In vivo, NF-L and MBP was observed in cerebrospinal fluid cells from patients with MS, suggesting neuronal debris is drained by this route after axonal damage. In summary, neuroaxonal debris is engulfed, phagocytosed, and degraded by HLA-DR(+) cells. Although uptake is essential for clearing neuronal debris, phagocytic cells could also play a role in augmenting autoimmunity to neuronal antigens.     

ED2-positive perivascular phagocytes produce interleukin-1β during delayed neuronal loss in the facial nucleus of the rat

Injection of Fluoro-Gold (FG) into the whisker pad of rats yields stable retrograde labeling of facial motoneurons. Subsequent removal of 10 mm from all facial nerve branches permanently deprives the FG-labeled motoneurons from their targets and the motoneurons gradually die. Neuronal debris is phagocytized by two types of neuronophages: parenchymal microglia (monoclonal antibody [MAb] OX42-positive, MAb ED2-negative) and perivascular phagocytes (OX42-negative, ED2-positive). Because both types of neuronophages express major histocompatibility complex (MHC) class II glycoproteins (MAb OX6-positive), they are considered to be the potential antigen-presenting cells of the brain. To check this hypothesis, we tested whether both types of neuronophages also synthetize the co-stimulatory cytokine interleukin-1β (IL-1β) immunocytochemically visualized by MAbs SILK-5/6. Employing combined fluorescent visualization of antigens (OX6, ED2, and SILK-5/6) in sections containing fluorescent (FG-prelabeled) neuronophages, we found that, during slowly occurring neuronal loss, the vast majority of IL-1β immunoreactive neuronophages were of perivascular (ED2-positive) origin. We concluded that, during delayed neuronal death “behind” an intact blood–brain barrier, the perivascular phagocytes were more likely to function as antigen-presenting cells than the parenchymal microglia. J. Neurosci. Res. 54:820–827, 1998. © 1998 Wiley-Liss, Inc.     

Complement-receptor-3 and scavenger-receptor-AI/II mediated myelin phagocytosis in microglia and macrophages

Microglia and macrophages express the alpha(M)/beta(2) integrin complement-receptor-3 (CR3/MAC-1; CD11b/CD18) and scavenger-receptor-AI/II (SRAI/II). Both can mediate myelin phagocytosis. We document that CR3/MAC-1 mediated myelin phagocytosis in microglia is modulated by complement and anti-CR3/MAC-1 mAbs. Complement augmented phagocytosis twofold. Anti-alpha(M) mAbs M1/70 and 5C6 inhibited and anti-beta(2) mAb M18/2 augmented myelin phagocytosis in the presence and absence of active complement. Active complement modulated phagocytosis inhibition by M1/70 and 5C6 and phagocytosis augmentation by M18/2. CR3/MAC-1 mediated myelin phagocytosis may thus be, at least partially, independent of but modulated by complement. Anti-beta(2) mAb Game-46 did not affect phagocytosis. However, combining M18/2 with Game-46 resulted in phagocytosis augmentation that was larger in magnitude than that induced by M18/2 alone. Thus, phagocytosis augmentation induced by one anti-beta(2) mAb was potentiated by another anti-beta(2) mAb. Combining M1/70 or 5C6 with M18/2 inhibited M18/2-induced augmentation. Overall, mAbs-induced phagocytosis modulation ranged three- to sevenfold from inhibition to augmentation. Anti-CR3/MAC-1 mAbs may reveal a mechanism by which native extracellular molecules bind to and modulate CR3/MAC-1 mediated myelin phagocytosis in microglia and macrophages. We further document SRAI/II mediated myelin phagocytosis in microglia and CR3/MAC-1 contributing to myelin phagocytosis two- to threefold more than SRAI/II when the two receptors function together.