Microglia Build Plaques to Protect the Brain

Could it be that microglia build plaques rather than bust them? In the April 15 Nature Immunology, researchers led by Greg Lemke at the Salk Institute for Biological Studies, La Jolla, California, add to the evidence for this idea. In a mouse model of amyloidosis, the researchers hobbled microglial phagocytosis by deleting the two primary receptors for this process, Axl and Mer. Surprisingly, fewer dense-core plaques formed. Instead, the Aβ accumulated in wispy cotton-wool plaques and around blood vessels. The data imply that microglia normally act like trash compactors, taking up amyloid and condensing it inside lysosomes before re-depositing it in an inert form. “We conclude that dense-core plaques don’t form spontaneously; they’re constructed by microglia,” Lemke told Alzforum. He believes this may be a mechanism the brain uses to limit the damage from Aβ by confining it.

  • When microglia cannot phagocytose, fewer dense plaques form in mice.
  • Instead, amyloid accumulates in wispy deposits and lines blood vessels.
  • Microglia may help contain amyloid by sequestering it into dense-core plaques.

The findings generated enthusiasm. “This is a landmark study,” Wei Cao, Manasee Gedam, and Hui Zheng at Baylor College of Medicine, Houston, wrote to Alzforum (full comment below). “The model the authors put forward is provocative, and it challenges our current understanding of microglia and plaque interaction.”

“The caliber of the work is terrific,” wrote Gary Landreth at Indiana University School of Medicine in Indianapolis (comment below). Henrik Zetterberg at the University of Gothenburg, Sweden, called the implications profound. “It’s an extremely exciting story, and really unexpected. It revises the amyloid cascade hypothesis, because it looks like microglia are the catalyst,” Zetterberg told Alzforum.

Pack It Tight. Microglia around plaques (red circle) take up loose amyloid (yellow fuzz) using TAM receptors (black and red), then condense it into dense deposits (red cross-hatches) inside their lysosomes. Then they expel this material, adding to dense-core plaques. [Courtesy of Huang et al., Nature Immunology.]

Specialized Phagocytic Machinery
Microglial phagocytosis is now well understood. The receptors Axl and Mer are part of a family of three receptor tyrosine kinases collectively known as TAMs (Lu and Lemke, 2001). Axl and Mer power microglial removal of damaged and dying cells, with Mer specialized to mop up apoptotic debris (Scott et al., 2001; Fourgeaud et al., 2016). In addition, signaling through these receptors suppresses pro-inflammatory cytokines, with the net effect of shifting microglia toward a phagocytic state (Rothlin et al., 2007). Microglia do not express the third TAM member, Tyro3, which is predominantly found in neurons.

The receptors have been linked previously to Alzheimer’s disease and amyloidosis. For example, Lemke and colleagues reported that microglia around plaques modestly boost Mer expression and massively upregulate Axl (Jul 2019 conference news). 

Super Star. Expression of Axl (green) lights up around dense-core plaques (white). [Courtesy of Huang et al., Nature Immunology.]

In the new paper, first author Youtong Huang further explored what these receptors are doing at plaques. She found that in 9.5-month-old APP/PS1 mice, which have abundant dense-core amyloid deposits, microglia around plaques expressed three times as much Mer protein, and 25 times as much Axl, as did microglia farther away. Meanwhile, immunostaining for the TAM ligand, Gas6, lit up all dense-core plaques. Gas6 constitutively binds TAMs, forming a kind of hybrid receptor, Lemke noted.

Gas6 alone does not activate signaling, however. For that, the TAM/Gas6 complex must bind phosphatidylserine, a phospholipid found on the inside of cell membranes. In dying or damaged cells, phosphatidylserine flips to the surface, acting as an “eat me” signal for microglia. The authors found that every plaque in APP/PS1 mice was coated with phosphatidylserine. Thus, plaques presented all the necessary stimuli for phagocytosis.

What happens when the system is disrupted? In APP/PS1 mice with both Axl and Mer knocked out, no Gas6 protein was present in the brain, even though its mRNA levels in microglia did not change. The data suggest that Gas6 is only stable when bound to Axl or Mer, Lemke noted. This protein-level regulation also explains why previous RNA-Seq studies did not detect any upregulation of Gas6 around plaques, he added. The finding highlights the importance of using proteomics as well as expression studies.

A New Paradigm: Microglia as Plaque Builders
Without Axl, Mer, and Gas6, Lemke expected to see far more plaques in the double-knockout mice. After all, microglia take up and degrade Aβ, right? To his surprise, he found about one-third fewer dense-core plaques at 12 months of age. By brain area, the difference appeared even more dramatic, with plaques largely wiped out, though the authors did not quantitate total amyloid burden (see image below). At the same time, the double knockouts had about 50 percent more diffuse cotton-wool plaques. They also developed twice as much cerebral amyloid angiopathy (CAA) as did control APP/PS1 mice. To Lemke, the findings suggest that microglial phagocytosis changes the nature of amyloid accumulation, helping pack the peptide into tight parenchymal deposits.

Phagocytosis Packs Plaque. In year-old APP/PS1 mice with phagocytosis receptors knocked out (right), few plaques (white) form compared to age-matched controls (left). [Courtesy of Huang et al., Nature Immunology.]

How do these cells do this? The authors followed fluorescently labeled microglia in 16-month-old APP/PS1 mice using live two-photon imaging. They watched microglia surround plaques and extend processes toward them. These microglia had a rounded, amoeboid shape that bespoke activation. They contained large quantities of internalized amyloid in their endosomes and lysosomes, making up almost 10 percent of their cell volume. On the other hand, when APP/PS1 mice lacked Axl and Mer, few microglia gathered around plaques, and these had more ramified processes that actively probed their environment, indicative of their normal surveillance phenotype. These microglia took up only a tenth as much amyloid as did their counterparts. Meanwhile, the halo of dystrophic neurites around plaques was 10 times larger than in control APP-PS1 mice.

To Lemke, the data suggest a model in which Axl- and Mer-positive microglia around plaques take up loosely organized amyloid and shunt it to lysosomes. In this acidic environment, amyloid packs together. Microglia then release this material either by exocytosis or cell death, resulting in the formation of dense-core plaques (see image at top of story).

“To me, this makes biological sense and fits with Aβ chemistry,” Zetterberg noted. He pointed out that amyloid aggregation is known to accelerate at a low pH like that found in lysosomes.

The model fits with other recent data implicating microglia in plaque formation. Jaime Grutzendler at Yale University, New Haven, Connecticut, found that microglia encase dense-core plaques, keeping them compact (May 2016 news). Michael Heneka at the University of Bonn, Germany, reported that microglia release protein complexes known as ASC specks that seed plaques (Dec 2017 news). Researchers led by Inhee Mook-Jung at the Seoul National University College of Medicine used two-photon imaging to figure out that microglia in 5XFAD mice promote plaque growth when they die and release accumulated Aβ into the extracellular space (Baik et al., 2016). 

Most dramatically, perhaps, Kim Green at the University of California, Irvine, killed off all microglia in young 5XFAD mice and found that almost no plaques formed, but massive CAA developed instead (Sep 2019 news). Green noted that the findings from the Axl and Mer double knockouts closely matched his data. “It’s almost a complete replication,” he told Alzforum. “It highlights how many of the functions of microglia might be mediated by just one or two genes.”

Then There's TREM2
Speaking of microglial genes, how do TAM receptors relate to TREM2? This receptor has broad effects on microglial survival, motility, and gene expression, and scientists believe it orchestrates the disease-associated microglial (DAM) phenotype found around plaques in mice. Some TREM2 genetic variants triple a person’s chances of getting AD (Nov 2012 news). Notably, DAM microglia turn up both Axl and Mer expression. Conversely, TREM2 knockouts fail to boost Axl, suggesting this receptor acts downstream of TREM2 (Keren-Shaul et al., 2017). 

Lemke’s data support this hierarchy. Microglia in the TAM double knockouts were still able to adopt a DAM phenotype, although expression of some of the characteristic genes was blunted. With regard to phagocytosis, however, the effect of TAM knockout was far greater than that of TREM2. Loss of Axl and Mer lowered phagocytosis 10-fold, compared to two- to threefold for TREM2 knockouts. Lemke believes these TAM receptors mediate the effects of TREM2 on phagocytosis.

Other data jibe with the idea that TREM2 and TAM receptors act in the same pathway. Knocking out TREM2 in mice leads to larger, more diffuse plaques similar to those seen in Axl/Mer double knockouts (Jan 2019 news). Conversely, TREM2 activation helps clear plaque halos and ameliorate axon and dendrite damage while leaving dense-core plaques intact (Mar 2020 news; Jun 2020 news). 

“The phenotype the authors describe is surprisingly similar to TREM2-deficient microglia, arguing that AXL/MERTK are essential executing molecules of the DAM program in amyloid plaque clearance,” Mikael Simons at Technical University Munich wrote to Alzforum (full comment below). Marco Colonna and Yingyue Zhou at Washington University in St. Louis agreed, “We are also intrigued by parallels and differences between the Axl-/- Mer-/- model and the TREM2-deficient model.”

To Remove Plaques or Not?
Do dense-core plaques help protect the brain? Evidence so far is limited. Huang and colleagues tested APP/PS1 mice in a fear-conditioning paradigm. They found the TAM double knockouts to be more forgetful than their counterparts with functioning Axl and Mer, hinting at some negative effects from having more diffuse Aβ and CAA. If that is the case, busting up dense-core plaques with, say, an antibody therapy might be counterproductive, Lemke noted. Researchers have debated for years if plaques are toxic or protective and if breaking them up would be a good strategy (Lee et al., 2004; Dec 2009 news). 

Alternatively, getting rid of plaques could simply be a waste of time. Charlie Glabe at UC Irvine believes the presence of phosphatidylserine on plaques fits with the idea that they are the remnants of dead neurons (Mar 2013 conference news). “If plaques represent ‘tombstone markers’ of antecedent pathology, targeting their removal maybe analogous to removing the trash after your house burns down,” he wrote to Alzforum (full comment below). Glabe noted that one way to test this idea would be to measure whether the remaining plaques in Axl/Mer double-knockout mice contain neuronal markers, which could suggest they arose through neuron death rather than microglial phagocytosis.

Another key question for AD therapy is how the presence of tau tangles would change the picture. The amyloidosis models used to date do not develop tangles. “We need to understand what happens when we have both pathologies present,” Green told Alzforum. Some data suggest microglial states are more complicated in tauopathy than in amyloidosis models (Dec 2020 news). 

It is unclear if the findings will translate to people. For example, the human brain does not have an exact parallel to mouse DAM microglia (May 2019 news; Dec 2020 news). However, Huang and colleagues found Gas6 coating plaques in postmortem sections from human AD brain. In the ADNI cohort, researchers have found that high amounts of Axl in cerebrospinal fluid correlated with amyloid deposition in the brain, suggesting the same phagocytic machinery may be at work in people (Mattsson et al., 2013). 

Zetterberg noted that this CSF Axl finding was unexpected and has not been pursued yet. “This is something we need to look into further,” he told Alzforum. Ideally, a future study would compare CSF Axl to longitudinal amyloid PET imaging to tease out Axl’s relationship to plaque deposition, he said.

A Heretical Idea
If dense-core plaques do protect the brain from amyloid toxicity, then stimulating plaque construction could be more beneficial than breaking them up, Lemke suggested. For example, the PPARγ agonist pioglitazone stimulates phagocytosis by boosting Mer expression (Dec 2012 news; Savage et al., 2015). However, pioglitazone and the related compound rosiglitazone have been tried as AD therapies with no success (Sep 2010 news; Jan 2018 news; Nov 2018 conference news). 

Likewise, the cancer drug bexarotene boosts Axl and Mer in microglia around plaques, but has not panned out as an AD therapy (Feb 2015 conference news; Feb 2016 news).—Madolyn Bowman Rogers

Comments

  1. Mer and Axl are phagocytic receptors expressed on microglia. During AD, Axl is highly upregulated in activated microglia, while Mer has a high baseline expression. In this exciting paper, Huang et al. presented provocative results showing that deficiency in Mer and Axl results in less microglia activation and fewer dense-core plaques in an AD mouse model. Contrary to our intuition that microglia help clear Aβ plaques, these results suggest that microglial phagocytosis promotes formation of dense-core plaque. Moreover, this study suggests that when it comes to therapeutics, interventions that reduce microglia phagocytosis may be effective in the treatment of AD.

    One interesting question raised by this study is which form of Aβ is neurotoxic. This study finds that dense-core plaques are neurotoxic; however, other studies found that fibrillar plaques are. We are also intrigued by the parallels and differences between the Axl-/- Mer-/- model and the TREM2-deficient model. We saw a similar phenotype of more diffuse plaques in the 5xFAD amyloid mouse model deficient in Trem2. Moreover, similar to the defective microglial clustering observed in Trem2-/- amyloid mouse models, Axl-/- Mer-/- microglia fail to encapsulate plaques in APP/PS1 mice. And yet, despite some reduction in DAM gene expression in the Axl-/- Mer-/- mice, the DAM transcription cluster is adequately present as revealed by single-cell RNA-Seq, unlike the almost complete absence of DAM clusters after TREM2 ablation.

    We also noted that TREM2 expression is not affected by genetic ablation of Axl and Mer.

    Altogether, these results suggest that TAM receptors and TREM2 have different functions, with TREM2 being necessary but not sufficient for activation of DAM signature. The fact that Axl-/- Mer-/- microglia are less activated compared to wild-type microglia in the APP/PS1 background is likely due to these microglia being farther away from plaques. Then the interesting question remains as to why the TAM receptor-deficient microglia are distant from plaques. What are the signals driving microglia migration toward the amyloid deposits?

    View all comments by Marco Colonna View all comments by Yingyue Zhou

  2. The TAM receptors Axl and Mer play an instrumental role in phagocytosis of cellular debris in the periphery of apoptotic cells generated during adult neurogenesis, as shown previously by Dr Lemke’s group (Fourgeaud et al., 2016). Axl is one of the hallmark genes upregulated in DAM/MGnD microglia subset in Aβ models (Keren-Shaul et al., 2017; Krasemann et al., 2017). We showed, in addition, that Axl protein is enriched in Stat1+ microglia surrounding plaques as part of innate interferon response, and that microglia wrapping around neuritic plaques in human brains also positively express AXL protein (Roy et al., 2020). 

    Here, Huang et al. set out to assess the role of TAM receptors in mouse models of AD. They show that Axl expression increases with age specifically in plaque-associated microglia in APP/PS1 mice. Along with the increase in Axl, they observe an upregulation of its ligand Gas6 and of the co-ligand phosphotidylserine in areas surrounding plaques. Using two photon in vivo live imaging, they show that APP/PS1;Axl-/-Mertk-/- microglia are deficient in detecting, binding, and phagocytizing amyloid plaques, highlighting the importance of these receptors for microglia to engage with plaques. Interestingly, between the two TAM receptors, Mer seems to be primarily accountable for plaque uptake and dense-core modification by microglia, which leaves an interesting question of any functional involvement of Axl around the plaques, besides promoting Gas6 deposition.

    Unexpectedly, the authors observe a drastic and specific reduction of compact plaques in APP/PS1 animals lacking the TAM receptors. Based on their single-cell transcriptomics data, they found genes related to DAM, lipid metabolism, and MHC class II antigen presentation were blunted in APP/PS1;Axl-/-Mertk-/- compared to APP/PS1. However, they didn’t observe any changes in cytokine and chemokine expression, emphasizing that the phenotypes they report are not a result of dampened neuroinflammation.

    Overall, the authors propose a model in which phosphotidylserine-Gas6-Axl complex activates TAM driving phagocytosis of plaques. Once internalized, the plaques are transferred to lysosomes, where the acidic environment promotes the aggregation of insoluble and protease resistant fibrils. These can be delivered via exocytosis or by cell death and contribute to formation and growth of dense-core plaques. In this regard, it would be interesting to understand if dead cells accrue preferentially in aged Mer-/-Axl-/- brain, thus contributing to certain aspects of the phenotypic observations in conjunction with neuritic plaques.

    This is a landmark study that links the core molecular machinery of phagocytes to amyloid pathology. The model the authors put forward is provocative and challenges our current understanding of microglia and plaque interaction, which surely will inspire further investigation and deeper dissection in the future.

    References:

    Fourgeaud L, Través PG, Tufail Y, Leal-Bailey H, Lew ED, Burrola PG, Callaway P, Zagórska A, Rothlin CV, Nimmerjahn A, Lemke G. TAM receptors regulate multiple features of microglial physiology. Nature. 2016 Apr 14;532(7598):240-244. Epub 2016 Apr 6 PubMed.

    Keren-Shaul H, Spinrad A, Weiner A, Matcovitch-Natan O, Dvir-Szternfeld R, Ulland TK, David E, Baruch K, Lara-Astaiso D, Toth B, Itzkovitz S, Colonna M, Schwartz M, Amit I. A Unique Microglia Type Associated with Restricting Development of Alzheimer's Disease. Cell. 2017 Jun 15;169(7):1276-1290.e17. Epub 2017 Jun 8 PubMed.

    Krasemann S, Madore C, Cialic R, Baufeld C, Calcagno N, El Fatimy R, Beckers L, O'Loughlin E, Xu Y, Fanek Z, Greco DJ, Smith ST, Tweet G, Humulock Z, Zrzavy T, Conde-Sanroman P, Gacias M, Weng Z, Chen H, Tjon E, Mazaheri F, Hartmann K, Madi A, Ulrich JD, Glatzel M, Worthmann A, Heeren J, Budnik B, Lemere C, Ikezu T, Heppner FL, Litvak V, Holtzman DM, Lassmann H, Weiner HL, Ochando J, Haass C, Butovsky O. The TREM2-APOE Pathway Drives the Transcriptional Phenotype of Dysfunctional Microglia in Neurodegenerative Diseases. Immunity. 2017 Sep 19;47(3):566-581.e9. PubMed.

    Roy ER, Wang B, Wan YW, Chiu G, Cole A, Yin Z, Propson NE, Xu Y, Jankowsky JL, Liu Z, Lee VM, Trojanowski JQ, Ginsberg SD, Butovsky O, Zheng H, Cao W. Type I interferon response drives neuroinflammation and synapse loss in Alzheimer disease. J Clin Invest. 2020 Apr 1;130(4):1912-1930. PubMed.

    View all comments by Wei Cao View all comments by Manasee Gedam View all comments by Hui Zheng

  3. Huang et al. report, in an elegant and detailed study, the central roles played by the TAM receptors Axl and Mer in amyloidogenic models of AD. There are a few surprises that will provoke re-evaluation of exactly how microglia act to actively influence the deposition and remodeling of amyloid plaques in the brains of murine models of AD. The most remarkable finding is that genetic inactivation of both Mer and Axl results in a paradoxical 35 percent decrease in the number of dense-core, thioS-positive plaques in 12-month-old mice. This effect is due principally to the action of Mer, and preferentially affects small plaques.

    The authors posit that dense-core plaque formation requires its “construction” and compaction from more diffusely organized fibrillar forms of Aβ. This is rather different from the conventional view of microglial phagocytic trimming and remodeling of diffuse plaques, which was well documented in Yuan et al. and other publications (Yuan et al., 2016). They note that it had been suggested that plaques might arise from death of microglia with deposition of their undigested phagocytic Aβ cargo, analogous to the earlier suggestion that neurons bearing intraneuronal Aβ found in these models die and then seed plaque formation (Moon et al., 2012). It remains unclear exactly how the receptor’s phagocytic functionality is modulated to drive compact plaque formation.

    The study clearly documents that loss of Axl and Mer dramatically reduces the ability of microglia to detect amyloid deposits and mount a complex cellular response, including changes in morphology, gene expression, and phagocytosis. Remarkably, the loss of Axl and Mer elicits effects on amyloid phagocytosis and plaque density that are much larger than that of TREM2.

    In addition, the manuscript adds interesting detail to the involvement of these TAM receptors in several aspects of disease pathogenesis in the animal models. These include the preferential expression of Axl in plaque-associated microglia, reduction in plaque-associated microglia, suppression of proliferation, increased neuritic dystrophy in the Axl/Mer-deficient mice, as well as an increase in cerebral amyloid angiopathy.

    Overall, this study adds considerable detail into the actions of the TAM receptors and reinforces an understanding of their importance in microglial biology in AD. The caliber of the work is terrific, a characteristic feature of the work from the Lemke lab. 

    References:

    Moon M, Hong HS, Nam DW, Baik SH, Song H, Kook SY, Kim YS, Lee J, Mook-Jung I. Intracellular Amyloid-β Accumulation in Calcium-Binding Protein-Deficient Neurons Leads to Amyloid-β Plaque Formation in Animal Model of Alzheimer's Disease. J Alzheimers Dis. 2012 Jan 1;29(3):615-28. PubMed.

    Yuan P, Condello C, Keene CD, Wang Y, Bird TD, Paul SM, Luo W, Colonna M, Baddeley D, Grutzendler J. TREM2 Haplodeficiency in Mice and Humans Impairs the Microglia Barrier Function Leading to Decreased Amyloid Compaction and Severe Axonal Dystrophy. Neuron. 2016 May 18;90(4):724-39. PubMed.

    View all comments by Gary Landreth

  4. This paper by Huang et al. reports a number of interesting observations that provide significant insights into the role of microglia in Alzheimer’s disease and into the mechanism of plaque biogenesis that have a number of implications for the amyloid hypothesis and amyloid-based therapeutic strategies.

    The authors report that the TAM system (Tyr3, Axl, Mer), which mediates the phagocytosis and degradation of cellular debris from dead and dying cells, is required for the recognition and recruitment of microglia to plaques, and that it promotes the formation of dense-core and neuritic plaques. Because phosphatidylserine (PS), an essential co-ligand for the TAM receptors, is only found on the cytosolic leaflet of normal cell membranes or on the surface of apoptotic and dying cells and intracellular vesicular debris, this suggests that a substantial amount of the aggregated precursor amyloid Aβ for neuritic plaques is contained in vesicles derived from dying neurons as we have previously suggested (Pensalfini et al., 2014; Sosna et al., 2018). This has a number of implications for the amyloid hypothesis, for the significance of neuritic plaques, and for the therapeutic targeting of amyloid plaques. 

    If neuritic plaques are derived from amyloid that had first aggregated within neurons and was released as a consequence of cell death, then plaque formation is the result of microglial accumulation and processing of “dystrophic neurites” that contain lysosome- and autophagosome-related vesicles packed with material reactive with aggregation-specific monoclonal antibodies for Aβ, and not the other way around. If neuritic plaques arise from dead and dying neurons, then these plaques really do represent “tombstone markers” of antecedent pathology, so targeting their removal maybe analogous to removing the trash after your house burns down. It looks much nicer, but it does not really help your living situation.

    A prediction of this model is that the remaining plaques in the APP/PS1Axl−/−Mertk−/- mice would contain more obvious neuronal remnants and markers in the “trash” that is not removed by microglia. If neuritic plaque amyloid aggregates initially within neurons, this could also explain why γ-secretase inhibitors (GSIs), which inhibit the secretion of Aβ from neurons, exacerbate cognitive decline in human clinical trials, since inhibiting Aβ secretion may lead to intraneuronal Aβ aggregation (Pensalfini et al., 2014). 

    Although these studies define a novel role for TAM receptors in neuritic and dense-core plaque formation, this may not be the only role of microglia in plaque biogenesis. In the double-knockout APP/PS1Axl−/−Mertk−/- mice, dense-core plaque formation was inhibited by less than 50 percent, but if microglia are ablated during the entire time period of intraneuronal amyloid accumulation and plaque formation, both intraneuronal amyloid and all types of plaques are inhibited by 90 percent (Sosna et al., 2018). 

    References:

    Pensalfini A, Albay R 3rd, Rasool S, Wu JW, Hatami A, Arai H, Margol L, Milton S, Poon WW, Corrada MM, Kawas CH, Glabe CG. Intracellular amyloid and the neuronal origin of Alzheimer neuritic plaques. Neurobiol Dis. 2014 Nov;71:53-61. Epub 2014 Aug 1 PubMed.

    Sosna J, Philipp S, Albay R 3rd, Reyes-Ruiz JM, Baglietto-Vargas D, LaFerla FM, Glabe CG. Early long-term administration of the CSF1R inhibitor PLX3397 ablates microglia and reduces accumulation of intraneuronal amyloid, neuritic plaque deposition and pre-fibrillar oligomers in 5XFAD mouse model of Alzheimer's disease. Mol Neurodegener. 2018 Mar 1;13(1):11. PubMed.

    View all comments by Charles Glabe

  5. Microglia respond to amyloid plaques by converting into a disease-associated state (DAM) that depends on TREM2. The DAM signature comprises a large set of genes including AXL, known to be involved in clearing apoptotic cells in which phosphatidylserine is exposed on the surface. Huang et al. now show that AXL/MERTK is involved not only in clearing cells undergoing apoptosis, but also in clearing amyloid plaques.

    They find that microglia phagocytose amyloid decorated with phosphatidylserine in an AXL/MERTK-dependent manner. Surprisingly, they find that AXL/MERTK-dependent phagocytosis of amyloid does not inhibit but rather promotes the formation of dense-core plaques. The phenotype the authors describe is surprisingly similar to TREM2-deficient microglia arguing that AXL/MERTK are essential executing molecules of the DAM program in amyloid plaque clearance. Previously, the increase of dense-core plaques has been shown to be a function of TREM2-dependent APOE secretion. However, in this study the authors propose that AXL/MERTK-dependent phagocytosis of Aβ may lead to aggregation within lysosomes, which is followed by the subsequent release of indigestible amyloid, which may form the core of the plaques.

    This is an important study not only for our understanding of phagocytic receptors in microglia, but in particular for our understanding their role in amyloid clearance. In future studies, it will be interesting to understand the role of phosphatidylserine in Aβ deposition, and to explore the role of seeding factors released or induced after phagocytosis.

    View all comments by Mikael Simons
    • User Profile Image Oleg Butovsky
      Brigham and Women's Hospital/Harvard Medical School
    • Posted:

    This is a very interesting study, demonstrating that TAM receptors, such as Axl and Mertk, are required for microglia phagocytosis of Aβ, resulting in the formation of dense-core plaques (for review describing differences between dense-core and diffuse plaques, see DeTure and Dickson, 2019). However, the identification of Mertk and not Axl as an essential receptor in microglial phagocytosis was recently described (Damisah et al., 2020). Moreover, we previously showed that expression of Mertk is significantly suppressed and expression of Axl is induced in Aβ-plaque associated microglia (see Krasemann et al., 2017, and mouse RNA-Seq data therein below). Thus, it is not clear why the authors mainly focused on Mertk/Axl double knockouts.

    In addition, there is no discrimination between the contribution of microglia and peripheral monocytes to pathology, as the authors used global genetic deletion. Furthermore, it seems that DAM genes are slightly downregulated in the double knockouts, although the authors interpreted that there was no change in DAM signature. Overall, the fact that microglial phagocytosis associated with increased plaque burden is an interesting observation that requires further investigation to determine whether microglial phagocytosis is beneficial or detrimental in AD.

    References:

    DeTure MA, Dickson DW. The neuropathological diagnosis of Alzheimer's disease. Mol Neurodegener. 2019 Aug 2;14(1):32. PubMed.

    Damisah EC, Hill RA, Rai A, Chen F, Rothlin CV, Ghosh S, Grutzendler J. Astrocytes and microglia play orchestrated roles and respect phagocytic territories during neuronal corpse removal in vivo. Sci Adv. 2020 Jun;6(26):eaba3239. Epub 2020 Jun 26 PubMed.

    Krasemann S, Madore C, Cialic R, Baufeld C, Calcagno N, El Fatimy R, Beckers L, O'Loughlin E, Xu Y, Fanek Z, Greco DJ, Smith ST, Tweet G, Humulock Z, Zrzavy T, Conde-Sanroman P, Gacias M, Weng Z, Chen H, Tjon E, Mazaheri F, Hartmann K, Madi A, Ulrich JD, Glatzel M, Worthmann A, Heeren J, Budnik B, Lemere C, Ikezu T, Heppner FL, Litvak V, Holtzman DM, Lassmann H, Weiner HL, Ochando J, Haass C, Butovsky O. The TREM2-APOE Pathway Drives the Transcriptional Phenotype of Dysfunctional Microglia in Neurodegenerative Diseases. Immunity. 2017 Sep 19;47(3):566-581.e9. PubMed.

    View all comments by Oleg Butovsky

  7. This elegant paper provides substantial evidence for a critical role of microglia in building up dense plaques in Aβ overproducing transgenic mice. Though the direct translatability of results obtained using the aggressive APP/PS1-based animal model to pathogenesis in sporadic AD might be limited, the results support the notion that plaque dissolution strategies, or impeding dense plaque formation, should not be a primary goal for therapies targeting Aβ.

    Indeed, early but often-overlooked studies have revealed poor correlation between cognitive decline and dense plaque load in AD brains (Terry et al., 1991), and the lack of clinical efficacy of merely dissolving plaques was established in the early AN1792 clinical trial (Bayer et al., 2005; Nicoll et al., 2019). 

    Based on these antecedents and on the current paper from the Lemke group, we are encouraged to focus our attention on early misfolded oligomeric species as the central molecular entities responsible for the pathogenic role of Aβ in AD (Cline et al., 2018). From a mechanistic perspective, the optimal profile for future Aβ therapies in prevention or treatment of spontaneous AD should be the neutralization of early misfolded oligomers with high selectivity, as opposed to physiological Aβ monomer turnover (Hillen, 2019) or plaque dissolution. 

    In the past, the main argument against developing those antibodies was the unavailability of practical biomarkers to take such an antibody to a decision point in early clinical studies with a  limited number of patients, time, and cost. With the breakthrough biomarker developments in the tau area (e.g., Barthélemy et al., 2020) during the last two years, we should now have reliable and easy blood-based readouts to test the mechanistic efficacy of moving these early aberrant tau species induced by Aβ oligomers via conformer-specific antibodies in a relevant clinical setting. Highly Aβ oligomer-specific antibodies and vaccines have been described preclinically (Hillen et al., 2010; Gibbs et al., 2019). 

    References:

    Terry RD, Masliah E, Salmon DP, Butters N, DeTeresa R, Hill R, Hansen LA, Katzman R. Physical basis of cognitive alterations in Alzheimer's disease: synapse loss is the major correlate of cognitive impairment. Ann Neurol. 1991 Oct;30(4):572-80. PubMed.

    Bayer AJ, Bullock R, Jones RW, Wilkinson D, Paterson KR, Jenkins L, Millais SB, Donoghue S. Evaluation of the safety and immunogenicity of synthetic Abeta42 (AN1792) in patients with AD. Neurology. 2005 Jan 11;64(1):94-101. PubMed.

    Nicoll JA, Buckland GR, Harrison CH, Page A, Harris S, Love S, Neal JW, Holmes C, Boche D. Persistent neuropathological effects 14 years following amyloid-β immunization in Alzheimer's disease. Brain. 2019 Jul 1;142(7):2113-2126. PubMed.

    Cline EN, Bicca MA, Viola KL, Klein WL. The Amyloid-β Oligomer Hypothesis: Beginning of the Third Decade. J Alzheimers Dis. 2018;64(s1):S567-S610. PubMed.

    Hillen H. The Beta Amyloid Dysfunction (BAD) Hypothesis for Alzheimer's Disease. Front Neurosci. 2019;13:1154. Epub 2019 Nov 7 PubMed.

    Barthélemy NR, Bateman RJ, Hirtz C, Marin P, Becher F, Sato C, Gabelle A, Lehmann S. Cerebrospinal fluid phospho-tau T217 outperforms T181 as a biomarker for the differential diagnosis of Alzheimer's disease and PET amyloid-positive patient identification. Alzheimers Res Ther. 2020 Mar 17;12(1):26. PubMed.

    Hillen H, Barghorn S, Striebinger A, Labkovsky B, Müller R, Nimmrich V, Nolte MW, Perez-Cruz C, van der Auwera I, van Leuven F, van Gaalen M, Bespalov AY, Schoemaker H, Sullivan JP, Ebert U. Generation and therapeutic efficacy of highly oligomer-specific beta-amyloid antibodies. J Neurosci. 2010 Aug 4;30(31):10369-79. PubMed.

    Gibbs E, Silverman JM, Zhao B, Peng X, Wang J, Wellington CL, Mackenzie IR, Plotkin SS, Kaplan JM, Cashman NR. A Rationally Designed Humanized Antibody Selective for Amyloid Beta Oligomers in Alzheimer's Disease. Sci Rep. 2019 Jul 8;9(1):9870. PubMed.

    View all comments by Heinz Hillen View all comments by Neil Cashman View all comments by Sergio Ferreira View all comments by William Klein
    • User Profile Image Steve Barger
      University of Arkansas for Medical Sciences
    • Posted:

    This is elegant and intriguing work. As one possible next step, it might be important to determine whether macrophage-like cells derived from circulating monocytes express the same constellation of TAM receptors, and behave in the same manner, toward diffuse Aβ deposits.

    View all comments by Steve Barger

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References

News Citations

  1. Do Microglia Finish Off Stressed Neurons Before Their Time?
  2. Barrier Function: TREM2 Helps Microglia to Compact Amyloid Plaques
  3. Do Microglia Spread Aβ Plaques?
  4. Are Microglia Plaque Factories?
  5. Enter the New Alzheimer’s Gene: TREM2 Variant Triples Risk
  6. Without TREM2, Plaques Grow Fast in Mice, Have Less ApoE
  7. Paper Alert: Mouse TREM2 Antibody Boosts Microglial Plaque Clean-Up
  8. In Mice, Activating TREM2 Tempers Plaque Toxicity, not Load
  9. Long Life With Tight Plaques—Repressing IGF-1 Protects AD Mice
  10. Like Star Born of Supernova, Plaque Born of Exploded Neuron?
  11. Microglia in Tauopathy: Not Just Homeostatic Versus DAM
  12. When It Comes to Alzheimer’s Disease, Do Human Microglia Even Give a DAM?
  13. Most Detailed Look Yet at Activation States of Human Microglia
  14. Can Phagocytosis, Memory Effects Revive Diabetes Meds?
  15. Trial Updates: B Vitamin Back in Vogue? Diabetes Drug Less Sweet
  16. There’s No Tomorrow for TOMMORROW
  17. Trials of Diabetes-Related Therapies: Mainly a Bust
  18. United in Confusion: TREM2 Puzzles Researchers in Taos
  19. Bexarotene—First Clinical Results Highlight Contradictions

Research Models Citations

  1. APPswe/PSEN1dE9 (line 85)
  2. 5xFAD (B6SJL)

Paper Citations

  1. Lu Q, Lemke G. Homeostatic regulation of the immune system by receptor tyrosine kinases of the Tyro 3 family. Science. 2001 Jul 13;293(5528):306-11. PubMed.
  2. Scott RS, McMahon EJ, Pop SM, Reap EA, Caricchio R, Cohen PL, Earp HS, Matsushima GK. Phagocytosis and clearance of apoptotic cells is mediated by MER. Nature. 2001 May 10;411(6834):207-11. PubMed.
  3. Fourgeaud L, Través PG, Tufail Y, Leal-Bailey H, Lew ED, Burrola PG, Callaway P, Zagórska A, Rothlin CV, Nimmerjahn A, Lemke G. TAM receptors regulate multiple features of microglial physiology. Nature. 2016 Apr 14;532(7598):240-244. Epub 2016 Apr 6 PubMed.
  4. Rothlin CV, Ghosh S, Zuniga EI, Oldstone MB, Lemke G. TAM receptors are pleiotropic inhibitors of the innate immune response. Cell. 2007 Dec 14;131(6):1124-36. PubMed.
  5. Baik SH, Kang S, Son SM, Mook-Jung I. Microglia contributes to plaque growth by cell death due to uptake of amyloid β in the brain of Alzheimer's disease mouse model. Glia. 2016 Dec;64(12):2274-2290. Epub 2016 Sep 23 PubMed.
  6. Keren-Shaul H, Spinrad A, Weiner A, Matcovitch-Natan O, Dvir-Szternfeld R, Ulland TK, David E, Baruch K, Lara-Astaiso D, Toth B, Itzkovitz S, Colonna M, Schwartz M, Amit I. A Unique Microglia Type Associated with Restricting Development of Alzheimer's Disease. Cell. 2017 Jun 15;169(7):1276-1290.e17. Epub 2017 Jun 8 PubMed.
  7. Lee HG, Casadesus G, Zhu X, Takeda A, Perry G, Smith MA. Challenging the amyloid cascade hypothesis: senile plaques and amyloid-beta as protective adaptations to Alzheimer disease. Ann N Y Acad Sci. 2004 Jun;1019:1-4. PubMed.
  8. Mattsson N, Insel P, Nosheny R, Zetterberg H, Trojanowski JQ, Shaw LM, Tosun D, Weiner M. CSF protein biomarkers predicting longitudinal reduction of CSF β-amyloid42 in cognitively healthy elders. Transl Psychiatry. 2013;3:e293. PubMed.
  9. Savage JC, Jay T, Goduni E, Quigley C, Mariani MM, Malm T, Ransohoff RM, Lamb BT, Landreth GE. Nuclear receptors license phagocytosis by trem2+ myeloid cells in mouse models of Alzheimer's disease. J Neurosci. 2015 Apr 22;35(16):6532-43. PubMed.

Further Reading

Primary Papers

  1. Huang Y, Happonen KE, Burrola PG, O'Connor C, Hah N, Huang L, Nimmerjahn A, Lemke G. Microglia use TAM receptors to detect and engulf amyloid β plaques. Nat Immunol. 2021 May;22(5):586-594. Epub 2021 Apr 15 PubMed.

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