Are microglia the masterminds of amyloid plaque formation? In the August 21 Nature Communications, researchers led by Kim Green at the University of California, Irvine, suggest these cells seed amyloid deposits. In a mouse model of amyloidosis, obliterating microglia at a young age all but prevented parenchymal plaques. Instead, Aβ ended up in blood vessels, inducing a cerebral amyloid angiopathy (CAA). Those few plaques that did form in the brain wreaked havoc on surrounding neurites, however, possibly because they were not encapsulated by microglia. “At early stages of Alzheimer’s disease, microglia have a complicated involvement, both protecting and harming the brain,” Green said. He presented some of these data at a recent Keystone symposium (Jul 2019 conference news).
- A second study reports that eliminating microglia abolishes plaque formation.
- Excess Aβ accumulates in blood vessels instead.
- Restoring microglia seeds new plaques, while vascular amyloid remains.
Other researchers said the findings elucidate the contribution of microglia to amyloidosis. “The authors present clear data that the presence of microglia is required to allow amyloid plaque deposition,” Christian Haass at the German Center for Neurodegenerative Diseases in Munich wrote to Alzforum (comment below). Mathias Jucker at the University of Tübingen, Germany, who did a similar study 10 years ago and found the opposite effect on plaques, focused on the vascular component. “To me, the most fascinating point is the increase in CAA … Quite possibly, vascular amyloid could be more damaging to the brain than parenchymal plaques, which would render microglial seeding of parenchymal deposits protective,” he wrote (comment below).
This study was made possible by the development of drugs to selectively kill off microglia. Five years ago, Green and colleagues, in collaboration with scientists at Plexxikon Inc., developed an inhibitor of colony-stimulating factor 1 receptor (CSF1R), an essential microglial survival factor. The inhibitor, PLX3397, almost completely wiped out these cells in wild-type mouse brain. After withdrawing the drug, microglia replenished themselves (Apr 2014 news).
Green and colleagues used PLX3397 to abolish microglia in 10-month-old, plaque-ridden 5xFAD mice. Their plaque burden did not budge, but more neurons and synapses survived and cognition improved (Spangenberg et al., 2016). Meanwhile, Charles Glabe and colleagues at UC Irvine fed PLX3397 to young, pre-plaque 5xFAD mice. This eliminated 80 percent of microglia and prevented 90 percent of plaques, while maintaining normal performance on a fear-conditioning test (Mar 2018 news).
However, PLX3397 inhibited not only CSF1R but also two homologous immune cell receptors. Hence Green’s group collaborated with Plexxikon to redesign it to be more specific and brain-penetrant. The new version, PLX5622, is 20-fold more selective for CSF1R than for homologous receptors, and 20 percent of it enters the brain. Indeed, it eliminated virtually all microglia from wild-type mice though, curiously, these mice maintained normal learning and memory six months after their microglia were gone.
Armed with this new pharmacological tool, Green and colleagues examined what microglial depletion would do to young 5xFAD mice. First author Elizabeth Spangenberg fed PLX5622 to 6-week-old animals until they were either 4 or 7 months old, then analyzed their brains. In the 4-month-old mice, microglia were absent from the cortex, but a few remained in the thalamus, retrosplenial cortex, and subiculum. Unexpectedly, amyloid plaques paralleled this distribution, with almost none in the cortex, but a few near remaining microglia in the other regions. “Because microglia are believed to phagocytose amyloid, we assumed that when we got rid of microglia, more plaques would form. But we saw the opposite,” Green said.
By 7 months of age, microglia hung on only in the subiculum. Still, some plaques persisted in the thalamus and retrosplenial cortex. These were more diffuse and irregularly shaped than plaques in untreated 5xFAD mice and contained little ApoE, consistent with prior reports that microglia are the main source of this protein in plaques (Jan 2019 news). The halo of dystrophic neurites that typically surrounds amyloid plaques was 50 percent larger in PLX5622 mice than in controls. This fits with the idea that microglia form a barrier around plaques that protects surrounding tissue (May 2016 news; Jun 2017).
Meanwhile, the scientists ascertained that the treated 5xFAD mice continued to produce as much amyloid precursor protein and its cleavage products as ever. In both 4- and 7-month-old 5xFAD mice on PLX5622, this excess Aβ peptide deposited in cerebral blood vessels. CAA can cause hemorrhages, and indeed, the researchers found microbleeds in these animals in the thalamus, where vessels are particularly leaky in mouse models of CAA (Davis et al., 2004). Notably, 5xFAD usually develop no CAA, and the untreated controls in this study did not, either. “The vascular pathology was immense,” Green said. “These results almost suggest that [one of] the functions of microglia in the normal brain is to protect us against CAA.”
Seven-month-old 5xFAD mice have no measurable defect in learning; however, they do spend more time in the open arm of an elevated plus maze, a sign of impairment in a normal anxiety behavior for these nocturnal animals. The 5xFAD mice without microglia spent even more time in the open arm.
What happens to the CAA mice if microglia return? Green and colleagues stopped feeding PLX5622 to 12 5xFAD mice at 4 months of age and, a month later, examined the brains. Lo and behold, they were speckled with just as many plaques as those of age-matched 5xFAD mice never exposed to the inhibitor, although their plaques were 25 percent smaller on average. And their CAA remained high, matching that in 5-month-old 5xFAD mice on PLX5622. “The fact that plaques come back is a new finding, and very intriguing,” commented Oleg Butovsky at Brigham and Women’s Hospital, Boston. Butovsky was a reviewer on Green’s paper.
The big question raised by this work is how microglia might promote plaque formation. Opinions vary. Green believes microglia take up extracellular Aβ and induce it to aggregate inside themselves, thus seeding plaques. In support of this, the authors found Aβ aggregates lurking inside microglial lysosomes in 5xFAD and 3xTg mice. These microglia were often located far from plaques, making it unlikely the cells were simply ingesting plaque material. Green and colleagues saw the same phenomenon in samples from postmortem human brains with AD pathology. Some earlier research also reported finding amyloid fibrils forming inside microglia, and suggested a role for these cells in seeding plaques (Wisniewski et al., 1990; Frackowiak et al., 1992; Stalder et al., 1999).
For his part, Glabe believes amyloid forms inside neurons, because plaques are typically surrounded by neuronal remnants (Mar 2013 conference news). Glabe previously suggested that crosstalk between microglia and neurons might stimulate intraneuronal plaque formation. Haass favors a third idea, that microglia secrete factors, such as ApoE or the protein complexes known as ASC specks, that facilitate extracellular Aβ fibrillization (Dec 2017 news).
Others raised caveats with the findings. Jucker noted that eliminating microglia for four weeks from 3-month-old APPPS1 mice, which more aggressively deposit amyloid than 5xFAD mice, did not dent plaque formation (Grathwohl et al., 2009). “In an aggressive model, microglia may play less of a role in deposition,” Jucker speculated. Butovsky pointed out that eliminating a cell type from the brain changes the milieu and the crosstalk between cells. Therefore, these experiments do not prove that plaque formation is a direct effect of microglia, he said.
Researchers led by Sangram Sisodia at the University of Chicago reported in the August 21 Journal of Neuroscience that PLX5622 eliminates microglia from mice carrying the pathogenic M146V mutation in presenilin 1. The inhibitor rescued neurogenesis, even though these 9-week-old animals had no evidence of amyloid plaques, and it reduced anxiety to wild-type levels, as seen by animals spending more time in lit areas, grooming less, and rearing up more to explore their surroundings. In both PS1 and 5xFAD mice, PLX5622 lowered anxiety.
One thing all the researchers agreed on is that eliminating microglia is no therapeutic strategy for Alzheimer’s disease. Mice are raised in a sterile environment, and so fare well without immune cells in the brain. In people, lack of microglia could lead to infections such as meningitis. Instead, the goal is to figure out what microglial phenotype best wards off Alzheimer’s pathology, Butovsky said. In future work, Green will try to answer this by studying how microglia contribute to plaque formation, and how these cells affect tau pathology.—Madolyn Bowman Rogers
- TREM2, Microglia Dampen Dangerous Liaisons Between Aβ and Tau
- Microglial Magic: Drug Wipes Them Out, New Set Appears
- Wiping Out Microglia Prevents Neuritic Plaques
- Without TREM2, Plaques Grow Fast in Mice, Have Less ApoE
- Barrier Function: TREM2 Helps Microglia to Compact Amyloid Plaques
- Hot DAM: Specific Microglia Engulf Plaques
- Like Star Born of Supernova, Plaque Born of Exploded Neuron?
- Do Microglia Spread Aβ Plaques?
Research Models Citations
- Spangenberg EE, Lee RJ, Najafi AR, Rice RA, Elmore MR, Blurton-Jones M, West BL, Green KN. Eliminating microglia in Alzheimer's mice prevents neuronal loss without modulating amyloid-β pathology. Brain. 2016 Apr;139(Pt 4):1265-81. Epub 2016 Feb 26 PubMed.
- Davis J, Xu F, Deane R, Romanov G, Previti ML, Zeigler K, Zlokovic BV, Van Nostrand WE. Early-onset and robust cerebral microvascular accumulation of amyloid beta-protein in transgenic mice expressing low levels of a vasculotropic Dutch/Iowa mutant form of amyloid beta-protein precursor. J Biol Chem. 2004 May 7;279(19):20296-306. Epub 2004 Feb 25 PubMed.
- Wisniewski HM, Vorbrodt AW, Wegiel J, Morys J, Lossinsky AS. Ultrastructure of the cells forming amyloid fibers in Alzheimer disease and scrapie. Am J Med Genet Suppl. 1990;7:287-97. PubMed.
- Frackowiak J, Wisniewski HM, Wegiel J, Merz GS, Iqbal K, Wang KC. Ultrastructure of the microglia that phagocytose amyloid and the microglia that produce beta-amyloid fibrils. Acta Neuropathol. 1992;84(3):225-33. PubMed.
- Stalder M, Phinney A, Probst A, Sommer B, Staufenbiel M, Jucker M. Association of microglia with amyloid plaques in brains of APP23 transgenic mice. Am J Pathol. 1999 Jun;154(6):1673-84. PubMed.
- Grathwohl SA, Kälin RE, Bolmont T, Prokop S, Winkelmann G, Kaeser SA, Odenthal J, Radde R, Eldh T, Gandy S, Aguzzi A, Staufenbiel M, Mathews PM, Wolburg H, Heppner FL, Jucker M. Formation and maintenance of Alzheimer's disease beta-amyloid plaques in the absence of microglia. Nat Neurosci. 2009 Nov;12(11):1361-3. PubMed.
- AD Genetic Risk Tied to Changes in Microglial Gene Expression
- TREM2, Microglia Dampen Dangerous Liaisons Between Aβ and Tau
- Gently Used: Can Recycled Microglia Receptors Prevent Plaque?
- When It Comes to Alzheimer’s Disease, Do Human Microglia Even Give a DAM?
- Parsing How Alzheimer’s Genetic Risk Works Through Microglia
- In Pathology Cascade, Microglia Rev Up After Plaques but Before Tangles
- Microglia Reveal Formidable Complexity, Deep Culpability in AD
- Spangenberg E, Severson PL, Hohsfield LA, Crapser J, Zhang J, Burton EA, Zhang Y, Spevak W, Lin J, Phan NY, Habets G, Rymar A, Tsang G, Walters J, Nespi M, Singh P, Broome S, Ibrahim P, Zhang C, Bollag G, West BL, Green KN. Sustained microglial depletion with CSF1R inhibitor impairs parenchymal plaque development in an Alzheimer's disease model. Nat Commun. 2019 Aug 21;10(1):3758. PubMed.
- Ortega-Martinez S, Palla N, Zhang X, Lipman E, Sisodia SS. Deficits in Enrichment-Dependent Neurogenesis and Enhanced Anxiety Behaviors Mediated by Expression of Alzheimer's Disease-Linked Ps1 Variants Are Rescued by Microglial Depletion. J Neurosci. 2019 Aug 21;39(34):6766-6780. Epub 2019 Jun 19 PubMed.