Cell: A novel mechanism by which microglia lead to increased Aβ deposition in Alzheimer's disease, neuropathological exacerbation, and cognitive deficits

Recent research has revealed the key role
of microglia and their receptors in influencing the accumulation of Aβ protein and myelin debris in neurodegenerative diseases.
But the key intracellular molecules that coordinate the neuroprotective function of microglia remain unclear
.

The team of John R.
Lukens from the Center for Brain Immunology and Glial Cells in the Department of Neuroscience at the University of Virginia published an article in the journal CELL in October 2022 entitled "SYK coordinates neuroprotective microglial responses in neurodegenerative disease", which pointed out that SYK regulates the activation and function of microglia.
Impaired function is associated
with worsening of neurodegenerative disease models such as Alzheimer's disease and multiple sclerosis.

The absence of SYK signal exacerbates the disease in AD and MS models, SYK regulates the proliferation and binding to Aβ of microgel, a key regulator of microgel activation and AKT/GSK3b signaling, and the phagocytosis clearance of neurotoxic substances by microglia depends on SYK
.

Figure 1: Microglial SYK deletion leads to increased Aβ burden and plaque composition changes in 5xFAD mice

1.
SYK signaling in microglia limits Aβ accumulation

The researchers used the transgenic mouse strategy to construct an AD model mouse with conditional knockout of small rubber SKY 5xFADSykΔDMG (Sykfl/flCx3cr1ERT2Cre 5×FADmice), and gave tamoxifen 2 weeks after weaning to induce SKY deletion, the results showed that the small rubber SKY signal played a key role in controlling Aβ aggregation and compactness, and the accumulation of Aβ protein in the cortex, hippocampus and thalamus of SYK signal deficient mice increased , the efficiency of compacting Aβ into plaques is lower, Aβ plaques are more filamentous and dense, and the sphericality of abnormal plaques is lower than that of the control group
.
ELISA results showed that the solubility levels of Aβ40 and Aβ42 in the model group were higher than those in the control group
.

Figure 2: SYK deletion of microglia in 5xFAD mice negatively affects neuronal health and aggravates AD-related behavior

2.
SYK deletion in microglia leads to deterioration of neuronal health and memory impairment in AD model mice

The researchers further assessed the health
of neurons using immunofluorescence staining.
SYK signal-deficient mice with increased dystrophic neurites associated with plaques in the cerebral cortex, hyperphosphorylated proteins around the plaques accumulate highly, producing neurotoxicity and increased
cell death.

Behavioral experiments suggest spatial learning disabilities and memory impairment in the Morris water maze test, and higher levels of risk-taking and exploratory behavior
in the elevated cross maze experiment.

Figure 3: SYK deficiency limits microglial proliferation and its association with Aβ plaques

3.
SYK regulates the proliferation of microglia and their binding to Aβ

Using immunofluorescence staining to label microglia, Aβ proteins, and related apoptotic proteins, the researchers observed that the plaque-associated microgels of SYK signal-deficient mice were damaged, and the number of microglia was lower than that of the control group, but apoptosis was not the main driver of the decrease in the number of microglia
.

Figure IV: Activation defects of SYK-deficient microglia in 5xFAD mice

4.
Aβ-induced microglial activation is impaired when SYK signal is missing

Sholl analysis was used to evaluate differences in microglial morphology, which showed a significant increase
in branch protrusions in non-plaque-associated microglia in the hippocampus and cortical regions of SYK signal-deficient mice.
Principal component analysis showed that the loss of SYK signal blocked the change
in the transcriptional pattern of microglia in the presence of Aβ.

Loss of SYK signaling leads to significant changes
in microglial genes.
RNA-SEQ results showed that the loss of SKY signal led to a significant decrease
in lipoprotein esterase expression in microglia.
KEGG analysis showed that many down-regulated genes were associated with neurodegeneration, with DAM stage 2 gene down-regulation being the most significant
.
PI3K/AKT signaling regulates processes and pathways
related to microglial activation in AD.
Magnetic bead sorting experiments showed that SKY signal loss led to a decrease
in the phosphorylation levels of AKT and GS3Kβ.
Figure 5: SYK plays a key role in microglial uptake and phagocytosis of antibodies

5.
SYK regulates the phagocytosis of Aβ

The researchers measured the phagocytosis of small gums using immunohistochemistry and found that the phagocytosis of small gums without SYK signal was impaired
.
Intraperitoneal injection of brain penetration dyes, labeling of fibrous antibodies, flow cytometry and the use of GSK3b inhibitors have once again confirmed that SYK promotes the phagocytosis of Aβ by small gums, and in the absence of SYK, GSK3β signaling dysregulation can lead to macrophage phagocytosis Aβ dysfunction
.

6.
Exogenous activation of the CLEC7A-SYK signaling pathway promotes Aβ clearance

CLEC7A is a receptor
that plays an important role in the activation of small gums in the pathological response to AD.
SYK is the key to phagocytic dysfunction, so the researchers injected Aβ-D-glucan and CLEC7A ligand into the hippocampal brain region by stereotactic injection of the brain, and detected the level of Aβ by immunofluorescence, and the results showed that the SYK signal stimulated by CLEC7A can enhance the phagocytic response
of microglia.

Figure 6: SYK deletion in microglia hinders EAE DAM formation

7.
SYK regulates DAM production in demyelinating neuroinflammatory diseases

The researchers found that SKY deletion mice experienced heavier paralysis and more severe demyelination
.
Using scRNA-seq, 6 unique populations of microcolloids were discovered
.
Based on the maturation state of the microgel and whether it was affected by SKY signaling, the researchers revealed 3 potential pathways:

1.
From homeostasis to high metabolism and M1 pathway;

2.
To CD36hi pathway;

3.
To the DAM and M2 pathways
.

When the SYK signal is missing, pathway 2 is followed, and pathway 3
is followed when SYK is present.

To better understand the biological processes driven by DAM clusters, plotKEGG and phagocytosis-related GO terms were used using DAM-defining genes, and common terms were found to include "phagosomes", "abnormal phagocytes morphology", and "microglial pathogen phagocytic pathways"
.

Figure 7: Disruption of microglial SYK signaling during demyelinating disease leads to impaired myelin debris accumulation and impaired oligodendrocytes development

8.
Deficient SYK signaling in demyelinating microglia, resulting in myelin debris accumulation and impaired oligodendrocytes proliferation

The damaged basic protein of myelin sheath in SYK-deficient mice increased significantly during demyelination and myelin reformation, and SYK deficiency in small gums had no significant effect
on the number of oligoprolites under homeostasis.
However, it has resulted in significant defects
in impaired myelin clearance in the demyelinating disease model.

Summary

In AD model mice, microglial SYK deletion leads to increased Aβ deposition, neuropathological exacerbation and cognitive deficits
in Alzheimer's disease.
The loss of SYK signal hinders the development of disease-associated microglia, alters AKT/GSK3β signaling, and limits the phagocytosis
of microglia.

In addition, in demyelinating diseases, SYK signaling is indispensable
for phagocytosis of microglia and acquisition of DAM.
Taken together, these results broaden the understanding of key innate immune signaling molecules that promote beneficial microglial function in response
to neurotoxic substances.