Microglia manipulation in Alzheimer’s disease models
The microglial/macrophage response is a key mediator of the immune response in the brain. Microglia can be activated following exogenous or endogenous stimulation by a variety of receptors. Stimulation of these receptors can induce activation of microglia into a ‘classical (M1)’ or ‘alternative (M2)’ phenotype. That microglia play a significant role in eliciting inflammation and clearing toxic products and damaged tissue cannot be disputed, but their direct role in disease progression is unclear. Near complete ablation of microglia, by crossing either APP23 or APP/PS1 mice with CD11b-TK mice, did not show differences in plaque formation and only a very small reduction in diffuse Aβ in the APP23/CD11b-TK model [
55], suggesting more subtle approaches to study their role are necessary.
A number of recent reviews have highlighted the current literature trends and debated the seemingly contradictory results relating to microglial involvement in AD [
56‐
58]. The activation state of microglia and their ability to phagocytose and clear amyloid in the brain seems to be a significant, but contentious, factor. Microglia and macrophages express a number of different receptors that can promote phagocytosis and clearance of Aβ that have been targeted. These include complement receptors, scavenger receptors, and cytokine/chemokine receptors that are associated with pathogen recognition (Table
2). These data can often seem incompatible and contradictory in many cases and yet yield some significant therapeutic targets and emphasize the multi-faceted and heterozygous nature of microglial response in AD from the beginning of the disease throughout its progression. Specific manipulation of signaling factors associated with a shift to the M2 phenotype is reported to promote clearance of Aβ and ameliorate other symptoms, as microglia exhibit a more anti-inflammatory, phagocytic phenotype. For example, suppression of fractalkine signaling, a negative regulator of microglial activation, is successful in reducing amyloid plaque burden and neuronal loss [
59‐
62]. In mouse models of other neurodegenerative disease such as Parkinson’s disease or ALS (amyotrophic lateral sclerosis), lack of CX3CR1 causes widespread neuron loss [
63], suggesting that the microglial activation profile seen here is an AD-specific effect. However, as APP mouse models do not exhibit significant neuron loss it is difficult to conclude if this is a tau-specific effect or relevant to human AD.
Table 2
Modulation of glia in Alzheimer’s disease mouse models
APP/PS11
| Scara1−/− | ↑Aβ, ↑mortality, ↓IDE, ↓Neprilysin | |
PDAPPSweInd line J202
| Scarb1−/− | ↑amyloid plaques, ↑CAA, ↔glial activation, ↑memory impairment | |
APP/PS13
| CD11b-TK | ↔Aβ, ↔amyloid plaques, ↑GFAP, ↓Iba1 | |
APP234
| CD11b-TK | ↓Aβ, ↓Iba1, ↔amyloid plaques | |
PDAPPSweInd line J202
| CxCR3-GFP ki | ↔Aβ, ↑microglial activation, ↑IL-6, ↑TNF-α, ↑p-tau, ↑memory impairment | |
TgCRND85
| CxCR3-GFP ki | ↓Aβ, ↓amyloid plaques, ↑ microglial phagocytosis, ↑microglial proliferation | |
APP/PS13
| CxCR3-GFP ki | ↓Aβ, ↓amyloid plaques, ↓microglia, ↑ microglial phagocytosis | |
R1.406
| CxCR3-GFP ki | ↓Aβ, ↓amyloid plaques | |
htau7
| CxCR3-GFP ki | ↑p-tau, ↑Gallyas-positive dystrophic neurites, ↓Iba1, ↑microglial activation (CD68+ and CD45+) | |
3xTg-AD8
| CxCR3-GFP ki | ↓neuronal loss | |
Tg25769
| Ccr2−/− | ↑Aβ, ↓NEP | |
APP/PS110
| Ccr2−/− | ↑soluble Aβ, ↑microglial activation, ↑memory impairment | |
APP/PS110
| NSE-COX2 | ↑Aβ, ↑PGE2 | |
Tg25769
| C1q−/− | ↔Aβ, ↓glial activation, ↑neuronal degeneration | |
Tg25769
| C1q−/− | ↔Aβ, ↓glial activation, ↓loss of synaptic markers | |
APP/PS111
| C1q−/− | ↔Aβ, ↓glial activation | |
TauP301L line JNLP312
| sCrry | ↑p-tau | |
Tg25769
| CD40L−/− | ↓p-tau | |
Tg25769
| CD40L−/− | ↓Aβ, ↓glial activation | |
APP/PS111
| CD40L−/− | ↓Aβ, ↓glial activation | |
APP/PS11
| Nlrp3−/− | ↓Aβ, ↓plaques, ↓IL-1β, ↓iNOS, ↑LTP, ↑spatial memory, ↑IDE | |
PDAPPSweInd line J202
| C3−/− | ↑Aβ, ↑amyloid plaques, ↑glial activation, ↑neuronal loss | |
APP/PS11
| CD14−/− | ↓Aβ, ↓amyloid plaques, ↓CD45+ activated microglia | |
APP/PS11
| CD33−/− | ↓Aβ, ↓plaques | |
Tg25769 (before plaque onset) | CD36−/− | ↔Aβ, ↔ROS | |
Tg25769 (old mice) | CD36−/− | ↓Aβ40, ↓CAA, ↑cognitive performance | |
APP/PS11
| CD45−/− | ↑Aβ, ↑amyloid plaques, ↑inflammatory microglia, ↑TNF-α, ↑IL-1β, ↑neuronal death | |
APP/PS13
| IRAK4KI/KI
| ↓Aβ, ↓amyloid plaques, ↓glial activation, ↑PPARγ, ↑IDE, ↑IFNγ, ↓iNOS | |
APP/PS11
| TLR4Lps-d
| ↑Aβ, ↑amyloid plaques | |
APP/PS11
| TLR4Lps-d
| ↑CD11b+ microglia, ↑GFAP | |
APP/PS11
| TLR4Lps-d
| ↑Aβ, ↑ amyloid plaques, ↓microglial activation, ↑cognitive impairment | |
APP/PS11
| MyD88−/− | ↓Aβ, ↓amyloid plaques, ↓CD11b+, CD45+ microglia | |
APP/PS110
| MyD88+/− | ↓amyloid plaques, ↑soluble Aβ, ↓IL-1β | |
APP/PS110
| TLR2−/− | Delayed plaque formation, ↑Aβ, ↑TGF-β, ↑memory impairment | |
Tg25769
| GFAP-MCP1 | ↑Aβ, ↑microglial activation | |
APP/PS11
| GFAP−/−Vim−/− | ↑Aβ, ↑amyloid plaques, ↑neurotic dystrophy, ↓activated astrocytes, ↑microglia, | |
In addition, targeting of the phagocytic phenotype of microglia has shown some promising results in AD mouse models. The complement pathway has been extensively studied in relation to AD and reports suggest that upregulating complement factors may target inflammatory processes by promoting migration and phagocytosis of inflammatory cells [
48,
71,
75].
Microglia and macrophages express a number of receptors that can promote clearance of Aβ, such as scavenger receptor class A1 (
Scara1) and class BI (
Scarb1). Knockout models for
Scarb1[
65] and
Scara1[
64] have shown alterations in Aβ load.
Additionally, Toll-like receptors (TLRs) and their co-receptors including MD-2, CD14, and CD36 [
90] are of great importance for the recognition of pathogens in the body and participate in the response of microglial cells to fibrillar forms of Aβ [
91]. Deletion of CD14, which acts as a co-receptor for LPS along with TLR2 and TLR4, in APP/PS1 mice reduced total microglial numbers, in particular CD45-positive microglia, attenuated AD pathology whilst also increasing the expression of TNF-α and IL-10, suggesting an induction of a shift of activation of microglia towards the M2b state [
76]. On the other hand, TLR2 deficiency accelerated spatial and contextual memory impairments, which correlated with increased levels of Aβ(1–42) and transforming growth factor-β in the brain of APP/PS1 mice [
87]. An essential adaptor protein for all TLR signaling, with the exception of TLR3, is the myeloid differentiation primary response protein 88 (MyD88). Decreasing the expression of MyD88 in APP/PS1 mice led to exacerbation of spatial memory deficits, increases in Aβ, reduced expression of the fractalkine receptor CX3CR1 and increased levels of APOE (Apolipoprotein E) together with reduced astrocyte and microglial activation [
85,
86]. These data indicate that TLR2 and TLR4 may be involved in Aβ clearance
in vivo and hence provide neuroprotection in AD [
92]. They also suggest that targeting specific glial activation states may prove fruitful in future clinical studies.
CD33 gene and TREM2, which are expressed in microglia, have been recently identified as genetic risks factors for AD [
93‐
96]. It was reported that CD33 is able to inhibit the uptake and clearance of Aβ42 in microglial cell cultures. This was confirmed by
in vivo results showing that brain levels of insoluble Aβ42 as well as amyloid plaque burden were markedly reduced in APP(Swe)/PS1(ΔE9)/CD33(−/−) mice. Therefore, CD33 inactivation appears to mitigate Aβ pathology [
77]. On the other hand, hypothesizing that the TREM2 risk variants impair TREM2 function, these new genetic studies suggest that reduced function of TREM2 causes reduced phagocytic clearance of amyloid proteins or cellular debris and thus impairs a protective mechanism in the brain [
94,
96].
There are a number of studies that attribute the clearance of amyloid in mouse models to infiltrating monocytes or perivascular macrophages [
97‐
100]. This is due to the evidence showing a reduced efficiency of microglia with age [
101] and bacterial and viral infections [
102]. However, the role of these peripheral monocytes in neurodegeneration remains unclear. One important aspect is the contribution of monocytes to resident macrophages, which is highly tissue-dependent and has been shown not to be relevant for brain microglia. However, recently it was suggested that, irrespective of their origin, macrophages/microglia can self-renew by local proliferation similar to that of stem cells [
103]. In fact, in animal models of prion disease it has been demonstrated that microglial proliferation is a major component in the evolution of chronic neurodegeneration [
104].
Many models that show peripheral monocytic infiltration use whole body irradiation which damages the blood–brain barrier itself, induces peripheral immune activation and can facilitate infiltration. Using this approach, it was recently published that microglia-depleted brain regions of CD11b-TK transgenic mice are repopulated with new Iba1-positive cells within 2 weeks, creating a niche for myeloid cells [
105]. However, using the technique of parabiosis (in which two mice share vasculature), GFP (Green Fluorescent Protein) -labelled monocytes from one mouse are not seen to infiltrate the brain of the other mouse, except following irradiation and bone marrow transplantation, which would suggest a pre-existing disease state is necessary in the brain for significant infiltration to occur [
102,
106]. In line with this, recent data provide strong evidence that the engraftment of myeloid cells in the brain parenchyma of AD transgenic mice does not occur normally during disease progression, but requires prior central nervous system conditioning to sufficiently attract bone marrow cells [
102]. These studies also highlight the importance of the chemokine receptor CCR2 in monocyte migration as the infiltrating cells following irradiation are characterized as CCR2
+. Interestingly, deletion of CCR2 in Tg2576 mice increased Aβ accumulation and reduced microglial recruitment into the brain, in particular phagocytic macrophages [
67]. In agreement with this, another study showed that restriction of CCR2 deficiency to perivascular myeloid cells drastically impaired Aβ clearance and amplified vascular Aβ deposition, while parenchymal plaque deposition remained unaffected [
102].
Furthermore, inflammatory IFNγ-secreting Th1 cells and IL-17-secreting Th17 cells have been shown to infiltrate the brain of older APP/PS1 mice [
107], supporting the observation of infiltrating T cells in the brain of AD patients [
108]. However, the role of these cells in the AD brain is still unknown.
Manipulation of astrocytes in animal models of Alzheimer’s disease
Astrocytes are becoming increasingly recognized as having key immune functions in the brain, and their role in Alzheimer’s disease progression has recently been investigated. Whilst currently falling behind the number of studies that are published assessing microglial function in AD, it is clear that astrocytes have a significant role to play in AD and therefore warrant significant future research.
Attenuation of astrocytic activation via deletion of
GFAP and
vimentin in APP/PS1 mice exacerbated amyloid plaque load independent of APP processing and Aβ production [
89], suggesting that astrocytes are important in amyloid clearance. Yet a previous study has shown that blocking astrocyte activation via AAV-
Gfa2 vectors in APP/PS1 mice also attenuates microglia activation, improves cognitive and synaptic function and reduces amyloid load [
109]. However these mice were analyzed at a considerably older age (16 to 18 months) when compared with the more recent study (8 to 12 months) which suggests that there may be a significant timing factor involved in targeting the immune response in AD.
Whether astrocytes are promoting amyloid clearance or exacerbating deposition is in debate; α
1-antichymotrypsin (α
1-ACT), an acute-phase protein that is overexpressed by activated astrocytes surrounding the amyloid plaques in human AD brains, has been proven to promote Aβ fibrillization. Confirming this, overexpression of a human transgene by astrocytes in the PDGF-APP
SweInd J9 or PDAPP mouse model promoted Aβ deposition and plaque formation [
36,
37]. It also affected tau phosphorylation and p-tau was increased both in single transgenic GFAP-α
1-ACT and in APP-GFAP-α
1-ACT mice [
38].