Crippling the complement cascade in a mouse model of AD leads to more plaques, but less neurodegeneration and better function, according to a new study from Cynthia Lemere and colleagues at Brigham and Women’s Hospital, Boston. Mice missing the complement protein C3 showed an altered immune response to Aβ: Microglia appeared less active, while levels of inflammatory cytokines did not climb as high as in the AD mice with C3. Without C3, synapses were preserved and the mice performed better on tests of learning and memory. The work appeared May 31 in Science Translational Medicine.
“This is a really important paper,” said Andrea Tenner, University of California, Irvine, noting that the rigorous quantitative data support the rationale for investigating therapeutics that inhibit complement activation-dependent processes for the treatment of AD.
Not Attracted: Plaques (green) in APP/PS1 mice lacking C3 (right) have far fewer infiltrating and more peripheral microglia (red) than plaques in C3-replete animals (left). [Courtesy of Science Translational Medicine/AAAS.]
The complement cascade plays a central role in innate immunity. In response to pathogens, dead cells, or debris, C3 and its fragments promote phagocytosis and inflammation. C3 also functions in nervous system development, where it works together with microglia to prune excess synapses, a novel function discovered by co-author Beth Stevens, Children’s Hospital Boston. Stevens and Lemere collaborated to uncover a role for C3 in age-dependent synapse loss in aged mice and in young APP transgenic animals that had yet to accumulate Aβ plaques (see Shi et al., 2015; Apr 2016 news).
What about older mice that have abundant plaques, ongoing complement activation, and inflammation revving up microglia? To address this, first author Qiaoqiao Shi crossed APPsw/PSEN1dE9 mice with C3 knockouts. When offspring were 16 months, old enough to develop plaques and neurodegeneration, she tested their learning and memory skills in a water maze. The C3 knockouts performed better than their C3-replete APP/PS1 controls, and just as well as wild-type mice of the same age. The C3 KOs also had a much higher plaque load than the transgenic controls. Aβ immunoreactivity, thioflavin-S positive plaques, and insoluble Aβ were all greater than in APP/PS1 mice, and the knockouts had many more large plaques.
Nevertheless, neurons fared better in the C3 knockouts. By 16 months, APP/PS1 mice had lost 40 percent of neurons in the CA3 region of the hippocampus, while APP/PS1 C3 knockouts only lost 10 percent. The CA3 is particularly vulnerable to complement-mediated synapse loss, and no neurodegeneration was seen in the CA1, or the dentate gyrus in any of the animals.
The researchers next looked at the astrocytes and microglia to see if their immune response to amyloid plaques might be different in the absence of C3. Indeed, while the numbers of microglia and astrocytes were similar in knockouts and controls, microglia morphology was not. In the knockouts the cells were smaller, with thinner processes, and they had lower levels of activation markers. They also migrated differently. Without complement, strikingly fewer microglia and astrocytes infiltrated the core of plaques, with more hanging back in the outskirts (see image above). “It seems that they come up to the edge but don’t go in,” Lemere said. “Instead of being recruited into the plaque, they seem to be less active and less phagocytic, which is probably why there is more amyloid deposition,” she told Alzforum. In the brain, C3 knockouts also produced less of the inflammatory cytokines TNFα, INFγ, and IL-12.
Lemere sees two possible explanations for the results. One is that C3 knockout suppresses a pro-inflammatory immune response against plaques that would otherwise cause downstream damage to the synapses and neurons. Another is that because plaques in the knockout grow so large, they sequester Aβ oligomers that would normally cause the downstream damage. The two are not mutually exclusive, she said.
However the knockout works, it seems that the mice cope with a higher plaque load, at least until they are 16 months old. “We think that’s because it’s not the plaques themselves that are toxic to the neurons, it’s the immune response to the plaques,” Lemere said.
Previously, Lemere and others reported that knocking out or inhibiting C3 in J20 mice expressing human APP increased not only plaque load but also neurodegeneration (Maier et al., 2008; Wyss-Coray, 2002). The protective effect of C3 knockout in the new study appears to contradict those results. Lemere chalks it up to strain differences in microglial activation. In the J20 mice, glia react less, and show no major alterations after C3 knockout. In the APP/PS1 mice, plaque-induced gliosis actually recapitulates human pathology more closely than in the J20 model, Lemere said.
Shane Liddelow, working in Ben Barres’ lab at Stanford University, agreed that the APP/PS1 mice likely model microglia involvement better than the J20 mice. “I thought the paper was quite interesting. We’ve known for a while that C3 knockouts have increased Aβ plaque load, but Lemere is now showing that even with this increased plaque load, you are seeing beneficial memory outcomes. That is the key finding,” said Liddelow.
Liddelow would like to know more about the glial phenotypes in the AD mice, and especially what the astrocytes do in the absence of C3. He and Barres recently published a study on a novel “killer astrocyte” phenotype, which arises when microglia pump out, among other things, the complement protein C1q (Jan 2017 news). Liddelow pointed out that astrocytes are the main producer of C3, and could be responsible for some of the neurotoxicity seen in these mice.
The results add to a growing enthusiasm for targeting the complement system to modulate inflammation and to treat AD and other neurodegenerative diseases. The mice in the current study never had any C3, so it remains to be seen what happens if C3 is removed or blocked only after older mice have accumulated plaques. Those studies are underway, Lemere told Alzforum. Her lab has a conditional C3 knockout and plans to cross that into the APP/PS1 mice.
William Hu, Emory University, Atlanta, who studies complement activation as a biomarker for AD progression, called the study exciting. “We know that complement activation in spinal fluid seems to be associated with the progression of Alzheimer’s, which presents a very nice human correlate of the phenomenon reported here in mice. What’s so enticing is that complement inhibitors are already approved by the FDA so these could be quickly tested in people,” he told Alzforum.—Pat McCaffrey
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Research Models Citations
- Shi Q, Colodner KJ, Matousek SB, Merry K, Hong S, Kenison JE, Frost JL, Le KX, Li S, Dodart JC, Caldarone BJ, Stevens B, Lemere CA. Complement C3-Deficient Mice Fail to Display Age-Related Hippocampal Decline. J Neurosci. 2015 Sep 23;35(38):13029-42. PubMed.
- Maier M, Peng Y, Jiang L, Seabrook TJ, Carroll MC, Lemere CA. Complement C3 deficiency leads to accelerated amyloid beta plaque deposition and neurodegeneration and modulation of the microglia/macrophage phenotype in amyloid precursor protein transgenic mice. J Neurosci. 2008 Jun 18;28(25):6333-41. PubMed.
- Wyss-Coray T, Yan F, Lin AH, Lambris JD, Alexander JJ, Quigg RJ, Masliah E. Prominent neurodegeneration and increased plaque formation in complement-inhibited Alzheimer's mice. Proc Natl Acad Sci U S A. 2002 Aug 6;99(16):10837-42. PubMed.
No Available Further Reading
- Shi Q, Chowdhury S, Ma R, Le KX, Hong S, Caldarone BJ, Stevens B, Lemere CA. Complement C3 deficiency protects against neurodegeneration in aged plaque-rich APP/PS1 mice. Sci Transl Med. 2017 May 31;9(392) PubMed.