Microglial Transcriptome Hints at Shortcomings of AD Model

From ALZFORUM – With the realization that microglia play a key role in neurodegenerative conditions, scientists have profiled their transcription patterns to see how they change during disease. A study published in the January 16 Cell Reports pulls together and analyzes 18 such data sets. Led by David Hansen at Genentech, South San Francisco, California, the researchers confirmed the presence of a neurodegeneration-related phenotype in mouse models of Alzheimer’s disease and spotted the signature in human Alzheimer’s tissue. They also found a set of inflammation-related genes expressed in AD brains, but not in mouse models. The researchers hope to accelerate microglial research by sharing the compiled data in an interactive database online.

“These data sets are one of the most valuable things for the community,” said Shane Liddelow, New York University, who has already mined the database for evidence of cross talk between microglia and astrocytes. Oleg Butovsky, Brigham and Women’s Hospital in Boston, also considers it an important resource but wondered how the authors’ analyses of the compiled data advances understanding of microglia.

Scientists have uncovered several microglial phenotypes associated with neurodegeneration and AD in particular, including a microglial neurodegenerative phenotype (MGnD) and “DAMs,” a.k.a disease-associated microglia. How they relate to each other, and what they do in neurodegeneration, still needs clarification.

Multifaceted Personality. Different disease models and other conditions (columns) induce different myeloid cell expression modules in the brain (rows). Microglial gene expression is down or unchanged in most models of conditions, whereas a neurodegeneration-related signature is up in those respective models. (Courtesy of Friedman et al., Cell Reports, 2018.)

In search of a more global view of microglia’s multiple roles in disease, Friedman collected transcriptional data from models of ischemia, infectious disease, inflammation, cancer, demyelination, tauopathy, and Alzheimer’s disease. Most of the data came from published reports, but the authors also added new RNA-seq data sets from the PS2APP model of AD, and from the P301L (Götz et al., 2001) and P301S tauopathy models.

To integrate all these data, Friedman used z-score normalization. For each study, he calculated how different disease conditions shifted expression of each gene from the average. Then he transformed the data into relative values that he could analyze together in one master gene expression matrix.

“One of the nice things about our paper is that it surveys a diverse set of microglial states. It’s trying to make sense of all those profiles, putting them all together,” said Hansen. Friedman added, “It’s also one of the first, maybe the first, to include a genome-wide expression data set of a tauopathy model.”

To identify common types of microglial activation in an unbiased way, Friedman selected 777 genes whose expression levels departed from the mean in the different models. He then used hierarchical clustering to group these genes into co-regulated modules. From 45 that emerged, the researchers focused on seven that were enriched for genes associated with cell proliferation, interferon signaling, responses to lipopolysaccharide (LPS), neurodegeneration, and functions of homeostatic microglia, macrophages, and neutrophils/monocytes (image above). The neurodegeneration profile proved similar to transcriptional changes seem in DAMs and MGnDs.

Reanalyzing single-cell data from the previous DAM study, Friedman and colleagues found 16 unique transcriptional signatures, suggesting discrete microglial states. Two of these signatures, one associated with DAMs and the other interferon-related, were more likely to show up in mouse models of AD than in controls. The interferon signature was roughly twice as abundant in 5xFAD mice as in controls.

The researchers also examined microglial expression in human AD tissue. They sequenced RNA from previously frozen samples of the fusiform gyrus from 33 Braak stage I-IV and 84 Braak stage III-VI cases from the Banner Research Institute tissue bank. The AD samples scored about 10 percent higher for the neurodegeneration module. The researchers also found an unexpected uptick in expression of the LPS module compared to mouse models, suggesting AD brains may have more inflammatory signaling and/or peripheral immune cell infiltration than do transgenic animals.

“It is common that when we look at animal models, they do not recapitulate well the glial aspects of disease,” said Liddelow.

Whether the postmortem findings truly reflect what happens in living AD brains remains to be seen, noted Michael Heneka, University of Bonn, Germany. “It is important to recognize that postmortem changes may be a strong confound, especially in AD, where microglia have been primed for decades.” Although the authors obtained tissue within five hours of death, microglial expression patterns can change in a matter of minutes (Jun 2017 news). “I think the most important next step is to get better expression data from human disease,” acknowledged Hansen.

Butovsky and Liddelow called for validation of the findings in situ. They would like to see how the microglia expression modules map to different locations in mouse and human brains, extending previous findings of disease-associated microglia. Indeed, Hansen is particularly interested in examining the distribution of cells expressing the interferon-related module. Heneka noted that it’s important to clarify the biological meaning of the transcriptional profiles, including the many untranslated transcripts. “Immune cells have a lot of bullets they never shoot,” he said.—Marina Chicurel

Source – ALZFORUM