NLRP3-mediated glutaminolysis regulates microglia in Alzheimer’s disease#

Authors#

Róisín M. Mcmanus, Max P. Komes, Angelika Griep, Francesco Santarelli, Stephanie Schwartz, Juan Ramón Perea, Michelle-Amirah Khalil, Johannes C.M. Schlachetzki, David Bouvier, Mario A. Lauterbach, Lea Heinemann, Titus Schlüter, Mehran Shaban Pour, Marta Lovotti, Rainer Stahl, Fraser Duthie, Juan F. Rodríguez-Alcázar, Susanne V. Schmidt, Jasper Spitzer, Peri Noori, Alberto Maillo, Andreas Boettcher, Brian Herron, John Mcconville, David Gomez-Cabrero, Jesper Tegnér, Christopher K. Glass, Karsten Hiller, Eicke Latz, Michael Heneka

Abstract#

The NLRP3 inflammasome plays a key role in the microglial innate immune response and upon activation leads to the caspase-1-dependent secretion of inflammatory cytokines such as IL-1β 1. We have previously shown that the NLRP3 inflammasome has an important role in the progression of Alzheimer’s disease (AD). It was activated in the post-mortem brain tissue of AD patients and the absence of NLRP3 in the APP/PS1 transgenic model protected mice from Aβ deposition, reduced neuroinflammation and rescued memory impairments 2. NLRP3 activation also causes the release of the protein ASC in speck formation, which rapidly enhances the aggregation of Aβ 3. Both fibrillar and oligomeric Aβ activate the NLRP3 inflammasome 4,5, thus triggering an ongoing cycle of IL-1β and ASC-mediated inflammation and Aβ deposition in the brain. Indeed blocking this with NLRP3-inhibition can rescue Aβ plaque deposition and cognitive impairment in APP/PS1 mice 6. This is not unique to Aβ as tau (which is found in AD, Frontotemporal dementia – FTD and tauopathies) also induces assembly of the NLRP3 inflammasome, NLRP3 is activated in the brains of patients with FTD and critically, depletion of NLRP3 protected mice from developing tau pathology 7. However, whether NLRP3 impacts changes in AD beyond these traditional pathways is unknown. Here, we set out to determine the genes and pathways under the influence of NLRP3, beyond the established release of IL-1β and ASC that could be contributing to the progression of AD. We found that the initial deposition of Aβ directly triggers activation of the NLRP3 inflammasome and development of a mRNA disease-associated microglial (DAM) signature. Absence of NLRP3 protects mice from developing AD-pathology and neuroinflammation with no impact on the DAM-phenotype. Single cell RNA sequencing (scRNAseq) revealed a unique cellular cluster enriched with cells from the APP/PS1.NLRP3-/- mice, which had an increase in pathways related to amino acid metabolism and phagocytosis. The most significantly changed gene in this cluster was Slc1a3, the glutamate/aspartate transporter. We verified this and found Slc1a3 was increased at a protein level in all NLRP3-/- mice and on NLRP3-/- CD11b+ microglial cells. Slc1a3 can be found on the mitochondria, lysosome, and nucleus and the NLRP3-/- cells had enhanced mitochondrial membrane potential, with a corresponding increase in oxidative phosphorylation (OXPHOS). Functionally, NLRP3-/- microglia had enhanced phagocytic uptake of Aβ, and this was strictly related to the utilisation of glutamine, glutamate and α-ketoglutarate (αKG). Critically, we found that chronic pharmacological inhibition of NLRP3 was able to mimic these changes in vitro and in the APP/PS1 model in vivo. Together this work highlights NLRP3 as a master regulator of cellular function and Aβ phagocytosis, through its ability to modulate mitochondrial activity and via regulation of metabolites, affect downstream canonical signalling pathways.