An immunological puzzle: The adaptive immune system fuels Alzheimer's disease pathology.

Alzheimer's disease (AD) is a devastating neurodegenerative disorder characterized by a concerning rise in prevalence. It is projected that the number of affected individuals will reach a staggering 150 million by 2050. While recent advancements in monoclonal antibodies targeting Aß have shown some clinical effects, there is an urgent need for improved therapies to effectively address the impeding surge of AD patients worldwide. To achieve this, a deeper understanding of the intricate mechanisms underlying the disease is crucial. In recent years, mounting evidence has underscored the vital role of the innate immune system in AD pathology. However, limited findings persist regarding the involvement of the adaptive immune system. Here, we report on the impact of the adaptive immune system on various aspects of AD by using AppNL-G-F mice crossed into a Rag2-/- background lacking mature adaptive immune cells. In addition, to simulate the continuous exposure to various challenges such as infections that is commonly observed in humans, the innate immune system was activated through the repetitive induction of peripheral inflammation. We observed a remarkably improved performance on complex cognitive tasks when a mature adaptive immune system is absent. Notably, this observation is pathologically associated with lower Aß plaque accumulation, reduced glial activation, and better-preserved neuronal networks in the mice lacking a mature adaptive immune system. Collectively, these findings highlight the detrimental role of the adaptive immune system in AD and underscore the need for effective strategies to modulate it for therapeutic purposes.

Limitations of PLX3397 as a microglial investigational tool: peripheral and off-target effects dictate the response to inflammation.

Microglia, the resident macrophages of the central nervous system (CNS), play a critical role in CNS homeostasis and neuroinflammation. Pexidartinib (PLX3397), a colony-stimulating factor 1 (CSF1) receptor inhibitor, is widely used to deplete microglia, offering flexible options for both long-term depletion and highly versatile depletion-repopulation cycles. However, the potential impact of PLX3397 on peripheral (immune) cells remains controversial. Until now, the microglia-specificity of this type of compounds has not been thoroughly evaluated, particularly in the context of peripherally derived neuroinflammation. Our study addresses this gap by examining the effects of PLX3397 on immune cells in the brain, liver, circulation and bone marrow, both in homeostasis and systemic inflammation models. Intriguingly, we demonstrate that PLX3397 treatment not only influences the levels of tissue-resident macrophages, but also affects circulating and bone marrow immune cells beyond the mononuclear phagocyte system (MPS). These alterations in peripheral immune cells disrupt the response to systemic inflammation, consequently impacting the phenotype irrespective of microglial depletion. Furthermore, we observed that a lower dose of PLX3397, which does not deplete microglia, demonstrates similar (non-)MPS effects, both in the periphery and the brain, but fails to fully replicate the peripheral alterations seen in the higher doses, questioning lower doses as a 'peripheral control' strategy. Overall, our data highlight the need for caution when interpreting studies employing this compound, as it may not be suitable for specific investigation of microglial function in the presence of systemic inflammation.

Experimental hepatic encephalopathy causes early but sustained glial transcriptional changes.

Hepatic encephalopathy (HE) is a common complication of liver cirrhosis, associated with high morbidity and mortality, for which no brain-targeted therapies exist at present. The interplay between hyperammonemia and inflammation is thought to drive HE development. As such, astrocytes, the most important ammonia-metabolizing cells in the brain, and microglia, the main immunomodulatory cells in the brain, have been heavily implicated in HE development. As insight into cellular perturbations driving brain pathology remains largely elusive, we aimed to investigate cell-type specific transcriptomic changes in the HE brain. In the recently established mouse bile duct ligation (BDL) model of HE, we performed RNA-Seq of sorted astrocytes and microglia at 14 and 28 days after induction. This revealed a marked transcriptional response in both cell types which was most pronounced in microglia. In both cell types, pathways related to inflammation and hypoxia, mechanisms commonly implicated in HE, were enriched. Additionally, astrocytes exhibited increased corticoid receptor and oxidative stress signaling, whereas microglial transcriptome changes were linked to immune cell attraction. Accordingly, both monocytes and neutrophils accumulated in the BDL mouse brain. Time-dependent changes were limited in both cell types, suggesting early establishment of a pathological phenotype. While HE is often considered a unique form of encephalopathy, astrocytic and microglial transcriptomes showed significant overlap with previously established gene expression signatures in other neuroinflammatory diseases like septic encephalopathy and stroke, suggesting common pathophysiological mechanisms. Our dataset identifies key molecular mechanisms involved in preclinical HE and provides a valuable resource for development of novel glial-directed therapeutic strategies.

Microdissection and Whole Mount Scanning Electron Microscopy Visualization of Mouse Choroid Plexus.

The choroid plexus (CP), a highly vascularized structure protruding into the ventricles of the brain, is one of the most understudied tissues in neuroscience. As it is becoming increasingly clear that this tiny structure plays a crucial role in health and disease of the central nervous system (CNS), it is of utmost importance to properly dissect the CP out of the brain ventricles in a way that allows downstream processing, ranging from functional to structural analysis. Here, isolation of the lateral and fourth brain ventricle mouse CP without the need for specialized tools or equipment is described. This isolation technique preserves the viability, function, and structure of cells within the CP. On account of its high vascularization, the CP can be visualized floating inside the ventricular cavities of the brain using a binocular microscope. However, transcardial perfusion required for downstream analysis can complicate the identification of the CP tissue. Depending on the further processing steps (e.g., RNA and protein analysis), this can be solved by visualizing the CP via transcardial perfusion with bromophenol blue. After isolation, the CP can be processed using several techniques, including RNA, protein, or single cell analysis, to gain further understanding on the function of this special brain structure. Here, scanning electron microscopy (SEM) on whole mount CP is used to get an overall view of the structure.

Microglial activation arises after aggregation of phosphorylated-tau in a neuron-specific P301S tauopathy mouse model.

Alzheimer's disease, progressive supranuclear palsy and frontotemporal dementia are characterized by neuronal expression of aberrant tau protein, tau hyperphosphorylation (pTAU), tau aggregation and neurofibrillary tangle formation sequentially culminating into neuronal cell death, a process termed tauopathy. Our aim was to address at which tauopathy stage neuroinflammation starts and to study the related microglial phenotype. We used Thy1-hTau.P301S (PS) mice expressing human tau with a P301S mutation specifically in neurons. Significant levels of cortical pTAU were present from 2 months onwards. Dystrophic morphological complexity of cortical microglia arose after pTAU accumulation concomitant with increased microglial lysosomal volumes and a significant loss of homeostatic marker Tmem119. Interestingly, we detected increases in neuronal pTAU and postsynaptic structures in the lysosomes of PS microglia. Moreover, the overall cortical postsynaptic density was decreased in 6-month-old PS mice. Together, our results indicate that microglia adopt a pTAU-associated phenotype, and are morphologically and functionally distinct from wild-type microglia after neuronal pTAU accumulation has initiated.