Progranulin Loss Results in Sex-Dependent Dysregulation of the Peripheral and Central Immune System

INTRODUCTION: Progranulin (PGRN) is a secreted glycoprotein, the expression of which is linked to several neurodegenerative diseases. Although its specific function is still unclear, several studies have linked it with lysosomal functions and immune system regulation. Here, we have explored the role of PGRN in peripheral and central immune system homeostasis by investigating the consequences of PGRN deficiency on adaptive and innate immune cell populations. METHODS: First, we used gene co-expression network analysis of published data to test the hypothesis that RESULTS: Male PGRN KO mice exhibited a lower abundance of microglial cells with higher MHC-II expression, increased CD44 expression on monocytes in the brain, and more CNS-associated CD8 DISCUSSION: Our data suggest that PGRN and GPNMB jointly regulate the peripheral and the central immune system in a sex-specific manner; thus, understanding their associated mechanisms could pave the way for developing new neuroprotective strategies to modulate central and peripheral inflammation to lower risk for neurodegenerative diseases and possibly delay or halt progression

Neurodegeneration and neuroinflammation are linked, but independent of a-synuclein inclusions, in a seeding/spreading mouse model of Parkinson's disease

A key pathological process in Parkinson's disease (PD) is the transneuronal spreading of α-synuclein. Alpha-synuclein (α-syn) is a presynaptic protein that, in PD, forms pathological inclusions. Other hallmarks of PD include neurodegeneration and microgliosis in susceptible brain regions. Whether it is primarily transneuronal spreading of α-syn particles, inclusion formation, or other mechanisms, such as inflammation, that cause neurodegeneration in PD is unclear. We used a model of spreading of α-syn induced by striatal injection of α-syn preformed fibrils into the mouse striatum to address this question. We performed quantitative analysis for α-syn inclusions, neurodegeneration, and microgliosis in different brain regions, and generated gene expression profiles of the ventral midbrain, at two different timepoints after disease induction. We observed significant neurodegeneration and microgliosis in brain regions not only with, but also without α-syn inclusions. We also observed prominent microgliosis in injured brain regions that did not correlate with neurodegeneration nor with inclusion load. Using longitudinal gene expression profiling, we observed early gene expression changes, linked to neuroinflammation, that preceded neurodegeneration, indicating an active role of microglia in this process. Altered gene pathways overlapped with those typical of PD. Our observations indicate that α-syn inclusion formation is not the major driver in the early phases of PD-like neurodegeneration, but that microglia, activated by diffusible, oligomeric α-syn, may play a key role in this process. Our findings uncover new features of α-syn induced pathologies, in particular microgliosis, and point to the necessity for a broader view of the process of α-syn spreading

An archaeal compound as a driver of Parkinson’s disease pathogenesis

Patients with Parkinson’s disease (PD) exhibit differences in their gut microbiomes compared to healthy individuals. Although differences have most commonly been described in the abundances of bacterial taxa, changes to viral and archaeal populations have also been observed. Mechanistic links between gut microbes and PD pathogenesis remain elusive but could involve molecules that promote α-synuclein aggregation. Here, we show that 2-hydroxypyridine (2-HP) represents a key molecule for the pathogenesis of PD. We observe significantly elevated 2-HP levels in faecal samples from patients with PD or its prodrome, idiopathic REM sleep behaviour disorder (iRBD), compared to healthy controls. 2-HP is correlated with the archaeal species Methanobrevibacter smithii and with genes involved in methane metabolism, and it is detectable in isolate cultures of M. smithii. We demonstrate that 2-HP is selectively toxic to transgenic α-synuclein overexpressing yeast and increases α-synuclein aggregation in a yeast model as well as in human induced pluripotent stem cell derived enteric neurons. It also exacerbates PD-related motor symptoms, α-synuclein aggregation, and striatal degeneration when injected intrastriatally in transgenic mice overexpressing human α-synuclein. Our results highlight the effect of an archaeal molecule in relation to the gut-brain axis, which is critical for the diagnosis, prognosis, and treatment of PD.

MICROGLIA IN PARKINSON´S DISEASE: IDENTITY, HETEROGENEITY AND THEIR CONTRIBUTION TO NEURODEGENERATION

Parkinson´s disease (PD) is the most common movement disorder caused by dopamine deficiency owing to a loss of dopaminergic neurons within the substantia nigra (SN). So far, there is no cure available, hence understanding the mechanisms by which dopaminergic neurons degenerate is essential for the development of future treatment strategies. Recently, a potential role of neuroinflammation, and especially the activation of microglial cells in PD was suggested, not being secondary to neuronal death, but rather primarily implicated in PD pathogenesis. Hence, we have ventured in to study neuroinflammation and microglia activation in the context of PD using in vivo and in vitro mouse models. Firstly, we addressed microglial heterogeneity in the healthy nigrostriatal pathway, the primary circuit affected in PD. By using single-cell RNA sequencing, we have identified four different microglial immune subsets within the midbrain and the striatum. Notably, we were able to distinguish a microglial subset with an immune alerted phenotype, which was mainly composed of microglial cells from the midbrain. The transcriptomic identity of this subset resembled partially to the one of inflammatory microglia. Additionally, in situ morphological studies, such as 3D reconstruction, revealed that microglia located within the midbrain is less complex than microglia with a striatal origin. Secondly, we studied the potential role of neuroinflammation and microglia in PD progression by using a PD-like mouse model of a-synuclein (a-syn) seeding and spreading. In this study, pre-formed fibrils (PFF) were injected into the mice striatum, and a combined neuropathological and transcriptomic analysis was performed at two time points that have distinct and increasing levels and distribution of a-syn pathology across different brain regions (13 and 90 days post-injection). Interestingly, neuropathological quantifications at 90 days post-injection uncovered that neuroinflammation and microglial reactivity are linked to neurodegeneration. However, pathology neither correlates with neurodegeneration nor with a-syn aggregation. Importantly, at 13 days post-injection, the transcriptomic analysis of the midbrain revealed the dysregulation of several inflammatory pathways and pointed to the overexpression of neurotoxic inflammatory mediators. Furthermore, at this time point, the presence of a-syn oligomers was detected in certain areas of the brain. Subsequently, we hypothesised that at early stages of PD pathogenesis, the presence of a-syn oligomeric forms induces a robust inflammatory response of microglia, which can be further associated with neurodegeneration. Thirdly, to understand if a-syn oligomers are the main inducers of microglial activation, we examined further the microglial inflammatory response to other a-syn conformations, monomers and fibrils (PFF1 and PFF2). For that, BV2 and primary microglial cells were exposed to the a-syn moieties at different concentrations and incubations times. Electron microscopy depicted some heterogeneity across the synthetic a-syn fibrils, suggesting that PFF1 and PFF2 were composed by different structures. Then, microglial reactivity to a-syn monomers and fibrils was investigated by RT-PCR, and no specific response of microglia to a-syn was encountered. Also, only one of the a-syn fibrils, the PFF1, decreased microglial phagocytic activity and reduced the expression of Il1b by microglia after LPS stimulation. Concomitant to the findings in the a-syn seeding and spreading model, we attempted to elucidate the molecular profile of microglia associated with neurodegeneration. In this particular study, RNA-sequencing was performed in isolated microglial cells in an early stage of pathology progression. In contrast with our previous results, no differences in the microglial profile were found between the PFF and the control mice. Lastly, we have investigated potential neuroprotective mechanisms associated with microglial reactivity counter-regulation. Considering previous observations that microglia express dopaminergic receptors, we investigated further whether apomorphine, a dopamine agonist with anti-oxidant properties, could govern microglial activation. The effect of apomorphine enantiomers was analysed within primary microglia cultures that were activated by exposure to mutated A53T monomeric a-syn. Herein, we demonstrated that microglial activation can be dampened by apomorphine, via the recruitment of Nrf2 to the nucleus, which results in a decreased release of proinflammatory mediators, such as TNFa or PGE2. Taken together, this study provides an additional characterisation of neuroinflammation and microglial cells in the context of PD, which ultimately contributes to a better understanding of their relationship with neurodegeneration

Single-Cell Transcriptional Profiling and Gene Regulatory Network Modeling in Tg2576 Mice Reveal Gender-Dependent Molecular Features Preceding Alzheimer-Like Pathologies

peer reviewedAlzheimer’s disease (AD) onset and progression is influenced by a complex interplay of several environmental and genetic factors, one of them gender. Pronounced gender differences have been observed both in the relative risk of developing AD and in clinical disease manifestations. A molecular level understanding of these gender disparities is still missing, but could provide important clues on cellular mechanisms modulating the disease and reveal new targets for gender-oriented disease-modifying precision therapies. We therefore present here a comprehensive single-cell analysis of disease-associated molecular gender differences in transcriptomics data from the neocortex, one of the brain regions most susceptible to AD, in one of the most widely used AD mouse models, the Tg2576 model. Cortical areas are also most commonly used in studies of post-mortem AD brains. To identify disease-linked molecular processes that occur before the onset of detectable neuropathology, we focused our analyses on an age with no detectable plaques and microgliosis. Cell-type specific alterations were investigated at the level of individual genes, pathways, and gene regulatory networks. The number of differentially expressed genes (DEGs) was not large enough to build context-specific gene regulatory networks for each individual cell type, and thus, we focused on the study of cell types with dominant changes and included analyses of changes across the combination of cell types. We observed significant disease-associated gender differences in cellular processes related to synapse organization and axonogenesis, and identified a limited set of transcription factors, including Egr1 and Klf6, as key regulators of many of the disease-associated and gender-dependent gene expression changes in the model. Overall, our analyses revealed significant celltype-specific gene expression changes in individual genes, pathways and subnetworks, including gender-specific and gender-dimorphic changes in both upstream transcription factors and their downstream targets, in the Tg2576 AD model before the onset of overt disease. This opens a window into molecular events that could determine gender-susceptibility to AD, and uncovers tractable target candidates for potential gender-specific precision medicine for AD.R-AGR-0621 - Dons Alzheimer Projekt (Dr. Glaab) (20151026-20480119) - SCHNEIDER Reinhar

Microglia phenotypes are associated with subregional patterns of concomitant tau, amyloid-β and α-synuclein pathologies in the hippocampus of patients with Alzheimer’s disease and dementia with Lewy bodies

The cellular alterations of the hippocampus lead to memory decline, a shared symptom between Alzheimer’s disease (AD) and dementia with Lewy Bodies (DLB) patients. However, the subregional deterioration pattern of the hippocampus differs between AD and DLB with the CA1 subfield being more severely affected in AD. The activation of microglia, the brain immune cells, could play a role in its selective volume loss. How subregional microglia populations vary within AD or DLB and across these conditions remains poorly understood. Furthermore, how the nature of the hippocampal local pathological imprint is associated with microglia responses needs to be elucidated. To this purpose, we employed an automated pipeline for analysis of 3D confocal microscopy images to assess CA1, CA3 and DG/CA4 subfields microglia responses in post-mortem hippocampal samples from late-onset AD (n = 10), DLB (n = 8) and age-matched control (CTL) (n = 11) individuals. In parallel, we performed volumetric analyses of hyperphosphorylated tau (pTau), amyloid-β (Aβ) and phosphorylated α-synuclein (pSyn) loads. For each of the 32,447 extracted microglia, 16 morphological features were measured to classify them into seven distinct morphological clusters. Our results show similar alterations of microglial morphological features and clusters in AD and DLB, but with more prominent changes in AD. We identified two distinct microglia clusters enriched in disease conditions and particularly increased in CA1 and DG/CA4 of AD and CA3 of DLB. Our study confirms frequent concomitance of pTau, Aβ and pSyn loads across AD and DLB but reveals a specific subregional pattern for each type of pathology, along with a generally increased severity in AD. Furthermore, pTau and pSyn loads were highly correlated across subregions and conditions. We uncovered tight associations between microglial changes and the subfield pathological imprint. Our findings suggest that combinations and severity of subregional pTau, Aβ and pSyn pathologies transform local microglia phenotypic composition in the hippocampus. The high burdens of pTau and pSyn associated with increased microglial alterations could be a factor in CA1 vulnerability in AD

An archaeal compound as a driver of Parkinson’s disease pathogenesis

Patients with Parkinson’s disease (PD) exhibit differences in their gut microbiomes compared to healthy individuals. Although differences have most commonly been described in the abundances of bacterial taxa, changes to viral and archaeal populations have also been observed. Mechanistic links between gut microbes and PD pathogenesis remain elusive but could involve molecules that promote α-synuclein aggregation. Here, we show that 2-hydroxypyridine (2-HP) represents a key molecule for the pathogenesis of PD. We observe significantly elevated 2-HP levels in faecal samples from patients with PD or its prodrome, idiopathic REM sleep behaviour disorder (iRBD), compared to healthy controls. 2-HP is correlated with the archaeal species Methanobrevibacter smithii and with genes involved in methane metabolism, and it is detectable in isolate cultures of M. smithii. We demonstrate that 2-HP is selectively toxic to transgenic α-synuclein overexpressing yeast and increases α-synuclein aggregation in a yeast model as well as in human induced pluripotent stem cell derived enteric neurons. It also exacerbates PD-related motor symptoms, α-synuclein aggregation, and striatal degeneration when injected intrastriatally in transgenic mice overexpressing human α-synuclein. Our results highlight the effect of an archaeal molecule in relation to the gut-brain axis, which is critical for the diagnosis, prognosis, and treatment of PD