Fra1 Controls Rheumatoid Factor Autoantibody Production by Bone Marrow Plasma Cells and the Development of Autoimmune Bone Loss.

Next to proinflammatory cytokines, autoimmunity has been identified as a key trigger for osteoclast activation and bone loss. IgG-rheumatoid factor (IgG-RF) immune complexes, which are present in patients with rheumatoid arthritis, were shown to boost osteoclast differentiation. To date, the regulation of IgG-RF production in the absence of inflammatory triggers is unknown. Herein, we describe Fra1 as a key checkpoint that controls IgG-RF production by plasma cells and regulates autoimmune-mediated bone loss. Fra1 deficiency in B cells (Fra1ΔBcell ) led to increased IgG1-producing bone marrow plasma cells, enhanced IgG-RF production, and increased bone loss associated with elevated osteoclast numbers after immunization. The effect of IgG-RF on osteoclasts in vitro and on osteoclasts associated with bone loss in vivo was dependent on FcγR, especially FcγR3. Furthermore, immunization of WT mice with T-cell-dependent antigens induced a significant and robust decrease in Fra1 expression in bone marrow B cells, which was followed by increased IgG1 production and the induction of osteoclast-mediated bone loss. Overall, these data identify Fra1 as a key mediator of IgG-RF production and autoimmune-mediated bone loss. © 2019 American Society for Bone and Mineral Research.

Extracellular aggregated alpha synuclein primarily triggers lysosomal dysfunction in neural cells prevented by trehalose.

Cell-to-cell propagation of aggregated alpha synuclein (aSyn) has been suggested to play an important role in the progression of alpha synucleinopathies. A critical step for the propagation process is the accumulation of extracellular aSyn within recipient cells. Here, we investigated the trafficking of distinct exogenous aSyn forms and addressed the mechanisms influencing their accumulation in recipient cells. The aggregated aSyn species (oligomers and fibrils) exhibited more pronounced accumulation within recipient cells than aSyn monomers. In particular, internalized extracellular aSyn in the aggregated forms was able to seed the aggregation of endogenous aSyn. Following uptake, aSyn was detected along endosome-to-lysosome and autophagosome-to-lysosome routes. Intriguingly, aggregated aSyn resulted in lysosomal activity impairment, accompanied by the accumulation of dilated lysosomes. Moreover, analysis of autophagy-related protein markers suggested decreased autophagosome clearance. In contrast, the endocytic pathway, proteasome activity, and mitochondrial homeostasis were not substantially affected in recipient cells. Our data suggests that extracellularly added aggregated aSyn primarily impairs lysosomal activity, consequently leading to aSyn accumulation within recipient cells. Importantly, the autophagy inducer trehalose prevented lysosomal alterations and attenuated aSyn accumulation within aSyn-exposed cells. Our study underscores the importance of lysosomes for the propagation of aSyn pathology, thereby proposing these organelles as interventional targets.

Alpha-synuclein activates BV2 microglia dependent on its aggregation state.

Synucleinopathies such as Parkinson's disease (PD), dementia with Lewy bodies (DLB), and multiple system atrophy (MSA) are defined by the presence of intracellular alpha-synuclein aggregates in neurons and/or oligodendrocytes. In addition, post mortem tissue analysis revealed profound changes in microglial morphology, indicating microglial activation and neuroinflammation. Thus, alpha-synuclein may directly activate microglia, leading to increased production of key pro-inflammatory cytokines like tumor necrosis factor-alpha (TNF-α) and interleukin-1beta (IL-1ß), which in turn modulates the disease progression. The distinct alpha-synuclein species, which mediates the activation of microglia, is not well defined. We hypothesized that microglial activation depends on a specific aggregation state of alpha-synuclein. Here, we show that primarily human fibrillar alpha-synuclein increased the production and secretion of pro-inflammatory cytokines by microglial BV2 cells compared to monomeric and oligomeric alpha-synuclein. BV2 cells also preferentially phagocytosed fibrillar alpha-synuclein compared to alpha-synuclein monomers and oligomers. Microglial uptake of alpha-synuclein fibrils and the consequent activation were time- and concentration-dependent. Moreover, the degree of fibrillization determined the efficiency of microglial internalization. Taken together, our study highlights the specific crosstalk of distinct alpha-synuclein species with microglial cells.

Infiltrating T lymphocytes reduce myeloid phagocytosis activity in synucleinopathy model.

BACKGROUND: Synucleinopathies comprise a group of neurodegenerative diseases associated with abnormal accumulation of α-synuclein. One of the key factors that contribute to the progression of synucleinopathies is neuroinflammation. However, the role of lymphocytes in synucleinopathies like Parkinson's disease (PD) remains largely unclear. METHODS: To investigate how lymphocytes impact synucleinopathies, human wild-type α-synuclein (WTS) transgenic mice were crossed with mice lacking mature lymphocytes (Rag2(-/-)). In this in vivo model, we quantified α-synuclein aggregation in the substantia nigra (SN) and striatum and determined the numbers of innate and adaptive immune cells in the central nervous system (CNS). The activation state of resident and infiltrated CNS myeloid cells (M1 vs. M2) was further classified by gene and protein expression analyses. The impact of T and B lymphocytes on the phagocytic activity of microglia in the presence of α-synuclein aggregates was addressed in BV2 microglia in vitro. RESULTS: Compared to WTS(+) Rag2(+/+) mice, where T but not B lymphocytes infiltrated the CNS, decreased amounts of α-synuclein aggregates were found in WTS(+) Rag2(-/-) mice devoid of mature lymphocytes. The presence of T lymphocytes did not alter the number of Iba1(+) microglia but increased the frequency of the CD11b(+) CD45(hi) population in the CNS, indicative of an increased number of infiltrated macrophages. Moreover, the M1 phenotype was more prominent in WTS(+) Rag2(+/+) mice, whereas the M2 activation state was dominating in the absence of lymphocytes in WTS(+) Rag2(-/-) mice. In vitro, in the presence of T but not B lymphocytes, significantly less α-synuclein was phagocytosed by BV2 microglia, further supporting the prevalence of the M1 phenotype in the presence of T lymphocytes. CONCLUSIONS: Peripheral T lymphocytes strongly contribute to increased α-synuclein pathology via modulation of CNS myeloid cell function. In the presence of T lymphocytes, microglia phagocytosis of aggregated α-synuclein is reduced, which increases the severity of synucleinopathy.

Posttranslational modification and mutation of histidine 50 trigger alpha synuclein aggregation and toxicity.

BACKGROUND: Aggregation and aggregation-mediated formation of toxic alpha synuclein (aSyn) species have been linked to the pathogenesis of sporadic and monogenic Parkinson's disease (PD). A novel H50Q mutation of aSyn, resulting in the substitution of histidine by glutamine, has recently been identified in PD patients. We have previously shown that the lipid peroxidation product 4-hydroxy-2-nonenal (HNE) induces the formation of HNE-aSyn adducts, thereby promoting aSyn oligomerization and increasing its extracellular toxicity to human dopaminergic neurons. Intriguingly, we identified histidine 50 (H50) of aSyn as one of the HNE modification target residues. These converging lines of evidence support the hypothesis that changes in H50 via posttranslational modification (PTM) and mutation trigger the formation of aggregated, toxic aSyn species, which interfere with cellular homeostasis. In the present study, we aim to elucidate 1) the role of H50 in HNE-mediated aSyn aggregation and toxicity, and 2) the impact of H50 mutation on aSyn pathology. Besides the PD-related H50Q, we analyze a PD-unrelated control mutation, in which H50 is replaced by an arginine residue (H50R). RESULTS: Analysis of HNE-treated aSyn revealed that H50 is the most susceptible residue of aSyn to HNE modification and is crucial for HNE-mediated aSyn oligomerization. Overexpression of aSyn with substituted H50 in H4 neuroglioma cells reduced HNE-induced cell damage, indicating a pivotal role of H50 in HNE modification-induced aSyn toxicity. Furthermore, we showed in vitro that H50Q/R mutations substantially increase the formation of high density and fibrillar aSyn species, and potentiate the oligomerization propensity of aSyn in the presence of a nitrating agent. Cell-based experiments also revealed that overexpression of H50Q aSyn in H4 cells promotes aSyn oligomerization. Importantly, overexpression of both H50Q/R aSyn mutants in H4 cells significantly increased cell death when compared to wild type aSyn. This increase in cell death was further exacerbated by the application of H2O2. CONCLUSION: A dual approach addressing alterations of H50 showed that either H50 PTM or mutation trigger aSyn aggregation and toxicity, suggesting an important role of aSyn H50 in the pathogenesis of both sporadic and monogenic PD.