Dectin-1 limits autoimmune neuroinflammation and promotes myeloid cell-astrocyte crosstalk via Card9-independent expression of Oncostatin M.

Pathologic roles of innate immunity in neurologic disorders are well described, but their beneficial aspects are less understood. Dectin-1, a C-type lectin receptor (CLR), is largely known to induce inflammation. Here, we report that Dectin-1 limited experimental autoimmune encephalomyelitis (EAE), while its downstream signaling molecule, Card9, promoted the disease. Myeloid cells mediated the pro-resolution function of Dectin-1 in EAE with enhanced gene expression of the neuroprotective molecule, Oncostatin M (Osm), through a Card9-independent pathway, mediated by the transcription factor NFAT. Furthermore, we find that the Osm receptor (OsmR) functioned specifically in astrocytes to reduce EAE severity. Notably, Dectin-1 did not respond to heat-killed Mycobacteria, an adjuvant to induce EAE. Instead, endogenous Dectin-1 ligands, including galectin-9, in the central nervous system (CNS) were involved to limit EAE. Our study reveals a mechanism of beneficial myeloid cell-astrocyte crosstalk regulated by a Dectin-1 pathway and identifies potential therapeutic targets for autoimmune neuroinflammation.

Detection limitations of prion seeding activities in blood samples from patients with sporadic prion disease.

BACKGROUND: Human prion diseases (HPDs) are fatal neurodegenerative disorders characterized by abnormal prion proteins (PrPSc). However, the detection of prion seeding activity in patients with high sensitivity remains challenging. Even though real-time quaking-induced conversion (RT-QuIC) assay is suitable for detecting prion seeding activity in a variety of specimens, it shows lower accuracy when whole blood, blood plasma, and blood-contaminated tissue samples are used. In this study, we developed a novel technology for the in vitro amplification of abnormal prion proteins in HPD to the end of enabling their detection with high sensitivity known as the enhanced quaking-induced conversion (eQuIC) assay. METHODS: Three antibodies were used to develop the novel eQUIC method. Thereafter, SD50 seed activity was analyzed using brain tissue samples from patients with prion disease using the conventional RT-QUIC assay and the novel eQUIC assay. In addition, blood samples from six patients with solitary prion disease were analyzed using the novel eQuIC assay. RESULTS: The eQuIC assay, involving the use of three types of human monoclonal antibodies, showed approximately 1000-fold higher sensitivity than the original RT-QuIC assay. However, when this assay was used to analyze blood samples from six patients with sporadic human prion disease, no prion activity was detected. CONCLUSION: The detection of prion seeding activity in blood samples from patients with sporadic prion disease remains challenging. Thus, the development of alternative methods other than RT-QuIC and eQuIC will be necessary for future research.

Formalin RT-QuIC assay detects prion-seeding activity in formalin-fixed brain samples from sporadic Creutzfeldt-Jakob disease patients.

BACKGROUND: The neuropathology of sporadic Creutzfeldt-Jakob disease (sCJD) is usually investigated using formalin-fixed and formic acid-treated brain tissue. However, formalin and formic acid treatment can interfere with immunostaining of abnormal prion protein. Therefore, there is a need for biochemical methods other than immunostaining to investigate abnormal prion protein in postmortem tissue. We developed RT-QuIC to quantitate the seeding activity (SD50) of sCJD brain tissue treated with formalin and formic acid. METHODS: We used endpoint RT-QuIC assays to analyze SD50 in formalin-fixed brain tissue from 19 sCJD patients (14 MM1 cases, 3 MM2-thalamic form [MM2T] cases and 2 MM2-cortical form [MM2C] cases) diagnosed according to Parchi's classification. We assessed SD50 in brains after incubation in formalin solution for over 1 month, and after treating formalin-fixed brain tissue with formic acid. We also examined how the SD50 values from formalin-fixed brain samples compared with neuropathological and immunohistochemical findings. RESULTS: The SD50 values of formalin-fixed brain samples from 14 MM1 cases, 2 MM2C cases, and 2 MM2T cases were 107.77±0.57/g tissue, 107.44±0.24/g tissue and 106.00±0.77/g tissue, respectively. The average SD50 value in MM1 unfixed brains decreased by 102.04 after formalin fixation for 1 month. In MM1 cases, after combined formalin and formic acid treatment, the SD50 value was reduced by approximately 105.16 compared with that of unfixed tissue. The SD50 values of formalin-fixed tissue showed a consistent pattern with the neuropathological findings in most brain regions examined. CONCLUSION: RT-QuIC enables the study of formalin-fixed brain tissue from sCJD patients that has not previously been amenable to analysis.

Postmortem Quantitative Analysis of Prion Seeding Activity in the Digestive System.

Human prion diseases are neurodegenerative disorders caused by prion protein. Although infectivity was historically detected only in the central nervous system and lymphoreticular tissues of patients with sporadic Creutzfeldt-Jakob disease, recent reports suggest that the seeding activity of Creutzfeldt-Jakob disease prions accumulates in various non-neuronal organs including the liver, kidney, and skin. Therefore, we reanalyzed autopsy samples collected from patients with sporadic and genetic human prion diseases and found that seeding activity exists in almost all digestive organs. Unexpectedly, activity in the esophagus reached a level of prion seeding activity close to that in the central nervous system in some CJD patients, indicating that the safety of endoscopic examinations should be reconsidered.

The AIM2 inflammasome is activated in astrocytes during the late phase of EAE.

Inflammasomes are a class of innate immune signaling platforms that activate in response to an array of cellular damage and pathogens. Inflammasomes promote inflammation under many circumstances to enhance immunity against pathogens and inflammatory responses through their effector cytokines, IL-1ß and IL-18. Multiple sclerosis and its animal model, experimental autoimmune encephalomyelitis (EAE), are autoimmune conditions influenced by inflammasomes. Despite work investigating inflammasomes during EAE, little remains known concerning the role of inflammasomes in the central nervous system (CNS) during the disease. Here, we used multiple genetically modified mouse models to monitor activated inflammasomes in situ based on oligomerization of apoptosis-associated speck-like protein containing a CARD (ASC) in the spinal cord. Using inflammasome reporter mice, we found heightened inflammasome activation in astrocytes after the disease peak. In contrast, microglia and CNS-infiltrated myeloid cells had few activated inflammasomes in the CNS during EAE. Astrocyte inflammasome activation during EAE was dependent on absent in melanoma 2 (AIM2), but low IL-1ß release and no significant signs of cell death were found. Thus, the AIM2 inflammasome activation in astrocytes may have a distinct role from traditional inflammasome-mediated inflammation.

[A case of combined central and peripheral demyelination].

A 22-year-old man had had difficulty running fast since about he was 10 years old. In June 2011, he was referred to us because of worsened unsteady gait. A neurological examination revealed mild ataxic speech, weakness of the four limbs, with spasticity, and pes cavus. Magnetic resonance T2-weighted images showed multiple high-intensity lesions in the bilateral periventricular white matter, brainstem, and thoracic spinal cord. Peripheral nerve conduction studies revealed marked motor conduction velocities were markedly reduced and sensory nerve velocities were not evoked in the upper and lower limbs. A sural nerve biopsy showed highly active demyelinating lesions. The patient was treated with high-dose steroid therapy (intravenous methylprednisolone, 1,000 mg/day × 3 days) followed by self-injection of interferon ß. With these treatments, his symptoms gradually improved. In this case, we could not detect the causative factors, and all autoantibodies tested, except for the anti-neurofascin antibody, were negative. The anti-neurofascin antibody might induce demyelination in the central and peripheral nervous systems. However, in the literature, the evidence of an association between this antibody and these clinical characteristics is not conclusive. We need more studies on the pathogenesis of combined central and peripheral demyelination to establish more effective therapies.