LaFrance‐Corey, Reghann G.

ABSTRACTCavin‐4 was identified as a potential autoantigen for immune‐mediated rippling muscle disease (iRMD). To validate this, we developed and tested various immunoassays, including a cell‐based assay (CBA), cavin‐4 recombinant protein ELISA, and multi‐peptide ELISA. Among 19 iRMD patients, all exhibited muscle rippling, and 13 had percussion‐induced mounding. All immunoassays demonstrated clinical and analytical specificities greater than 95%. The protein ELISA had the highest sensitivity (94.7%) and specificity (99.9%), outperforming CBA (sensitivity 89.5%, specificity 99.6%) and the multi‐peptide ELISA (sensitivity 79.0%, specificity 97.2%). Our results suggest that the cavin‐4 protein ELISA is a promising tool for high‐throughput clinical testing in iRMD.

Abstract The immunological response to CNS viral infection must balance the need for effective viral clearance with the need to preserve irreplaceable neurons. Failure to achieve this balance may result in death or severe cognitive impairment. We have observed hippocampal injury, compromised cognitive function, and seizures in C57BL/6 mice following acute infection with TMEV. We established that an initial innate immune response controlled by inflammatory monocyte infiltration into the brain is responsible for this neural injury. To better understand the pathogenic mechanisms involved in monocyte recruitment to the CNS we asked whether reducing the initial viral inoculum impacted the evolution of neural injury. We found that mice infected with standard (2×105 PFU) or low inoculum (1.25×104 PFU) TMEV exhibited a graded inflammatory response. Notably, a 16-fold reduction in viral inoculum resulted in 80% attenuation of the inflammatory monocyte response, concomitant with a 90% attenuation in acute production of CCL2 and complete abrogation of acute increases in TNFα and IL-6 in the hippocampus. Cognitive performance was preserved in low inoculum mice and seizure burden was highly attenuated. Despite the introduction of 16-times less virus, viral load in the brain was equivalent at 3 dpi between groups, indicating that the amount of replicating virus was not associated with reduced neuropathology. Our findings support a model in which the earliest host response to pathogen in the CNS shapes the size of the inflammatory response and constrains the downstream pathological sequelae.

Abstract MR imaging suggests an anatomic correspondence between WM and GM lesions in progressing MS patients, suggesting an underlying neuronal-signaling or neurodegenerative element to the progressive disease state. While retrograde signals evolving from stressed axons that alter soma gene expression have been previously described in models of axonal transection or crush injury, those resulting from inflammation and demyelination are unknown. Using microfluidic chambers that segregate neuronal soma and axons, we have previously demonstrated increased axonal MHC I following treatment with IFNg, a cytokine prominent in demyelinating MS lesions. Furthermore, expression of antigen-loaded MHC I predisposed axons to injury by antigen-specific CD8+ T cells in vitro. Here we report that neurons also upregulate expression of MHC class Ib mRNAs in response to axonal stimulation with IFNg. Genes most highly upregulated following cytokine treatment included H2-M3, H2-Q5, H2-Q6, H2-Q7, H2-Q8, H2-T10 , and H2-T23. We further report the development of an in vivo model system for measuring axonal vulnerability to antigen-specific immune attack following demyelination using an AAV vector to induce neuron-specific expression of axonally-targeted neoantigen (ovalbumin) tagged with cleavable eGFP. We also present optimized methods using AAV vectors in mice for infecting retinal, motor cortex, and hippocampal neurons to induce target gene expression in axons projecting to optic nerve and corpus collosum.

ObjectiveWe recently reported successful treatment of a child with febrile infection‐related epilepsy syndrome (FIRES), a subtype of new onset refractory status epilepticus, with the recombinant interleukin‐1 (IL1) receptor antagonist (IL1RA) anakinra. On this basis, we tested whether endogenous IL1RA production or function is deficient in FIRES patients.MethodsLevels of IL1β and IL1RA were measured in serum and cerebrospinal fluid (CSF). The inhibitory activity of endogenous IL1RA was assessed using a cell‐based reporter assay. IL1RN gene variants were identified by sequencing. Expression levels for the secreted and intracellular isoforms of IL1RA were measured in patient and control cells by real‐time polymerase chain reaction.ResultsLevels of endogenous IL1RA and IL1β were elevated in the serum and CSF of patients with FIRES (n = 7) relative to healthy controls (n = 10). Serum from FIRES patients drove IL1R signaling activity and potentiated IL1R signaling in response to exogenous IL1β in a cell‐based reporter assay. Functional assessment of endogenous IL1RA activity in 3 FIRES patients revealed attenuated inhibition of IL1R signaling. Sequencing of IL1RN in our index patient revealed multiple variants. This was accompanied by reduced expression of intracellular but not secreted isoforms of IL1RA in the patient's peripheral blood mononuclear cells.InterpretationOur findings suggest that FIRES is associated with reduced expression of intracellular IL1RA isoforms and a functional deficiency in IL1RA inhibitory activity. These observations may provide insight into disease pathogenesis for FIRES and other inflammatory seizure disorders and may provide a valuable biomarker for therapeutic decision‐making. Ann Neurol 2019;85:526–537

SummaryObjectivesAlthough secondary hemophagocytic lymphohistiocytosis (HLH) has been reported in children with critical illness of various etiologies, it has not been reported in patients with febrile infection–related epilepsy syndrome (FIRES). We describe a series of patients with concurrent HLH and FIRES in an effort to establish common pathophysiologic abnormalities.MethodsFive patients with FIRES who were assessed for HLH were identified from a neurocritical care database. All were previously healthy and had extensive diagnostic testing. All had clinical deterioration with multiorgan dysfunction prompting HLH screening 20‐29 days after hospitalization. Markers for inflammatory dysregulation were assessed in cerebrospinal fluid (CSF) and serum at various time points. Outcomes were assessed 6 months after presentation.ResultsThree patients met clinical criteria for secondary HLH. Elevation of specific cytokines/chemokines was variable. CSF neopterin, high mobility group box 1 (HMGB1), and C‐X‐C motif chemokine ligand 8 (CXCL8) were significantly elevated in all. Interleukin‐1β (IL‐1β) and IL‐18 were not elevated in any of the samples. Treatment and outcomes were variable.SignificanceWe describe 3 patients with HLH and FIRES. The co‐occurrence of these 2 rare disorders suggests the possibility of a common immune dysregulation phenotype prolonging epileptogenesis. HLH screening in critically ill patients with FIRES may yield a broader understanding of shared inflammatory processes.