A Noncanonical CD56dimCD16dim/- NK Cell Subset Indicative of Prior Cytotoxic Activity Is Elevated in Patients with Autoantibody-Mediated Neurologic Diseases.

Neuromyelitis optica spectrum disorder (NMOSD), myelin oligodendrocyte glycoprotein Ab disease, and autoimmune myasthenia gravis (MG) are autoantibody-mediated neurologic conditions where autoantibodies can induce Ab-dependent cellular cytotoxicity (ADCC), a NK cell-mediated effector function. However, whether ADCC is a pathogenic mechanism in patients with these conditions has not been confirmed. We sought to characterize circulatory NK cells using functional assays, phenotyping, and transcriptomics to elucidate their role in pathology. NK cells from NMOSD patients and MG patients with elevated disease burden exhibited reduced ADCC and CD56dimCD16hi NK cells, along with an elevated frequency of CD56dimCD16dim/- NK cells. We determined that ADCC induces a similar phenotypic shift in vitro. Bulk RNA sequencing distinguished the CD56dimCD16dim/- population from the canonical CD56dimCD16hi cytotoxic and CD56hiCD16- immunomodulatory subsets, as well as CD56hiCD16+ NK cells. Multiparameter immunophenotyping of NK cell markers, functional proteins, and receptors similarly showed that the CD56dimCD16dim/- subset exhibits a unique profile while still maintaining expression of characteristic NK markers CD56, CD94, and NKp44. Notably, expression of perforin and granzyme is reduced in comparison with CD56dimCD16hi NK cells. Moreover, they exhibit elevated trogocytosis capability, HLA-DR expression, and many chemokine receptors, including CCR7. In contrast with NMOSD and MG, myelin oligodendrocyte glycoprotein Ab disease NK cells did not exhibit functional, phenotypic, or transcriptomic perturbations. In summary, CD56dimCD16dim/- NK cells are a distinct peripheral blood immune cell population in humans elevated upon prior cytotoxic activity by the CD56dimCD16hi NK cell subset. The elevation of this subset in NMOSD and MG patients suggests prior ADCC activity.

Remission of severe myasthenia gravis after autologous stem cell transplantation.

OBJECTIVE: Myasthenia gravis (MG) is an autoantibody-mediated neuromuscular junction disorder involving the acetylcholine receptors on the motor endplate. The safety and response to high-dose chemotherapy (HDIT) and autologous hematopoietic cell transplantation (HCT) were assessed in a patient with severe refractory MG. METHODS: As part of a pilot study of HDIT/HCT for patients with treatment-resistant autoimmune neurological disorders, a patient with severe refractory MG underwent treatment. After mobilization of hematopoietic stem cells with rituximab, prednisone, and G-CSF, the patient had HDIT consisting of carmustine, etoposide, cytarabine, melphalan, and rabbit antithymocyte globulin, followed by autologous HCT. The effect of treatment on the autoantibody to the acetylcholine receptor (AChR) was assessed. RESULTS: The patient had been diagnosed with AChR antibody-positive MG 14 years before HDIT/HCT and had failed thymectomy, therapeutic plasma exchange, and multiple immunomodulatory agents. The Myasthenia Gravis Foundation of America (MGFA) clinical classification was IVb before HDIT/HCT. She tolerated HDIT/HCT well and started to improve clinically within days of treatment. At both 1 and 2 years after HDIT/HCT, patients remained symptom-free. After HDIT/HCT, AChR-binding autoantibodies persisted, and the relative frequency of immune cell subtypes shifted. INTERPRETATION: HDIT/HCT induced a complete response of disease activity in a patient with severe refractory MG. This response may suggest that a cell-mediated etiology may be a significant contributing factor in refractory MG cases. A phase 2 clinical trial is warranted to establish if HDIT/HCT can be an effective therapy for severe refractory MG and to gain a further understanding of disease pathogenesis.

MOGAD patient autoantibodies induce complement, phagocytosis, and cellular cytotoxicity.

Myelin oligodendrocyte glycoprotein (MOG) antibody-associated disease (MOGAD) is an inflammatory demyelinating CNS condition characterized by the presence of MOG autoantibodies. We sought to investigate whether human MOG autoantibodies are capable of mediating damage to MOG-expressing cells through multiple mechanisms. We developed high-throughput assays to measure complement activity (CA), complement-dependent cytotoxicity (CDC), antibody-dependent cellular phagocytosis (ADCP), and antibody-dependent cellular cytotoxicity (ADCC) of live MOG-expressing cells. MOGAD patient sera effectively mediate all of these effector functions. Our collective analyses reveal that (a) cytotoxicity is not incumbent on MOG autoantibody quantity alone; (b) engagement of effector functions by MOGAD patient serum is bimodal, with some sera exhibiting cytotoxic capacity while others did not; (c) the magnitude of CDC and ADCP is elevated closer to relapse, while MOG-IgG binding is not; and (d) all IgG subclasses can damage MOG-expressing cells. Histopathology from a representative MOGAD case revealed congruence between lesion histology and serum CDC and ADCP, and we identified NK cells, mediators of ADCC, in the cerebrospinal fluid of relapsing patients with MOGAD. Thus, MOGAD-derived autoantibodies are cytotoxic to MOG-expressing cells through multiple mechanisms, and assays quantifying CDC and ADCP may prove to be effective tools for predicting risk of future relapses.

Lost in post-translational modification-Dengue virus writes its own sequel.

Elevated frequency of afucosylated IgG1 antibodies during dengue virus infection is associated with prior infection and predicts severe disease.

High-throughput investigation of molecular and cellular biomarkers in NMOSD.

OBJECTIVE: To identify candidate biomarkers associated with neuromyelitis optica spectrum disorder (NMOSD) using high-throughput technologies that broadly assay the concentrations of serum analytes and frequencies of immune cell subsets. METHODS: Sera, peripheral blood mononuclear cells (PBMCs), and matched clinical data from participants with NMOSD and healthy controls (HCs) were obtained from the Collaborative International Research in Clinical and Longitudinal Experience Study NMOSD biorepository. Flow cytometry panels were used to measure the frequencies of 39 T-cell, B-cell, regulatory T-cell, monocyte, natural killer (NK) cell, and dendritic cell subsets in unstimulated PBMCs. In parallel, multiplex proteomics assays were used to measure 46 serum cytokines and chemokines in 2 independent NMOSD and HC cohorts. Multivariable regression models were used to assess molecular and cellular profiles in NMOSD compared with HC. RESULTS: NMOSD samples had a lower frequency of CD16+CD56+ NK cells. Both serum cohorts and multivariable logistic regression revealed increased levels of B-cell activating factor associated with NMOSD. Interleukin 6, CCL22, and CCL3 were also elevated in 1 NMOSD cohort of the 2 analyzed. Multivariable linear regression of serum analyte levels revealed a correlation between CX3CL1 (fractalkine) levels and the number of days since most recent disease relapse. CONCLUSIONS: Integrative analyses of cytokines, chemokines, and immune cells in participants with NMOSD and HCs provide congruence with previously identified biomarkers of NMOSD and highlight CD16+CD56+ NK cells and CX3CL1 as potential novel biomarker candidates.

Does pedicle screw fixation of the subaxial cervical spine provide adequate stabilization in a multilevel vertebral body fracture model? An in vitro biomechanical study.

BACKGROUND: Cervical vertebral body fractures generally are treated through an anterior-posterior approach. Cervical pedicle screws offer an alternative to circumferential fixation. This biomechanical study quantifies whether cervical pedicle screws alone can restore the stability of a three-column vertebral body fracture, making standard 360° reconstruction unnecessary. METHODS: Range of motion (2.0 Nm) in flexion-extension, lateral bending, and axial rotation was tested on 10 cadaveric specimens (five/group) at C2-T1 with a spine kinematics simulator. Specimens were tested for flexibility of intact when a fatigue protocol with instrumentation was used to evaluate construct longevity. For a C4-6 fracture, spines were instrumented with 360° reconstruction (corpectomy spacer + plate + lateral mass screws) (Group 1) or cervical pedicle screw reconstruction (C3 and C7 only) (Group 2). FINDINGS: Results are expressed as percentage of intact (100%). In Group 1, 360° reconstruction resulted in decreased motion during flexion-extension, lateral bending, and axial rotation, to 21.5%, 14.1%, and 48.6%, respectively, following 18,000 cycles of flexion-extension testing. In Group 2, cervical pedicle screw reconstruction led to reduced motion after cyclic flexion-extension testing, to 38.4%, 12.3%, and 51.1% during flexion-extension, lateral bending, and axial rotation, respectively. INTERPRETATION: The 360° stabilization procedure provided the greatest initial stability. Cervical pedicle screw reconstruction resulted in less change in motion following cyclic loading with less variation from specimen to specimen, possibly caused by loosening of the shorter lateral mass screws. Cervical pedicle screw stabilization may be a viable alternative to 360° reconstruction for restoring multilevel vertebral body fracture.

Biomechanical Assessment of Stabilization of Simulated Type II Odontoid Fracture with Case Study.

STUDY DESIGN: Researchers created a proper type II dens fracture (DF) and quantified a novel current posterior fixation technique with spacers at C1-C2. A clinical case study supplements this biomechanical analysis. PURPOSE: Researchers explored their hypothesis that spacers combined with posterior instrumentation (PI) reduce range of motion significantly, possibly leading to better fusion outcomes. OVERVIEW OF LITERATURE: Literature shows that the atlantoaxial joint is unique in allowing segmental rotary motion, enabling head turning. With no intervertebral discs at these joints, multiple ligaments bind the axis to the skull base and to the atlas; an intact odontoid (dens) enhances stability. The most common traumatic injury at these strong ligaments is a type II odontoid fracture. METHODS: Each of seven specimens (C0-C3) was tested on a custom-built six-degrees-of-freedom spine simulator with constructs of intact state, type II DF, C1-C2 PI, PI with joint capsulotomy (PIJC), PI with spacers (PIS) at C1-C2, and spacers alone (SA). A bending moment of 2.0 Nm (1.5°/sec) was applied in flexion-extension (FE), lateral bending (LB), and axial rotation (AR). One-way analysis of variance with repeated measures was performed. RESULTS: DF increased motion to 320%, 429%, and 120% versus intact (FE, LB, and AR, respectively). PI significantly reduced motion to 41%, 21%, and 8%. PIJC showed negligible changes from PI. PIS reduced motion to 16%, 14%, and 3%. SA decreased motion to 64%, 24%, and 54%. Reduced motion facilitated solid fusion in an 89-year-old female patient within 1 year. CONCLUSIONS: Type II odontoid fractures can lead to acute or chronic instability. Current fixation techniques use C1-C2 PI or an anterior dens screw. Addition of spacers alongside PI led to increased biomechanical rigidity over intact motion and may offer an alternative to established surgical fixation techniques.

Imaging Axonal Degeneration and Repair in Preclinical Animal Models of Multiple Sclerosis.

Multiple sclerosis (MS) is a central nervous system (CNS) disease characterized by chronic neuroinflammation, demyelination, and axonal damage. Infiltration of activated lymphocytes and myeloid cells are thought to be primarily responsible for white matter damage and axonopathy. Over time, this neurologic damage manifests clinically as debilitating motor and cognitive symptoms. Existing MS therapies focus on symptom relief and delay of disease progression through reduction of neuroinflammation. However, long-term strategies to remyelinate, protect, or regenerate axons have remained elusive, posing a challenge to treating progressive forms of MS. Preclinical mouse models and techniques, such as immunohistochemistry, flow cytometry, and genomic and proteomic analysis have provided advances in our understanding of discrete time-points of pathology following disease induction. More recently, in vivo and in situ two-photon (2P) microscopy has made it possible to visualize continuous real-time cellular behavior and structural changes occurring within the CNS during neuropathology. Research utilizing 2P imaging to study axonopathy in neuroinflammatory demyelinating disease has focused on five areas: (1) axonal morphologic changes, (2) organelle transport and health, (3) relationship to inflammation, (4) neuronal excitotoxicity, and (5) regenerative therapies. 2P imaging may also be used to identify novel therapeutic targets via identification and clarification of dynamic cellular and molecular mechanisms of axonal regeneration and remyelination. Here, we review tools that have made 2P accessible for imaging neuropathologies and advances in our understanding of axonal degeneration and repair in preclinical models of demyelinating diseases.