Aquaporin-4–binding autoantibodies in patients with neuromyelitis optica impair glutamate transport by down-regulating EAAT2

Neuromyelitis optica (NMO)-immunoglobulin G (IgG) is a clinically validated serum biomarker that distinguishes relapsing central nervous system (CNS) inflammatory demyelinating disorders related to NMO from multiple sclerosis. This autoantibody targets astrocytic aquaporin-4 (AQP4) water channels. Clinical, radiological, and immunopathological data suggest that NMO-IgG might be pathogenic. Characteristic CNS lesions exhibit selective depletion of AQP4, with and without associated myelin loss; focal vasculocentric deposits of IgG, IgM, and complement; prominent edema; and inflammation. The effect of NMO-IgG on astrocytes has not been studied. In this study, we demonstrate that exposure to NMO patient serum and active complement compromises the membrane integrity of CNS-derived astrocytes. Without complement, astrocytic membranes remain intact, but AQP4 is endocytosed with concomitant loss of Na+-dependent glutamate transport and loss of the excitatory amino acid transporter 2 (EAAT2) . Our data suggest that EAAT2 and AQP4 exist in astrocytic membranes as a macromolecular complex. Transport-competent EAAT2 protein is up-regulated in differentiating astrocyte progenitors and in nonneural cells expressing AQP4 transgenically. Marked reduction of EAAT2 in AQP4-deficient regions of NMO patient spinal cord lesions supports our immunocytochemical and immunoprecipitation data. Thus, binding of NMO-IgG to astrocytic AQP4 initiates several potentially neuropathogenic mechanisms: complement activation, AQP4 and EAAT2 down-regulation, and disruption of glutamate homeostasis

Perylene Diimide Nanoprobes for In Vivo Tracking of Mesenchymal Stromal Cells Using Photoacoustic Imaging.

Noninvasive bioimaging techniques are critical for assessing the biodistribution of cellular therapies longitudinally. Among them, photoacoustic imaging (PAI) can generate high-resolution images with a tissue penetration depth of ∼4 cm. However, it is essential and still highly challenging to develop stable and efficient near-infrared (NIR) probes with low toxicity for PAI. We report here the preparation and use of perylene diimide derivative (PDI) with NIR absorbance (around 700 nm) as nanoprobes for tracking mesenchymal stromal cells (MSCs) in mice. Employing an in-house synthesized star hyperbranched polymer as a stabilizer is the key to the formation of stable PDI nanoparticles with low toxicity and high uptake by the MSCs. The PDI nanoparticles remain within the MSCs as demonstrated by in vitro and in vivo assessments. The PDI-labeled MSCs injected subcutaneously on the flanks of the mice are clearly visualized with PAI up to 11 days postadministration. Furthermore, bioluminescence imaging of PDI-labeled luciferase-expressing MSCs confirms that the administered cells remain viable for the duration of the experiment. These PDI nanoprobes thus have good potential for tracking administered cells in vivo using PAI

Comparison of MRI lesion evolution in different central nervous system demyelinating disorders

Background and Objective: There are few studies that compare lesion evolution across different CNS demyelinating diseases, yet knowledge of this may be important for diagnosis and understanding differences in disease pathogenesis. We sought to compare MRI T2-lesion evolution in myelin-oligodendrocyte-glycoprotein-IgG-associated disorder (MOGAD), aquaporin-4-IgG-positive neuromyelitis optica spectrum disorder (AQP4-IgG-NMOSD), and multiple sclerosis (MS). Methods: In this descriptive study, we retrospectively identified Mayo Clinic patients with MOGAD, AQP4-IgG-NMOSD, or MS and: 1) brain or myelitis attack; 2) available attack MRI within 6 weeks; and 3) follow-up MRI beyond 6 months without interval relapses in that region. Two neurologists identified the symptomatic or largest T2-lesion for each patient (index lesion). MRIs were then independently reviewed by two neuroradiologists blinded to diagnosis to determine resolution of T2-lesions by consensus. The index T2-lesion area was manually outlined acutely and at follow-up to assess variation in size. Results: We included 156 patients (MOGAD, 38; AQP4-IgG-NMOSD, 51; MS, 67) with 172 attacks (brain, 81; myelitis, 91). The age (median [range]) differed between MOGAD (25 [2-74]), AQP4-IgG-NMOSD (53 [10-78]) and MS (37 [16-61]) (p<0.01) and female sex predominated in the AQP4-IgG-NMOSD (41/51 [80%]) and MS (51/67 [76%]) groups but not among those with MOGAD (17/38 [45%]). Complete resolution of the index T2-lesion was more frequent in MOGAD (brain, 13/18[72%]; spine, 22/28[79%]) than AQP4-IgG-NMOSD (brain, 3/21[14%]; spine, 0/34[0%]) and MS (brain, 7/42[17%]; spine, 0/29[0%]), p<0.001. Resolution of all T2-Lesions occurred most often in MOGAD (brain, 7/18[39%]; spine, 22/28[79%]) than AQP4-IgG-NMOSD (brain, 2/21[10%]; spine, 0/34[0%]), and MS (brain, 2/42[5%]; spine, 0/29[0%]), p< 0.01. There was a larger median (range) reduction in T2-lesion area in mm2 on follow-up axial brain MRI with MOGAD (213[55-873]) than AQP4-IgG-NMOSD (104[0.7-597]) (p=0.02) and MS, 36[0-506]) (p< 0.001) and the reductions in size on sagittal spine MRI follow-up in MOGAD (262[0-888]) and AQP4-IgG-NMOSD (309[0-1885]) were similar (p=0.4) and greater than MS (23[0-152]) (p<0.001)

Heterozygous Variants in KMT2E Cause a Spectrum of Neurodevelopmental Disorders and Epilepsy.

We delineate a KMT2E-related neurodevelopmental disorder on the basis of 38 individuals in 36 families. This study includes 31 distinct heterozygous variants in KMT2E (28 ascertained from Matchmaker Exchange and three previously reported), and four individuals with chromosome 7q22.2-22.23 microdeletions encompassing KMT2E (one previously reported). Almost all variants occurred de novo, and most were truncating. Most affected individuals with protein-truncating variants presented with mild intellectual disability. One-quarter of individuals met criteria for autism. Additional common features include macrocephaly, hypotonia, functional gastrointestinal abnormalities, and a subtle facial gestalt. Epilepsy was present in about one-fifth of individuals with truncating variants and was responsive to treatment with anti-epileptic medications in almost all. More than 70% of the individuals were male, and expressivity was variable by sex; epilepsy was more common in females and autism more common in males. The four individuals with microdeletions encompassing KMT2E generally presented similarly to those with truncating variants, but the degree of developmental delay was greater. The group of four individuals with missense variants in KMT2E presented with the most severe developmental delays. Epilepsy was present in all individuals with missense variants, often manifesting as treatment-resistant infantile epileptic encephalopathy. Microcephaly was also common in this group. Haploinsufficiency versus gain-of-function or dominant-negative effects specific to these missense variants in KMT2E might explain this divergence in phenotype, but requires independent validation. Disruptive variants in KMT2E are an under-recognized cause of neurodevelopmental abnormalities

Investigation of hospital discharge cases and SARS-CoV-2 introduction into Lothian care homes

Background The first epidemic wave of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) in Scotland resulted in high case numbers and mortality in care homes. In Lothian, over one-third of care homes reported an outbreak, while there was limited testing of hospital patients discharged to care homes. Aim To investigate patients discharged from hospitals as a source of SARS-CoV-2 introduction into care homes during the first epidemic wave. Methods A clinical review was performed for all patients discharges from hospitals to care homes from 1st March 2020 to 31st May 2020. Episodes were ruled out based on coronavirus disease 2019 (COVID-19) test history, clinical assessment at discharge, whole-genome sequencing (WGS) data and an infectious period of 14 days. Clinical samples were processed for WGS, and consensus genomes generated were used for analysis using Cluster Investigation and Virus Epidemiological Tool software. Patient timelines were obtained using electronic hospital records. Findings In total, 787 patients discharged from hospitals to care homes were identified. Of these, 776 (99%) were ruled out for subsequent introduction of SARS-CoV-2 into care homes. However, for 10 episodes, the results were inconclusive as there was low genomic diversity in consensus genomes or no sequencing data were available. Only one discharge episode had a genomic, time and location link to positive cases during hospital admission, leading to 10 positive cases in their care home. Conclusion The majority of patients discharged from hospitals were ruled out for introduction of SARS-CoV-2 into care homes, highlighting the importance of screening all new admissions when faced with a novel emerging virus and no available vaccine

SARS-CoV-2 Omicron is an immune escape variant with an altered cell entry pathway

Vaccines based on the spike protein of SARS-CoV-2 are a cornerstone of the public health response to COVID-19. The emergence of hypermutated, increasingly transmissible variants of concern (VOCs) threaten this strategy. Omicron (B.1.1.529), the fifth VOC to be described, harbours multiple amino acid mutations in spike, half of which lie within the receptor-binding domain. Here we demonstrate substantial evasion of neutralization by Omicron BA.1 and BA.2 variants in vitro using sera from individuals vaccinated with ChAdOx1, BNT162b2 and mRNA-1273. These data were mirrored by a substantial reduction in real-world vaccine effectiveness that was partially restored by booster vaccination. The Omicron variants BA.1 and BA.2 did not induce cell syncytia in vitro and favoured a TMPRSS2-independent endosomal entry pathway, these phenotypes mapping to distinct regions of the spike protein. Impaired cell fusion was determined by the receptor-binding domain, while endosomal entry mapped to the S2 domain. Such marked changes in antigenicity and replicative biology may underlie the rapid global spread and altered pathogenicity of the Omicron variant

Evaluation of Cytotoxicity and Bioimaging of Nitrogen-Vacancy Nanodiamonds

Nanodiamonds, due to their chemical inertness and biocompatibility, have found extensive uses in drug delivery and biomedical applications. Fluorescent nanodiamonds with fluorescent properties generated by nitrogen-vacancy defects have been intensively investigated for bioimaging, due to their high quantum yield and high photobleaching stability. In addition, the surface properties and particle size of nanodiamonds have significant impacts on cellular uptake and imaging quality. In this study, nitrogen-vacancy nanodiamonds with different particle sizes (40 nm and 90 nm) have been physicochemically characterised and investigated for their cytotoxicity and potential in fluorescence imaging. The nanodiamonds (with concentrations up to 100 µg/mL) showed cell viability &gt;70% with mesenchymal stromal cells. The number of nanodiamonds was observed to have a larger impact on cell viability than the mass of nanodiamonds. Larger nanodiamonds (90 nm) exhibited a lower level of cytotoxicity, higher cellular uptake and fluorescence intensity. The results indicate the potential of using fluorescent nanodiamonds as a nanoprobe for effective bioimaging and cell tracking.</jats:p

Modification of nanodiamonds for fluorescence bioimaging.

Non-invasive bioimaging is essential in enhancing pre-clinical diagnosis and therapy. Developing efficient imaging probes with high stability, low toxicity, and the potential of offering high resolution images is a very important aspect of developing non-invasive bioimaging techniques. Fluorescent nanodiamonds, which are produced by high energy beam irradiation and high temperature/pressure treatment, have been extensively investigated. In this study, we report the chemical modification of common nanodiamonds (prepared by detonation and high-pressure high-temperature milling) using a stable fluorophore (perylene diimide derivative) via carbodiimide coupling. The resulting nanodiamonds show good biocompatibility, cellular uptake and fluorescent imaging potential with mesenchymal stromal cells. This method provides an efficient alternative approach to the preparation and the use of fluorescent nanodiamonds for bioimaging, with the potential benefit of chemically adjusting the structure of perylene diimide for optimized emission/absorbance wavelength