Biology subjects

Mathias, A.

Publications and source records attributed to Mathias, A..

5 recordsLinked to original sources

bioRxiv2024

KIR3DL1 and Tox identify clonally expanded encephalitogenic neuron-specific CD8+ T cells in autoimmune encephalitis

Autoreactive CD8+ T cells are the principal suspects in autoimmune encephalitis (AIE) with antibodies targeting intracellular neuronal antigens So far, the search for neuron-autoreactive CD8+ T cells has been focused on a few autoantigens and did not yield convincing results. Here, we leveraged natural antigen presentation by hiPSC-derived neurons to look at the global autoreactive CD8+ T cell response, independently of pre-conceived hypothesis of the autoantigens involved in the disease. This unbiased approach allowed for the identification of rare polyclonal neuron-reactive CD8+ T cells in healthy donors, and contrastingly, expanded clonotypes in two patients with anti-Ri AIE. Detailed ex vivo phenotypic characterization of these clonotypes revealed a specific transcriptional program suggestive of a pathogenic potential. In particular, this subset can be identified by the expression of KIR3DL1 and TOX. Strikingly, we could also demonstrate that CD8+ T cells found in the brain of an anti-Ri AIE patient display a similar phenotype associated with cytotoxicity and encephalitogenic features.

immunology↗

bioRxiv2024

Human stem cell derived neurons and astrocytes to detect auto-reactive IgG in neurological diseases

Background and objectivesUp to 46% of patients with presumed autoimmune limbic encephalitis are seronegative for all currently known CNS antigens. We developed a cell-based assay (CBA) to screen for novel neural antibodies in serum and CSF using neurons and astrocytes derived from human induced pluripotent stem cells (hiPSC). MethodsHuman iPSC-derived astrocytes or neurons were incubated with serum/CSF from 99 patients (42 with inflammatory neurological diseases (IND) and 57 with non-IND (NIND)). The IND group included 11 patients with previously established neural antibodies, six with seronegative neuromyelitis optica spectrum disorder (NMOSD), 12 with suspected autoimmune encephalitis/paraneoplastic syndrome (AIE/PNS), and 13 with other IND (OIND). IgG binding to fixed CNS cells was detected using fluorescently-labelled antibodies and analyzed through automated fluorescence measures. IgG neuronal/astrocyte reactivity was further analyzed by flow cytometry. Peripheral blood mononuclear cells (PBMC) were used as CNS-irrelevant control target cells. Reactivity profile was defined as positive using a Robust regression and Outlier removal test with a false discovery rate at 10% following each individual readout. ResultsUsing our CBA, we detected antibodies recognizing hiPSC-derived neural cells in 19/99 subjects. Antibodies bound specifically to astrocytes in nine cases, to neurons in eight cases and to both cell types in two cases, as confirmed by microscopy single-cell analyses. Highlighting the significance of our novel 96-well CBA assay, neural-specific antibody binding was more frequent in IND (15/42) than in NIND patients (4/57) (Fisher test, p=0.0005). Three of three patients with astrocyte- reactive (2 AQP4+ NMO, 1 GFAP astrocytopathy), and 3/4 with intracellular neuron-reactive antibodies (2 Hu+, 1 Ri+ AIE/PNS), as identified in diagnostic laboratories, were also positive with our CBA. Most interestingly, we showed antibody-reactivity in 2/6 seronegative NMOSD, 6/12 probable AIE/PNS, and 1/13 OIND. Flow cytometry using hiPSC-derived CNS cells or PBMC detected antibody binding in 13 versus 0 patients, respectively, establishing the specificity of the detected antibodies for neural tissue. DiscussionOur unique hiPSC-based CBA allows for the screening of novel neuron-/astrocyte-reactive antibodies in patients with suspected immune-mediated neurological syndromes, and negative testing in established routine laboratories, opening new perspectives in establishing early diagnosis of such complex diseases.

neuroscience↗

bioRxiv2023

Cell-binding IgM in CSF is distinctive of multiple sclerosis and targets the iron transporter SCARA5

Intrathecal IgM production in multiple sclerosis (MS) is associated with a worse disease course. To investigate pathogenic relevance of autoreactive IgM in MS, CSF from two independent cohorts, including MS patients and controls, were screened for antibody binding to induced pluripotent stem cell-derived neurons and astrocytes, and a panel of CNS-related cell lines. IgM binding to a primitive neuro-ectodermal tumour cell line discriminated 10% of MS donors from controls. Transcriptomes of single IgM producing CSF B cells from patients with cell-binding IgM were sequenced and used to produce recombinant monoclonal antibodies for characterisation and antigen identification. We produced 5 cell-binding recombinant IgM antibodies, of which one, cloned from an HLA-DR+ plasma-like B cell, mediated antigen-dependent complement activation. Immunoprecipitation and mass spectrometry, and biochemical and transcriptome analysis of the target cells identified the iron transport scavenger protein SCARA5 as the antigen target of this antibody. Intrathecal injection of a SCARA5 antibody led to an increased T cell infiltration in an EAE model. CSF IgM might contribute to CNS inflammation in MS by binding to cell surface antigens like SCARA5 and activating complement, or by facilitating immune cell migration into the brain.

immunology↗

bioRxiv2023

Unique proteomic signature of JCPyV-infected human astrocytes: from cells to extracellular vesicles

JC polyomavirus (JCPyV) is an opportunistic virus that remains in a latent state in the kidneys of more than half of the human adult population. In rare cases of severe immune suppression, the virus is able to establish a lytic infection of glial cells in the brain, resulting in a debilitating, demyelinating disease known as progressive multifocal leukoencephalopathy (PML). Because of the exceptional species and tissue specificity of the virus, appropriate models of JCPyV infection in the brain are lacking, thus hampering progress towards the development of novel antiviral strategies and biomarkers of disease activity. While PML has traditionally been characterized as a lytic infection of oligodendrocytes, more recent findings suggest an important role for astrocytes during the initial stages of disease. Here, using human induced pluripotent stem cell (hiPSC) derived-astrocytes coupled with a multiparametric approach, we show that 1. JCPyV readily infects and replicates in astrocytes, 2. JCPyV strongly dysregulates the cell biology and 3. these findings adequately reflect ex vivo findings. We perform an in-depth characterization of the effect of JCPyV on the cell proteome over time, demonstrating a strong dysregulation of the cell cycle and activation of the DNA damage response. Furthermore, we show that the proteomic signature observed for infected astrocytes is extended to excreted vesicles, underlining their potential to gain valuable insights into JCPyV propagation in the brain.

neuroscience↗

bioRxiv2023

Deletion of smooth muscle O-GlcNAc transferase prevents development of atherosclerosis in western diet-fed hyperglycemic ApoE-/- mice in vivo

Accumulating evidence highlights protein O-GlcNAcylation as a putative pathogenic contributor of diabetic vascular complications. We previously reported that elevated protein O-GlcNAcylation correlates with increased atherosclerotic lesion formation and VSMC proliferation in response to hyperglycemia. However, the role of O-GlcNAc transferase (OGT), regulator of O-GlcNAc signaling, in evolution of diabetic atherosclerosis remains elusive. The goal of this study was to determine whether smooth muscle OGT (smOGT) plays a direct role in hyperglycemia-induced atherosclerotic lesion formation and SMC de-differentiation. Using tamoxifen-inducible Myh11-CreERT2 and OGTfl/fl mice, we generated smOGTWT and smOGTKO mice, with and without ApoE-null backgrounds. Following STZ-induced hyperglycemia, smOGTWT and smOGTKO mice were kept on standard laboratory diet for study duration. In a parallel study, smOGTWTApoE-/- and smOGTKOApoE-/- were kept on Western diet beginning 8-wks-age. Animals harvested at 14-16-wks-age were used for plasma and tissue collection. Loss of smOGT augmented SM contractile marker expression in aortic vessels of STZ-induced hyperglycemic smOGTKO mice. Consistently, smOGT deletion attenuated atherosclerotic lesion lipid burden (Oil red O), plaque area (H&E), leukocyte (CD45) and smooth muscle cell (ACTA2) abundance in Western diet-induced hyperglycemic smOGTKOApoE-/- mice. This was accompanied with increased SM contractile markers, and reduced inflammatory and proliferative marker expression. Further, smOGT deletion attenuated YY1 and SRF expression (transcriptional regulators of SM contractile genes) in hyperglycemic smOGTKOApoE-/- and smOGTKO mice. These data uncover an atheroprotective outcome of smOGT loss-of-function and suggest a direct regulatory role of OGT-mediated O-GlcNAcylation in VSMC de-differentiation in hyperglycemia.

pathology↗