Investigating anti-inflammatory and immunomodulatory properties of brivaracetam and lacosamide in experimental autoimmune encephalomyelitis (EAE).

PURPOSE: Inflammation plays a role in drug-resistant epilepsy (DRE). We have previously reported an increased proportion of CD4 T cells displaying a pro-inflammatory profile in the peripheral blood of adults with DRE. Specific anti-epileptic drugs (AEDs) exhibit immunomodulatory properties that could increase the risk of infections but also contribute to their beneficial impact on DRE and other neurological diseases. The impact of novel generation AEDs on the profile of immune cells and on neuroinflammatory processes remains unclear. METHODS: We compared the influence of brivaracetam and lacosamide on the activation of human and murine peripheral immune cells in vitro and in vivo in active experimental autoimmune encephalomyelitis (EAE), a common mouse model of central nervous system inflammation. RESULTS: We found that brivaracetam and lacosamide at 2.5 µg/ml did not impair the survival and activation of human immune cells, but a higher dose of 25 µg/ml decreased mitogen-induced proliferation of CD8 T cells in vitro. Exposure to high doses of brivaracetam, and to a lesser extent lacosamide, reduced the proportion of CD25+ and CD107a+ CD8+ human T cells in vitro, and the frequency of CNS-infiltrating CD8+ T cells at EAE onset and CD11b+ myeloid cells at peak in vivo. Prophylactic administration of brivaracetam or lacosamide did not delay EAE onset but significantly improved the clinical course in the chronic phase of EAE compared to control. CONCLUSION: Novel generation AEDs do not impair the response to immunization with MOG peptide but improve the course of EAE, possibly through a reduction of neuroaxonal damage.

Contact-Dependent Granzyme B-Mediated Cytotoxicity of Th17-Polarized Cells Toward Human Oligodendrocytes.

Multiple sclerosis (MS) is characterized by the loss of myelin and of myelin-producing oligodendrocytes (OLs) in the central nervous system (CNS). Pro-inflammatory CD4+ Th17 cells are considered pathogenic in MS and are harmful to OLs. We investigated the mechanisms driving human CD4+ T cell-mediated OL cell death. Using fluorescent and brightfield in vitro live imaging, we found that compared to Th2-polarized cells, Th17-polarized cells show greater interactions with primary human OLs and human oligodendrocytic cell line MO3.13, displaying longer duration of contact, lower mean speed, and higher rate of vesicle-like structure formation at the sites of contact. Using single-cell RNA sequencing, we assessed the transcriptomic profile of primary human OLs and Th17-polarized cells in direct contact or separated by an insert. We showed that upon close interaction, OLs upregulate the expression of mRNA coding for chemokines and antioxidant/anti-apoptotic molecules, while Th17-polarized cells upregulate the expression of mRNA coding for chemokines and pro-inflammatory cytokines such as IL-17A, IFN-γ, and granzyme B. We found that secretion of CCL3, CXCL10, IFN-γ, TNFα, and granzyme B is induced upon direct contact in cocultures of human Th17-polarized cells with human OLs. In addition, we validated by flow cytometry and immunofluorescence that granzyme B levels are upregulated in Th17-polarized compared to Th2-polarized cells and are even higher in Th17-polarized cells upon direct contact with OLs or MO3.13 cells compared to Th17-polarized cells separated from OLs by an insert. Moreover, granzyme B is detected in OLs and MO3.13 cells following direct contact with Th17-polarized cells, suggesting the release of granzyme B from Th17-polarized cells into OLs/MO3.13 cells. To confirm granzyme B-mediated cytotoxicity toward OLs, we showed that recombinant human granzyme B can induce OLs and MO3.13 cell death. Furthermore, pretreatment of Th17-polarized cells with a reversible granzyme B blocker (Ac-IEPD-CHO) or a natural granzyme B blocker (serpina3N) improved survival of MO3.13 cells upon coculture with Th17 cells. In conclusion, we showed that human Th17-polarized cells form biologically significant contacts with human OLs and exert direct toxicity by releasing granzyme B.

Identification of SARS-CoV-2-specific immune alterations in acutely ill patients.

Dysregulated immune profiles have been described in symptomatic patients infected with SARS-CoV-2. Whether the reported immune alterations are specific to SARS-CoV-2 infection or also triggered by other acute illnesses remains unclear. We performed flow cytometry analysis on fresh peripheral blood from a consecutive cohort of (a) patients hospitalized with acute SARS-CoV-2 infection, (b) patients of comparable age and sex hospitalized for another acute disease (SARS-CoV-2 negative), and (c) healthy controls. Using both data-driven and hypothesis-driven analyses, we found several dysregulations in immune cell subsets (e.g., decreased proportion of T cells) that were similarly associated with acute SARS-CoV-2 infection and non-COVID-19-related acute illnesses. In contrast, we identified specific differences in myeloid and lymphocyte subsets that were associated with SARS-CoV-2 status (e.g., elevated proportion of ICAM-1+ mature/activated neutrophils, ALCAM+ monocytes, and CD38+CD8+ T cells). A subset of SARS-CoV-2-specific immune alterations correlated with disease severity, disease outcome at 30 days, and mortality. Our data provide an understanding of the immune dysregulation specifically associated with SARS-CoV-2 infection among acute care hospitalized patients. Our study lays the foundation for the development of specific biomarkers to stratify SARS-CoV-2-positive patients at risk of unfavorable outcomes and to uncover candidate molecules to investigate from a therapeutic perspective.

On the structure-function of MHC class II molecules and how single amino acid polymorphisms could alter intracellular trafficking.

Classical HLA class II molecules are highly polymorphic heterodimeric transmembrane proteins encoded by a polygenic cluster on chromosome 6. Polymorphic residues in the membrane-distal domains ensure that a large collection of microbial peptides can be bound in the human population. Still, the HLA-DR, -DP and -DQ isotypes show a high degree of conservation in their overall tertiary and quaternary structures, in line with their common function in T cell receptor activation. Interestingly, the primary structure of the intracellular domains are highly divergent between isotypes and they also show allotypic variations. The functional impact of these differences remains to be fully appreciated. Here, we address the role of the MHC class II cytoplasmic tails in intracellular trafficking. First, the emphasis will be on the interplay between the cytoplasmic domains of classical human MHC class II molecules and those of the invariant chain chaperone (CD74) isoforms. Then, we will examine the importance of the highly conserved ß-chain cytoplasmic lysine residue in the ubiquitin-driven trafficking of MHC class II molecules. These considerations should help understand the potential functional impact of sequence variations that may arise in the cytoplasmic tails and transmembrane domains of MHC class II molecules.