Tolerance Induction of IgG+ Memory B Cells by T Cell-Independent Type II Antigens

Vesicular stomatitis virus (VSV) induces an early T cell-independent neutralizing lgM response that is followed by a long-lived, T cell-dependent lgG response. We used the specific amplification factor of several 100x of VSV-virions for immunohistology to analyze the localization of VSV-specific B cells at different time points after immunization. At the peak of the IgM response (day 4), VSV-specific B cells were predominantly present in the red pulp and marginal zone but not in the T area. These B cells were mostly stained in the cytoplasm, characterizing them as antibody secreting cells. By day 6 after immunization, germinal centers (GC) containing surface-stained VSV-specific B cells became detectable and were fully established by day 12. At the same time, large VSV-specific B cell aggregates were present in the red pulp. High numbers of VSV-specific GC associated with persisting antigen were present 1 mo after immunization and later, i.e., considerably longer than has been observed for haptens. Some GC, exhibiting follicular dendritic cells and containing VSV-specific, proliferating B cells were still detectable up to 100 d after immunization. Long-lived GC were also observed after immunization with recombinant VSV-glycoprotein in absence of adjuvants. Thus some anti-virally protective (memory) B cells are cycling and locally proliferate in long-lived GC in association with persisting antigen and therefore seem responsible for long-term maintenance of elevated antibody levels. These observations extend earlier studies with carrier hapten antigens in adjuvant depots or complexed with specific IgG; they are the first to show colocalization of antigen and specific memory B cells and to analyze a protective neutralizing antibody response against an acute viral infection.

Download Full-text

COVID-19 convalescents exhibit deficient humoral and T cell responses to variant of concern Spike antigens at 12 month post-infection

10.1101/2021.11.08.21266035 ◽

2021 ◽

Author(s):

Pablo Garcia-Valtanen ◽

Christopher Martin Hope ◽

Makutiro Ghislain Masavuli ◽

Arthur Eng Lip Yeow ◽

Harikrishnan Balachandran ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

B Cells ◽

Post Infection ◽

T Cell Responses ◽

Memory B Cells ◽

Pseudotyped Virus ◽

Cell Responses ◽

Specific Memory ◽

Cell Functionality

Background The duration and magnitude of SARS-CoV-2 immunity after infection, especially with regard to the emergence of new variants of concern (VoC), remains unclear. Here, immune memory to primary infection and immunity to VoC was assessed in mild-COVID-19 convalescents one year after infection and in the absence of viral re-exposure or COVID-19 vaccination. Methods Serum and PBMC were collected from mild-COVID-19 convalescents at ~6 and 12 months after a COVID-19 positive PCR (n=43) and from healthy SARS-CoV-2-seronegative controls (n=15-40). Serum titers of RBD and Spike-specific Ig were quantified by ELISA. Virus neutralisation was assessed against homologous, pseudotyped virus and homologous and VoC live viruses. Frequencies of Spike and RBD-specific memory B cells were quantified by flow cytometry. Magnitude of memory T cell responses was quantified and phenotyped by activation-induced marker assay, while T cell functionality was assessed by intracellular cytokine staining using peptides specific to homologous Spike virus antigen and four VoC Spike antigens. Findings At 12 months after mild-COVID-19, >90% of convalescents remained seropositive for RBD-IgG and 88.9% had circulating RBD-specific memory B cells. Despite this, only 51.2% convalescents had serum neutralising activity against homologous live-SARS-CoV-2 virus, which decreased to 44.2% when tested against live B.1.1.7, 4.6% against B.1.351, 11.6% against P.1 and 16.2%, against B.1.617.2 VoC. Spike and non-Spike-specific T cells were detected in >50% of convalescents with frequency values higher for Spike antigen (95% CI, 0.29-0.68% in CD4+ and 0.11-0.35% in CD8+ T cells), compared to non-Spike antigens. Despite the high prevalence and maintenance of Spike-specific T cells in Spike 'high-responder' convalescents at 12 months, T cell functionality, measured by cytokine expression after stimulation with Spike epitopes corresponding to VoC was severely affected. Interpretations SARS-CoV-2 immunity is retained in a significant proportion of mild COVID-19 convalescents 12 months post-infection in the absence of re-exposure to the virus. Despite this, changes in the amino acid sequence of the Spike antigen that are present in current VoC result in virus evasion of neutralising antibodies, as well as evasion of functional T cell responses.

Download Full-text

Vaccine-elicited CD4 T cells prevent the deletion of antiviral B cells in chronic infection

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.2108157118 ◽

2021 ◽

Vol 118 (46) ◽

pp. e2108157118

Author(s):

Kerstin Narr ◽

Yusuf I. Ertuna ◽

Benedict Fallet ◽

Karen Cornille ◽

Mirela Dimitrova ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

B Cells ◽

B Cell ◽

Cd4 T Cells ◽

Memory B Cells ◽

Type I ◽

Clonal Deletion ◽

Cell Immunity ◽

B Cell Immunity

Chronic viral infections subvert protective B cell immunity. An early type I interferon (IFN-I)–driven bias to short-lived plasmablast differentiation leads to clonal deletion, so-called “decimation,” of antiviral memory B cells. Therefore, prophylactic countermeasures against decimation remain an unmet need. We show that vaccination-induced CD4 T cells prevented the decimation of naïve and memory B cells in chronically lymphocytic choriomeningitis virus (LCMV)-infected mice. Although these B cell responses were largely T independent when IFN-I was blocked, preexisting T help assured their sustainability under conditions of IFN-I–driven inflammation by instructing a germinal center B cell transcriptional program. Prevention of decimation depended on T cell–intrinsic Bcl6 and Tfh progeny formation. Antigen presentation by B cells, interactions with antigen-specific T helper cells, and costimulation by CD40 and ICOS were also required. Importantly, B cell–mediated virus control averted Th1-driven immunopathology in LCMV-challenged animals with preexisting CD4 T cell immunity. Our findings show that vaccination-induced Tfh cells represent a cornerstone of effective B cell immunity to chronic virus challenge, pointing the way toward more effective B cell–based vaccination against persistent viral diseases.

Download Full-text

Potential anti-EBV effects associated with elevated interleukin-21 levels: a case report

BMC Infectious Diseases ◽

10.1186/s12879-020-05609-z ◽

2020 ◽

Vol 20 (1) ◽

Author(s):

Kristian Assing ◽

Christian Nielsen ◽

Marianne Jakobsen ◽

Charlotte B. Andersen ◽

Kristin Skogstrand ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

B Cells ◽

B Cell ◽

Plasma Cell ◽

Germinal Center ◽

Plasma Cells ◽

Cell Formation ◽

Memory B Cells ◽

Memory B Cell

Abstract Background Germinal center derived memory B cells and plasma cells constitute, in health and during EBV reactivation, the largest functional EBV reservoir. Hence, by reducing germinal center derived formation of memory B cells and plasma cells, EBV loads may be reduced. Animal and in-vitro models have shown that IL-21 can support memory B and plasma cell formation and thereby potentially contribute to EBV persistence. However, IL-21 also displays anti-viral effects, as mice models have shown that CD4+ T cell produced IL-21 is critical for the differentiation, function and survival of anti-viral CD8+ T cells able to contain chronic virus infections. Case presentation We present immunological work-up (flow-cytometry, ELISA and genetics) related to a patient suffering from a condition resembling B cell chronic active EBV infection, albeit with moderately elevated EBV copy numbers. No mutations in genes associated with EBV disease, common variable immunodeficiency or pertaining to the IL-21 signaling pathway (including hypermorphic IL-21 mutations) were found. Increased (> 5-fold increase 7 days post-vaccination) CD4+ T cell produced (p < 0.01) and extracellular IL-21 levels characterized our patient and coexisted with: CD8+ lymphopenia, B lymphopenia, hypogammaglobulinemia, compromised memory B cell differentiation, absent induction of B-cell lymphoma 6 protein (Bcl-6) dependent peripheral follicular helper T cells (pTFH, p = 0.01), reduced frequencies of peripheral CD4+ Bcl-6+ T cells (p = 0.05), compromised plasmablast differentiation (reduced protein vaccine responses (p < 0.001) as well as reduced Treg frequencies. Supporting IL-21 mediated suppression of pTFH formation, pTFH and CD4+ IL-21+ frequencies were strongly inversely correlated, prior to and after vaccination, in the patient and in controls, Spearman’s rho: − 0.86, p < 0.001. Conclusions To the best of our knowledge, this is the first report of elevated CD4+ IL-21+ T cell frequencies in human EBV disease. IL-21 overproduction may, apart from driving T cell mediated anti-EBV responses, disrupt germinal center derived memory B cell and plasma cell formation, and thereby contribute to EBV disease control.

Download Full-text

Ectopic Lymphoid Follicles in Multiple Sclerosis: Centers for Disease Control?

Frontiers in Neurology ◽

10.3389/fneur.2020.607766 ◽

2020 ◽

Vol 11 ◽

Author(s):

Austin Negron ◽

Olaf Stüve ◽

Thomas G. Forsthuber

Keyword(s):

Multiple Sclerosis ◽

T Cells ◽

T Cell ◽

B Cells ◽

B Cell ◽

T Cell Responses ◽

Memory B Cells ◽

Lymphoid Follicles ◽

Cell Responses ◽

B Cell Responses

While the contribution of autoreactive CD4+ T cells to the pathogenesis of Multiple Sclerosis (MS) is widely accepted, the advent of B cell-depleting monoclonal antibody (mAb) therapies has shed new light on the complex cellular mechanisms underlying MS pathogenesis. Evidence supports the involvement of B cells in both antibody-dependent and -independent capacities. T cell-dependent B cell responses originate and take shape in germinal centers (GCs), specialized microenvironments that regulate B cell activation and subsequent differentiation into antibody-secreting cells (ASCs) or memory B cells, a process for which CD4+ T cells, namely follicular T helper (TFH) cells, are indispensable. ASCs carry out their effector function primarily via secreted Ig but also through the secretion of both pro- and anti-inflammatory cytokines. Memory B cells, in addition to being capable of rapidly differentiating into ASCs, can function as potent antigen-presenting cells (APCs) to cognate memory CD4+ T cells. Aberrant B cell responses are prevented, at least in part, by follicular regulatory T (TFR) cells, which are key suppressors of GC-derived autoreactive B cell responses through the expression of inhibitory receptors and cytokines, such as CTLA4 and IL-10, respectively. Therefore, GCs represent a critical site of peripheral B cell tolerance, and their dysregulation has been implicated in the pathogenesis of several autoimmune diseases. In MS patients, the presence of GC-like leptomeningeal ectopic lymphoid follicles (eLFs) has prompted their investigation as potential sources of pathogenic B and T cell responses. This hypothesis is supported by elevated levels of CXCL13 and circulating TFH cells in the cerebrospinal fluid (CSF) of MS patients, both of which are required to initiate and maintain GC reactions. Additionally, eLFs in post-mortem MS patient samples are notably devoid of TFR cells. The ability of GCs to generate and perpetuate, but also regulate autoreactive B and T cell responses driving MS pathology makes them an attractive target for therapeutic intervention. In this review, we will summarize the evidence from both humans and animal models supporting B cells as drivers of MS, the role of GC-like eLFs in the pathogenesis of MS, and mechanisms controlling GC-derived autoreactive B cell responses in MS.

Download Full-text

Naive T-cell receptor transgenic T cells help memory B cells produce antibody

Immunology ◽

10.1111/j.1365-2567.2006.02446.x ◽

2006 ◽

Vol 119 (3) ◽

pp. 376-384 ◽

Cited By ~ 9

Author(s):

Darragh Duffy ◽

Chun-Ping Yang ◽

Andrew Heath ◽

Paul Garside ◽

Eric B. Bell

Keyword(s):

T Cells ◽

T Cell ◽

B Cells ◽

T Cell Receptor ◽

Cell Receptor ◽

Memory B Cells ◽

Naïve T Cell ◽

Naive T Cell

Download Full-text

Induction of ‘Tolerance’ in a T-Cell Clone from a Mild Hemophilia Patient

Blood ◽

10.1182/blood.v112.11.3526.3526 ◽

2008 ◽

Vol 112 (11) ◽

pp. 3526-3526

Author(s):

David W. Scott ◽

Elizabeth Kadavil ◽

Ai-Hong Zhang ◽

Ruth A. Ettinger ◽

Kathleen Pratt

Keyword(s):

T Cell ◽

B Cells ◽

Hemophilia A ◽

Cell Clone ◽

Tolerance Induction ◽

Hla Dr ◽

Cell Clones ◽

T Cell Clone ◽

Pre Treatment

Abstract A major obstacle in the treatment of Hemophilia A is that patients can develop an inhibitory immune response to therapeutic doses of coagulation factor VIII (fVIII). Over the last decade, we have developed a B-cell delivered gene therapy approach to prevent the development of inhibitory antibodies (“inhibitors”) in fVIII knockout mice (see Lei and Scott, Blood105: 4865, 2005). In our murine platform, activated primary spleen B cells or bone marrow cells are transduced with a retroviral vector encoding the fVIII A2 and/or C2 domain fused to an IgG heavy chain, and these cells are injected systemically into immunocompetent fVIII knockout animals. The recipients are rendered specifically tolerant to the encoded C2 and A2 domains, as evidenced by a >90% reduction of inhibitor titers, even in primed animals. To help evaluate the potential of this approach for translation, we are developing in vitro models for tolerance induction using human T-cell clones isolated from subjects with mild hemophilia A. The clones are isolated by single-cell sorting of CD4+ cells that are labeled by fluorescent HLA-DR tetramers complexed with peptides containing fVIII epitopes, followed by expansion with HLA-DR mismatched peripheral blood mononuclear cells (PBMC), phytohaemagglutinin, and interleukin-2. Our initial model utilizes a T-cell clone from an individual with mild hemophilia A due to fVIII missense genotype A2201P, which recognizes an HLA-DRA-DRB1*0101-restricted epitope within a synthetic peptide corresponding to fVIII residues 2194–2213. All of the antigen-specific T-cell clones isolated from this subject secreted interferon-gamma (IFN-γ) when stimulated by fVIII2194–2213 presented by irradiated HLA-DR-matched PBMCs or with plate-bound anti-CD3. Because of their robust response to a clinically relevant epitope in fVIII, one of these clones that expanded well in culture was chosen for initial testing of a modified gene therapy platform similar to that developed using the murine hemophilia A model. HLA-matched peripheral blood B cells were activated with antibodies to IgM or with CD40L-expressing fibroblasts and then transduced with a modified retroviral vector containing the human C2 domain sequence in-frame with the IgG sequence. These B cells were cultured with the hemophilic T-cell clone. After pre-treatment (“tolerance-induction step”), the cells were washed and then stimulated by plate-bound anti-CD3. The subsequent IFN-γ response (measured by ELIspots and ELISA) was dramatically reduced compared to the response of same T-cell clone cultured with mock-transduced B cells. The post-treatment reduction in IFN-γ secretion was equivalent to that induced after soluble anti-CD3 pre-treatment, a known method to induce T-cell anergy in vitro. Interestingly, IL-10 was produced during the tolerance induction (pre-treatment) phase, most likely from the activated B cells. Preliminary, parallel experiments with B cells transduced with a “gutless” adenovirus vector expressing C2-Ig did not result in a similar down-regulation of the T-cell response, suggesting that this non-integrating method of expressing antigens for tolerance is not effective, at least in this system. These results are the first to demonstrate in vitro modulation of cytokine responses using DR-restricted, fVIII-specific T cells from a hemophilia A subject. Further investigations using T-cell clones from hemophilic subjects with and without anti-fVIII antibodies will allow us to explore mechanisms of tolerance and may also suggest novel approaches to reduce inhibitor titers.

Download Full-text