B cells in the pneumococcus-infected lung are heterogeneous and require CD4+ T cell help including CD40L to become resident memory B cells.

Recovery from respiratory pneumococcal infections generates lung-localized protection against heterotypic bacteria, mediated by resident memory lymphocytes. Optimal protection in mice requires re-exposure to pneumococcus within days of initial infection. Serial surface marker phenotyping of B cell populations in a model of pneumococcal heterotypic immunity revealed that bacterial re-exposure stimulates the immediate accumulation of dynamic and heterogeneous populations of B cells in the lung, and is essential for the establishment of lung resident memory B (BRM) cells. The B cells in the early wave were activated, proliferating locally, and associated with both CD4+ T cells and CXCL13. Antagonist- and antibody-mediated interventions were implemented during this early timeframe to demonstrate that lymphocyte recirculation, CD4+ cells, and CD40 ligand (CD40L) signaling were all needed for lung BRM cell establishment, whereas CXCL13 signaling was not. While most prominent as aggregates in the loose connective tissue of bronchovascular bundles, morphometry and live lung imaging analyses showed that lung BRM cells were equally numerous as single cells dispersed throughout the alveolar septae. We propose that CD40L signaling from antigen-stimulated CD4+ T cells in the infected lung is critical to establishment of local BRM cells, which subsequently protect the airways and parenchyma against future potential infections.

Signatures of CD4+ T and B cells are associated with distinct stages of chronic chagasic cardiomyopathy.

Introduction: Chagas disease is a neglected parasitic disease caused by Trypanosoma cruzi. While most patients are asymptomatic, around 30% develop Chronic Chagasic Cardiomyopathy (CCC). Methods: Here, we employed high-dimensional flow cytometry to analyze CD4+ T and B cell compartments in patients during the chronic phase of Chagas disease, presenting the asymptomatic and mild or moderate/severe cardiac clinical forms. Results: Effector CD27-CD4+ T cells were expanded in both CCC groups, and only mild CCC patients showed higher frequencies of effector memory and T follicular helper (Tfh) cells than healthy donors (CTL) and asymptomatic patients. Unsupervised analysis confirmed these findings and further revealed the expansion of a specific subpopulation composed of Tfh, transitional, and central memory CD4+ T cells bearing a phenotype associated with strong activation, differentiation, and exhaustion in patients with mild but not moderate/severe CCC. In contrast, patients with mild and moderate/severe CCC had lower frequencies of CD4+ T cells expressing lower levels of activation markers, suggesting resting status, than CTL. Regarding the B cell compartment, no alterations were found in naïve CD21-, memory cells expressing IgM or IgD, marginal zone, and plasma cells in patients with Chagas disease. However, expansion of class-switched activated and atypical memory B cells was observed in all clinical forms, and more substantially in mild CCC patients. Discussion: Taken together, our results showed that T. cruzi infection triggers changes in CD4+ T and B cell compartments that are more pronounced in the mild CCC clinical form, suggesting an orchestrated cellular communication during Chagas disease. Conclusion: Overall, these findings reinforce the heterogeneity and complexity of the immune response in patients with chronic Chagas disease and may provide new insights into disease pathology and potential markers to guide clinical decisions.

Causal relationship between immune cells and atrial fibrillation: A Mendelian randomization study.

Atrial fibrillation (AF) is a prevalent cardiac arrhythmia, with recent research indicating a correlation between immune system characteristics and the development of AF. However, it remains uncertain whether the immunological response is the primary underlying component or a secondary consequence of AF. Initially, we investigated the effect of immune cells on AF by performing forward Mendelian randomization (MR) analyses with immune cells as the exposure variable and their associated genetic variants as instrumental variables. Subsequently, we performed reverse MR analyses with AF as the exposure variable and immune cells as the outcome variable to exclude the interference of reverse causality, to distinguish between primary and secondary effects, and to further elucidate the causal relationship between the immune system and AF. We discovered that membrane proteins on specific immune cells, such as CD25 on memory B cells-which functions as a part of the interleukin-2 receptor-may be risk factors for AF development, with odds ratios of 1.0233 (95% confidence interval: 1.0012-1.0458, P = .0383). In addition, certain immune cell counts, such as the CD4 regulatory T cell Absolute Count, play a protective factor in the development of AF (odds ratio: 0.9513, 95% confidence interval: 0.9165-0.9874; P = .0086). More detailed results are elaborated in the main text. Our MR study has yielded evidence that substantiates a genetically inferred causal association between the immune system and AF. Identifying the risk factors associated with AF is vital to facilitate the development of innovative pharmaceutical treatments.

Interleukin-21 receptor signaling promotes metabolic dysfunction-associated steatohepatitis-driven hepatocellular carcinoma by inducing immunosuppressive IgA+ B cells.

BACKGROUND: Dysregulation of immune surveillance is tightly linked to the development of metabolic dysfunction-associated steatohepatitis (MASH)-driven hepatocellular carcinoma (HCC); however, its underlying mechanisms remain unclear. Herein, we aimed to determine the role of interleukin-21 receptor (IL-21R) in MASH-driven HCC. METHODS: The clinical significance of IL-21R was assessed in human HCC specimens using immunohistochemistry staining. Furthermore, the expression of IL-21R in mice was assessed in the STAM model. Thereafter, two different MASH-driven HCC mouse models were applied between IL-21R-deficient mice and wild type controls to explore the role of IL-21R in MASH-driven HCC. To further elucidate the potential mechanisms by which IL-21R affected MASH-driven HCC, whole transcriptome sequencing, flow cytometry and adoptive lymphocyte transfer were performed. Finally, flow cytometry, enzyme-linked immunosorbent assay, immunofluorescent staining, chromatin immunoprecipitation assay and western blotting were conducted to explore the mechanism by which IL-21R induced IgA+ B cells. RESULTS: HCC patients with high IL-21R expression exhibited poor relapse-free survival, advanced TNM stage and severe steatosis. Additionally, IL-21R was demonstrated to be upregulated in mouse liver tumors. Particularly, ablation of IL-21R impeded MASH-driven hepatocarcinogenesis with dramatically reduction of lipid accumulation. Moreover, cytotoxic CD8+ T lymphocyte activation was enhanced in the absence of IL-21R due to the reduction of immunosuppressive IgA+ B cells. Mechanistically, the IL-21R-STAT1-c-Jun/c-Fos regulatory axis was activated in MASH-driven HCC and thus promoted the transcription of Igha, resulting in the induction of IgA+ B cells. CONCLUSIONS: IL-21R plays a cancer-promoting role by inducing IgA+ B cells in MASH-driven hepatocarcinogenesis. Targeting IL-21R signaling represents a potential therapeutic strategy for cancer therapy.

Patients and mice with deficiency in the SNARE protein SYNTAXIN-11 have a secondary B cell defect.

SYNTAXIN-11 (STX11) is a SNARE protein that mediates the fusion of cytotoxic granules with the plasma membrane at the immunological synapses of CD8 T or NK cells. Autosomal recessive inheritance of deleterious STX11 variants impairs cytotoxic granule exocytosis, causing familial hemophagocytic lymphohistiocytosis type 4 (FHL-4). In several FHL-4 patients, we also observed hypogammaglobulinemia, elevated frequencies of naive B cells, and increased double-negative DN2:DN1 B cell ratios, indicating a hitherto unrecognized role of STX11 in humoral immunity. Detailed analysis of Stx11-deficient mice revealed impaired CD4 T cell help for B cells, associated with disrupted germinal center formation, reduced isotype class switching, and low antibody avidity. Mechanistically, Stx11-/- CD4 T cells exhibit impaired membrane fusion leading to reduced CD107a and CD40L surface mobilization and diminished IL-2 and IL-10 secretion. Our findings highlight a critical role of STX11 in SNARE-mediated membrane trafficking and vesicle exocytosis in CD4 T cells, important for successful CD4 T cell-B cell interactions. Deficiency in STX11 impairs CD4 T cell-dependent B cell differentiation and humoral responses.

Potential role of IGF-1R in the interaction between orbital fibroblasts and B lymphocytes: an implication for B lymphocyte depletion in the active inflammatory phase of thyroid-associated ophthalmopathy.

BACKGROUND: Thyroid eye disease (TED) is an inflammatory process involving lymphocyte-mediated immune response and orbital tissue damage. The anti-insulin-like growth factor-1 receptor (IGF-1R) antibodies produced by B lymphocytes are involved in the activation of orbital fibroblasts and the inflammatory process of orbital tissue damage in TED. The purpose of this study was to explore the role of IGF-1R in the mechanistic connection between orbital fibroblasts and B lymphocytes in TED. METHODS: Orbital fibroblasts sampled from orbital connective tissues and peripheral B lymphocytes isolated from peripheral blood, which were obtained from 15 patients with TED and 15 control patients, were co-cultured at a ratio of 1:20. The level of IGF-1R expression in orbital fibroblasts was evaluated by flow cytometry and confocal microscopy. Transient B lymphocyte depletion was induced with anti-CD20 monoclonal antibody rituximab, while the IGF-1R pathway was blocked by the IGF-1R binding protein. The expression levels of interleukin-6 (IL-6) and regulated upon activation, normal T cell expressed and secreted (RANTES) in the co-culture model were quantified via ELISA. RESULTS: IGF-1R expression was significantly elevated in TED orbital fibroblasts compared to that of controls. A 24-h co-culture of orbital fibroblasts with peripheral B lymphocytes induced elevated expression levels of IL-6 and RANTES in each group (TED patients and controls), with the highest levels occurring in TED patients (T + T group). Rituximab and IGF-1R binding protein significantly inhibited increased levels of IL-6 and RANTES in the co-culture model of TED patients. CONCLUSIONS: IGF-1R may mediate interaction between orbital fibroblasts and peripheral B lymphocytes; thus, blocking IGF-1R may reduce the local inflammatory response in TED. Rituximab-mediated B lymphocyte depletion played a role in inhibiting inflammatory responses in this in vitro co-culture model, providing a theoretical basis for the clinical application of anti-CD20 monoclonal antibodies in TED.

Double-negative B cells and DNASE1L3 colocalise with microbiota in gut-associated lymphoid tissue.

Intestinal homeostasis is maintained by the response of gut-associated lymphoid tissue to bacteria transported across the follicle associated epithelium into the subepithelial dome. The initial response to antigens and how bacteria are handled is incompletely understood. By iterative application of spatial transcriptomics and multiplexed single-cell technologies, we identify that the double negative 2 subset of B cells, previously associated with autoimmune diseases, is present in the subepithelial dome in health. We show that in this location double negative 2 B cells interact with dendritic cells co-expressing the lupus autoantigens DNASE1L3 and C1q and microbicides. We observe that in humans, but not in mice, dendritic cells expressing DNASE1L3 are associated with sampled bacteria but not DNA derived from apoptotic cells. We propose that fundamental features of autoimmune diseases are microbiota-associated, interacting components of normal intestinal immunity.

The STING inhibitor (ISD-017) reduces glomerulonephritis in 129.B6.Fcgr2b-deficient mice.

The absence of stimulator of interferon genes (STING) in 129.B6.Fcgr2b-deficient mice rescue lupus phenotypes. The administration of a STING inhibitor (ISD017) into the young 129.B6.Fcgr2b-deficient mice prevents lupus nephritis development. This study mainly aimed to evaluate the effects of STING inhibition (ISD107) on established SLE in mice to prove that ISD017 could be a good therapeutic drug to reverse the already set-up autoimmunity and kidney impairment. Twenty-four-week-old Fcgr2b-deficient mice were treated with cyclophosphamide (25 mg/kg, intraperitoneal, once per week), ISD017 (10 mg/kg, intraperitoneal, three times per week), or control vehicle for 8 weeks, and were analyzed for phenotypes. Both ISD017 and cyclophosphamide treatment increased long-term survival and reduced the severity of glomerulonephritis in Fcgr2b-deficient mice. While cyclophosphamide reduced activated B cells (B220+GL-7+), ISD017 decreased activated T cells (CD4+CD69+) and neutrophils (Ly6c+Ly6g+) in Fcgr2b-deficient mice. In addition, ISD017 reduced IL-1ß and interferon-inducible genes. In summary, ISD017 treatment in symptomatic 129.B6.Fcgr2b-deficient mice reduced the severity of glomerulonephritis and increased long-term survival. ISD017 worked comparably to cyclophosphamide for treating lupus nephritis in 129.B6.Fcgr2b-deficient mice. ISD017 reduced activated T cells and neutrophils, while cyclophosphamide targeted activated B cells. These results suggested that STING inhibitors can potentially be a new therapeutic drug for treating lupus.

Simultaneous deletion of ORMDL1 and ORMDL3 proteins disrupts immune cell homeostasis.

ORMDL3 is a prominent member of a family of highly conserved endoplasmic reticulum resident proteins, ORMs (ORM1 and ORM2) in yeast, dORMDL in Drosophila and ORMDLs (ORMDL1, ORMDL2, and ORMDL3) in mammals. ORMDL3 mediates feedback inhibition of de novo sphingolipid synthesis. Expression levels of ORMDL3 are associated with the development of inflammatory and autoimmune diseases including asthma, systemic lupus erythematosus, type 1 diabetes mellitus and others. It has been shown that simultaneous deletions of other ORMDL family members could potentiate ORMDL3-induced phenotypes. To understand the complex function of ORMDL proteins in immunity in vivo, we analyzed mice with single or double deletions of Ormdl genes. In contrast to other single and double knockouts, simultaneous deletion of ORMDL1 and ORMDL3 proteins disrupted blood homeostasis and reduced immune cell content in peripheral blood and spleens of mice. The reduced number of splenocytes was not caused by aberrant immune cell homing. A competitive bone marrow transplantation assay showed that the development of Ormdl1-/-/Ormdl3-/- B cells was dependent on lymphocyte intrinsic factors. Highly increased sphingolipid production was observed in the spleens and bone marrow of Ormdl1-/-/Ormdl3-/- mice. Slight, yet significant, increase in some sphingolipid species was also observed in the spleens of Ormdl3-/- mice and in the bone marrow of both, Ormdl1-/- and Ormdl3-/- single knockout mice. Taken together, our results demonstrate that the physiological expression of ORMDL proteins is critical for the proper development and circulation of lymphocytes. We also show cell-type specific roles of individual ORMDL family members in the production of different sphingolipid species.

Unravelling B cell heterogeneity: insights into flow cytometry-gated B cells from single-cell multi-omics data.

Introduction: B cells play a pivotal role in adaptive immunity which has been extensively characterised primarily via flow cytometry-based gating strategies. This study addresses the discrepancies between flow cytometry-defined B cell subsets and their high-confidence molecular signatures using single-cell multi-omics approaches. Methods: By analysing multi-omics single-cell data from healthy individuals and patients across diseases, we characterised the level and nature of cellular contamination within standard flow cytometric-based gating, resolved some of the ambiguities in the literature surrounding unconventional B cell subsets, and demonstrated the variable effects of flow cytometric-based gating cellular heterogeneity across diseases. Results: We showed that flow cytometric-defined B cell populations are heterogenous, and the composition varies significantly between disease states thus affecting the implications of functional studies performed on these populations. Importantly, this paper draws caution on findings about B cell selection and function of flow cytometric-sorted populations, and their roles in disease. As a solution, we developed a simple tool to identify additional markers that can be used to increase the purity of flow-cytometric gated immune cell populations based on multi-omics data (AlliGateR). Here, we demonstrate that additional non-linear CD20, CD21 and CD24 gating can increase the purity of both naïve and memory populations. Discussion: These findings underscore the need to reconsider B cell subset definitions within the literature and propose leveraging single-cell multi-omics data for refined characterisation. We show that single-cell multi-omics technologies represent a powerful tool to bridge the gap between surface marker-based annotations and the intricate molecular characteristics of B cell subsets.

[Research progress on ferroptosis in tumor immunity].

Ferroptosis is a novel form of cell death that is induced by excessive accumulation of ferrous ions and lipid peroxides. It triggers the release of damage-associated molecular patterns through autophagy-dependent mechanisms, serving as an adjunct to immunogenic cell death and activating both adaptive and innate immunity. In the tumor microenvironment, the regulation and influence of tumor cells and immune cells undergoing ferroptosis are regulated by various factors, which plays a crucial role in tumor development, treatment, and prognosis. This article provides an overview of the biological effects of ferroptosis on immune cells such as T cells, macrophages, neutrophils and B cells and tumor cells in the tumor microenvironment.

[Research progress on the differences between T and B cell responses to three different forms of human papilloma virus (HPV) vaccination].

One of the most prevalent malignancies in women is cervical cancer. Cervical cancer is mostly brought on by chronic high-risk human papillomavirus 16 (HPV16) and HPV18 infection. Currently, the widely used HPV vaccines are the bivalent Cervarix, the tetravalent Gardasil, and the 9-valent Gardasil-9.There are differences in T cell effector molecule changes, B cell antibody level, duration, age and the injection after vaccination of the three vaccines.

Exploring the depths of IgG4: insights into autoimmunity and novel treatments.

IgG4 subclass antibodies represent the rarest subclass of IgG antibodies, comprising only 3-5% of antibodies circulating in the bloodstream. These antibodies possess unique structural features, notably their ability to undergo a process known as fragment-antigen binding (Fab)-arm exchange, wherein they exchange half-molecules with other IgG4 antibodies. Functionally, IgG4 antibodies primarily block and exert immunomodulatory effects, particularly in the context of IgE isotype-mediated hypersensitivity reactions. In the context of disease, IgG4 antibodies are prominently observed in various autoimmune diseases combined under the term IgG4 autoimmune diseases (IgG4-AID). These diseases include myasthenia gravis (MG) with autoantibodies against muscle-specific tyrosine kinase (MuSK), nodo-paranodopathies with autoantibodies against paranodal and nodal proteins, pemphigus vulgaris and foliaceus with antibodies against desmoglein and encephalitis with antibodies against LGI1/CASPR2. Additionally, IgG4 antibodies are a prominent feature in the rare entity of IgG4 related disease (IgG4-RD). Intriguingly, both IgG4-AID and IgG4-RD demonstrate a remarkable responsiveness to anti-CD20-mediated B cell depletion therapy (BCDT), suggesting shared underlying immunopathologies. This review aims to provide a comprehensive exploration of B cells, antibody subclasses, and their general properties before examining the distinctive characteristics of IgG4 subclass antibodies in the context of health, IgG4-AID and IgG4-RD. Furthermore, we will examine potential therapeutic strategies for these conditions, with a special focus on leveraging insights gained from anti-CD20-mediated BCDT. Through this analysis, we aim to enhance our understanding of the pathogenesis of IgG4-mediated diseases and identify promising possibilities for targeted therapeutic intervention.

A new perspective on therapies involving B-cell depletion in autoimmune diseases.

It has been rediscovered in the last fifteen years that B-cells play an active role in autoimmune etiology rather than just being spectators. The clinical success of B-cell depletion therapies (BCDTs) has contributed to this. BCDTs, including those that target CD20, CD19, and BAFF, were first developed to eradicate malignant B-cells. These days, they treat autoimmune conditions like multiple sclerosis and systemic lupus erythematosus. Particular surprises have resulted from the use of BCDTs in autoimmune diseases. For example, even in cases where BCDT is used to treat the condition, its effects on antibody-secreting plasma cells and antibody levels are restricted, even though these cells are regarded to play a detrimental pathogenic role in autoimmune diseases. In this Review, we provide an update on our knowledge of the biology of B-cells, examine the outcomes of clinical studies employing BCDT for autoimmune reasons, talk about potential explanations for the drug's mode of action, and make predictions about future approaches to targeting B-cells other than depletion.

Everlasting first impressions in B cells.

Initial imprinting by type 1 interferons shapes memory B cell generation in chronic viral infection.

Oxidative phosphorylation regulates B cell effector cytokines and promotes inflammation in multiple sclerosis.

Dysregulated B cell cytokine production contributes to pathogenesis of immune-mediated diseases including multiple sclerosis (MS); however, the underlying mechanisms are poorly understood. In this study we investigated how cytokine secretion by pro-inflammatory (GM-CSF-expressing) and anti-inflammatory (IL-10-expressing) B cells is regulated. Pro-inflammatory human B cells required increased oxidative phosphorylation (OXPHOS) compared with anti-inflammatory B cells. OXPHOS reciprocally modulated pro- and anti-inflammatory B cell cytokines through regulation of adenosine triphosphate (ATP) signaling. Partial inhibition of OXPHOS or ATP-signaling including with BTK inhibition resulted in an anti-inflammatory B cell cytokine shift, reversed the B cell cytokine imbalance in patients with MS, and ameliorated neuroinflammation in a myelin oligodendrocyte glycoprotein (MOG)-induced experimental autoimmune encephalitis mouse model. Our study identifies how pro- and anti-inflammatory cytokines are metabolically regulated in B cells and identifies ATP and its metabolites as a "fourth signal" that shapes B cell responses and is a potential target for restoring the B cell cytokine balance in autoimmune diseases.

Altered X-chromosome inactivation predisposes to autoimmunity.

In mammals, males and females show marked differences in immune responses. Males are globally more sensitive to infectious diseases, while females are more susceptible to systemic autoimmunity. X-chromosome inactivation (XCI), the epigenetic mechanism ensuring the silencing of one X in females, may participate in these sex biases. We perturbed the expression of the trigger of XCI, the noncoding RNA Xist, in female mice. This resulted in reactivation of genes on the inactive X, including members of the Toll-like receptor 7 (TLR7) signaling pathway, in monocyte/macrophages and dendritic and B cells. Consequently, female mice spontaneously developed inflammatory signs typical of lupus, including anti-nucleic acid autoantibodies, increased frequencies of age-associated and germinal center B cells, and expansion of monocyte/macrophages and dendritic cells. Mechanistically, TLR7 signaling is dysregulated in macrophages, leading to sustained expression of target genes upon stimulation. These findings provide a direct link between maintenance of XCI and female-biased autoimmune manifestations and highlight altered XCI as a cause of autoimmunity.

The ion channel TRPV5 regulates B-cell signaling and activation.

Introduction: B-cell activation triggers the release of endoplasmic reticulum calcium stores through the store-operated calcium entry (SOCE) pathway resulting in calcium influx by calcium release-activated calcium (CRAC) channels on the plasma membrane. B-cell-specific murine knockouts of SOCE do not impact humoral immunity suggesting that alternative channels may be important. Methods: We identified a member of the calcium-permeable transient receptor potential (TRP) ion channel family, TRPV5, as a candidate channel expressed in B cells by a quantitative polymerase chain reaction (qPCR) screen. To further investigate the role of TRPV5 in B-cell responses, we generated a murine TRPV5 knockout (KO) by CRISPR-Cas9. Results: We found TRPV5 polarized to B-cell receptor (BCR) clusters upon stimulation in a PI3K-RhoA-dependent manner. TRPV5 KO mice have normal B-cell development and mature B-cell numbers. Surprisingly, calcium influx upon BCR stimulation in primary TRPV5 KO B cells was not impaired; however, differential expression of other calcium-regulating proteins, such as ORAI1, may contribute to a compensatory mechanism for calcium signaling in these cells. We demonstrate that TRPV5 KO B cells have impaired spreading and contraction in response to membrane-bound antigen. Consistent with this, TRPV5 KO B cells have reduced BCR signaling measured through phospho-tyrosine residues. Lastly, we also found that TRPV5 is important for early T-dependent antigen specific responses post-immunization. Discussion: Thus, our findings identify a role for TRPV5 in BCR signaling and B-cell activation.

B-cell immune dysregulation with low soluble CD22 levels in refractory seronegative myasthenia gravis.

Myasthenia gravis (MG), primarily caused by acetylcholine receptor (AChR) autoantibodies, is a chronic autoimmune disorder causing severe muscle weakness and fatigability. In particular, seronegative MG constitutes 10%-15% of MG cases and presents diagnostic challenges especially in early-onset female patients who often show severe disease and resistance to immunosuppressive therapy. Furthermore, the immunopathology of seronegative MG remains unclear. Thus, in this study, we aimed to elucidate the pathogenic mechanism of seronegative MG using scRNA-seq analysis and plasma proteome analysis; in particular, we investigated the relationship between immune dysregulation status and disease severity in refractory seronegative MG. Employing single-cell RNA-sequencing and plasma proteome analyses, we analyzed peripheral blood samples from 30 women divided into three groups: 10 healthy controls, 10 early-onset AChR-positive MG, and 10 refractory early-onset seronegative MG patients, both before and after intravenous immunoglobulin treatment. The disease severity was evaluated using the MG-Activities of Daily Living (ADL), MG composite (MGC), and revised 15-item MG-Quality of Life (QOL) scales. We observed numerical abnormalities in multiple immune cells, particularly B cells, in patients with refractory seronegative MG, correlating with disease activity. Notably, severe MG cases had fewer regulatory T cells without functional abnormalities. Memory B cells were found to be enriched in peripheral blood cells compared with naïve B cells. Moreover, plasma proteome analysis indicated significantly lower plasma protein levels of soluble CD22, expressed in the lineage of B-cell maturation (including mature B cells and memory B cells), in refractory seronegative MG patients than in healthy donors or patients with AChR-positive MG. Soluble CD22 levels were correlated with disease severity, B-cell frequency, and RNA expression levels of CD22. In summary, this study elucidates the immunopathology of refractory seronegative MG, highlighting immune disorders centered on B cells and diminished soluble CD22 levels. These insights pave the way for novel MG treatment strategies focused on B-cell biology.