B-cell Activation Research Articles

Raman spectroscopy of lymphocytes from patients with the Epstein–Barr virus infection

In this study, Raman spectroscopy is applied to trace lymphocytes activation following contact with the Epstein–Barr virus (EBV) of the herpesvirus family. The biomarker of cell activation is found to be the 520 cm−1 band, indicating formation of immunoglobulins. The blood samples are obtained from patients diagnosed with infectious mononucleosis and treated at the University Hospital in Kraków. The lymphocytes’ Raman spectra are collected using a mapping technique, exciting samples with a 514.5 nm line of Ar + laser. Measurements are performed on the 1st, 4th, 6th, 12th and 30th day of hospitalization, until the patient has recovered. The highest intensity of the immunoglobulin marker is observed on the 4th day of hospitalization, while the results of the blood count of patients show the greatest increase in the number of lymphocytes at the beginning of hospitalization. No activated lymphocytes were observed in the blood of healthy volunteers. Some information is provided by the evaluation of B-cell activation by estimating the activated areas in the cells, which are determined by the presence of the Ig marker. The 900 cm−1 band and band around 1450 cm−1 are also analyzed as markers of the presence of the latent membrane protein, LMP2A (and 2B), of the EBV viral protein. The anomalous degree of depolarization observed in B-cells in the course of EBV infection appears to be due to the influence of a virus protein, disrupting BCR signal transduction.

B-Cell Activation Biomarkers in Salivary Glands Are Related to Lymphomagenesis in Primary Sjögren's Disease: A Pilot Monocentric Exploratory Study.

Primary Sjögren's disease is primarily driven by B-cell activation and is associated with a high risk of developing non-Hodgkin's lymphoma (NHL). Over the last few decades, microRNA-155 (miR-155) has arisen as a key regulator of B-cells. Nevertheless, its role in primary Sjögren's disease remains elusive. Thus, the purpose of this study was (i) to explore miR-155, B-cell activating factor (BAFF)-receptor (BAFF-R), and Interleukin 6 receptor (IL-6R) expression in the labial salivary glands (LSG) of patients with primary Sjögren's disease, aiming to identify potential B-cell activation biomarkers related to NHL development. Twenty-four patients with primary Sjögren's disease, and with available tissue blocks from a LSG biopsy performed at diagnosis, were enrolled. Among them, five patients developed B-cell NHL during follow-up (7.3 ± 3.1 years). A comparison group of 20 individuals with sicca disease was included. Clinical and laboratory parameters were recorded and the LSG biopsies were evaluated to assess local inflammation in terms of miR-155/BAFF-R and IL-6R expression. Stratifying the primary Sjögren's disease cohort according to lymphomagenesis, miR-155 was upregulated in primary Sjögren's disease patients who experienced NHL, more so than those who did not experience NHL. Moreover, miR-155 expression correlated with the focus score (FS), as well as BAFF-R and IL-6R expression, which were increased in primary Sjögren's disease patients and in turn related to neoplastic evolution. In conclusion, epigenetic modulation may play a crucial role in the aberrant activation of B-cells in primary Sjögren's disease, profoundly impacting the risk of NHL development.

The therapeutic potential of RANTES/CCL5 across diverse infections and its synergistic enhancement by ezrin peptide RepG3 for long COVID

The amplification of anti-infective immunity against a wide spectrum of acute and chronic infections caused by various pathogens is mediated by RANTES/CCL5. This chemokine controls infections caused by viruses, bacteria, fungi, and protozoans. In addition, RANTES/CCL5 exhibits anti-cancer effects by increasing NK-cell activity and targeting tumors. RANTES/CCL5 acts by amplifying antigen-specific immunity on mucosal surfaces, programmed T-cell responses, cytotoxic T lymphocytes (CTL), B-cell activation, and antibody production. RANTES/CCL5 exerts its effects by binding to C-C receptors, thereby triggering JAK/STAT signaling and inducing the migration of lymphocytes, NK cells, and monocytes. In the brain, RANTES/CCL5 activates astrocytes and upregulates anti-inflammatory interleukin (IL)-10 expression. Inflammatory cytokines rapidly induce RANTES/CCL5 expression in fibroblasts, epithelial cells, and monocytes/macrophages. In T-cells, RANTES/CCL5 expression is mediated by translational control of the transcription factor RFLAT-1/KLF13, which is responsible for a 3-day delay in RANTES/CCL5 secretion after T-cell activation. A cell membrane multi-protein complex containing CFTR, EBP50, ezrin, and PKC is a dominant regulator of both RANTES/CCL5 and inflammatory cytokine expression. Treatment of a volunteer patient suffering from long COVID/vaccine injury with the ezrin peptide RepG3 alleviated symptoms, substantially reduced serum proinflammatory cytokines to normal levels, and enhanced the expression of RANTES/CCL5. The immune amplification activities of RANTES/CCL5 and the ezrin peptide RepG3 exhibit striking similarities. In contrast, the ezrin peptide RepG3 differs from RANTES/CCL5 in its ability to significantly inhibit the expression of proinflammatory cytokines IL-1β, IL-6, IL-8, IL-13, TNF-α, and proinflammatory chemokines MIP-1α and MIP-1β. The mechanism through which the ezrin peptide RepG3 enhances adaptive immunity likely involves its induction of systemic elevation of RANTES/CCL5 expression and the simultaneous inhibition of proinflammatory cytokine expression.

A Brazilian Rare-Disease Center's Experience with Glucosylsphingosine (lyso-Gb1) in Patients with Gaucher Disease: Exploring a Novel Correlation with IgG Levels in Plasma and a Biomarker Measurement in CSF.

Gaucher disease (GD, OMIM 230800) is one of the most common lysosomal disorders, being caused by the deficient activity of the enzyme acid β-glucocerebrosidase (Gcase). Three clinical forms of Gaucher's disease (GD) are classified based on neurological involvement. Type 1 (GD1) is non-neuronopathic, while types 2 (GD2) and 3 (GD3) are neuronopathic forms. Gcase catalyzes the conversion of glucosylceramide (GlcCer) into ceramide and glucose. As GlcCer accumulates in lysosomal macrophages, it undergoes deacylation to become glycosylsphingosine (lyso-Gb1), which has shown to be a useful and reliable biomarker for the diagnosis and monitoring of treated and untreated patients with GD. Multiple myeloma (MM) is one of the leading causes of cancer-related death among patients with GD and monoclonal gammopathy of undetermined significance (MGUS) is a non-neoplastic condition that can be a telltale sign of a B clonal proliferation caused by the chronic activation of B cells. This study aimed to quantify Lyso-Gb1 levels in dried blood spots (DBS) and cerebrospinal fluid (CSF) as biomarkers for Gaucher disease (GD) and discuss the association of this biomarker with other clinical parameters. This is a mixed-methods study incorporating both cross-sectional and longitudinal elements within a cohort design with a convenience-sampling strategy. Data collection took place from January 2012 to March 2023. Lyso-Gb1 extraction from DBS involved the use of a methanol-acetonitrile-water mixture, followed by incubation and centrifugation. Analysis was performed using UPLC-MS/MS with MassLynx software version 4.2 and the control group for the DBS measurements included general newborns. CSF Lyso-Gb1 was extracted using ethyl acetate, analyzed by UPLC-MS/MS with a calibration curve, and expressed in pmol/L. Lysosomal activity in CSF was assessed by measuring chitotriosidase (Cht), and other lysosomal enzyme activities were assessed as previously described in the literature. Patients with metachromatic leukodystrophy (MLD) were used as controls. Thirty-two treated patients (twenty-nine GD1 and three GD3, all on ERT except for one GD type on SRT with eliglustat) and three untreated patients (one GD1, one GD2, and one GD3) were included. When analyzing only the treated GD1 group, a significant correlation was found between lyso-Gb1 and age (rho = -0.447, p = 0.001), ChT, and IgG levels (rho = 0.73, p < 0.001; and rho = 0.36, p = 0.03, respectively). Five GD1 patients (three females, mean age 40 years) also had their CSF collected and analyzed. The average measurement of lyso-Gb1 in CSF was 94 pmol/L (range: 57.1-157.9 pmol/L) versus <6.2 pmol/L in the control group (MLD). This is the first time, to the best of our knowledge, that lyso-Gb1 has been associated with IgG levels. While this finding reflects a risk for MGUS or MM and not only chronic plasma B-cell activation, it still requires further studies. Moreover, the analysis of CSF lyso-Gb1 levels in GD1 patients was demonstrated to be significantly higher than the control group. This raises the hypothesis that CSF lyso-Gb1 may serve as a valuable indicator for neurological involvement in GD, providing insights into the potential implications for neurological manifestations in GD, including GD1. The correlation between lyso-Gb1 and ChT levels in treated GD1 patients further underscores the interconnectedness of lysosomal markers and their relevance in monitoring.

Implications of IFNγ SNP rs2069705 in primary Sjögren's syndrome: transcriptional activation and B cell infiltration.

Primary Sjögren's syndrome (pSS) is characterized by its autoimmune nature. This study investigates the role of the IFNγ SNP rs2069705 in modulating the susceptibility to pSS. Differential expression of IFNγ and BAFF was analyzed using the GEO database's mRNA microarray GSE84844. Genotyping of the IFNγ SNP rs2069705 was conducted via the dbSNP website. The JASPAR tool was used for predicting transcription factor bindings. Techniques such as dual-luciferase reporter assays, Chromatin immunoprecipitation, and analysis of a pSS mouse model were applied to study gene and protein interactions. A notable increase in the mutation frequency of IFNγ SNP rs2069705 was observed in MNCs from the exocrine glands of pSS mouse models. Bioinformatics analysis revealed elevated levels of IFNγ and BAFF in pSS samples. The model exhibited an increase in both CD20+ B cells and cells expressing IFNγ and BAFF. Knocking down IFNγ resulted in lowered BAFF expression and less lymphocyte infiltration, with BAFF overexpression reversing this suppression. Activation of the Janus kinase (JAK)/STAT1 pathway was found to enhance transcription in the BAFF promoter region, highlighting IFNγ's involvement in pSS. In addition, rs2069705 was shown to boost IFNγ transcription by promoting interaction between its promoter and STAT4. SNP rs2069705 in the IFNγ gene emerges as a pivotal element in pSS susceptibility, primarily by augmenting IFNγ transcription, activating the JAK/STAT1 pathway, and leading to B-lymphocyte infiltration in the exocrine glands.NEW & NOTEWORTHY The research employed a combination of bioinformatics analysis, genotyping, and experimental models, providing a multifaceted approach to understanding the complex interactions in pSS. We have uncovered that the rs2069705 SNP significantly affects the transcription of IFNγ, leading to altered immune responses and B-lymphocyte activity in pSS.

Successful use of tacrolimus for treatment-refractory neuroblastoma-associated opsoclonus-myoclonus-ataxia syndrome: A case series.

Opsoclonus-myoclonus-ataxia syndrome (OMAS) is an autoimmune central nervous system disorder, primarily manifesting as a paraneoplastic sequalae to neuroblastoma, and characterized by motor disorders and behavioral disturbances. OMAS is typified by aberrant B-cell and T-cell activation. Current treatment involves immunosuppression using corticosteroids, intravenous immunoglobulin, and rituximab. However, these approaches often lead to treatment-related toxicities and symptomatic recurrences with chronic neurocognitive impairment. We treated three children with refractory neuroblastoma-associated OMAS with tacrolimus, a T-cell-targeting calcineurin inhibitor, effectively controlling symptoms within a month and enabling the discontinuation of immunosuppression with minimal side effects. Tacrolimus shows promise as a therapeutic option for refractory OMAS.

Suppression of B-Cell Activation by Human Cord Blood-Derived Stem Cells (CB-SCs) through the Galectin-9-Dependent Mechanism.

We developed the Stem Cell Educator therapy among multiple clinical trials based on the immune modulations of multipotent cord blood-derived stem cells (CB-SCs) on different compartments of immune cells, such as T cells and monocytes/macrophages, in type 1 diabetes and other autoimmune diseases. However, the effects of CB-SCs on the B cells remained unclear. To better understand the molecular mechanisms underlying the immune education of CB-SCs, we explored the modulations of CB-SCs on human B cells. CB-SCs were isolated from human cord blood units and confirmed by flow cytometry with different markers for their purity. B cells were purified by using anti-CD19 immunomagnetic beads from human peripheral blood mononuclear cells (PBMCs). Next, the activated B cells were treated in the presence or absence of coculture with CB-SCs for 7 days before undergoing flow cytometry analysis of phenotypic changes with different markers. Reverse transcription-polymerase chain reaction (RT-PCR) was utilized to evaluate the levels of galectin expressions on CB-SCs with or without treatment of activated B cells in order to find the key galectin that was contributing to the B-cell modulation. Flow cytometry demonstrated that the proliferation of activated B cells was markedly suppressed in the presence of CB-SCs, leading to the downregulation of immunoglobulin production from the activated B cells. Phenotypic analysis revealed that treatment with CB-SCs increased the percentage of IgD+CD27- naïve B cells, but decreased the percentage of IgD-CD27+ switched B cells. The transwell assay showed that the immune suppression of CB-SCs on B cells was dependent on the galectin-9 molecule, as confirmed by the blocking experiment with the anti-galectin-9 monoclonal antibody. Mechanistic studies demonstrated that both calcium levels of cytoplasm and mitochondria were downregulated after the treatment with CB-SCs, causing the decline in mitochondrial membrane potential in the activated B cells. Western blot exhibited that the levels of phosphorylated Akt and Erk1/2 signaling proteins in the activated B cells were also markedly reduced in the presence of CB-SCs. CB-SCs displayed multiple immune modulations on B cells through the galectin-9-mediated mechanism and calcium flux/Akt/Erk1/2 signaling pathways. The data advance our current understanding of the molecular mechanisms underlying the Stem Cell Educator therapy to treat autoimmune diseases in clinics.

The role of BAFF and APRIL in IgA nephropathy: pathogenic mechanisms and targeted therapies.

Immunoglobulin A nephropathy (IgAN), characterized by mesangial deposition of galactose-deficient-IgA1 (Gd-IgA1), is the most common biopsy-proven primary glomerulonephritis worldwide. Recently, an improved understanding of its underlying pathogenesis and the substantial risk of progression to kidney failure has emerged. The "four-hit hypothesis" of IgAN pathogenesis outlines a process that begins with elevated circulating levels of Gd-IgA1 that trigger autoantibody production. This results in the formation and deposition of immune complexes in the mesangium, leading to inflammation and kidney injury. Key mediators of the production of Gd-IgA1 and its corresponding autoantibodies are B-cell activating factor (BAFF), and A proliferation-inducing ligand (APRIL), each playing essential roles in the survival and maintenance of B cells and humoral immunity. Elevated serum levels of both BAFF and APRIL are observed in patients with IgAN and correlate with disease severity. This review explores the complex pathogenesis of IgAN, highlighting the pivotal roles of BAFF and APRIL in the interplay between mucosal hyper-responsiveness, B-cell activation, and the consequent overproduction of Gd-IgA1 and its autoantibodies that are key features in this disease. Finally, the potential therapeutic benefits of inhibiting BAFF and APRIL in IgAN, and a summary of recent clinical trial data, will be discussed.

Construction of Peptide Vaccine Candidate Based on β-Cell Epitopes of Indonesian Monkeypox Virus (MPXV) Virulence Protein:A Reverse Vaccinology

Infection with a DNA virus called monkeypox virus (MPXV) in humans has been identified in the Congo since 1970. Antiviral drugs are not effective for preventing MPXV infection. MPXV infection cases in Indonesia are very low but MPXV has the potential to become a global pandemic so it is very important to do prevention such as vaccine development. This study aims to construct a B cell epitope-based peptide vaccine candidate in Indonesian MPXV through an in silico approach.The development of the MPXV vaccine can be performed through a computational approach for preliminary studies. In silico-based construction of vaccines using B cell epitopes, antigenicity, allergenicity, docking, and molecular dynamics analysis have been used by researchers and scientists in solving viral infection cases. We recommend Pep A and Pep D as vaccine candidates because they allow recognition by B cells, antigenic peptides, non-allergenic and non-toxin. Peptide vaccine candidate can trigger B-cell activation to produce IgM isotype-specific antibodies through BCR interaction. In summary, the results of this study can be used for an initial study of MPXV vaccine development in Indonesia.

Abstract A028: Multiomic modelling of pancreatic IPMN stroma reveals distinct tertiary lymphoid structure distribution: Mapping the transcriptomic landscape via regional bulk, single-cell and subcellular approaches

Abstract Background Intraductal Papillary Mucinous Neoplasms (IPMN) remain the largest subtype of cystic pancreatic cancer precursors. Cancer risk has been observed to vary between histological grades and histological subtypes. Spatial characterisation has revealed variation in immune cell distribution disease subtypes. Methods A cohort of 14 surgically resected IPMN tumors across a range of histological grades (Low-grade LG, high-grade HG and invasive IPMN cancers IPMN-PDAC) and subtypes (gastric, intestinal, pancreaticobiliary) underwent multimodal spatial interrogation. After expert histopathological annotation; tissue sections underwent regional whole transcriptome analysis with TempO-seq and 10x Visium. Extensive region analysis with NanoString GeoMx employing segmentation using staining for epithelium (PanCK) fibroblast stroma (aSMA) was performed. A tissue microarray was then constructed, and representative cores underwent single-cell and subcellular spatial transcriptomic analysis with NanoString CosMx. Raw count data and digital images were exported and analysed using Seurat, Giotto, SPATA2 and custom R pipelines. Region deconvolution, Harmony integration, Leiden Clustering, Gene Set Enrichment Analysis, Trajectory Mapping, and Moran’s I Analysis were performed. Results TempO-seq revealed up-regulation of canonical oncogene expression in all IPMN-PDAC cases as compared to LG and HG IPMN. There were no statistically significant differences in expression comparing LG and HG IPMN lesions. Visium differential gene expression and clustering models identified cancer epithelium, stroma and lymphatic components that correlated with histopathological annotations. Immune cell rich spots were identified via gene ontology of top 50 marker genes. The spatial trajectory of B-cell expression signatures in IPMN-PDAC showed greater concentration and less variability than HG tumors, Moran’s I -0.34 vs 0.12 (P = 0.021). B-cells in HG IPMN were showed greater concentration and less variability than LG IPMN, Moran’s I -0.15 vs 0.02 (P=0.05). Much of this concentration was in identified lymphoid aggregates or tertiary lymphoid structures. The spatial trajectory of T-cell expression. The spatial trajectory of macrophage expression showed greater concentration and less variability than in LG and HG tumors. GeoMx analysis of stromal regions identified up regulation of B cell signatures in IPMN PDAC and HG IPMN when compared to LG IPMN. Markers associated with TLS formation were found to be significantly upregulated including CXCL13, CXCR5 and LAMP3. Pancreaticobiliary subtypes were found to have the least concentrated B-cell distribution across subtypes. Conclusions Immune cell composition was found to vary across histological grades and subtypes of IPMN. A paradoxical relationship was observed between T-cell infiltrates and B-cell and macrophage populations. The greater proportion of tertiary lymphoid structures, and high levels of TLS specific gene expression, may indicate a propensity for b-cell activation in IPMN tumors which progress to malignancy. Citation Format: Andrew J. Cameron, Assya Legrini, Colin S. Wood, Craig Nourse, Yoana Doncheva, Claire Kennedy Dietrich, Colin Nixon, Jennifer Hay, Fraser Duthie, Pawel Herzyk, Jennifer Morton, Nigel B. Jamieson. Multiomic modelling of pancreatic IPMN stroma reveals distinct tertiary lymphoid structure distribution: Mapping the transcriptomic landscape via regional bulk, single-cell and subcellular approaches [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Pancreatic Cancer; 2023 Sep 27-30; Boston, Massachusetts. Philadelphia (PA): AACR; Cancer Res 2024;84(2 Suppl):Abstract nr A028.

Specific Features of Juvenile Idiopathic Arthritis Patients' Cytokine Profile.

Juvenile idiopathic arthritis (JIA) is a systemic autoimmune disease that affects the joints, leading to disability. Cytokines and signaling molecules expressed by the immune system cells play a key role in JIA pathogenesis. Understanding how their content changes during pathology development can open up new opportunities for its diagnosis and treatment. The blood plasma of 30 patients with JIA (14 males and 16 females with a mean age of 12.2 ± 4.1) and 20 relatively healthy individuals (10 males and 10 females with a mean age of 10.20 ± 5.85) was analyzed to determine the levels of cytokines using the MILLIPLEX® kit. An increase in interleukins (IL)-1α, 1β, 2, 4, 5, 6, 7, 8, 9, 10, 13, 15, 17F, 22, and 27 and a decrease in IL-3 levels have been shown in patients with JIA. Levels of cytokines, which are important for B-cell activation and proliferation, are increased, while levels of T-cell activating factors remained similar to the control group. Based on our results, it can be assumed that the use of combination therapy aimed at inhibiting both nonspecific interleukins and cytokines that activate B-cells will be more effective for the treatment of JIA.

An overlapping case of IgG4-related disease and systemic lupus erythematosus treated with belimumab: a case-based review.

IgG4-related disease (IgG4-RD) is a systemic condition in which IgG4+ plasma cell infiltration and fibrosis cause organ swelling and lead to diverse clinical manifestations. Although IgG4-RD typically responds to glucocorticoids (GCs), relapse during tapering occurs and an early GC-sparing approach might therefore be beneficial. Systemic lupus erythematosus (SLE) is a chronic inflammatory disease with multiple symptoms that is also treated with GCs as a first-line therapy. Recently, belimumab, a recombinant human IgG-1λ monoclonal antibody that inhibits B-cell activating factor, was approved, but reports of use for IgG4-RD are scarce. Here, we present a rare case of IgG4-RD complicated with SLE which was successfully treated with belimumab. A 67-year-old man was diagnosed with IgG4-RD based on a high serum IgG4 level and histopathological findings. Furthermore, he had pericardial effusion on echocardiography, and laboratory tests revealed thrombocytopenia, autoimmune hemolysis, positive anti-nuclear antibodies, positive anti-DNA antibodies, and hypocomplementemia. These data led to an SLE diagnosis. Treatment was started with prednisolone at 40mg/day, plus hydroxychloroquine, which initially improved both the SLE and IgG4-RD symptoms. During the GC tapering, belimumab was added and clinical symptoms resolved completely. Our case and the literature review summarize reported rare overlapping cases of IgG4-RD and SLE and suggest that belimumab is a promising candidate for the treatment of IgG4-RD.

Impaired B-cell function in ERCC2 deficiency.

Trichothiodystrophy-1 (TTD1) is an autosomal-recessive disease and caused by mutations in ERCC2, a gene coding for a subunit of the TFIIH transcription and nucleotide-excision repair (NER) factor. In almost half of these patients infectious susceptibility has been reported but the underlying molecular mechanism leading to immunodeficiency is largely unknown. The aim of this study was to perform extended molecular and immunological phenotyping in patients suffering from TTD1. Cellular immune phenotype was investigated using multicolor flow cytometry. DNA repair efficiency was evaluated in UV-irradiation assays. Furthermore, early BCR activation events and proliferation of TTD1 lymphocytes following DNA damage induction was tested. In addition, we performed differential gene expression analysis in peripheral lymphocytes of TTD1 patients. We investigated three unrelated TTD1 patients who presented with recurrent infections early in life of whom two harbored novel ERCC2 mutations and the third patient is a carrier of previously described pathogenic ERCC2 mutations. Hypogammaglobulinemia and decreased antibody responses following vaccination were found. TTD1 B-cells showed accumulation of γ-H2AX levels, decreased proliferation activity and reduced cell viability following UV-irradiation. mRNA sequencing analysis revealed significantly downregulated genes needed for B-cell development and activation. Analysis of B-cell subpopulations showed low numbers of naïve and transitional B-cells in TTD1 patients, indicating abnormal B-cell differentiation in vivo. In summary, our analyses confirmed the pathogenicity of novel ERCC2 mutations and show that ERCC2 deficiency is associated with antibody deficiency most likely due to altered B-cell differentiation resulting from impaired BCR-mediated B-cell activation and activation-induced gene transcription.

Pre-vaccination transcriptomic profiles of immune responders to the MUC1 peptide vaccine for colon cancer prevention.

Self-antigens abnormally expressed on tumors, such as MUC1, have been targeted by therapeutic cancer vaccines. We recently assessed in two clinical trials in a preventative setting whether immunity induced with a MUC1 peptide vaccine could reduce high colon cancer risk in individuals with a history of premalignant colon adenomas. In both trials, there were immune responders and non-responders to the vaccine. Here we used PBMC pre-vaccination and 2 weeks after the first vaccine of responders and non-responders selected from both trials to identify early biomarkers of immune response involved in long-term memory generation and prevention of adenoma recurrence. We performed flow cytometry, phosflow, and differential gene expression analyses on PBMCs collected from MUC1 vaccine responders and non-responders pre-vaccination and two weeks after the first of three vaccine doses. MUC1 vaccine responders had higher frequencies of CD4 cells pre-vaccination, increased expression of CD40L on CD8 and CD4 T-cells, and a greater increase in ICOS expression on CD8 T-cells. Differential gene expression analysis revealed that iCOSL, PI3K AKT MTOR, and B-cell signaling pathways are activated early in response to the MUC1 vaccine. We identified six specific transcripts involved in elevated antigen presentation, B-cell activation, and NF-κB1 activation that were directly linked to finding antibody response at week 12. Finally, a model using these transcripts was able to predict non-responders with accuracy. These findings suggest that individuals who can be predicted to respond to the MUC1 vaccine, and potentially other vaccines, have greater readiness in all immune compartments to present and respond to antigens. Predictive biomarkers of MUC1 vaccine response may lead to more effective vaccines tailored to individuals with high risk for cancer but with varying immune fitness.

Mustard seed major allergen Sin a1 activates intestinal epithelial cells and also dendritic cells that drive type 2 immune responses.

Mustard seeds belong to the food category of mandatory labelling due to the severe reactions they can trigger in allergic patients. However, the mechanisms underlying allergic sensitization to mustard seeds are poorly understood. The aim of this work is to study type 2 immune activation induced by the mustard seed major allergen Sin a1 via the intestinal mucosa, employing an in vitro model mimicking allergen exposure via the intestinal epithelial cells (IECs). Sin a1 was isolated from the total protein extract and exposed to IEC, monocyte derived dendritic cells (DCs) or IEC/DC co-cultures. A system of consecutive co-cultures was employed to study the generic capacity of Sin a1 to induce type 2 activation leading to sensitization: IEC/DC, DC/T-cell, T/B-cell and stem cell derived mast cells (MCs) derived from healthy donors. Immune profiles were determined by ELISA and flow cytometry. Sin a1 activated IEC and induced type-2 cytokine secretion in IEC/DC co-culture or DC alone (IL-15, IL-25 and TSLP), and primed DC induced type 2 T-cell skewing. IgG secretion in the T-cell/B-cell phase was enhanced in the presence of Sin a1 in the first stages of the co-culture. Anti-IgE did not induce degranulation but promoted IL-13 and IL-4 release by MC primed with the supernatant from B-cells co-cultured with Sin a1-IEC/DC or -DC primed T-cells. Sin a1 enhanced the release of type-2 inflammatory mediators by epithelial and dendritic cells; the latter instructed generic type-2 responses in T-cells that resulted in B-cell activation, and finally MC activation upon anti-IgE exposure. This indicates that via activation of IEC and/or DC, mustard seed allergen Sin a1 is capable of driving type 2 immunity which may lead to allergic sensitization.

Characteristics and impact of infiltration of B-cells from systemic sclerosis patients in a 3D healthy skin model.

In systemic sclerosis (SSc), B-cells are activated and present in the skin and lung of patients where they can interact with fibroblasts. The precise impact and mechanisms of the interaction of B-cells and fibroblasts at the tissular level are poorly studied. We investigated the impact and mechanisms of B-cell/fibroblast interactions in cocultures between B-cells from patients with SSc and 3-dimensional reconstituted healthy skin model including fibroblasts, keratinocytes and extracellular matrix. The quantification and description of the B-cell infiltration in 3D cocultures were performed using cells imagery strategy and cytometry. The effect of coculture on the transcriptome of B-cells and fibroblasts was studied with bulk and single-cell RNA sequencing approaches. The mechanisms of this interaction were studied by blocking key cytokines like IL-6 and TNF. We showed a significant infiltration of B-cells in the 3D healthy skin model. The amount but not the depth of infiltration was higher with B-cells from SSc patients and with activated B-cells. B-cell infiltrates were mainly composed of naïve and memory cells, whose frequencies differed depending on B-cells origin and activation state: infiltrated B-cells from patients with SSc showed an activated profile and an overexpression of immunoglobulin genes compared to circulating B-cells before infiltration. Our study has shown for the first time that activated B-cells modified the transcriptomic profile of both healthy and SSc fibroblasts, toward a pro-inflammatory (TNF and IL-17 signaling) and interferon profile, with a key role of the TNF pathway. B-cells and 3D skin cocultures allowed the modelization of B-cells infiltration in tissues observed in SSc, uncovering an influence of the underlying disease and the activation state of B-cells. We showed a pro-inflammatory effect on skin fibroblasts and pro-activation effect on infiltrating B-cells during coculture. This reinforces the role of B-cells in SSc and provide potential targets for future therapeutic approach in this disease.

Advancements and prospects of novel biologicals for myasthenia gravis: toward personalized treatment based on autoantibody specificities.

Myasthenia gravis (MG) is an antibody-mediated autoimmune disease with a prevalence of 150-250 cases per million individuals. Autoantibodies include long-lived antibodies against the acetylcholine receptor (AChR), mainly of the IgG1 subclass, and IgG4, produced almost exclusively by short-lived plasmablasts, which are prevalent in muscle-specific tyrosine kinase (MuSK) myasthenia gravis. Numerous investigations have demonstrated that MG patients receiving conventional medication today still do not possess satisfactory symptom control, indicating a substantial disease burden. Subsequently, based on the type of the autoantibody and the pathogenesis, we synthesized the published material to date and reached a conclusion regarding the literature related to personalized targeted therapy for MG. Novel agents for AChR MG have shown their efficacy in clinical research, such as complement inhibitors, FcRn receptor antagonists, and B-cell activating factor (BAFF) inhibitors. Rituximab, a representative drug of anti-CD20 therapy, has demonstrated benefits in treatment of MuSK MG patients. Due to the existence of low-affinity antibodies or unidentified antibodies that are inaccessible by existing methods, the treatment for seronegative MG remains complicated; thus, special testing and therapy considerations are necessary. It may be advantageous to initiate the application of novel biologicals at an early stage of the disease. Currently, therapies can also be combined and individualized according to different types of antibodies. With such a wide range of drugs, how to tailor treatment strategies to patients with various conditions and find the most suitable solution for each MG profile are our necessary and urgent aims.

Heterologous booster vaccination enhances antibody responses to SARS-CoV-2 by improving Tfh function and increasing B-cell clonotype SHM frequency.

Heterologous prime-boost has broken the protective immune response bottleneck of the COVID-19 vaccines. however, the underlying mechanisms have not been fully elucidated. Here, we investigated antibody responses and explored the response of germinal center (GC) to priming with inactivated vaccines and boosting with heterologous adenoviral-vectored vaccines or homologous inactivated vaccines in mice. Antibody responses were dramatically enhanced by both boosting regimens. Heterologous immunization induced more robust GC activation, characterized by increased Tfh cell populations and enhanced helper function. Additionally, increased B-cell activation and antibody production were observed in a heterologous regimen. Libra-seq was used to compare the differences of S1-, S2- and NTD-specific B cells between homologous and heterologous vaccination, respectively. S2-specific CD19+ B cells presented increased somatic hypermutations (SHMs), which were mainly enriched in plasma cells. Moreover, a heterologous booster dose promoted the clonal expansion of B cells specific to S2 and NTD regions. In conclusion, the functional role of Tfh and B cells following SARS-CoV-2 heterologous vaccination may be important for modulating antibody responses. These findings provide new insights for the development of SARS-CoV-2 vaccines that induce more robust antibody response.

Safety and efficacy of telitacicept in refractory systemic lupus erythematosus patients who failed treatment with belimumab : Acase series.

This study aimed to determine the effect and safety of telitacicept, an antagonist of BLyS/APRIL-mediated Bcell activation, in patients with systemic lupus erythematosus (SLE) who failed treatment with belimumab and in whom telitacicept was administered combined with conventional therapy. Areview of published reports on telitacicept for SLE was also performed. Aretrospective review was performed of the records of patients seen in the Department of Rheumatology at the Wuhan Hospital of Chinese and Western Medicine, Wuhan, China, with refractory SLE who had failed treatment with belimumab. The terms "systemic lupus erythematosus" and "telitacicept" were used to identify patients reported in the English medical literature. Identified were 14refractory SLE patients, 3males (21%) and 11females(79%). The median age was 32.9years. The median disease duration was 8.9years. Patients in this cohort received telitacicept for an average of 34.1weeks (17-62weeks) and the total SLE responder index4 (SRI-4) response rate was 78.9% (n = 11). The mean SLE Disease Activity Index (SLEDAI) score declined from 8.6 at baseline (95% confidence interval [CI] 7.87-9.28) to 4.29 at the endpoint (95% CI 3.4-5.16). All cases (100%) had hypocomplementemia at baseline, and 7cases (50%) reported normal C3 and C4 levels at the follow-up endpoint. At the observation endpoint, the 24‑h urinary protein value of the 13cases with proteinuria (baseline 24‑h urinary protein > 0.5 g/d) displayed areduction, and 3values turned negative. Although some patients had low serum total immunoglobulin (Ig) levels, subnormal IgG levels, and absolute counts of peripheral blood lymphocytes after treatment, no serious infection was reported. One case was refractory lupus hepatitis confirmed by liver pathology, and upon change to change to telitacicept treatment, liver function returned to normal. This is the first case series in SLE patients who accepted telitacicept treatment after failed treatment with belimumab. Our case series and review of the literature show that telitacicept combined with the original standard treatment may significantly improve disease activity while reducing prednisone use. No major safety issues were seen in this group of patients. Telitacicept may be apromising drug for the treatment of refractory lupus hepatitis.

Interleukin-37 contributes to endometrial regenerative cell–mediated immunotherapeutic effect on chronic allograft vasculopathy

Background aimsChronic allograft vasculopathy (CAV) remains a predominant contributor to late allograft failure after organ transplantation. Several factors have already been shown to facilitate the progression of CAV, and there is still an urgent need for effective and specific therapeutic approaches to inhibit CAV. Human mesenchymal-like endometrial regenerative cells (ERCs) are free from the deficiencies of traditional invasive acquisition methods and possess many advantages. Nevertheless, the exact immunomodulation mechanism of ERCs remains to be elucidated. MethodsC57BL/6 (B6) mouse recipients receiving BALB/c mouse donor abdominal aorta transplantation were treated with ERCs, negative control (NC)-ERCs and interleukin (IL)-37−/−ERCs (ERCs with IL-37 ablation), respectively. Pathologic lesions and inflammatory cell infiltration in the grafts, splenic immune cell populations, circulating donor-specific antibody levels and cytokine profiles were analyzed on postoperative day (POD) 40. The proliferative capacities of Th1, Th17 and Treg subpopulations were assessed in vitro. ResultsAllografts from untreated recipients developed typical pathology features of CAV, namely endothelial thickening, on POD 40. Compared with untreated and IL-37−/−ERC-treated groups, IL-37–secreting ERCs (ERCs and NC-ERCs) significantly reduced vascular stenosis, the intimal hyperplasia and collagen deposition. IL-37–secreting ERCs significantly inhibited the proliferation of CD4+T cells, reduced the proportions of Th1 and Th17 cells, but increased the proportion of Tregs in vitro. Furthermore, in vitro results also showed that IL-37–secreting ERCs significantly inhibited Th1 and Th17 cell responses, abolished B-cell activation, diminished donor-specific antibody production and increased Treg proportions. Notably, IL-37–secreting ERCs remarkably downregulated the levels of pro-inflammatory cytokines (interferon-γ, tumor necrosis factor-α, IL-1β, IL-6 and IL-17A) and increased IL-10 levels in transplant recipients. ConclusionsThe knockdown of IL-37 dramatically abrogates the therapeutic ability of ERCs for CAV. Thus, this study highlights that IL-37 is indispensable for ERC-mediated immunomodulation for CAV and improves the long-term allograft acceptance.