Follicular Helper Research Articles

Identification of necroptosis genes and characterization of immune infiltration in non-alcoholic steatohepatitis

BackgroundThe most common progressive form of non-alcoholic fatty liver disease (NAFLD) is non-alcoholic steatohepatitis (NASH), which is characterized by the development of cirrhosis, and requires liver transplantation. We screened for the differentially expressed necroptosis-related genes in NASH in this study, and analyzed immune infiltration through microarray and bioinformatics analysis to identify potential biomarkers, and explore the molecular mechanisms involved in NASH.MethodsThe GSE24807 microarray dataset of NASH patients and healthy controls was downloaded, and we identified the differentially expressed genes (DEGs). Necroptosis-related differential genes (NRDEGs) were extracted from these DEGs, and functionally annotated by enrichment analyses. The core genes were obtained by constructing gene co-expression networks using weighted gene co-expression network analysis (WGCNA). Finally, the transcription factor (TF) regulatory network and the mRNA-miRNA network were constructed, and the infiltrating immune cell populations were analyzed with CIBERSORT.ResultsWe identified six necroptosis-related genes (CASP1, GLUL, PYCARD, IL33, SHARPIN, and IRF9), and they are potential diagnostic biomarkers for NASH. In particular, PYCARD is a potential biomarker for NAFLD progression. Analyses of immune infiltration showed that M2 macrophages, γδ T cells, and T follicular helper cells were associated with the immune microenvironment of NASH, which is possibly regulated by CASP1, IL33, and IRF9.ConclusionsWe identified six necroptosis-related genes in NASH, which are also potential diagnostic biomarkers. Our study provides new insights into the molecular mechanisms and immune microenvironment of NASH.

A novel inactivated oral rabies vaccine with the incorporation of U-OMP19 enhances the immunogenicity by reducing viral proteins degradation and activating dendritic cells in a mouse model

Currently, dogs, especially stray dogs, and/or wild animals are the main sources of rabies transmission, and oral vaccination is the most practical way to control rabies in these animals. Safety and efficacy are two key criteria for developing oral vaccines. Concerning the efficacy of oral vaccines, degradation of immunogens by gastrointestinal fluid is a major challenge, resulting in suboptimal immune responses after vaccination. For safety reasons, inactivated vaccines are the most optimal choice. In the present study, a recombinant rabies virus (RABV) with un-lipidated outer membrane protein 19 (U-OMP19) of Brucella spp incorporated into RABV virions, designated as LBNSE-OMP19-G, was constructed and rescued. We found that U-OMP19 was incorporated into LBNSE-OMP19-G virion, which could protect RABV G protein from digestion by gastrointestinal fluids in vitro. Moreover, the immunogenicity of LBNSE-OMP19-G as an inactivated oral vaccine was evaluated, and the inactivated LBNSE-OMP19-G could activate more dendritic cells (DCs) and promote the generation of follicular helper T (TFH) cells, germinal center (GC) B cells, and plasma cells in immunized mice compared with those in mice immunized with parent virus LNBSE, which consequently induced a higher level of virus neutralizing antibody and provided better protection after a lethal challenge of rabies. These data indicate that LBNSE-OMP19-G, which has good safety and immunogenicity, could be a potential inactivated oral rabies vaccine candidate.

CXCL13/CXCR5 axis facilitates TFH expansion and correlates with disease severity in adults with immune thrombocytopenia

BackgroundImmune thrombocytopenia (ITP) is an autoimmune bleeding disorder defined by a diminished platelet count. ITP pathogenesis involves intricate changes to both cellular and humoral immunity. The pivotal roles of follicular helper T (TFH) cells in the maturations of B cells and the production of antibodies are well-established. However, the specific role of TFH to the immunopathogenesis of ITP remain incompletely understood. This study aimed to clarify the association of CXCL13/CXCR5 axis with TFH in adults with ITP. MethodsA total of 97 ITP patients and 41 healthy controls were enrolled. CD4+CXCR5+ TFH, CD4+CXCR5+PD-1+ TFH, CD4+CXCR5+Foxp3+ follicular regulatory T cells (TFR), and desialylated platelets in peripheral blood were measured by flow cytometry. Plasma cytokines were assessed by enzyme-linked immunosorbent assay. CD4+ T cells cocultured with chemokine CXCL13 in vitro was performed for the measurement of TFH proliferation. Intracellular production of reactive oxygen species (ROS) was examined by dichlorodihydrofluorescein diacetate (DCFH-DA) probe staining. ResultsWe observed a significant increase in circulating TFH and a marked decrease in circulating TFR in the entire ITP cohort. The ratio of TFH/TFR was elevated, accompanied by heightened levels of platelet desialylation, cytokines BAFF, HMGB1, and IL-21, while levels of IL-10 were downregulated in adults with ITP. Notably, patients with ITP exhibiting platelet count below 50 × 109/L had dramatically elevated levels in both chemokine CXCL13 and its receptor CXCR5+ TFH compared to those with platelet count above 100 × 109/L. High frequencies of TFH correlated with poor therapeutic response. Furthermore, in vitro CD4+ T cell proliferation assay demonstrated a CXCL13 dose-dependent increase in the frequencies in both CD4+CXCR5+ TFH and CD4+CXCR5+PD-1+ TFH from ITP patients. Intriguingly, DCFH-DA assay illustrated a significant enhancement in intracellular ROS generation in CXCR5+ T cell subsets, especially in CD4+CXCR5+PD-1+ TFH from 4 patients with ITP. ConclusionsThese results underscore the pivotal role of CXCL13/CXCR5 axis-drived TFH expansion in the pathogenesis of ITP, providing a potential disease severity biomarker.

Interleukin 9 mediates T follicular helper cell activation to promote antibody responses.

Antigen-specific humoral responses are orchestrated through complex interactions among immune cells in lymphoid tissues, including the collaboration between B cells and T follicular helper (Tfh) cells. Accumulating evidence indicates a crucial role for interleukin-9 (IL-9) in the formation of germinal centers (GCs), enhancing the generation of class-switched high-affinity antibodies. However, the exact function of IL-9 in Tfh cell regulation remains unclear. In this study, we examined the humoral immune responses of CD4Cre/+Il9rafl/fl mice, which lack an IL-9-specific receptor in Tfh cells. Upon intraperitoneal immunization with sheep red blood cells (SRBCs), CD4Cre/+Il9rafl/fl mice displayed diminished levels of SRBC-specific IgG antibodies in their sera, along with reduced levels of GC B cells and plasma cells. Notably, Il9ra-deficient Tfh cells in the spleen exhibited decreased expression of their signature molecules such as B-cell lymphoma 6, C-X-C chemokine receptor 5, IL-4, and IL-21 compared to control mice. In models of allergic asthma induced by house dust mite (HDM) inhalation, CD4Cre/+Il9rafl/fl mice failed to elevate serum levels of HDM-specific IgE and IgG. This was accompanied by reductions in Tfh cells, GC B cells, and plasma cells in mediastinal lymph nodes. Furthermore, group 2 innate lymphoid cells (ILC2s) were identified as producers of IL-9 under immunizing conditions, possibly induced by leukotrienes released by activated IgD+ B cells around the T-B border. These observations may indicate the critical role of IL-9 receptor signaling in the activation of Tfh cells, with ILC2s potentially capable of supplying IL-9 in organized lymphoid tissues.

Tofacitinib alleviated salivary gland inflammation and reduced the percentages of effector T cells in murine Sjögren's disease.

The Janus kinases-signal transducer and activator of transcription (JAK-STAT) signalling pathway plays a crucial role in autoimmunity and the signalling pathways of many cytokines in Sjögren's disease (SjD). Therefore, the aim of this study was to investigate both the therapeutic and immunomodulatory effects of the oral JAK3/JAK2/JAK1 inhibitor tofacitinib in a murine model of SjD. Tofacitinib or vehicle was administered orally to the mice with SjD for 6 weeks. Salivary flow rate was measured every three weeks. Pathological changes of salivary gland were detected by haematoxylin-eosin staining, and the percentages of subsets of CD4+ T cells and B cells in the cervical lymph nodes (cLNs) and spleen was determined by flow cytometry. Tofacitinib significantly ameliorated submandibular gland inflammation compared to the control group, as evidenced by reduced lymphocytic infiltration. Salivary flow rates improved significantly in tofacitinib treated mice compared to controls, indicating restored salivary gland function. The treatment also led to a substantial decrease in follicular helper T (Tfh) cells and the Tfh/Treg ratio in both the spleen and cLNs. Additionally, the frequencies of T helper 1 (Th1) and T helper 17 (Th17) cells were reduced in the spleen and cLNs. Our data indicated that tofacitinib reduced percentages of effector T cells in an animal model of SjD. In addition, tofacitinib alleviated salivary gland inflammation and hypofunction, offering new insights into the clinical management of SjD.

Exploring the prognostic value of T follicular helper cell levels in chronic lymphocytic leukemia

Chronic lymphocytic leukemia (CLL) presents with heterogeneous clinical outcomes, suggesting varied underlying pathogenic mechanisms. This study aims to elucidate the impact of T follicular helper (Tfh) cells on CLL progression and prognosis. Gene expression profile data for CLL were collected from GSE22762 and GSE39671 datasets. Patients were divided into high and low groups using Tfh levels using the optimal cutoff value based on overall survival (OS) and time-to-first treatment (TTFT). Differential expression analysis was performed between these groups, followed by co-expression network analysis and single-sample Gene Set Enrichment Analysis (ssGSEA). Marker genes of Tfh cells were used to construct prognostic models. Additionally, 40 CLL patients were recruited and categorized based on median Tfh levels. Marker gene expression was assessed using RT-qPCR and Western Blot, and immune cell levels were determined through flow cytometry. The high group showed better prognosis compared to the low group. Among the 1121 differentially expressed genes identified, five co-expression networks were constructed, with the turquoise module showing the highest correlation with Tfh cells. Genes within this module significantly participate in cytokine-cytokine receptor interaction, PI3K-Akt signaling pathway, and natural killer cell mediated cytotoxicity. Tfh cells were significantly negatively correlated with activated B cells and positively correlated with Tregs. The Random Survival Forest (RSF) model identified 10 marker genes, and further analysis using Lasso regression and nomogram selected CLEC4A, RAE1, CD84, and PRDX1 as prognostic markers. In the high group, levels of CLEC4A and RAE1 were higher than in the low group, whereas CD84 and PRDX1 were lower. Flow cytometry revealed that the level of activated B cells in the high Tfh group was significantly lower than in the low Tfh group, while the level of Tregs is significantly higher in the high Tfh group. This study seeks to contribute to a more detailed understanding of the pathogenesis of CLL, delving into the prognostic significance of Tfh.

Integrated bioinformatics analysis identifies PCSK9 as a prognosticator correlated with lipid metabolism in pancreatic adenocarcinoma

BackgroundPancreatic adenocarcinoma (PAAD) is the most frequent kind of pancreatic cancer (PC). Recent studies suggest that lipid metabolism facilitates tumorigenesis, disease progression, and resistance to therapy by promoting lipid synthesis, accumulation, and breakdown. Thus, exploring the lipid metabolism network could unveil novel therapeutic avenues for early detection, precision medicine, and prognostication in PAAD. This project intends to develop new lipid metabolism-related biomarkers for PAAD diagnosis and investigate the link between important genes and immune cell infiltration (ICI).MethodsTissue samples from 20 PAAD patients and 20 healthy controls were obtained. Analysis were focused on the datasets GSE71729 and GSE16515, which include samples of PAAD (n = 161) and those from healthy human tissue (n = 61), derived from the GEO database. Knockdown of PCSK9 on PC cells were conducted by si-RNA and sh-RNA. Migration and cell functional experiments were performed to assess the role of PCSK9 in cell multiplication. Furthermore, a xenograft mouse model was employed to confirm PCSK9's function in vivo.ResultsThe expression level of Proprotein convertase subtilisin/kexin type 9 (PCSK9) is significantly elevated in tissues affected by PAAD when compared to normal tissues. Survival analyses indicated that increased PCSK9 levels are inversely related to overall and disease-free survival (DFS). PCSK9’s functional annotation associated it with the cell cycle and metabolism, especially energy metabolism. Examination of ICI data determined that PCSK9 expression demonstrated an unambiguous association with the M0 macrophages, T follicular helper cells (Tfh), gamma delta T cells and activated DC, and an inverse relationship with Monocytes, CD8+ T cells, memory B cells, resting CD4+ memory T cells, activated NK cells and resting DC abundance. PCSK9 expression knockdown has the ability to impede PC cells’ migration and proliferation.ConclusionOur study identified PCSK9 as a critical gene in PAAD. Expression levels of PCSK9 varied between PAAD and normal samples. ROC analysis verified PCSK9's strong capacity to differentiate PC from normal samples. Importantly, PCSK9 expression was considerably elevated in PC cell lines and tissues. Furthermore, PCSK9 stimulates the migration and proliferation of tumor cells in vivo and vitro.

Identification of key hub genes in knee osteoarthritis through integrated bioinformatics analysis.

Knee osteoarthritis (KOA) is a common chronic joint disease globally. Synovial inflammation plays a pivotal role in its pathogenesis, preceding cartilage damage. Identifying biomarkers in osteoarthritic synovial tissues holds promise for early diagnosis and targeted interventions. Gene expression profiles were obtained from the Gene Expression Omnibus database. Subsequent analyses included differential expression gene (DEG) analysis and weighted gene co-expression network analysis (WGCNA) on the combined datasets. We performed functional enrichment analysis on the overlapping genes between DEGs and module genes and constructed a protein-protein interaction network. Using Cytoscape software, we identified hub genes related to the disease and conducted gene set enrichment analysis on these hub genes. The CIBERSORT algorithm was employed to evaluate the correlation between hub genes and the abundance of immune cells within tissues. Finally, Mendelian randomization analysis was utilized to assess the potential of these hub genes as biomarkers. We identified 46 differentially expressed genes (DEGs), comprising 20 upregulated and 26 downregulated genes. Using WGCNA, we constructed a gene co-expression network and selected the most relevant modules, resulting in 24 intersecting genes with the DEGs. KEGG enrichment analysis of the intersecting genes identified the IL-17 signaling pathway, associated with inflammation, as the most significant pathway. Cytoscape software was utilized to rank the candidate genes, with JUN, ATF3, FOSB, NR4A2, and IL6 emerging as the top five based on the Degree algorithm. A nomogram model incorporating these five genes, supported by ROC curve analysis, validated their diagnostic efficacy. Immune infiltration and correlation analysis revealed that macrophages were significantly associated with JUN (p < 0.01), FOSB (p < 0.01), and NR4A2 (p < 0.05). Additionally, T follicular helper cells showed significant associations with ATF3 (p < 0.05), FOSB (p < 0.05), and JUN (p < 0.05). Mendelian randomization analysis provided strong evidence linking JUN (IVW: OR = 0.910, p = 0.005) and IL6 (IVW: OR = 1.024, p = 0.026) with KOA. Through the utilization of various bioinformatics analysis methods, we have pinpointed key hub genes relevant to knee osteoarthritis. These findings hold promise for advancing pre-symptomatic diagnostic strategies and enhancing our understanding of the biological underpinnings behind knee osteoarthritis susceptibility genes.

How type-2 dendritic cells induce Th2 differentiation: Instruction, repression, or fostering T cell-T cell communication?

Allergic disease is caused by the activation of allergen-specific CD4+ type-2 T follicular helper cells (Tfh2) and T helper 2 (Th2) effector cells that secrete the cytokines IL-4, IL-5, IL-9, and IL-13 upon allergen encounter, thereby inducing IgE production by B cells and tissue inflammation. While it is accepted that the priming and differentiation of naïve CD4+ T cells into Th2 requires allergen presentation by type 2 dendritic cells (DC2s), the underlying signals remain unidentified. In this review we focus on the interaction between allergen-presenting DC2s and naïve CD4+ T cells in lymph node (LN), and the potential mechanisms by which DC2s might instruct Th2 differentiation. We outline recent advances in characterizing DC2 development and heterogeneity. We review mechanisms of allergen sensing and current proposed mechanisms of Th2 differentiation, with specific consideration of the role of DC2s and how they might contribute to each mechanism. Finally, we assess recent publications reporting a detailed analysis of DC-T cell interactions in LNs and how they support Th2 differentiation. Together, these studies are starting to shape our understanding of this key initial step of the allergic immune response.

BAFF promotes follicular helper T cell development and germinal center formation through BR3 signal.

T follicular helper (Tfh) cells represent an important subset of CD4+ T cells that is crucial to the maturation and differentiation of B cells and the production of high-affinity antibodies. Since BAFF, a vital B cell survival factor, is also crucial to B cell maturation and differentiation, we assessed the effects of BAFF on Tfh cell development and function. We demonstrate that deficiency of BAFF, but not of APRIL, markedly inhibits Tfh cell development, germinal center (GC) formation, and antigen-specific antibody production. The promoting effect of BAFF on Tfh cell development is dependent on expression of BR3 on T cells, and its promoting effect on GC formation is dependent on expression of BR3 on both T cells and B cells. BAFF directly promotes expression of the Tfh cell-characteristic genes via NF-κB signaling. This effect does need BR3 expression. Thus, BAFF not only has direct effects on B cells, but it also has direct effects on Tfh cell differentiation via engagement of BR3 which collectively promote GC formation and production of high-affinity antibodies. This dual effect of BAFF on B cells and Tfh cells may help explain the clinical utility of BAFF antagonists in the management of certain autoimmune diseases.

Establish a novel immune-related gene prognostic risk index (IRGPRI) associated with CD8+ cytotoxic T lymphocytes in non-small-cell lung cancer (NSCLC)

BackgroundThe aim of this study is to create an index called IRGPRI (immune-related gene prognostic risk index) that can be utilized for predicting the prognosis and assessing the efficacy of immune checkpoint inhibitors (ICIs) therapy in patients with non-small-cell lung cancer (NSCLC). MethodsDistinguishing gene expression patterns (DEGs) were detected in CD8+ cytotoxic T lymphocytes (CTLs) compared to other cellular types such as CD4 T cells, B cells, plasma cells, and CD8 Tex using the advanced technology of Single-cell RNA Sequencing (scRNA-seq). The construction of IRGPRI was accomplished by employing LASSO Cox regression analysis. We conducted a comparative analysis on clinical characteristics and molecular features, such as pathway enrichment and gene mutation, among the distinct subgroups of IRGPRI. Furthermore, we explored the correlation between immunological characteristics and IRGPRI subgroups to comprehensively assess the effectiveness of ICIs in NSCLC patients. ResultsA total of 109 genes were identified by intersecting immune-related genes with DEGs obtained from single-cell RNA sequencing data (GSE131907), specifically comparing CTLs to other cell types. From these, we selected 7 prognosis-related genes, namely TRBC1, HLA-DMA, CTSH, RAC1, CTSL, ANXA2, and CEBPB. These genes were used to construct the IRGPRI. The prognosis of patients diagnosed with NSCLC was found to be significantly better in the low-risk group compared to the high-risk group, as demonstrated by Kaplan-Meier (K-M) survival analysis. This observation was further confirmed through the utilization of data from the GEO cohort. The low-risk group demonstrated an increase in pathways linked with immune response, whereas the high-risk group exhibited a higher prevalence of pathways related to cancer. Furthermore, it was noted in the TCGA cohort that there existed a significant rise in the mutation frequency of every gene within the high-risk group as opposed to the low-risk group. Missense variation emerged as the most prevalent form of mutation. According to the analysis of immune cell infiltration and function, the comprehensive findings suggest that the group with a low risk is characterized by an increased presence of plasma cells, CTLs, T cells follicular helper, Tregs, and Dendritic cell resting. Additionally, they exhibit a higher score in terms of immune function for B cells, CD8+ T cells, checkpoint activity, T cell inhibition and stimulation. Moreover, this low-risk group demonstrates greater efficacy when treated with ICIs therapy compared to the high-risk group. ConclusionsOur research effectively developed and verified a unique IRGPRI, showcasing its association with immune-related characteristics. As a result, the potential of IRGPRI as a valuable biomarker for predicting prognosis and evaluating the effectiveness of ICIs treatment in cancer is evident.

Variations in the germinal center response revealed by genetically diverse mouse strains.

The humoral response is complex and involves multiple cellular populations and signaling pathways. Bacterial and viral infections, as well as immunization regimens, can trigger this type of response, promoting the formation of microanatomical cellular structures called germinal centers (GCs). GCs formed in secondary lymphoid organs support the differentiation of high-affinity plasma cells and memory B cells. There is growing evidence that the quality of the humoral response is influenced by genetic variants. Using 12 genetically divergent mouse strains, we assessed the impact of genetics on GC cellular traits. At steady state, in the spleen, lymph nodes and Peyer's patches, we quantified GC B cells, plasma cells and follicular helper Tcells. These traits were also quantified in the spleen of mice following immunization with a foreign antigen, namely, sheep red blood cells, in addition to the number and size of GCs. We observed both strain- and organ-specific variations in cell type abundance, as well as for GC number and size. Moreover, we find that some of these traits are highly heritable. Importantly, the results of this study inform on the impact of genetic diversity in shaping the GC response and identify the traits that are the most impacted by genetic background.

Compromiso gástrico por enfermedad relacionada con IgG4

Introduction: IgG4-related disease (IgG4-RD) is a recently described entity, characterized by IgG4+ infiltration of plasma cells and storiform pattern sclerosis mediated by CD4+ lymphocytes and follicular helper T-cells (Tfhs). Most patients present with adenopathy and weight loss. Among the most affected sites are the pancreas, salivary and lacrimal glands, aorta, and retroperitoneum. Gastric involvement is rare. Diagnosis is based on histopathological examination of the involved organ, clinical manifestations, and serological results. Glucocorticoid therapy is the first line management. Case report: a 38-year-old male patient with symptoms suggestive of upper gastrointestinal (GI) bleeding, upper GI endoscopy revealed a gastric corpus ulcer, suspicious of an ulcerated gastric subepithelial lesion, possibly a gastrointestinal stromal tumor (GIST). Consequently, laparoscopic subtotal gastrectomy was performed. Histology was compatible with an ulcerated IgG4-related fibrosclerosing tumor. Glucocorticoid therapy was started with patient´s favorable response. Conclusion: in this paper, we will review the cases of IgG4-RD with gastric involvement reported in the literature to describe the most important features of this condition.

A single dose of an ALVAC vector-based RABV virus-like particle candidate vaccine induces a potent immune response in mice, cats and dogs

Rabies, caused by the Rabies virus (RABV), is a highly fatal zoonotic disease. Existing rabies vaccines have demonstrated good immune efficacy, but the complexity of immunization procedures and high cost has impeded the elimination of RABV, particularly in the post-COVID-19 era. There is a pressing need for safer and more effective rabies vaccines that streamline vaccination protocols and reduce expense. To meet this need, we have developed a potential rabies vaccine candidate called ALVAC-RABV-VLP, utilizing CRISPR/Cas9 gene editing technology. This vaccine employs a canarypox virus vector (ALVAC) to generate RABV virus-like particles (VLPs). In mice, a single dose of ALVAC-RABV-VLP effectively activated dendritic cells (DCs), follicular helper T cells (Tfh), and the germinal center (GC)/plasma cell axis, resulting in durable and effective humoral immune responses. The survival rate of mice challenged with lethal RABV was 100%. Similarly, in dogs and cats, a single immunization with ALVAC-RABV-VLP elicited a stronger and longer-lasting antibody response. ALVAC-RABV-VLP induced superior cellular and humoral immunity in both mice and beagles compared to the commercial inactivated rabies vaccine. In conclusion, ALVAC-RABV-VLP induced robust protective immune responses in mice, dogs and cats, offering a novel, cost-effective, efficient, and promising approach for herd prevention of rabies.

In vitro modulation of T cells in myasthenia gravis by low-dose IL-2.

Follicular helper (Tfh), peripheral helper (Tph), and regulatory (Treg) T cells are involved in myasthenia gravis (MG) pathogenesis, an autoimmune disorder arising from autoantibodies targeting neuromuscular junction proteins. This study explores the impact of low-dose IL-2 on Tfh, Tph, and Treg cells in vitro in MG. Acetylcholine-receptor antibody-positive MG (AChR-MG), muscle-specific kinase antibody-positive MG (MuSK-MG) patients, and healthy controls (HC) were studied. Blood cells were cultured with/without IL-2 and compared by the ratios of IL-2 stimulated/unstimulated cultures. In both AChR-MG and MuSK-MG patients, CD25+FoxP3+Tregs were lower, while CXCR5+PD-1+ or ICOS+Tfh and CXCR5-PD-1+ or ICOS+Tph cells were higher compared with HC. Among the MG group, the FoxP3+ Treg cells in AChR-MG patients were even lower compared with MuSK-MG patients. In vitro IL-2 stimulation increased Tregs in all groups while decreasing PD-1+/ICOS+Tfh and PD-1+/ICOS+Tph populations. The fold-increase ratio of Tregs and the fold-decrease ratio of PD-1+ or ICOS+Tfh and ICOS+Tph cells in AChR-MG and MuSK-MG patients were greater than in HCs. Low-dose IL-2 treatment may balance Tfh, Tph, and Treg cells in MG patients, offering a potential opportunity for disease modulation.

Ipsilateral or contralateral boosting of mice with mRNA vaccines confers equivalent immunity and protection against a SARS-CoV-2 Omicron strain.

Boosting with mRNA vaccines encoding variant-matched spike proteins has been implemented to mitigate their reduced efficacy against emerging SARS-CoV-2 variants. Nonetheless, in humans, it remains unclear whether boosting in the ipsilateral or contralateral arm with respect to the priming doses impacts immunity and protection. Here, we boosted K18-hACE2 mice with either monovalent mRNA-1273 (Wuhan-1 spike) or bivalent mRNA-1273.214 (Wuhan-1 + BA.1 spike) vaccine in the ipsilateral or contralateral leg after a two-dose priming series with mRNA-1273. Boosting in the ipsilateral or contralateral leg elicited equivalent levels of serum IgG and neutralizing antibody responses against Wuhan-1 and BA.1. While contralateral boosting with mRNA vaccines resulted in the expansion of spike-specific B and T cells beyond the ipsilateral draining lymph node (DLN) to the contralateral DLN, administration of a third mRNA vaccine dose at either site resulted in similar levels of antigen-specific germinal center B cells, plasmablasts/plasma cells, T follicular helper cells, and CD8+ T cells in the DLNs and the spleen. Furthermore, ipsilateral and contralateral boosting with mRNA-1273 or mRNA-1273.214 vaccines conferred similar homologous or heterologous immune protection against SARS-CoV-2 BA.1 virus challenge with equivalent reductions in viral RNA and infectious virus in the nasal turbinates and lungs. Collectively, our data show limited differences in B and T cell immune responses after ipsilateral and contralateral site boosting by mRNA vaccines that do not substantively impact protection against an Omicron strain.IMPORTANCESequential boosting with mRNA vaccines has been an effective strategy to overcome waning immunity and neutralization escape by emerging SARS-CoV-2 variants. However, it remains unclear how the site of boosting relative to the primary vaccination series shapes optimal immune responses or breadth of protection against variants. In K18-hACE2 transgenic mice, we observed that intramuscular boosting with historical monovalent or variant-matched bivalent vaccines in the ipsilateral or contralateral limb elicited comparable levels of serum spike-specific antibody and antigen-specific B and T cell responses. Moreover, boosting on either side conferred equivalent protection against a SARS-CoV-2 Omicron challenge strain. Our data in mice suggest that the site of intramuscular boosting with an mRNA vaccine does not substantially impact immunity or protection against SARS-CoV-2 infection.

Neoadjuvant nivolumab or nivolumab plus ipilimumab in early-stage triple-negative breast cancer: a phase 2 adaptive trial.

Immune checkpoint inhibition (ICI) with chemotherapy is now the standard of care for stage II-III triple-negative breast cancer; however, it is largely unknown for which patients ICI without chemotherapy could be an option and what the benefit of combination ICI could be. The adaptive BELLINI trial explored whether short combination ICI induces immune activation (primary end point, twofold increase in CD8+ T cells or IFNG), providing a rationale for neoadjuvant ICI without chemotherapy. Here, in window-of-opportunity cohorts A (4 weeks of anti-PD-1) and B (4 weeks of anti-PD-1 + anti-CTLA4), we observed immune activation in 53% (8 of 15) and 60% (9 of 15) of patients, respectively. High levels of tumor-infiltrating lymphocytes correlated with response. Single-cell RNA sequencing revealed that higher pretreatment tumor-reactive CD8+ T cells, follicular helper T cells and shorter distances between tumor and CD8+ T cells correlated with response. Higher levels of regulatory T cells after treatment were associated with nonresponse. Based on these data, we opened cohort C for patients with high levels of tumor-infiltrating lymphocytes (≥50%) who received 6 weeks of neoadjuvant anti-PD-1 + anti-CTLA4 followed by surgery (primary end point, pathological complete response). Overall, 53% (8 of 15) of patients had a major pathological response (<10% viable tumor) at resection, with 33% (5 of 15) having a pathological complete response. All cohorts met Simon's two-stage threshold for expansion to stage II. We observed grade ≥3 adverse events for 17% of patients and a high rate (57%) of immune-mediated endocrinopathies. In conclusion, neoadjuvant immunotherapy without chemotherapy demonstrates potential efficacy and warrants further investigation in patients with early triple-negative breast cancer. ClinicalTrials.gov registration: NCT03815890 .

Diverse Clinical and Immunological Profiles in Patients with IPEX Syndrome: a Multicenter Analysis from Turkey.

Immunodysregulation, Polyendocrinopathy, Enteropathy, and X-linked syndrome (IPEX), caused by pathogenic FOXP3 variants, is a rare autoimmune disorder with diverse clinical features, including early-onset diabetes, eczema, and enteropathy. Atypical cases show milder symptoms and unique signs, requiring different treatments. Therefore, there are ambiguities in the accurate diagnosis and management of IPEX. We sought to present clinical, genetic, and immunological assessments of 12 IPEX patients with long-term follow-up to facilitate the diagnosis and management of the disease. Clinical findings and treatment options of the patients were collected over time. Lymphocyte subpopulations, protein expressions, regulatory T (Treg) and circulating T follicular helper (cTFH) cells, and T-cell proliferation were analyzed. Predominant presentations included autoimmunity (91.6%), failure to thrive (66.7%), and eczema (58.3%). There were four classical and eight atypical IPEX individuals. Allergic manifestations were more common in atypical patients. Notably, chronic diarrhea demonstrated heightened severity compared to other manifestations. Four patients (33.3%) demonstrated eosinophilia, and nine (75%) showed high serum IgE levels. Most patients exhibited normal percentages of Treg cells with reduced CD25, FOXP3, and CTLA-4 expressions, corrected after hematopoietic stem cell transplantation (HSCT). Compared to healthy controls, the TH2-like skewing accompanied by reduced TH17-like responses was observed in cTFH and Treg cells of patients. Overall, nine patients (75%) received immunosuppressants (ISs), and six (50%) underwent HSCT, which was the only treatment revealing sustained control. Sirolimus was used in six patients and showed better control than other ISs. The first cohort from Turkey with long-term follow-up results, comparing typical and atypical cases, provides insights into the outcomes of different therapeutic modalities and T- cell subtype changes in IPEX syndrome.

In vivo CAR T cell therapy against angioimmunoblastic T cell lymphoma

BackgroundFor angioimmunoblastic T cell lymphoma (AITL), a rare cancer, no specific treatments are available and survival outcome is poor. We previously developed a murine model for AITL that mimics closely human disease and allows to evaluate new treatments. As in human AITL, the murine CD4+ follicular helper T (Tfh) cells are drivers of the malignancy. Therefore, chimeric antigen receptor (CAR) T cell therapy might represent a new therapeutic option.MethodsTo prevent fratricide among CAR T cells when delivering an CD4-specific CAR, we used a lentiviral vector (LV) encoding an anti-CD4 CAR, allowing exclusive entry into CD8 T cells.ResultsThese anti-CD4CAR CD8-targeted LVs achieved in murine AITL biopsies high CAR-expression levels in CD8 T cells. Malignant CD4 Tfh cells were eliminated from the mAITL lymphoma, while the CAR + CD8 T cells expanded upon encounter with the CD4 receptor and were shaped into functional cytotoxic cells. Finally, in vivo injection of the CAR + CD8-LVs into our preclinical AITL mouse model carrying lymphomas, significantly prolonged mice survival. Moreover, the in vivo generated functional CAR + CD8 T cells efficiently reduced neoplastic T cell numbers in the mAITL tumors.ConclusionThis is the first description of in vivo generated CAR T cells for therapy of a T cell lymphoma. The strategy described offers a new therapeutic concept for patients suffering from CD4-driven T cell lymphomas.

Identification of mitochondria-related genes as diagnostic biomarkers for diabetic nephropathy and their correlation with immune infiltration: New insights from bioinformatics analysis

BackgroundDiabetic nephropathy (DN) is a common and severe microvascular complication of diabetes. Mitochondrial dysfunction and immune inflammation are important factors in the pathogenesis of DN. However, the specific mechanisms and their intricate interactions in DN remain unclear. Besides, there are no effective specific predictive or diagnostic biomarkers for DN so far. Therefore, this study aims to elucidate the role of mitochondrial-related genes and their possibility as predictive or diagnostic biomarkers, as well as their crosstalk with immune infiltration in the progression of DN. MethodsBased on the GEO database and limma R package, the differentially expressed genes (DEGs) of DN were identified. Mitochondrial-related DEGs (MitoDEGs) were then obtained by intersecting these DEGs with mitochondria-related genes from the MitoCarta 3.0 database. Subsequently, the candidate hub genes were further screened by gene co-expression network analysis (WGCNA), and verified mRNA levels of these genes by real-time quantitative PCR (qRT-PCR) in high-glucose-treated human proximal tubular (HK-2) cells. The verified hub genes were utilized to construct a combined diagnostic model for DN, with its diagnostic efficacy assessed across the GSE30122 and GSE96804 datasets. Additionally, the immune infiltration pattern in DN was assessed with the CIBERSORT algorithm, and the Nephroseq v5 database was used to analyze the correlation between hub genes and clinical features of DN. ResultsSeven mitochondria-related candidate hub genes were screened from 56 MitoDEGs. Subsequently, the expression levels of six of them, namely EFHD1, CASP3, AASS, MPC1, NT5DC2, and BCL2A1, exhibited significant inter-group differences in the HK-2 cell model. The diagnostic model based on the six genes demonstrated good diagnostic efficacy in both training and validation sets. Furthermore, correlation analysis indicated that EFHD1 and AASS, downregulated in DN, are positively correlated with eGFR and negatively with serum creatinine. Conversely, CASP3, NT5DC2, and BCL2A1, upregulated in DN, show opposite correlations. In addition, spearman analysis revealed that the six hub genes were significantly associated with the infiltration of immune cells, including M1 and M2 macrophages, mast cells, resting NK cells, gamma delta T cells, and follicular helper T cells. ConclusionThis study elucidated the characteristics of mitochondria-related genes and their correlation with immune cell infiltration in DN, providing new insights for exploring the pathogenesis of DN and facilitating the identification of new potential biomarkers and therapeutic targets.