The immunomodulatory effect of koumine on B cells under dependent and independent responses by T cells.

Dysregulated activation of polyclonal B cells and production of pathogenic antibodies are involved in the development of rheumatoid arthritis (RA). Therefore, targeted B cell therapy is effective against RA. Gelsemium elegans (Gardn. & Champ.) Benth., a toxic plant widely distributed in Southeast Asia, has been used for treating rheumatoid pain, neuropathic pain, spasticity, skin ulcers, and cancers for many years in traditional Chinese medicine. Koumine, an alkaloid monomer from Gelsemium elegans Benth., exerts therapeutic effects against RA. However, whether koumine affects B cells remains unknown. In this study, the effect of koumine on B cells under T cell-independent (TI) and T cell-dependent (TD) immune responses is investigated in vitro and in vivo. Mouse primary B cells were obtained by immunomagnetic bead sorting, and immunomodulatory effects of koumine on the activation, proliferation, and differentiation of B cells were determined in TI and TD models induced by lipopolysaccharide (LPS) and anti-CD40 antibodies in vitro, respectively. The humoral immune responses of TI and TD were established using NP-AECM-FICOLL and NP-CGG in C57BL/6J mice, respectively. We found that koumine inhibited B cell differentiation in the TI model and inhibited B cell activation and proliferation in the TD model in vitro. Koumine also inhibited antibody secretion in TI immune response, TD initial immune response, and in TD secondary immune response. Our results reveal that koumine has a direct and indirect immune regulatory effect on B cells, showing that it can directly inhibit the differentiation and secretion of autoantibodies after abnormal activation of B cells, and indirectly inhibit the activation and proliferation of TD B cells to reduce the secretion of antibodies. It may be an important mechanism for its anti-RA effect in mice, providing a rationale and laboratory data support for the application of koumine in anti-human RA therapy.

Dysregulation of TFH-B-TRM lymphocyte cooperation is associated with unfavorable anti-PD-1 responses in EGFR-mutant lung cancer.

Patients with non-small cell lung cancer (NSCLC) with epidermal growth factor receptor (EGFR) mutations exhibit an unfavorable response to PD-1 inhibitor through unclear mechanisms. Hypothesizing that EGFR mutations alter tumor-immune interactions, we compare tumor-infiltrating lymphocytes between EGFR mutant (EGFR-MT) and wild type (EGFR-WT) tumors through single-cell transcriptomic analysis. We find that B cells, CXCL13-producing follicular helper CD4+ T (TFH)-like cells, and tissue-resident memory CD8+ T (TRM)-like cells decreased in EGFR-MT tumors. The NOTCH-RBPJ regulatory network, which is vital for persistence of TRM state, is perturbed, and the interactions between TFH and B cells through the CXCL13-CXCR5 axis disappear in EGFR-MT tumors. Notably, the proportion of TRM-like cells is predictive for anti-PD-1 response in NSCLC. Our findings suggest that the impairment of TFH-B-TRM cooperation in tertiary lymphoid structure formation, accompanied by the dysregulation of TRM homeostasis and the loss of TFH-B crosstalk, underlies unfavorable anti-PD-1 response in EGFR-MT lung tumors.

PD-1 blockade restores helper activity of tumor-infiltrating, exhausted PD-1hiCD39+ CD4 T cells.

Tumor antigen-specific CD4 T cells accumulate at tumor sites, evoking their involvement in antitumor effector functions in situ. Contrary to CD8 cytotoxic T lymphocyte exhaustion, that of CD4 T cells remains poorly appreciated. Here, using phenotypic, transcriptomic, and functional approaches, we characterized CD4 T cell exhaustion in patients with head and neck, cervical, and ovarian cancer. We identified a CD4 tumor-infiltrating lymphocyte (TIL) population, defined by high PD-1 and CD39 expression, which contained high proportions of cytokine-producing cells, although the quantity of cytokines produced by these cells was low, evoking an exhausted state. Terminal exhaustion of CD4 TILs was instated regardless of TIM-3 expression, suggesting divergence with CD8 T cell exhaustion. scRNA-Seq and further phenotypic analyses uncovered similarities with the CD8 T cell exhaustion program. In particular, PD-1hiCD39+ CD4 TILs expressed the exhaustion transcription factor TOX and the chemokine CXCL13 and were tumor antigen specific. In vitro, PD-1 blockade enhanced CD4 TIL activation, as evidenced by increased CD154 expression and cytokine secretion, leading to improved dendritic cell maturation and consequently higher tumor-specific CD8 T cell proliferation. Our data identify exhausted CD4 TILs as players in responsiveness to immune checkpoint blockade.

Autoreactive T-Lymphocytes in Inflammatory Skin Diseases.

The presence of one or several autoantigen(s) and a response by the adaptive immune system are the key criteria to classify a pathology as an autoimmune disease. The list of entities fulfilling this criterion is currently growing in the light of recent advancements in the pathogenetic understanding of a number of important dermatoses. The role of autoreactive T-lymphocytes differs amongst these pathologies. While they are directly involved as effector cells attacking and sometimes killing their respective target in some diseases (e.g., vitiligo), they provide help to B-lymphocytes, which in turn produce the pathogenic autoreactive antibodies in others (pemphigus and pemphigoid). Atopic dermatits is a chimera in this regard, as there is evidence for both functions. Psoriasis is an example for an entity where autoantigens were finally identified, suggesting that at least a subgroup of patients should be classified as suffering from a true autoimmune rather than autoinflammatory condition. Identification of resident memory T-lymphocytes (TRM) helped to understand why certain diseases relapse at the same site after seemingly effective therapy. Therefore, the in-depth characterization of autoreactive T-lyphocytes goes way beyond an academic exercise and opens the door toward improved therapies yielding durable responses. TRM are particularly suitable targets in this regard, and the clinical efficacy of some established and emerging therapeutic strategies such as the inhibition of Janus Kinase 3 or interleukin 15 may rely on their capacity to prevent TRM differentiation and maintenance. Research in this field brings us closer to the ultimate goal in the management of autoimmunity at large, namely resetting the immune system in order to restore the state of tolerance.

The Epitope-Specific Anti-human CD4 Antibody MAX.16H5 and Its Role in Immune Tolerance.

T cell modulation in the clinical background of autoimmune diseases or allogeneic cell and organ transplantations with concurrent preservation of their natural immunological functions (e.g., pathogen defense) is the major obstacle in immunology. An anti-human CD4 antibody (MAX.16H5) was applied intravenously in clinical trials for the treatment of autoimmune diseases (e.g., rheumatoid arthritis) and acute late-onset rejection after transplantation of a renal allograft. The response rates were remarkable and no critical allergic problems or side effects were obtained. During the treatment of autoimmune diseases with the murine MAX.16H5 IgG1 antibody its effector mechanisms with effects on lymphocytes, cytokines, laboratory and clinical parameters, adverse effects as well as pharmacodynamics and kinetics were studied in detail. However, as the possibility of developing immune reactions against the murine IgG1 Fc-part remains, the murine antibody was chimerized, inheriting CD4-directed variable domains of the MAX.16H5 IgG1 connected to a human IgG4 backbone. Both antibodies were studied in vitro and in specific humanized mouse transplantation models in vivo with a new scope. By ex vivo incubation of an allogeneic immune cell transplant with MAX.16H5 a new therapy strategy has emerged for the first time enabling both the preservation of the graft-vs.-leukemia (GVL) effect and the permanent suppression of the acute graft-vs.-host disease (aGVHD) without conventional immunosuppression. In this review, we especially focus on experimental data and clinical trials obtained from the treatment of autoimmune diseases with the murine MAX.16H5 IgG1 antibody. Insights gained from these trials have paved the way to better understand the effects with the chimerized MAX.16H5 IgG4 as novel therapeutic approach in the context of GVHD prevention.

Two populations of circulating PD-1hiCD4 T cells with distinct B cell helping capacity are elevated in early rheumatoid arthritis.

OBJECTIVE: A novel population of B helper cells, phenotypically CD4+CXCR5-PD-1hi, has been described in the synovial tissues and peripheral blood of seropositive RA patients, and termed 'peripheral helper T' (Tph) cells. Contrary to CD4+CXCR5+PD-1hi follicular helper T (Tfh), Tph cells are not located in lymphoid organs but accumulate in inflamed tissues. Our objective was to study the frequency of circulating Tph (cTph) and circulating Tfh cell counterparts (cTfh) in patients with early RA (eRA). METHODS: Freshly isolated peripheral blood mononuclear cells from 56 DMARD-naïve eRA patients and 56 healthy controls were examined by flow cytometry. Autologous cocultures of naïve or memory B cells were established with isolated peripheral blood Tph or Tfh cells. RESULTS: Seropositive (RF+ and/or ACPA+, n = 38) but not seronegative eRA patients (n = 18) demonstrated increased frequencies and absolute numbers of cTph and cTfh cells. cTph but not cTfh cells expressed CCR2. Those eRA patients who experienced a significant clinical improvement at 12 months demonstrated a marked decrease of their cTph cell numbers whereas their cTfh cell numbers remained unchanged. Both isolated Tph and isolated Tfh cells were able to induce maturation of memory B cells, whereas only Tfh cells could differentiate naïve B cells. CONCLUSION: Two populations of PD-1hiCD4 T cells with distinct phenotype and B cell helping capacity are increased in the peripheral blood of seropositive eRA patients. Whereas cTph cells are present only in patients with an active disease, cTfh cells seem to be constitutively elevated.

The Post-GWAS Era: How to Validate the Contribution of Gene Variants in Lupus.

PURPOSE OF REVIEW: Systemic lupus erythematosus (SLE) is a complex autoimmune disease with strong genetic associations. Here, we provide an update on recent advancements in validating SLE candidate genes and risk variants identified in genome-wide association studies (GWAS). RECENT FINDINGS: A pairing of computational biology with new and emerging techniques has significantly increased our understanding of SLE associated variants. Specifically, generation of mutations within mice and examination of patient samples has been the dominant mechanisms for variant validation. While progress has been made in validating some genes, the number of associated genes is growing with minimal exploration of the effects of individual variants on SLE. This indicates that further examination of SLE risk variants in a cell-type-specific manner is required for better understanding of their contributions to SLE disease mechanisms.

Linked help from bacterial proteins drives autoantibody production in small vessel vasculitis.

The small vessel vasculitides granulomatosis with polyangiitis (GPA) and microscopic polyangiitis are associated with autoantibodies to neutrophil cytoplasm antigens (ANCA), principally proteinase-3 (PR3) and myeloperoxidase (MPO). There is an association between GPA and nasal carriage of Staphylococcus aureus. The recent finding that S. aureus produces proteins that bind tightly to and block the function of both PR3 and MPO suggests a mechanism for ANCA formation. The bacterial protein-autoantigen conjugate is recognised by B cells with ANCA specificity, internalised, and the bacterial protein processed and presented to T cells with specificity for bacterial peptides. The T cell can then provide help to the B cell, allowing class switching, affinity maturation and the production of pathogenic ANCA. This mechanism predicts that T cells with this specificity will be found in patients, and that the bacterial protein-autoantigen conjugate will be particularly efficient at eliciting ANCA production.

Viral subversion of B cell responses within secondary lymphoid organs.

Antibodies play a crucial role in virus control. The production of antibodies requires virus-specific B cells to encounter viral antigens in lymph nodes, become activated, interact with different immune cells, proliferate and enter specific differentiation programmes. Each step occurs in distinct lymph node niches, requiring a coordinated migration of B cells between different subcompartments. The development of multiphoton intravital microscopy has enabled researchers to begin to elucidate the precise cellular and molecular events by which lymph nodes coordinate humoral responses. This Review discusses recent studies that clarify how viruses interfere with antibody responses, highlighting how these mechanisms relate to our topological and temporal understanding of B cell activation within secondary lymphoid organs.

Relationship of mean platelet volume to lymphocyte ratio and coronary collateral circulation in patients with stable angina pectoris.

BACKGROUND: In patients with coronary artery disease, coronary collateral circulation (CCC) develops as an adaptation to ischemia and contributes toward reduction of cardiovascular events. Recently, the mean platelet volume-to-lymphocyte ratio (MPVLR) has emerged as a novel and readily available marker of inflammation and thrombosis. This study aimed to investigate the relationship between MPVLR and development of CCC. PATIENTS AND METHODS: A total of 332 patients with stable angina pectoris undergoing coronary arteriography were enrolled and divided on the basis of the development of CCC into two groups: group with adequate CCC (n=243) and group with impaired CCC (n=89). Routine complete blood count parameters and high-sensitivity C-reactive protein (hsCRP) were measured before coronary arteriography. RESULTS: Both MPVLR and hsCRP levels were higher in the impaired CCC group (P<0.001 and P=0.007, respectively). Multivariate logistic regression analysis indicated that MPVLR was associated independently with impaired CCC [odds ratio (OR): 1.706, 95% confidence interval (CI): 1.328-2.192, P<0.001]. In addition to MPVLR, hsCRP (OR: 1.144, P=0.030) and fasting blood glucose (OR: 1.007, P=0.049) were also associated independently with impaired CCC. In receiver operating characteristics curve analysis, an optimal cut-off point for MPVLR (4.47) was found to predict the presence of good CCC with a sensitivity of 75.3% and a specificity of 71.2% (P<0001). CONCLUSION: Our findings suggest that measurement of MPVLR may predict the development of CCC in patients with stable coronary artery disease. An increased MPVLR is associated independently with impaired CCC in these patients.

Crosstalk between type II NKT cells and T cells leads to spontaneous chronic inflammatory liver disease.

BACKGROUND & AIM: Natural killer T (NKT) cells are CD1d-restricted innate-like T cells that modulate innate and adaptive immune responses. Unlike the well-characterized invariant/type I NKT cells, type II NKT cells with a diverse T cell receptor repertoire are poorly understood. This study defines the pathogenic role of type II NKT cells in the etiology of chronic liver inflammation. METHODS: Transgenic mice with the Lck promoter directing CD1d overexpression on T cells in Jα18 wild-type (Lck-CD1dTgJα18+; type I NKT cell sufficient) and Jα18-deficient (Lck-CD1dTgJα18o, type I NKT cell deficient) mice were analyzed for liver pathology and crosstalk between type II NKT cells and conventional T cells. CD1d expression on T cells in peripheral blood samples and liver sections from autoimmune hepatitis patients and healthy individuals were also examined. RESULTS: Lck-CD1dTgJα18o and Lck-CD1dTgJα18+ mice developed similar degrees of liver pathology resembling chronic autoimmune hepatitis in humans. Increased CD1d expression on T cells promoted the activation of type II NKT cells and other T cells. This resulted in Th1-skewing and impaired Th2 cytokine production in type II NKT cells. Dysfunction of type II NKT cells was accompanied by conventional T cell activation and pro-inflammatory cytokine production, leading to a hepatic T/B lymphocyte infiltration, elevated autoantibodies and hepatic injury in Lck-CD1dTg mice. A similar mechanism could be extended to humans as CD1d expression is upregulated on activated human T cells and increased presence of CD1d-expressing T cells was observed in autoimmune hepatitis patients. CONCLUSIONS: Our data reveals enhanced crosstalk between type II NKT cells and conventional T cells, leading to a Th1-skewed inflammatory milieu, and consequently, to the development of chronic autoimmune liver disease. Lay summary: CD1d overexpression on T cells enhances crosstalk between type II NKT cells and T cells, resulting in their aberrant activation and leading to the development of chronic autoimmune liver disease.

[Immunological alterations in common variable immunodeficiency]. / Alteraciones inmunológicas en la inmunodeficiencia común variable.

Common variable immunodeficiency (CVID) is the largest group of symptomatic primary immune deficiencies; it is characterized by hypogammaglobulinemia, poor response to vaccines and increased susceptibility to infections. Cellular phenotypes and abnormalities have been described both in adaptive and innate immune response. Several classifications of common variable immunodeficiency are based on defects found on T and B cells, which have been correlated with clinical manifestations. In recent years, significant progress has been made in elucidating the genetic mechanisms that result in a IDCV phenotype. Massive sequencing technologies have favored the description of mutations in several genes, but only in 2 % to 10 % of patients. These monogenetic defects are: ICOS, TNFRSF13B (TACI), TNFRS13C (BAFFR), TNRFSF12 (TWEAK), CD19, CD81, CR2 (CD21), MS4A1 (CD20), (CD27), LRBA, CTLA4, PRKCD, PLCG2, NFKB1, NFKB2, PIK3CD, PIK3R, VAV1, RAC1, BLK, IKZF1 (IKAROS) and IRF2BP2. These findings have provided a possible explanation for the pathogenesis of IDCV, since these molecules play an important role in the co-operation between B and T cells in the germinal center, as well as in intrinsic signaling pathways of both.

La inmunodeficiencia común variable constituye el mayor grupo de inmunodeficiencias primarias sintomáticas; se caracterizan por hipogammaglobulinemia, pobre respuesta a las vacunas y susceptibilidad aumentada a las infecciones. Se han descrito fenotipos celulares y anormalidades tanto en la respuesta inmune adaptativa como en la innata. Varias de las clasificaciones se basan en los defectos encontrados en las células T y B, que se han correlacionado con las manifestaciones clínicas. En los últimos años se ha progresado significativamente en el desentrañamiento de los mecanismos genéticos que resultan en un fenotipo de inmunodeficiencia común variable. Las tecnologías de secuenciación masiva han favorecido la descripción de mutaciones en varios genes, pero solo en 2 a 10 % de los pacientes. Estos defectos monogénicos son ICOS, TNFRSF13B (TACI), TNFRS13C (BAFFR), TNRFSF12 (TWEAK), CD19, CD81, CR2 (CD21), MS4A1 (CD20), TNFRSF7 (CD27), LRBA, CTLA4, PRKCD, PLCG2, NFKB1, NFKB2, PIK3CD, PIK3R, VAV1, RAC1, BLK, IKZF1 (IKAROS) y IRF2BP2. Los anteriores hallazgos han proporcionado una posible explicación para la patogénesis de la inmunodeficiencia común variable, ya que esas moléculas desempeñan un papel importante en la cooperación entre las células B y T en el centro germinal, así como en las vías de señalización intrínseca de ambas.

Positive and negative functions of B lymphocytes in tumors.

Accumulating evidence indicated that B lymphocytes exerted complex functions in tumor immunity. On the one hand, B lymphocytes can inhibit tumor development through antibody generation, antigen presentation, tumor tissue interaction, and direct killing. On the other hand, B lymphocytes have tumor-promoting functions. A typical type of B lymphocytes, termed regulatory B cells, is confirmed to attenuate immune response in a tumor environment. In this paper, we summarize the current understanding of B-cell functions in tumor immunology, which may shed light on potential therapeutic strategies against cancer.

Circulating T follicular helper cells with increased function during chronic graft-versus-host disease.

Chronic graft-versus-host disease (cGVHD) remains a major late complication of allogeneic hematopoietic stem cell transplantation (HSCT). Previous studies have established that both donor B and T cells contribute to immune pathology in cGVHD but the mechanisms responsible for coordinated B- and T-cell responses directed against recipient antigens have not been understood. T follicular helper cells (TFH) play an important role in the regulation of B-cell immunity. We performed extensive phenotypic and functional analysis of circulating TFH (cTFH) and B cells in 66 patients after HSCT. Patients with active cGVHD had a significantly lower frequency of cTFH compared with patients without cGVHD. This was associated with higher CXCL13 plasma levels suggesting increased homing of TFH to secondary lymphoid organs. In patients with active cGVHD, cTFH phenotype was skewed toward a highly activated profile with predominance of T helper 2 (Th2)/Th17 subsets. Activated cTFH in patients with cGVHD demonstrated increased functional ability to promote B-cell immunoglobulin secretion and maturation. Moreover, the activation signature of cTFH was highly correlated with increased B-cell activation and plasmablast maturation in patients after transplant. These studies provide new insights into the immune pathogenesis of human cGVHD and identify TFH as a key coordinating element supporting B-cell involvement in this disease.

Local T/B cooperation in inflamed tissues is supported by T follicular helper-like cells.

Autoimmune diseases and other inflammatory conditions are characterized by large lymphocytic tissue infiltrates in which T and B cells can be found in close contact. Here, using a murine airway inflammation model, we compare antigen-specific T and B cells in lung tissue versus lung-draining lymph node. In the lung we identify a B-cell population exhibiting a classical germinal centre phenotype without being organized into ectopic lymphoid tissue. By contrast, classical CXCR5(+) Bcl-6(+) T follicular helper cells are not present. Nevertheless, lung-infiltrating T cells exhibit follicular helper-like properties including the potential to provide help to naive B cells. The lung tissue is also a survival niche for memory T and B cells remaining in residual peribronchial infiltrates after resolution of inflammation. Collectively, this study shows the importance of T/B cooperation not only in lymph nodes but also in inflamed peripheral tissues for local antibody responses to infection and autoimmunity.

T Follicular Helper Cell-Dependent Clearance of a Persistent Virus Infection Requires T Cell Expression of the Histone Demethylase UTX.

Epigenetic changes, including histone methylation, control T cell differentiation and memory formation, though the enzymes that mediate these processes are not clear. We show that UTX, a histone H3 lysine 27 (H3K27) demethylase, supports T follicular helper (Tfh) cell responses that are essential for B cell antibody generation and the resolution of chronic viral infections. Mice with a T cell-specific UTX deletion had fewer Tfh cells, reduced germinal center responses, lacked virus-specific immunoglobulin G (IgG), and were unable to resolve chronic lymphocytic choriomeningitis virus infections. UTX-deficient T cells showed decreased expression of interleukin-6 receptor-α and other Tfh cell-related genes that were associated with increased H3K27 methylation. Additionally, Turner Syndrome subjects, who are predisposed to chronic ear infections, had reduced UTX expression in immune cells and decreased circulating CD4(+) CXCR5(+) T cell frequency. Thus, we identify a critical link between UTX in T cells and immunity to infection.

Mechanisms of sickle cell alloimmunization.

Red blood cell (RBC) alloimmunization can be a life-threatening complication for patients with sickle cell disease (SCD) receiving therapeutic transfusions. Despite provision of extended antigen-matched donor RBCs, patients continue to develop antibodies due to high degree of polymorphisms in the immunogenic antigens in individuals of African ancestry. Identification of biomarkers of alloimmunization in this patient population is therefore of great interest and will help to identify in advance patients most likely to make antibodies in response to transfusion. We have recently identified altered T cell responses and innate immune abnormalities in alloimmunized SCD patients. In this paper, we summarize this work and propose our working model of how innate immune abnormalities can contribute to pathogenic T cell responses in alloimmunized SCD patients. We believe that unravelling the basis of such altered interactions at the cellular and molecular level will help future identification of biomarkers of alloimmunization with the goal that this information will ultimately help guide therapy in these patients.

Imaging Tolerance Induction in the Classic Medawar Neonatal Mouse Model: Active Roles of Multiple F1-Donor Cell Types.

The immature immune system is uniquely susceptible to tolerance induction and thus an attractive target for immunomodulation strategies for organ transplantation. Newborn mice injected with adult semi-allogeneic lymphohematopoietic cells accept transplants without immunosuppressive drugs. Early in vivo/in situ events leading to neonatal tolerance remain poorly understood. Here, we show by whole body/organ imaging that injected cells home to lymphoid organs and liver where various F1-donor cell types selectively alter neonatal immunity. In host thymus, F1-donor dendritic cells (DC) interact with developing thymocytes and regulatory T cells suggesting a role in negative selection. In spleen and lymph nodes, F1-donor regulatory T/B cells associate with host alloreactive cells and by themselves prolong cardiac allograft survival. In liver, F1-donor cells give rise to albumin-containing hepatocyte-like cells. The neonatal immune system is lymphopenic, Th-2 immunodeviated and contains immature DC, suggesting susceptibility to regulation by adult F1-donor cells. CD8a T cell inactivation greatly enhances chimerism, suggesting that variable emerging neonatal alloreactivity becomes a barrier to tolerance induction. This comprehensive qualitative imaging study systematically shows contribution of multiple in vivo processes leading simultaneously to robust tolerance. These insights into robust tolerance induction have important implications for development of strategies for clinical application.