Cell Activating Factor Receptor Research Articles

Single-cell analysis of human nasal mucosal IgE antibody secreting cells reveals a newly minted phenotype

Immunoglobulin (Ig) E is central to the pathogenesis of allergic conditions, including allergic fungal rhinosinusitis. However, little is known about IgE antibody secreting cells (ASCs). We performed single-cell RNA sequencing from cluster of differentiation (CD)19+ and CD19- ASCs of nasal polyps from patients with allergic fungal rhinosinusitis (n=3). Nasal polyps were highly enriched in CD19+ ASCs. Class-switched IgG and IgA ASCs were dominant (95.8%), whereas IgE ASCs were rare (2%) and found only in the CD19+ compartment. Through Ig gene repertoire analysis, IgE ASCs shared clones with IgD-CD27- "double-negative" B cells, IgD+CD27+ unswitched memory B cells, and IgD-CD27+ switched memory B cells, suggesting ontogeny from both IgD+ and memory B cells. Transcriptionally, mucosal IgE ASCs upregulate pathways related to antigen presentation, chemotaxis, B cell receptor stimulation, and survival compared with non-IgE ASCs. Additionally, IgE ASCs have a higher expression of genes encoding lysosomal-associated protein transmembrane 5 (LAPTM5) and CD23, as well as upregulation of CD74 (receptor for macrophage inhibitory factor), store-operated Calcium entry-associated regulatory factor (SARAF), and B cell activating factor receptor (BAFFR), which resemble an early minted ASC phenotype. Overall, these findings reinforce the paradigm that human ex vivo mucosal IgE ASCs have a more immature plasma cell phenotype than other class-switched mucosal ASCs and suggest unique functional roles for mucosal IgE ASCs in concert with Ig secretion.

B cell-specific mAb–siRNA conjugates improve experimental myasthenia

Myasthenia gravis (MG) is a debilitating autoimmune disease characterized by muscle fatigue and weakness caused by autoantibody- and complement-mediated damage to the neuromuscular junction. This study sought to compare the efficacy of unique sets of monoclonal antibody–siRNA conjugates, individually (mono) or in combination (duo), against the crucial receptors predominantly or solely expressed on two subsets of B cells—plasma B cells and their precursor (transitional mature B) cells in a mouse model of MG. At the optimized doses, the conjugates, likely due to the combined activities of mAb and siRNA, substantially decreased the expression levels of CD268 (B cell-activating factor receptor) in mature B cells and CD269 (B-cell maturation antigen) in plasma cells concomitantly with reducing the levels of acetylcholine receptor (AChR)-specific autoantibodies. PEGylation, but not pretreatment with an antibody against type 1 interferon receptor, further improved duoconjugate-induced reduction in the autoantibody levels. Our results show that the duoconjugate treatment significantly improved the clinical symptoms of MG, consistent with the preservation of bungarotoxin-bound functional AChRs. In the future, developing similar target-specific combination molecules can potentially turn into a new and effective therapeutic approach for MG.

Pooled screening of CAR T cells identifies diverse immune signaling domains for next-generation immunotherapies.

Chimeric antigen receptors (CARs) repurpose natural signaling components to retarget T cells to refractory cancers but have shown limited efficacy in persistent, recurrent malignancies. Here, we introduce "CAR Pooling," a multiplexed approach to rapidly identify CAR designs with clinical potential. Forty CARs with signaling domains derived from a range of immune cell lineages were evaluated in pooled assays for their ability to stimulate critical T cell effector functions during repetitive stimulation that mimics long-term tumor antigen exposure. Several domains were identified from the tumor necrosis factor (TNF) receptor family that have been primarily associated with B cells. CD40 enhanced proliferation, whereas B cell-activating factor receptor (BAFF-R) and transmembrane activator and CAML interactor (TACI) promoted cytotoxicity. These functions were enhanced relative to clinical benchmarks after prolonged antigen stimulation, and CAR T cell signaling through these domains fell into distinct states of memory, cytotoxicity, and metabolism. BAFF-R CAR T cells were enriched for a highly cytotoxic transcriptional signature previously associated with positive clinical outcomes. We also observed that replacing the 4-1BB intracellular signaling domain with the BAFF-R signaling domain in a clinically validated B cell maturation antigen (BCMA)-specific CAR resulted in enhanced activity in a xenotransplant model of multiple myeloma. Together, these results show that CAR Pooling is a general approach for rapid exploration of CAR architecture and activity to improve the efficacy of CAR T cell therapies.

CVID-Associated B Cell Activating Factor Receptor Variants Change Receptor Oligomerization, Ligand Binding, and Signaling Responses

PurposeBinding of the B cell activating factor (BAFF) to its receptor (BAFFR) activates in mature B cells many essential pro-survival functions. Null mutations in the BAFFR gene result in complete BAFFR deficiency and cause a block in B cell development at the transition from immature to mature B cells leading therefore to B lymphopenia and hypogammaglobulinemia. In addition to complete BAFFR deficiency, single nucleotide variants encoding BAFFR missense mutations were found in patients suffering from common variable immunodeficiency (CVID), autoimmunity, or B cell lymphomas. As it remained unclear to which extent such variants disturb the activity of BAFFR, we performed genetic association studies and developed a cellular system that allows the unbiased analysis of BAFFR variants regarding oligomerization, signaling, and ectodomain shedding.MethodsIn addition to genetic association studies, the BAFFR variants P21R, A52T, G64V, DUP92-95, P146S, and H159Y were expressed by lentiviral gene transfer in DG-75 Burkitt’s lymphoma cells and analyzed for their impacts on BAFFR function.ResultsBinding of BAFF to BAFFR was affected by P21R and A52T. Spontaneous oligomerization of BAFFR was disturbed by P21R, A52T, G64V, and P146S. BAFF-dependent activation of NF-κB2 was reduced by P21R and P146S, while interactions between BAFFR and the B cell antigen receptor component CD79B and AKT phosphorylation were impaired by P21R, A52T, G64V, and DUP92-95. P21R, G64V, and DUP92-95 interfered with phosphorylation of ERK1/2, while BAFF-induced shedding of the BAFFR ectodomain was only impaired by P21R.ConclusionAlthough all variants change BAFFR function and have the potential to contribute as modifiers to the development of primary antibody deficiencies, autoimmunity, and lymphoma, P21R is the only variant that was found to correlate positively with CVID.

Modulation of Immune Responses Against HA1 Influenza Vaccine Candidate by B-lymphocyte Stimulator Cytokine in Mice.

Utilizing subunit vaccines is one of the strategies to address influenza infection. Recent innovations have focused on high doses of vaccine antigens and immune enhancers or adjuvant to induce more robust and long-lasting immune responses. Here, an effect of the B cell-activating factor receptor (BAFF-R) to increase the magnitude and durability of immune responses of the recombinant HA1 (rHA1) protein against the H1N1 influenza virus was studied. The HA1 protein and the effector domain of BAFF-R were expressed in the pET-28a (+) vector. Eight-week-old BALB/c mice were equally grouped into five groups (n=20). The 15 and 25 μg/μL of rHA1 were mixed with 2 μg/μL of rBAFF-R and injected three times for vaccinated groups. Three control groups were received normal saline and two concentrations of rHA1. The ability of rBAFF-R in eliciting HA-specific antibody response and stimulating T lymphocyte proliferation to induce the cell-mediated immunity was assayed. Induction of protection was evaluated following the challenge with PR8 strain. Analysis of immune responses showed that the co-administration of rBAFF-R with rHA1 boosted HI responses to the antigen in mice, whilst it was not able to promote the T cell proliferation responses against influenza. Compared to rHA1alone, the rBAFF-R/rHA1 generated efficient protection for the animals. There were no significant differences in eliciting the immune responses in mice immunized with the lower dose of rHA1 than that with the higher dose. The data indicate the rBAFF-R can enhance the primary and memory immune responses to protect against influenza infection.

Aberrant B Cell Receptor Signaling in Naïve B Cells from Patients with Idiopathic Pulmonary Fibrosis.

Idiopathic pulmonary fibrosis (IPF) is a chronic and ultimately fatal disease in which an impaired healing response to recurrent micro-injuries is thought to lead to fibrosis. Recent findings hint at a role for B cells and autoimmunity in IPF pathogenesis. We previously reported that circulating B cells from a fraction of patients, compared with healthy controls, express increased levels of the signaling molecule Bruton’s tyrosine kinase (BTK). However, it remains unclear whether B cell receptor (BCR) signaling is altered in IPF. Here, we show that the response to BCR stimulation is enhanced in peripheral blood B cells from treatment-naïve IPF patients. We observed increased anti-immunoglobulin-induced phosphorylation of BTK and its substrate phospholipase Cγ2 (PLCγ2) in naïve but not in memory B cells of patients with IPF. In naïve B cells of IPF patients enhanced BCR signaling correlated with surface expression of transmembrane activator and calcium-modulator and cyclophilin ligand interactor (TACI) but not B cell activating factor receptor (BAFFR), both of which provide pro-survival signals. Interestingly, treatment of IPF patients with nintedanib, a tyrosine kinase inhibitor with anti-fibrotic and anti-inflammatory activity, induced substantial changes in BCR signaling. These findings support the involvement of B cells in IPF pathogenesis and suggest that targeting BCR signaling has potential value as a treatment option.

Increased let-7b-5p is associated with enhanced BAFF-R expression and B cell survival in immune thrombocytopenia

BackgroundB cells play a key role in the pathogenesis of immune thrombocytopenia (ITP) by producing platelet autoantibodies. Accumulating evidence suggest that microRNA (miRNA) is a critical regulator in B cells. The contribution of miRNA to B cell dysfunction in ITP has not been described. The aim of this study was to examine the expression of miRNA let-7b-5p in B cells of ITP patients and investigate its possible association with B cell function in ITP. MethodsThe CD19+ cells were isolated from peripheral mononuclear cells of ITP patients and healthy controls using immunomagnetic microbeads. B cell survival in vitro was evaluated by cell counting. The level of let-7b-5p was quantified by quantitative PCR. The surface expression of B cell activating factor receptor (BAFF-R) was detected by flow cytometry. The role of let-7b-5p was examined in isolated B cells by transfecting miRNA mimics or inhibitors. ResultsThe results showed that let-7b-5p in B cells was elevated, and B cell survival was enhanced in ITP patients compared with healthy controls. BAFF and B cell receptor stimulation can induce the expression of let-7b-5p in vitro. Overexpression of let-7b-5p in B cells enhanced the expression of surface BAFF-R and promoted B cell survival. Moreover, let-7b-5p enhanced the phosphorylation of NF-κB2 p100 and upregulated the expression of survival factor Bcl-xL after BAFF induction. ConclusionLet-7b-5p is a pro-survival miRNA in B cells and increased let-7b-5p is associated with enhanced surface BAFF-R in ITP.

BAFF receptor antibody for mantle cell lymphoma therapy

ABSTRACT Mantle cell lymphoma (MCL) is an aggressive form of B cell non-Hodgkin’s lymphoma and remains incurable under current treatment modalities. One of the main reasons for treatment failure is the development of drug resistance. Accumulating evidence suggests that B cell activating factor (BAFF) and BAFF receptor (BAFF-R) play an important role in the proliferation and survival of malignant B cells. High serum BAFF levels are often correlated with poor drug response and relapse in MCL patients. Our study shows that BAFF-R is expressed on both MCL patient cells and cell lines. BAFF-R knockdown leads to MCL cell death showing the importance of BAFF-R signaling in MCL survival. Moderate knockdown of BAFF-R in MCL cells did not affect its viability, but sensitized them to cytarabine treatment in vitro and in vivo, with prolonged mice survival. Anti-BAFF-R antibody treatment promoted drug-induced MCL cell death. Conversely, the addition of recombinant BAFF (rhBAFF) to MCL cells protected them from cytarabine-induced apoptosis. We tested the efficacy of a humanized defucosylated ADCC optimized anti-BAFF-R antibody in killing MCL. Our data show both in vitro and in vivo efficacy of this antibody for MCL therapy. To conclude, our data indicate that BAFF/BAFF-R signaling is crucial for survival and involved in drug resistance of MCL. Targeting BAFF-R using BAFF-R antibody might be a promising therapeutical strategy to treat MCL patients resistant to chemotherapy.

Phase Ib Study of Ianalumab (VAY736) and Ibrutinib in Patients with Chronic Lymphocytic Leukemia (CLL) on Ibrutinib Therapy

Phase Ib Study of Ianalumab (VAY736) and Ibrutinib in Patients with Chronic Lymphocytic Leukemia (CLL) on Ibrutinib Therapy

LT-K63 Enhances B Cell Activation and Survival Factors in Neonatal Mice That Translates Into Long-Lived Humoral Immunity.

Adjuvants enhance magnitude and duration of immune responses induced by vaccines. In this study we assessed in neonatal mice if and how the adjuvant LT-K63 given with a pneumococcal conjugate vaccine, Pnc1-TT, could affect the expression of tumor necrosis factor receptor (TNF-R) superfamily members, known to be involved in the initiation and maintenance of antibody responses; B cell activating factor receptor (BAFF-R) and B cell maturation antigen (BCMA) and their ligands, BAFF, and a proliferation inducing ligand (APRIL). Initially we assessed the maturation status of different B cell populations and their expression of BAFF-R and BCMA. Neonatal mice had dramatically fewer B cells than adult mice and the composition of different subsets within the B cell pool differed greatly. Proportionally newly formed B cells were most abundant, but they had diminished BAFF-R expression which could explain low proportions of marginal zone and follicular B cells observed. Limited BCMA expression was also detected in neonatal pre-plasmablasts/plasmablasts. LT-K63 enhanced vaccine-induced BAFF-R expression in splenic marginal zone, follicular and newly formed B cells, leading to increased plasmablast/plasma cells, and their enhanced expression of BCMA in spleen and bone marrow. Additionally, the induction of BAFF and APRIL expression occurred early in neonatal mice immunized with Pnc1-TT either with or without LT-K63. However, BAFF+ and APRIL+ cells in spleens were maintained at a higher level in mice that received the adjuvant. Furthermore, the early increase of APRIL+ cells in bone marrow was more profound in mice immunized with vaccine and adjuvant. Finally, we assessed, for the first time in neonatal mice, accessory cells of the plasma cell niche in bone marrow and their secretion of APRIL. We found that LT-K63 enhanced the frequency and APRIL expression of eosinophils, macrophages, and megakaryocytes, which likely contributed to plasma cell survival, even though APRIL+ cells showed a fast decline. All this was associated with enhanced, sustained vaccine-specific antibody-secreting cells in bone marrow and persisting vaccine-specific serum antibodies. Our study sheds light on the mechanisms behind the adjuvanticity of LT-K63 and identifies molecular pathways that should be triggered by vaccine adjuvants to induce sustained humoral immunity in early life.

Dynamic regulation of hypoxia-inducible factor-1α activity is essential for normal B cell development.

B lymphocyte development and selection are central to adaptive immunity and self-tolerance. These processes require B cell receptor (BCR) signaling and occur in bone marrow, an environment with variable hypoxia, but whether hypoxia-inducible factor (HIF) is involved is unknown. We show that HIF activity is high in human and murine bone marrow pro-B and pre-B cells and decreases at the immature B cell stage. This stage-specific HIF suppression is required for normal B cell development because genetic activation of HIF-1α in murine B cells led to reduced repertoire diversity, decreased BCR editing and developmental arrest of immature B cells, resulting in reduced peripheral B cell numbers. HIF-1α activation lowered surface BCR, CD19 and B cell-activating factor receptor and increased expression of proapoptotic BIM. BIM deletion rescued the developmental block. Administration of a HIF activator in clinical use markedly reduced bone marrow and transitional B cells, which has therapeutic implications. Together, our work demonstrates that dynamic regulation of HIF-1α is essential for normal B cell development.

Immunohistochemical identification of T and B lymphocytes in formalin-fixed paraffin-embedded chicken tissues using commercial antibodies

Immunohistochemical method to detect avian lymphocytes is an efficient and reliable tool for accurate diagnosis, and immunological analysis of avian diseases. However, there are scarce studies reporting immunohistochemistry (IHC) using commercially available antibodies against lymphocytes in formalin-fixed paraffin-embedded (FFPE) chicken tissues. In the present study, we established an immunohistochemical method to identify chicken T and B lymphocytes in FFPE chicken tissues using commercial antibodies against chicken or human antigens. For this IHC method, the five tested anti-T lymphocyte antibodies reacted with chicken T lymphocytes on the FFPE sections. Further, 10 commercial anti-B lymphocyte antibodies were tested; of these, three successfully detected chicken B lymphocytes for IHC. In particular, anti-human CD3 (clone F7.2.38) antibody was most suitable for the detection of chicken T lymphocytes, whereas anti-chicken B cell activating factor receptor (BAFF-R) antibody (clone 2C4) was most suitable for the detection of chicken B lymphocytes under our IHC staining conditions. These two antibodies reacted with numerous lymphocytes of all representative lymphoid tissues without problematic background staining and nonspecific reactions. Our results indicate that T and B lymphocytes in FFPE chicken tissues can be immunohistochemically detected using commercial antibodies.

Factors associated with refractory autoimmune necrotizing myopathy with anti-signal recognition particle antibodies

BackgroundAutoimmune necrotizing myopathy with anti-signal recognition particle antibodies (ANM-SRP) is regarded as refractory myositis, whereby some patients respond poorly to conventional immunosuppression and require B cell depletion treatment. This study aimed to evaluate factors associated with refractory ANM-SRP.ResultsClinical and pathological data from 48 patients with ANM-SRP were collected. We followed up clinical symptoms and image changes over 12 months. Univariate and multivariate analyses were undertaken to determine the associations between variables of interest and poor response to therapy. Refractory ANM-SRP appeared in 32.5% of patients who showed no or minimal improvement after 12 months of steroid therapy. The clinical risk factors for refractory patients were being male (OR, 19.57; P < 0.001), severe muscle weakness (OR, 7.51; P < 0.001) and concurrent interstitial lung disease (OR, 39.70; P < 0.001). The imaging refractory-related factor was the fatty infiltration rate of thigh muscles over 3 months (P = 0.022) and the pathological factor associated with refractory ANM-SRP was the high expression of B cell activating factor receptor (BAFF-R) in muscle (P = 0.036).ConclusionBeing male, severe muscle weakness, concurrent interstitial lung disease, quick development of muscle fatty infiltration and more BAFF-R and B lymphocyte infiltration in muscle indicate a poor response to immunosuppressive therapy in patients with ANM-SRP.

The efficacy and mechanism for action of iguratimod in primary Sjögren's syndrome patients.

Primary Sjögren's syndrome (pSS) has been proven as a systemic autoimmune disorder (such as Sjogren's syndrome dry eye). This research aimed to evaluate potential treating effects of Iguratimod on pSS. Fifty pSS patients were enrolled and randomly divided into Conventional group and Iguratimod group. Improvement in pSS was evaluated every 4weeks. pSS disease activity was evaluated with European League Against Rheumatism (EULAR) Sjögren's syndrome disease activity index (ESSDAI). Symptoms were evaluated by determining EULAR Sjögren's syndrome patient-reported index (ESSPRI), platelet (PLT), IgG and Schirmer I test. Peripheral blood B cell molecules (CD135, IgD, CD38, CD20) and human B cell-activating factor-receptor (BAFF-R) were analyzed with flow cytometry. After treating for 12-weeks, pSS patients in Iguratimod and Conventional group showed a significant decrease in disease activity (ESSPRI, ESSDAI, PLT, IgG and Schirmer I test) comparing with baselines. Patients' ESSPRI (2.92 ± 0.19) and disease activity of ESSDAI (4.32 ± 0.29), PLT (95.64 ± 1.86), IgG (13.0 ± 0.45) and Schirmer I test (4.67 ± 0.31) in Iguratimod group were significantly lower compared to Conventional group (4.64 ± 0.15, 5.8 ± 2.08, 77.44 ± 1.41, 16.5 ± 0.44 and 2.25 ± 0.11) (p < 0.0001). Changes of ESSPRI, ESSDAI, PLT, IgG and Schirmer I test were remarkable observed between two groups (p < 0.001). Iguratimod and Conventional treatment demonstrated a significant reduction in total B cells in pSS patients compared with pre-treatment. The pSS patients from Iguratimod and Conventional group showed a significant decreased BAFF-R (61.82 ± 1.52, 74.07 ± 1.11) and CD38+IgD+ (48.08 ± 0.92, 62.66 ± 1.12) on B cells after treatment compared with baseline (92.26 ± 0.32, 91.53 ± 0.45, 84.39 ± 0.59, 85.04 ± 0.46) (p < 0.001). After treating 12 weeks, BAFF-R, CD38+IgD+ expression in Iguratimod group decreased significantly compared to Conventional group (p < 0.001). Iguratimod alleviated symptoms and mediated adaptive-immunity balance by suppressing BAFF-R positive B cell in pSS patients.

Differences of B cells, plasma cells, and related cytokines expression in gingival tissues between periodontitis and periodontal healthy subjects

This study aimed to compare the differences of B cells, plasma cells, and related cytokines expression in gingival tissues between periodontitis and periodontal healthy subjects. Gingival tissues were collected from periodontal healthy subjects (periodontal healthy group, n=12) and periodontitis patients (periodontitis group, n=15). Hematoxylin-eosin (HE) staining was used for histopathological examination. Immunohistochemical staining (CD19, CD38, and CD138) was applied to detect the expression of B cells and plasma cells. B cell-activating factor (BAFF) and soluble receptor activator of nuclear factor-κB ligand (sRANKL) were detected by enzyme-linked immunosorbent assay. Extensive inflam-matory cell infiltration was found in the gingival tissues of the periodontitis group. The number of CD19(+), CD38(+), and CD138(+) cells of the periodontitis group was significantly higher than that of the periodontal healthy group (P<0.000 1). BAFF and sRANKL levels of the periodontitis group were higher than those of the periodontal healthy group (P<0.01, P< 0.001, respectively). The expression of B cells, plasma cells, and their related BAFF and sRANKL cytokines were significantly higher in periodon-titis patients than those in the periodontal healthy subjects, sug-gesting that B cells and plasma cells may be involved in the development of periodontitis.

Decreased of BAFF-R expression and B cells maturation in patients with hepatitis B virus-related hepatocellular carcinoma.

BACKGROUNDRecent evidence has indicated the role of B cells and B cell-activating factor (BAFF) in the development of hepatocellular carcinoma (HCC).AIMTo characterize circulating BAFF receptor expression and B cell subpopulations in patients with hepatitis B virus (HBV)-related HCC.METHODSPeripheral blood samples collected from 41 patients with chronic HBV infection (25 patients without HCC and 16 patients with HCC) and 9 healthy controls were assessed for BAFF receptors [BAFF-R(B cell-activating factor receptor), transmembrane activator and cyclophilin ligand interactor, B-cell maturation antigen] and B cell subpopulations by multicolor flow cytometry.RESULTSThe frequency of BAFF-R expressing B cells to total B cells was significantly lower in patients with HCC (3.39% ± 2.12%) compared with the non-HCC group (5.37% ± 1.90%) and healthy controls (6.23% ± 2.32%), whereas there was no difference in transmembrane activator and cyclophilin ligand interactor and B-cell maturation antigen. The frequencies of CD27+IgD+ memory B cells, CD27+IgD- class-switched memory B cells and plasmablasts were significantly lower in the patients with HCC compared to patients without HCC (1.23 ± 1.17 vs 3.09 ± 1.55, P = 0.001, 0.60 ± 0.44 vs 1.69 ± 0.86, P < 0.0001 and 0.16 ± 0.12 vs 0.37 ± 0.30, P = 0.014, respectively). However, the ratio of naïve and transitional B cell did not differ significantly between the three groups. In addition, decreased BAFF-R expression on B cells was significantly correlated with large tumor size and advanced tumor stage.CONCLUSIONOur data demonstrated BAFF-R expression was reduced in B cells that involved with the frequencies of B cells maturation in patients with HCC. The depletion of BAFF-R might play an important role in the development of HCC in patients with chronic HBV infection.

Increased Ileal Immunoglobulin A Production and Immunoglobulin A-Coated Bacteria in Diarrhea-Predominant Irritable Bowel Syndrome.

OBJECTIVES:Immune activation and intestinal microbial dysbiosis could induce diarrhea-predominant irritable bowel syndrome (IBS-D). We examined the roles of ileal immunoglobulin A (IgA) and IgA-coated bacteria in IBS-D pathogenesis.METHODS:Peripheral blood, fecal samples, and ileal and cecal biopsies were collected from 32 healthy volunteers and 44 patients with IBS-D. Quantitative reverse transcriptase polymerase chain reaction was used to assess differential gene expression. IgA levels in the blood and fecal samples were quantified by an enzyme-linked immunosorbent assay. IgA+ cells were assessed by immunofluorescence imaging. Flow-cytometry-based IgA+ bacterial cell sorting and 16S rRNA gene sequencing (IgA-SEQ) was used to isolate and identify fecal IgA+ bacteria.RESULTS:Fecal IgA, particularly IgA1, was upregulated in patients with IBS-D. IgA class switch and B cell–activating factor-receptor were increased in the terminal ileum of patients. The intestinal microbiota composition was altered in patients compared with that in controls. IgA-SEQ showed that the proportion of fecal IgA-coated bacteria was increased significantly in patients with IBS-D. IgA+ bacteria in patients with IBS-D showed higher abundances of Escherichia–Shigella, Granulicatella, and Haemophilus compared with healthy controls and IgA− bacteria in patients with IBS-D. The Escherichia–Shigella IgA coating index was positively correlated with anxiety and depression. The Escherichia–Shigella relative abundance, luminal IgA activity, and some altered IgA-coated bacteria were positively associated with the clinical manifestations of IBS-D.DISCUSSION:Microbial dysbiosis may promote the terminal ileal mucosa to produce higher levels of IgA, increasing the proportion of IgA-coated bacteria by activating IgA class switching, which might regulate local inflammation and clinical manifestations in IBS-D. IgA may mediate the effects of microbial dysbiosis on the pathogenesis of IBS-D.

Abstract P1108: Serum Immunoglobulins are Not Necessary for Experimental Hypertension

The role of B cells and serum immunoglobulins (Ig) in hypertension is unclear. Elevated serum IgG has been observed in some hypertensive patients and in animal models of hypertension. Mice with a severe B cell deficiency due to lack of B cell activating factor receptor (BAFF-R) and mice lacking the cytokine interleukin 21 (IL21) - which is a potent inducer of B cell Ig class switching and high affinity Ig production - are protected from Ang II-induced hypertension and exhibit reduced serum IgG levels. Thus, we sought to determine whether hypertensive IgGs play a causative role in hypertension and end-organ dysfunction. First, we purified serum IgG from normotensive and hypertensive wild type (WT) mice and adoptively transferred equal amounts of IgG by intraperitoneal injection into normotensive recipient mice followed by infusion of a sub-pressor dose of angiotensin II (Ang II) for 2 weeks. Flow cytometric analysis of aortas revealed a modest increase in F4/80+ macrophages (p=0.04) and CD8+ T cells (p=0.02) in mice that received hypertensive IgG. However, there was no difference in endothelial dependent relaxation, renal inflammation, albuminuria, and blood pressure (BP) between the two groups. As an alternative method to investigate the role of class-switched high affinity Igs, we studied mice with genetic deletion of activation-induced deaminase (AID) which is an enzyme necessary for somatic hypermutation (to generate high affinity Igs) and isotype class switching. AID -/- mice exhibit undetectable IgG and higher levels of IgM in serum compared to WT mice. Ang II infusion for 4 weeks revealed no difference in BP, aortic inflammation, renal inflammation, and albuminuria. We also studied mice with deficiency of the IgM heavy chain (uMT). These mice exhibit undetectable levels of all immunoglobulins and are severely deficient in mature B cells. uMT-/- mice are also not protected from hypertension induced by Ang II infusion or deoxycorticosterone acetate (DOCA-salt) treatment. Together, these results demonstrate that while elevated serum immunoglobulins seen in hypertensive animals and humans may represent a biomarker of aberrant immune activation, they are likely not playing a causal role in hypertension pathophysiology.

Tumor necrosis factor receptor associated factor 3 (TRAF3) regulates B cell antigen receptor (BCR) signaling

Abstract Tumor necrosis factor receptor associated factor 3 (TRAF3) is an intracellular adaptor protein that plays multiple cell type- and context-dependent roles. In B cells, TRAF3 negatively regulates signaling by several receptors important to survival and activation, including the B cell activating factor receptor (BAFF-R), CD40, toll-like receptors and the IL-6 receptor. Deletions or inactivating mutations of TRAF3 are common in the human B cell malignancies multiple myeloma and B cell lymphoma (BCL). Enhanced homeostatic B cell survival in B cell-specific TRAF3-deficient mice (B-Traf3−/−) causes enlarged secondary lymphoid organs, spontaneous germinal centers, and autoantibody production. Highlighting the role TRAF3 plays as a B cell tumor suppressor, B-Traf3−/− mice develop BCL at an increased rate as they age. The B cell antigen receptor (BCR) is crucial for B cell antigen recognition, survival, proliferation, and antibody production. Defects in BCR signaling can lead to malignancy, and many BCLs are dependent upon BCR signaling for survival. TRAF3 regulates the signaling of several pathways that interact with the BCR pathway, but the role of TRAF3 in regulating BCR signals is unknown, a knowledge gap we are currently addressing. Our initial studies show that TRAF3 associates with CD79b of the BCR complex. Phosphorylation of Syk, a proximal BCR signaling target, is increased after BCR stimulation in the absence of TRAF3, with increased downstream phosphorylation and activation of Btk, PLCγ2, Akt, Erk, and canonical NF-κB. This suggests that TRAF3 plays an important role as a regulator of BCR signaling. Ongoing studies are examining the mechanisms of this regulation.

Cirrhosis and Autoimmune Liver Disease

Autoimmune hepatitis (AIH), primary biliary cholangitis (PBC), and primary sclerosing cholangitis (PSC) constitute the most frequently observed forms of autoimmune liver diseases. Each of these autoimmune liver diseases might present with cirrhosis at diagnosis, and a significant proportion of patients develop cirrhosis during follow-up. This manuscript provides a review that addresses how to monitor and manage patients with cirrhosis secondary to autoimmune liver diseases. For patients with PBC, the farnesoid X receptor (FXR) agonist, obeticholic acid (OCA), is the first approved drug since ursodeoxycholic acid (UDCA) and is licensed for non-responders to or those intolerant of UDCA. Bezafibrate has been shown to be effective in non-responders to UDCA, but is not licensed for this clinical indication at present. For patients with AIH, rituximab, a monoclonal antibody against the protein CD20 is a potential option for patients with suboptimal response to corticosteroids. New treatment options are currently being investigated for AIH and PSC that include the anti-B cell-activating factor receptor monoclonal antibodies, nor-UDCA, amongst others, but the efficacy in patients with cirrhosis has not been fully established. Cirrhosis is present in 30–50% of patients at the time of diagnosis of AIH, in 20% of patients with PBC, and in 35% of patients with PSC. Therefore, cirrhosis constitutes one of the main complications of autoimmune liver disease and continues to develop at a frequency of approximately 3–6% per year. Patients with compensated AIH cirrhosis and histological inflammatory activity benefit of corticosteroid treatment. Patients with PBC and compensated cirrhosis should receive UDCA. OCA should be considered in non-responders but used with very close monitoring in patients with decompensated cirrhosis. There is no effective treatment option available for patients suffering from PSC. Liver transplantation is indicated for patients who have progressed to decompensated cirrhosis, and those with intractable symptoms or hepatocellular carcinoma within transplant criteria. Standard monitoring for cirrhosis-related complications is highly recommended.