The TLR7 agonist imiquimod selectively inhibits IL-4-induced IgE production by suppressing IgG1/IgE class switching and germline ε transcription through the induction of BCL6 expression in B cells.

Imiquimod (IMQ) is a selective toll-like receptor 7 (TLR7) agonist. TLR7 activation leads to the production of IFN-γ and pro-inflammatory cytokines by innate immune cells. However, the role of TLR7 in B cells is not fully understood. In this study, we investigated the direct effect of in vitro stimulation with IMQ on Ab production and isotype switching in B cells. IMQ selectively diminished IL-4-induced IgE and IgG1 production in anti-CD40-activated mouse B cells. IMQ also inhibited germline ε transcripts (GLTε)/GLTγ1 and post-switch ε transcripts (PSTε)/PSTγ1 expression, while enhancing GLTγ2c and PSTγ2c expression in anti-CD40/IL-4-stimulated B cells. Interestingly, IMQ abrogated IL-4-induced circle transcripts ε-γ1 (CTε-γ1) expression, indicative of sequential switching from IgG1 to IgE. Furthermore, IMQ repressed IL-4-induced surface IgE/IgG1 expression while increasing surface IgG2c expression. The selective inhibition of IgE synthesis was not due to IMQ-induced production of IFN-γ or IL-12 in the same culture. IMQ also enhanced BCL6 expression, a transcriptional repressor for the GLTε promoter, in anti-CD40/IL-4-stimulated B cells. In addition, BCL6 siRNA restored IMQ-mediated suppression of GLTε transcription. Therefore, these results indicate that TLR7 engagement by IMQ inhibits IL-4-induced GLTε transcription by enhancing BCL6 expression and inhibits IL-4-induced sequential switching from IgM to IgE via IgG1, thus resulting in the downregulation of IgE production by B cells.

Heat-Killed Saccharomyces cerevisiae, A Dectin-1 Agonist, Selectively Induces IgG4 Production by Human B Cells.

Dectin-1 is a major receptor that recognizes fungal cell wall ß-glucan. We previously reported that heat-killed Saccharomyces cerevisiae (HKSC), a Dectin-1 agonist, selectively induces IgG1 class switching in mouse B cells. Dectin-1 is also expressed on human B cells; however, Dectin-1 function in human B cells remains unknown. This study aimed to investigate the direct effect of in vitro stimulation using HKSC on Ig class switching in human B cells. HKSC selectively induced the expression of germline γ4 transcripts (GLTγ4) by human B cell line 2E2, and HKSC significantly augmented GLTγ4 promoter activity. Moreover, HKSC selectively enhanced GLTγ4 expression and IgG4 production by anti-CD40-activated human tonsillar resting B cells. Thus, these results suggest that Dectin-1 maybe involved in selective IgG4 class switching by human B cells.

Aberrant expression of interleukin-10 and activation-induced cytidine deaminase in B cells from patients with Behçet's disease.

Despite extensive studies, the pathogenesis of Behçet's disease (BD) remains unclear. In particular, the roles of B cells in patients with BD have not been elucidated. Activation-induced cytidine deaminase (AID) is a critical enzyme for immunoglobulin (Ig) heavy chain class switching and somatic hypermutation in B cells and the abnormal expression of AID in various immune conditions has previously been studied. B10 cells, an interleukin (IL)-10-secreting subset of regulatory B cells, function to downregulate inflammation and autoimmunity. Thus, in the present study, the relevance of B cells in patients with BD was investigated. The plasma levels of IL-10 and IgA and the proportions of cluster of differentiation (CD)43+ B cells, excluding naïve B cells, were measured in 16 patients with BD and 16 age- and sex-matched healthy controls (HCs). Additionally, the mRNA levels of IL-10 and AID were assessed in B cells from fresh peripheral blood samples of the BD patients and HCs. The plasma level of IL-10 in patients with BD did not differ significantly from that in HCs. Similarly, there was no significant difference in the plasma level of IgA, although a slight increase was observed in patients with BD compared with that in HCs. There were no differences in CD43+CD19+ B cell numbers between patients with BD and HCs. However, IL-10 mRNA levels were significantly reduced (P<0.05), while AID mRNA levels were significantly increased (P<0.01) in the B cells of patients with BD compared with those in HCs. These results provide insight into the role of B cells in patients with BD.

Expression of activation-induced cytidine deaminase splicing variants in patients with ankylosing spondylitis.

To investigate the expression patterns of activation-induced cytidine deaminase (AID) variants in peripheral blood mononuclear cells (PBMCs) of patients with ankylosing spondylitis (AS) and examine their clinical implications, we isolated PBMCs from healthy controls (HC, n = 33) and patients with AS (n = 62), and measured mRNA expression of AID variants and translesion synthesis (TLS) polymerases using quantitative real-time polymerase chain reaction. The proportion of patients with AS in whom AID splicing variant (sv) 2 was expressed was significantly higher than that of HC (p = .031). 80.7% of AS patients were treated with tumour necrosis factor inhibitor (TNFi). Significantly higher proportion of the TNFi-treated group expressed sv2 compared to the TNF-naïve group (p = .037). And we compared the level of AID variants expression between the TNFi-treated group and the TNF-naïve group. The expression levels of AID full-length (FL) and sv1 were significantly lower in the TNFi-treated group than the TNF-naïve group (FL: p = .002, sv1: p = .045). In addition, we investigated mRNA expression levels of translesion synthesis (TLS) polymerases in PBMCs from patients with AS and HC. The expression level of TLS pol ι was significantly lower in patients with AS than in HC (p = .007). In conclusion, AS patients expressed significantly higher levels of sv2 than HC. TNFi treatment restored the gene expression of the AID variants (FL, sv1, and sv2) in patients with AS. A clear understanding of the underlying cellular and molecular mechanisms will help to identify the pathogenesis of AS better and to develop novel therapeutic targets.

Dectin-1 agonist selectively induces IgG1 class switching by LPS-activated mouse B cells.

Heat-killed Saccharomyces cerevisiae (HKSC) is an agonist for Dectin-1, a major fungal cell wall ß-glucan receptor. We previously reported that HKSC selectively enhances IgG1 production by LPS-activated mouse B cells. To determine if this IgG1 selectivity is caused by selective IgG1 class switching, we performed RT-PCRs for measuring germline transcripts (GLTs), flow cytometric analyses for detecting Ig-expressing cells, and ELISPOT assays for measuring the number of Ig-secreting cells in HKSC/LPS-stimulated mouse B cell cultures. HKSC selectively enhanced expression of GLTγ1, the number of IgG1-expressing cells, and the number of IgG1-secreting B cells in the presence of LPS stimulation. In addition, HKSC induced the expression of CD69, an activation marker for B lymphocytes, and the expression of surface Dectin-1. Two Dectin-1 antagonists, laminarin and a neutralizing Dectin-1 antibody, selectively diminished HKSC-reinforced IgG1 production by LPS-stimulated B cells. Furthermore, depleted zymosan (dzn), a Dectin-1 agonist with increased selectivity, also selectively enhanced GLTγ1 transcription. The Dectin-1 antagonists blocked dzn-induced IgG1 production by LPS-activated B cells. Collectively, these results suggest that Dectin-1 agonists selectively induce IgG1 class switching by direct stimulation of Dectin-1 on LPS-activated B cells resulting in selective production of IgG1.

Ginsenoside Rg1 and 20(S)-Rg3 Induce IgA Production by Mouse B Cells.

Ginsenosides are the major components of ginseng, which is known to modulate blood pressure, metabolism, and immune function, and has been used to treat various diseases. It has been reported that ginseng and several ginsenosides have immunoregulatory effects on the innate and T cell-mediated immune response. However, their effects on the humoral immune response have not been fully explored. The present study examined the direct effects of red ginseng extract (RGE) and ginsenosides on mouse B cell proliferation and on antibody production and the expression of germline transcripts (GLT) by mouse B cells in vitro. RGE slightly reduced B cell proliferation, but increased IgA production by LPS-stimulated B cells. Furthermore, ginsenoside Rg1 and 20(S)-Rg3 selectively induced IgA production and expression of GLTα transcripts by LPS-stimulated B cells. Collectively, these results suggest that ginsenoside Rg1 and 20(S)-Rg3 can drive the differentiation of B cells into IgA-producing cells through the selective induction of GLTα expression.

Population pharmacokinetics of multiple alcohol intake in humans.

The objective of this study was to determine population-based pharmacokinetics parameters for ethanol following multiple intake and to identify the factors influencing the pharmacokinetics. Three different solutions of alcoholic liquor (ethanol 55.39 ± 0.45 g) with different dissolved oxygen concentrations were administered, and blood alcohol concentration was determined in 59 healthy subjects using a breath analyzer. Samples (n = 2955) were collected at various time points. Population pharmacokinetic modeling was performed to describe the pharmacokinetics of ethanol. The influence of individuals' demography and dissolved oxygen concentration was investigated, and Visual Predictive Check and bootstrapping were conducted for internal evaluation. The developed model was used to perform simulations to visualize the effects of covariates on individuals. A one-compartment model with Michaelis-Menten elimination kinetics described the multiple ethanol intake data. Population pharmacokinetic estimates of V(max) and K(m) were 3.256 mmol min(-1) and 0.8183 mmol L(-1), respectively. V(d)/F was estimated to be 77.0 L, and K(a) was 0.0767 min(-1). Body weight, age, and the dissolved oxygen concentration were confirmed to be significant covariates. The mean estimates from the developed population pharmacokinetic model were very similar to those from 500 bootstrap samples, and Visual Predictive Check showed that approximately 94% of the observed data fit well within the 5th-95th percentile. A one-compartment model with nonlinear elimination kinetics for multiple ethanol intake was developed and the significant covariates were determined. The robustness of the developed model was evaluated by bootstrap and Visual Predictive Check. The final model and implanted covariates explained well the variability and underlying mechanism of ethanol PK.